Landscape Review and Evidence Map of Gene Therapy, Part I: Adenovirus, Adeno-associated Virus, and Clustered Regularly Interspaced Short Palindromic Repeats

Andrea Richardson, Eric Apaydin, Sangita Baxi, Jerry Vockley, Olamigoke Akinniranye, Jody Larkin, Aneesa Motala, and Susanne Hempel

RAND Corporation

March 2019

Revision [February 2020] Since the completion of this report a gene therapy herein described as AVXS-101 has been approved by the FDA as Zolgensma therefore the report was revised in November 2019 to reflect that change.

All statements, findings, and conclusions in this publication are solely those of the authors and do not necessarily represent the views of the Patient-Centered Outcomes Research Institute (PCORI) or its Board of Governors. This publication was developed through a contract to support PCORI's work. Questions or comments may be sent to PCORI at [email protected] or by mail to Suite 900, 1828 L Street, NW, Washington, DC 20036.

©2019 Patient-Centered Outcomes Research Institute. For more information see www.pcori.org

EMERGING TECHNOLOGIES AND THERAPEUTICS REPORT 2 Table of Contents Executive Summary ...... 5 Introduction of Report Series ...... 7 History of Gene Therapies ...... 7 Aim of the Report ...... 7 Methodology Overview ...... 7 Data Sources ...... 8 Figure 1. Content and Data Sources ...... 9 Organization of the Report Series and Organization of This Report ...... 9 Description of Interventions/ Technologies ...... 11 Table 1. Intervention Descriptions ...... 11 Table 2. FDA-approved Therapies ...... 12 Table 3. Potential Pipeline Gene Therapies ...... 13 Adenoviral, AAV, and CRISPR Gene Therapy ...... 16 Adenoviral Therapy Indications ...... 16 AAV Therapy Indications ...... 17 CRISPR Therapy Indications ...... 18 Key Question 1a. What different types or modalities of the intervention have been used in clinical practices and clinical research studies? ...... 18 Key Question 1b. What are the potential/presumed advantages and disadvantages in contrast to current practices? ...... 19 Key Question 1c. What are the safety issues or expected adverse events? ...... 21 Context in Which Gene Therapy Is Used ...... 23 Key Question 2a. What is the approval process and current approval/certification status for gene therapies? ...... 23 Key Question 2b. Are any additional accompanying resources or technologies required? ...... 23 Key Question 2c. What is the current state of adoption in practice and settings? ...... 24 Key Question 2d. What are the operator factors, such as training and staffing? ...... 24 Ongoing Premarket and Postmarket Gene Therapy Studies ...... 25 Ongoing Trial Characteristics ...... 25 Table 4. Ongoing Adenovirus, AAV, and CRISPR Trials ...... 25 Key Question 3a. What are the indication/patient inclusion criteria in ongoing trials? ...... 31 Key Question 3b. What are the types of interventions in ongoing trials? ...... 31 Key Question 3c. What are the study designs/sizes of ongoing trials? ...... 31 Key Question 3d. What are the comparators in ongoing trials? ...... 31 Key Question 3e. What are the previous/concurrent treatments in ongoing trials? ...... 32 Key Question 3f. What is the length of follow-up in ongoing trials? ...... 32 Key Question 3g. What outcomes are measured in ongoing trials? ...... 32 Current Evidence Supporting Gene Therapy ...... 33 Characteristics of Published Trials ...... 33 Table 5. Summary of Published Adenovirus and AAV Studies ...... 33

EMERGING TECHNOLOGIES AND THERAPEUTICS REPORT 3 Key Question 4a. What are the indication/patient inclusion criteria in published studies? ...... 34 Key Question 4b. What are the types of interventions in published studies? ...... 34 Key Question 4c. What are the study designs/sizes of published studies? ...... 34 Key Question 4d. What are the comparators in published studies? ...... 35 Key Question 4e. What are the previous/concurrent treatments in published studies? ...... 35 Key Question 4f. What is the length of follow-up in published studies? ...... 35 Key Question 4g. What outcomes are measured in published studies? ...... 35 Key Question 4h. What are the adverse events in published studies? ...... 36 Evidence Map of Ongoing and Completed Gene Therapy Trials ...... 37 Figure 2. Gene Therapy Evidence Map: Adenovirus, AAV, and CRISPR ...... 38 Important Issues Raised by Gene Therapy ...... 39 Key Question 5a. What are the implications of the current level of adoption and future diffusion, given the current level of evidence: efficacy/safety, ethical, disparity, resource allocation, and decision making? ...... 39 Key Question 5b. What are the key issues pertaining to decisional uncertainty? ...... 40 Key Question 5c. What are potential areas of research focus for PCORI and others? ...... 40 References ...... 42 Appendix ...... 72 Appendix A: Report Methodology ...... 72 Literature Searches ...... 72 Literature Review Procedure ...... 73 Inclusion Criteria ...... 73 Data Extraction ...... 74 Evidence Map ...... 74 Evidence Tables ...... 74 Appendix B: Search Strategies ...... 75 Published Literature ...... 75 Grey Literature ...... 79 Appendix C: Literature Flow Diagram ...... 82 Appendix D: Stakeholder and Key Informant Interview Information ...... 83 Interviews ...... 83 Recruitment ...... 83 Procedure...... 83 Interview Data Collection and Analysis Strategy ...... 85 Appendix E: Evidence Maps Stratified by Intervention ...... 86 Appendix Evidence Map 1. Adenovirus Trials ...... 86 Appendix Evidence Map 2. AAV Trials ...... 87 Appendix Evidence Map 3. CRISPR ...... 88 Appendix F: Evidence Tables Published Trials ...... 89 Appendix Table A1. Published Adenovirus Trials ...... 89 Appendix Table A2. Published AAV Trials ...... 161

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Executive Summary Human gene therapy has evolved rapidly and may be poised to affect mainstream medicine. Gene therapy initially targeted incurable genetic diseases (eg, metabolic diseases) but is now most commonly used to treat certain cancers. In response to the increase in approved gene therapy approaches, the Patient-Centered Outcomes Research Institute (PCORI) commissioned a landscape review. The purpose of this report is to better understand the evidence supporting currently approved gene therapies and those that may be available in the near term in the United States. This is the first of 2 reports and it focuses on adenovirus, adeno-associated virus vector-based (AAV) gene therapy, and clustered regularly interspaced short palindromic repeats (CRISPR). In November 2018, we reviewed scientific journal and grey literature to identify evaluations of therapies that meet the US Food and Drug Administration’s (FDA) definition of gene therapy. Our overall aim was to provide a current view of the existing evidence on gene therapy and to identify gaps that need to be addressed for the field to move forward in the United States. We reviewed both completed and ongoing trials involving human participants, including both controlled and uncontrolled trials. The literature results are documented in visualizations (ie, evidence maps) and comprehensive evidence tables. We also conducted semistructured interviews with a diverse set of stakeholders to elicit feedback on challenges, hopes, and important issues related to gene therapy. We selected key informants who provided a range of perspectives, including patients and patient advocates, clinicians, payers, insurers, public policymaker representatives, and industry representatives. This report addresses the description of the interventions, the context in which gene therapy is used, ongoing premarket and postmarket gene therapy studies, and current evidence for the use of gene therapy. The FDA has approved 10 interventions that meet the definition of gene therapy; 2 are included in this report. Luxturna (Voretigene neparvovec) was the first AAV-based therapy, approved in 2017 to treat biallelic RPE65 mutation-associated retinal dystrophy. Zolgensma* (Onasemnogene abeparvovec-xioi) is an AAV-based therapy that was approved in 2019 for the treatment of spinal muscular atrophy. Our search of the grey literature identified 76 therapies that meet the FDA’s definition of gene therapy and that have not yet been approved by the FDA but may be close to approval. A wide variety of modalities are being tested to treat patients. The most common type of therapy is based on AAV. The identified therapies are being used to treat multiple disease types, including vision disorders (eg, choroidemia), muscular dystrophy (eg, Duchenne), blood disorders (eg, hemophilia), Sanfilippo syndrome, cancer, HIV, and Parkinson’s disease. In our literature search, we screened 3859 citations and assessed 1817 full-text publications for eligibility. Of these, we identified 251 completed trials that tested adenovirus or AAV interventions. We also identified 19 ongoing trials, including a trial using CRISPR, that have completed patient recruitment. The indications in the identified completed and ongoing trials were cancer (49%), ocular disorders (9%), cardiovascular disease (9%), neurodegenerative disorders (9%), respiratory conditions (4%), blood disorders (4%), immune deficiency (4%), muscular conditions (3%), inflammatory conditions (1%), and other indications that were not easily categorized (eg, venous leg ulcer disease, 9%). Pharmaceutical companies claim that certain gene therapies have the potential to offer a one-time administration that will cure disease. Results from small trials are promising; however, true durability

* This report was updated on February 3, 2020 to reflect the approval of Zolgensma (formerly AVXS-101) for spinal muscular atrophy (approved on 5/24/19).

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of treatment remains unclear on the basis of current published results. Further, most existing gene therapy trials are based on single-arm trials or trials with multiple groups who were treated with different levels of the same therapy. However, randomized controlled trials (RCTs) are increasingly being implemented in cancer, cardiovascular disease, immune deficiency, and otherwise categorized indication trials. Sample sizes remain relatively small across the studies, likely owing to the rarity of the indications addressed by gene therapy. We observed larger sample sizes in the RCTs for more common diseases, such as cardiovascular conditions, or for vaccine trials. Most studies evaluated safety and indicators of biodistribution, and relatively few studies tested clinical efficacy. The potential and presumed advantage of gene therapies is, first and foremost, cures for previously incurable and fatal diseases. However, the potential disadvantages of gene therapies relative to current treatment practices include many unknowns related to the following: (1) how clinically effective treatments are compared with the risk for adverse outcomes; (2) how gene therapies will be implemented in health care systems in which providers may be unfamiliar with requisite procedures and patients are not well informed about what gene therapies are; (3) the ethical implications of altering the genome; and (4) how patients and payers will pay for potentially expensive gene therapy. While adenoviral gene therapies have been tested extensively, AAV therapies may be more promising, given their popularity in trials of therapies in the pipeline. CRISPR’s promise remains to be thoroughly tested, but findings should soon shed light on the future of this therapy. To address the gaps in knowledge that might support the approval and use of gene therapies, several areas of research may warrant future work. Suggested future research includes examining how Centers of Excellence can ensure that high-quality manufacturing processes and health care delivery standards are followed. In addition, patient registries with long-term follow-up will reduce uncertainty about the durability of gene therapy effects. Other research could examine how training and education programs for providers and patients might improve provider familiarity with procedures and patient understanding of how the therapy works and the risks of unintended effects. Finally, microsimulation modeling of different scenarios of patient access and treatment efficacy might address the uncertainty about how gene therapy will affect disease.

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Introduction of Report Series The FDA defines gene therapy as products “that mediate their effects by transcription and/or translation of transferred genetic material and/or by integrating into the host genome and that are administered as nucleic acids, viruses, or genetically engineered microorganisms. The products may be used to modify cells in vivo or transferred to cells ex vivo prior to administration to the recipient.”1 Gene therapies operate by doing one of the following: • Replacing a disease-causing gene with a healthy copy of the gene • Inactivating a disease-causing gene that is not functioning properly • Introducing a new or modified gene into the body to help treat a disease1

History of Gene Therapies Human gene therapy evolved from bacterial and viral cell transformation studies.2 Experimentation in humans started in the 1990s when the FDA approved a gene therapy trial in 2 children with adenosine deaminase deficiency, a monogenic disease that leads to severe immunodeficiency.3 Some immune responses improved in this small study, and the results suggested that the treatment might be safe and effective. Despite some tragic setbacks, gene therapy research has continued to expand and improve. Gene therapy initially targeted relatively rare incurable genetic diseases (eg, metabolism diseases) but is now most commonly used to treat certain cancers, including some that are inherited and some that are not. Cancer comprises 65% of indications addressed by current gene therapy clinical trials, followed by monogenic diseases (11%), infectious diseases (7%), and cardiovascular diseases (7%).4 The FDA has approved 16 products that are either cellular products or gene therapies: cord blood products (50%), autologous or allogenic cellular products (25%), and disease-specific therapies (25%).5 Nine of the products meet the criteria in the FDA’s definition of a gene therapy, and they employ multiple intervention modalities to treat specific cancers and vision loss.

Aim of the Report Given the increase in approved gene therapy products and techniques, we sought to better understand the evidence base regarding ongoing and published trials, safety, and efficacy results for currently approved gene therapies and those that might be available in the near term (~5 years) in the United States and to identify gaps that need to be addressed for the field to move forward. This report addresses the existing evidence on gene therapy and outlines evidence gaps.

Methodology Overview Our methods are described in detail in Appendix A. Briefly, we established a landscape review of multiple data sources to cover critical aspects of current and future gene therapy approaches. Specifically, we describe the evidence base for FDA-approved gene therapies that are currently used to treat or cure conditions. In addition, we describe ongoing clinical trials that are testing applications that might be poised to gain FDA approval, as well as the conditions that are relevant for future applications. The search strategy is documented in Appendix B. We applied explicit inclusion and exclusion criteria, and the literature flow is shown in Appendix C. The FDA has approved gene therapies on the basis of the results of single-arm trials when a life-threatening condition has no alternative treatment and its severity6 justifies not performing controlled trials. Thus, we included both controlled and uncontrolled trials in our literature review. The literature results are documented in a central figure and comprehensive evidence tables.

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We also conducted interviews with stakeholders who provided a variety of perspectives on gene therapies. We elicited feedback about challenges, hopes, and important issues related to gene therapy. We included patients and patient advocates, clinicians, payers, insurers, public policymaker representatives, and industry representatives and analysts. In semistructured interviews, these key informants contributed to the development of our search strategy and the identification of interventions that could be approved within the next 5 years. We also elicited their perspectives on aspects of beneficial and harmful outcomes, important issues to patients, gene therapy–related challenges and gaps, hopes for future progress, and their thoughts on how to move the field forward. More detail is available in Appendix D. We interviewed the following key informants: Patient advocates • Sharon Terry, president and chief executive officer, Genetic Alliance • Ben Wakana, executive director, Patients for Affordable Drugs Clinicians • Flora Lum, MD, vice president of quality and data science, American Academy of Ophthalmology • Cary Harding, MD, professor of molecular and medical genetics, Oregon Health & Science University Insurers • Geoff Crawford, MD, medical director, Anthem • John Yao, MD, staff vice president, Anthem • Naomi Aronson, MD, executive director, Technology Evaluation Center, Blue Shield of California Public policymakers • Katherine B. Szarama, presidential management fellow, Centers for Medicare and Medicaid Services Industry representatives and analysts • Michael Ciarametaro, vice president of research, National Pharmaceutical Council • Daryl Pritchard, PhD, senior vice president, science policy, Personalized Medicine Coalition

Data Sources The report is based on peer-reviewed literature, grey literature, and key informant interviews. Figure 1 maps data sources to the report’s content; it indicates which sources we used to collect information on each topic. For example, we used the information we gathered from the research databases to inform the content related to the types of studies, safety, and efficacy for each intervention. (For more details on the methodology, see Appendix A and Appendix B.)

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Figure 1. Content and Data Sources

Organization of the Report Series and Organization of This Report This is the first of 2 reports on the evidence base for FDA-approved therapies that meet the definition for gene therapy. This report also covers interventions that are not yet approved but that we identified multiple times in our search for therapies in the research pipeline. We focus on adenovirus, adeno- associated virus and clustered regularly interspaced short palindromic repeats gene therapy. A second report will address chimeric antigen receptor T cell (CAR-T), autologous cell, zinc finger nuclease (ZFN), antisense, ribonucleic acid interference (RNAi) therapies and oncolytic viral therapy, which uses a replication-competent, attenuated derivative of herpes simplex virus type 1. The sheer number of studies, spanning multiple interventions and conditions, necessitates the use of broad summaries. However, we do highlight specific issues that the authors or the key informants believe are especially relevant to the report. For each report, the findings are organized by key questions.

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Description of Interventions/Technologies • 1a. What different types or modalities of the intervention have been used in clinical practices and clinical research studies? • 1b. What are the potential/presumed advantages and disadvantages in contrast to current practices? • 1c. What are the safety issues or expected adverse events?

Context in Which Gene Therapy Is Used • 2a. What is the approval process and current approval/certification status for gene therapies? • 2b. Are any additional accompanying resources or technologies required? • 2c. What is the current state of adoption in practice and settings? • 2d. What are the operator factors, such as training and staffing?

Ongoing Premarket and Postmarket Gene Therapy Studies • 3a. What are the indication/patient inclusion criteria in ongoing trials? • 3b. What are the types of interventions in ongoing trials? • 3c. What are the study designs/sizes of ongoing trials? • 3d. What are the comparators in ongoing trials? • 3e. What are the previous/concurrent treatments in ongoing trials? • 3f. What is the length of follow-up in ongoing trials? • 3g. What outcomes are measured in ongoing trials?

Current Evidence Supporting Gene Therapy • 4a. What are the indication/patient inclusion criteria in published studies? • 4b. What are the types of interventions in published studies? • 4c. What are the study designs/sizes of published studies? • 4d. What are the comparators in published studies? • 4e. What are the previous/concurrent treatments in published studies? • 4f. What is the length of follow-up in published studies? • 4g. What outcomes are measured in published studies? • 4h. What are the adverse events in published studies? Furthermore, for each report, we also document the available evidence in comprehensive evidence tables, which list key characteristics for all included studies. We stratify the evidence by gene therapy type and by completed and published studies versus potential pipeline therapies in ongoing studies. We also provide an evidence map—a visualization of the evidence. Evidence maps are useful as signposts for health care practitioners, policymakers, and funding agencies. They are ideally suited to providing many different stakeholders with an overview of a broad research area.

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Description of Interventions/Technologies The FDA has currently approved 16 cellular or gene therapies. Table 1 describes the 7 intervention types that meet the FDA’s definition of gene therapy.

Table 1. Intervention Descriptions Intervention Type Description Adeno-associated virus vector The virus infects patient cells to deliver a healthy copy of a gene so that it will be properly expressed, thereby curing the disease. Antisense Complementary messenger RNA binds to and silences a disease-causing gene. Chimeric antigen receptor T cell T cells are removed from the patient, genetically modified to attack specific cancer cells, and infused back into the patient. Genetically modified oncolytic viral therapy using a The virus infects and replicates in cancer cells to replication-competent, attenuated derivative of herpes express protein (eg, granulocyte-macrophage colony- simplex virus type 1 stimulating factor), which results in cell lysis and increased antitumor immunity. RNA interference Double-stranded RNA degrades RNA that encodes disease-causing protein. Autologous cell Recovered cells or tissue from a patient are genetically modified, then reintroduced. Zinc finger nuclease DNA-binding proteins are engineered to create double- strand breaks in DNA at specific targeted locations.

Abbreviations: DNA, deoxyribonucleic acid; RNA, ribonucleic acid.

Table 2 lists the 10 FDA-approved therapies (as of November, 2019) that meet the definition of gene therapy. In January 2013, the FDA approved Kynamro (Mipomersen sodium), the first therapy within the scope of this report to be approved. Kynamro is based on antisense technology and is used to treat homozygous familial hypercholesterolemia. In 2015, the FDA approved Imlygic (Talimogene laherparepvec) to treat melanoma. Imlygic is a genetically modified oncolytic viral therapy that uses a replication-competent, attenuated derivative of herpes simplex virus type 1. The next 3 approved therapies use antisense technology to treat Duchenne muscular dystrophy (Eteplirsen, Exondys 51), spinal muscular atrophy (Spinraza, Nusinersen), and polyneuropathy of hereditary transthyretin amyloidosis (Tegsedi, Inotersen). Luxturna, approved in 2017, was the first AAV-based therapy to treat biallelic RPE65 mutation-associated retinal dystrophy. Subsequently in 2017, the FDA approved 2 CAR-T therapies. Yescarta is used to treat adult patients with relapsed or refractory large B-cell lymphoma, and Kymriah treats children and young adults with refractory B-cell precursor acute lymphoblastic leukemia. In 2018 Kymriah gained additional approval to treat adult patients with relapsed or refractory large B-cell lymphoma. In 2018 the FDA approved an RNA interference drug to treat hereditary transthyretin amyloidosis with polyneuropathy. A key aspect of gene therapies is their potential to cure disease with one application. For instance, the CAR-T therapies Kymriah and Yescarta are not expected to need repeated procedures, and the AAV therapy Luxturna is expected to be curative with one procedure. However, there has not yet been sufficient follow-up to know whether these treatments are truly curative—the number of treated patients is small, and researchers need to follow patients for several more years. Long-term follow-up trials are under way for Luxturna7; so far, 12-month follow-up results support maximum treatment benefit.8 Similarly, Yescarta has been shown to have durable response after 15 months, as

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documented in the CAR-T evidence table in Appendix F.9 Trial results also suggest that Kymriah has a durable response after a median follow-up of 13 months; this trial is ongoing (see section entitled Ongoing Premarket and Postmarket Gene Therapy Studies.10 Most key informants mentioned the need for longer-term studies, spanning at least 10 years, to assess durability.

Table 2. FDA-approved Therapies Modality/Type Name (Brand First FDA Indication Approval Are Repeat of Gene Therapy Name if Approval Date Date for Procedures Approach Applicable) Specific Expected? Indication Antisense Kynamro 1/29/13 Homozygous familial 1/29/13 Yes (Mipomersen hypercholesterolemia sodium) Genetically Imlygic 10/27/15 Melanoma 10/27/15 Yes modified oncolytic (Talimogene viral therapy laherparepvec) (replication- competent, attenuated derivative of herpes simplex virus type 1) Antisense Eteplirsen 9/19/16 Duchenne muscular 9/19/16 Yes (Exondys 51) dystrophy Antisense Nusinersen 12/23/16 Spinal muscular 12/23/16 Yes (Spinraza) atrophy Antisense Tegsedi 10/5/18 Polyneuropathy of 10/5/18 Yes (Inotersen) hereditary transthyretin amyloidosis AAV Luxturna 12/19/17 Biallelic RPE65 12/19/17 No (Voretigene mutation-associated neparvovec) retinal dystrophy AAV Zolgensma* 5/24/19 Spinal muscular 5/24/19 No (Onasemnogene atrophy abeparvovec-xioi) CAR-T: CD19- Yescarta 10/18/17 Relapsed or 10/18/17 No directed (Axicabtagene refractory large B- genetically ciloleucel) cell lymphoma modified autologous T-cell immunotherapy CAR-T: CD19- Kymriah 8/30/17 Patients up to 25 8/30/17 No directed (Tisagenlecleucel) years of age with genetically refractory B-cell modified precursor acute autologous T-cell lymphoblastic immunotherapy leukemia Adult patients with 5/1/18 No relapsed or refractory large B- cell lymphoma

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Modality/Type Name (Brand First FDA Indication Approval Are Repeat of Gene Therapy Name if Approval Date Date for Procedures Approach Applicable) Specific Expected? Indication RNA interference; Patisiran 8/10/18 Hereditary 8/10/18 Yes specifically, (Onpattro, ALN- transthyretin double-stranded TTR02) amyloidosis with small interfering polyneuropathy ribonucleic acid, formulated as lipid nanoparticles for delivery to hepatocytes

Abbreviations: AAV, adeno-associated virus vector; CAR-T, chimeric antigen receptor T cell; FDA, US Food and Drug Administration; RNA, ribonucleic acid. Note: Shading indicates adenovirus and AAV gene therapy. *This table was updated on February 3, 2020 to reflect the approval of Zolgensma (formerly AVXS-101) for spinal muscular atrophy (approved on 5/24/19)

Table 3 lists the identified therapies that have not yet been approved but might be close to approval. To identify these therapies, we searched the grey literature, including a database maintained by the Journal of Gene Medicine and Pink Sheet of Pharma Intelligence (see Appendix B). The table presents the intervention modality, name/brand, FDA designation, and indication associated with the therapies that have yet to be approved. Adenovirus, AAV, and CRISPR therapies are shaded; the unshaded therapies are addressed in the second report.

Table 3. Potential Pipeline Gene Therapies Modality/Type of Intervention Name of Intervention Indication (Brand Name if Applicable) AAV A001 (AAV2/5-OPTIRPE65) Leber congenital amaurosis AAV A002 (AAV2/8- hCARp.hCNGB3) Achromatopsia AAV AAV2.REP1 (AAV-mediated REP1 ) Choroideremia AAV NSR-REP1 (AAV2-REP1) Choroideremia AAV rAAV2.REP1 Choroideremia AAV SPK-7001 (AAV2-hCHM) Choroideremia AAV rAAV2tYF-PR1.7-hCNGB3 Achromatopsia AAV AAV-RPGR X-linked retinitis pigmentosa AAV ABO-101 (rAAV9.CMV.hNAGLU) Sanfilippo syndrome type B AAV ABO-102 (scAAV9.U1a.hSGSH) Sanfilippo syndrome type A AAV BIIB087 (rAAV2tYF-CB-hRS1) X-linked retinoschisis AAV BIIB088 X-linked retinitis pigmentosa AAV SB-525 Hemophilia A AAV SHP654/BAX 888 Hemophilia A AAV SPK-8011 Hemophilia A AAV Valoctocogene Roxaparvovec (BMN Hemophilia A 270) AAV AMT-060 Hemophilia B AAV AMT-061 (AAV5-hFIXco-Padua) Hemophilia B AAV Fidanacogene elaparvovec (SPK- Hemophilia B 9001, PF-06838435)

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Modality/Type of Intervention Name of Intervention Indication (Brand Name if Applicable) AAV PF-06939926 (BMB-D001) Duchenne muscular dystrophy AAV SGT-001 Duchenne muscular dystrophy AAV AAVrh74.MHCK7.micro-Dystrophin Duchenne muscular dystrophy AAV AVXS-101* Spinal muscular atrophy AAV MYO-101 Limb-girdle muscular dystrophy, (scAAVrh74.MHCK7.hSGCB) type 2E AAV MYO-102 (scAAVrh74.tMCK.hSGCA) Limb-girdle muscular dystrophy, type 2D AAV MYO-201 Limb-girdle muscular dystrophy, (rAAVrh74.MHCK7.DYSF.DV) type 2B /Miyoshi Myopathy AAV rAAV1.tMCK.human-alpha- Limb-girdle muscular dystrophy, sarcoglycan type 2D AAV VY-AADC Parkinson's disease Adenovirus, oncolytic Tasadenoturev (DNX-2401) Brain cancer Adenovirus, vaccine Ad26.ZEBOV Ebola CRISPR-Cas9–CAR-T-cell therapy Anti-mesothelin CAR-T cells Solid tumor cancers CRISPR-Cas9–CAR-T-cell therapy CD7.CAR/28zeta CAR-T cells T-cell acute lymphoblastic leukemia, T-cell non-Hodgkin lymphoma CRISPR-Cas9–CAR-T-cell therapy PD-1 knockout EBV-CTL Gastric carcinoma, nasopharyngeal carcinoma, T-cell lymphoma, adult Hodgkin lymphoma, diffuse large B- cell lymphoma CRISPR-Cas9–CAR-T-cell therapy PD-1 knockout T cells Prostate cancer, bladder cancer, non-small-cell lung cancer, renal cell carcinoma, esophageal cancer CRISPR-Cas9–CAR-T-cell therapy UCART019 B-cell leukemia or lymphoma CRISPR-Cas9–CAR-T-cell therapy Universal dual specificity CD19 and B-cell leukemia or lymphoma CD20 or CD22 CAR-T cells CRISPR-Cas9–autologous CD34+ CTX001 Beta-thalassemia human hematopoietic stem and progenitor cells Antisense IONIS-MAPTRx Alzheimer's disease Antisense RG6042 (IONIS-HTTRx) Huntington's disease Antisense Volanesorsen Familial chylomicronemia syndrome, familial partial lipodystrophy Autologous cell therapy Strimvelis (GSK2696273) Severe combined immunodeficiency Autologous cell therapy Lenti-D Cerebral adrenoleukodystrophy (Lorenzo’s Oil disease) Autologous cell therapy LentiGlobin (BB305) Beta-thalassemia Autologous cell therapy OTL-101 Adenosine deaminase severe combined immunodeficiency Autologous cell therapy OTL-200 (GSK2696274) Metachromatic leukodystrophy Autologous cell therapy EB-101 Recessive dystrophic epidermolysis bullosa Autologous cell therapy FCX-007 Recessive dystrophic epidermolysis bullosa Autologous cell therapy FCX-013 Moderate to severe localized scleroderma

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Modality/Type of Intervention Name of Intervention Indication (Brand Name if Applicable) CAR-T-cell therapy bb2121 B-cell maturation antigen-expressing multiple myeloma, multiple myeloma CAR-T-cell therapy BCMA (EGFRt/BCMA-41BBz CAR-T Multiple myeloma cell) CAR-T-cell therapy JCAR014 Non-Hodgkin lymphoma CAR-T-cell therapy JCAR016 Acute myeloid leukemia, non-small- cell lung cancer, mesothelioma CAR-T-cell therapy JCAR018 Non-Hodgkin lymphoma, pediatric acute lymphoblastic leukemia CAR-T-cell therapy JCAR020 Ovarian cancer CAR-T-cell therapy JCAR023 Pediatric neuroblastoma CAR-T-cell therapy JCAR024 ROR1+ cancers (solid tumors, blood cancers) CAR-T-cell therapy JCARH125 B-cell maturation antigen target in late-line myeloma CAR-T-cell therapy Lisocabtagene maraleucel (Liso-Cel, Non-Hodgkin lymphoma, diffuse JCAR017) large B-cell lymphoma, follicular lymphoma, mantle-cell lymphoma, primary mediastinal B-cell lymphoma Injectable retroviral replicating Vocimagene amiretrorepvec (Toca Glioma vector that encodes a prodrug 511 & Toca FC) activator enzyme, cytosine deaminase RNAi ALN-TTRsc02 Acquired transthyretin amyloidosis RNAi AMG 890 (ARO-LPA) Apolipoprotein A reduction RNAi ARO-AAT Alpha-1 antitrypsin deficiency liver disease RNAi ARO-HBV Chronic hepatitis B infection RNAi Cemdisiran (ALN-CC5) Complement mediated disease RNAi Fitusiran (ALN-AT3sc) Hemophilia A, B RNAi Givosiran (ALN-AS1) Acute hepatic porphyrias RNAi Inclisiran (ALN-PCSsc) Hypercholesterolemia RNAi Lumasiran (ALN-GO1) Primary hyperoxaluria Type 1 RNAi Revusiran (ALN-TTRsc) Transthyretin-mediated amyloidosis, familial amyloidotic polyneuropathy, ATTR amyloidosis, familial amyloid neuropathies, mediated familial amyloidotic cardiomyopathy Wyeth strain vaccinia virus/ Pexastimogene Devacirepvec (Pexa- Cancer (various) granulocyte-macrophage colony Vec, JX-594) stimulating factor expression ZFN SB-318 MPS I (Scheie, Hurler-Scheie, and Hurler syndromes) ZFN SB-913 MPS II (Hunter syndrome) ZFN SB-FIX Hemophilia B ZFN ST-400 Beta-thalassemia ZFN–CAR-T-cell therapy SB-728-T HIV ZFN–stem cells SB-728-HSPC HIV

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Abbreviations: AAV, adeno-associated virus vector; CAR-T, chimeric antigen receptor T cell; CRISPR, clustered regularly interspaced short palindromic repeats; CRISPR-Cas, CRISPR-associated protein; MPS, Mucopolysaccharidosis; RNA, ribonucleic acid; RNAi, RNA interference; ZFN, zinc finger nuclease. Note: Shading indicates adenovirus and AAV gene therapy. *AVXS-101 (Avexis/Novartis) to treat spinal muscular atrophy was approved on May 24, 2019 as Zolgensma

Our search identified 76 therapies that meet the FDA’s definition of gene therapy and that have not yet been approved (ie, pipeline therapies). The most common type of therapy is AAV (37%). The next most common types are CAR-T-cell therapy (13%), RNAi (13%), autologous cell therapy (11%), CRISPR-Cas9–CAR-T-cell therapy (8%), ZFN (5%), antisense (4%), adenovirus (3%), CRISPR- Cas9–autologous CD34+ human hematopoietic stem and progenitor cells (1%), injectable retroviral replicating vector that encodes a prodrug activator enzyme, cytosine deaminase (1%), Wyeth strain vaccinia virus/granulocyte-macrophage colony stimulating factor expression (1%), ZFN–CAR-T-cell therapy (1%), and ZFN–stem cell therapy (1%). The identified therapies are being used to treat multiple disease types, including vision disorders (eg, choroidemia), muscular dystrophy (eg, Duchenne), blood disorders (eg, hemophilia), Sanfilippo syndrome, cancer, HIV, and Parkinson’s disease. We asked key informants to name any pipeline therapies that are likely to gain approval in the next 5 years. They identified 6 therapies, which are included in Table 3: • AVXS-101 (Avexis/Novartis) to treat spinal muscular atrophy (2 nominations) was approved on May 24, 2019 as Zolgensma • Valoctocogene roxaparvovec (Biomarin) to treat hemophilia A (1 nomination) • SPK-9001(Spark Therapeutics) to treat hemophilia A (2 nominations) • SPK-8011 (Spark Therapeutics) to treat hemophilia B (1 nomination) • AMT-061 (uniQure) to treat hemophilia B (1 nomination) • LentiGlobin (Bluebird bio) to treat beta-thalassemia (2 nominations) One key informant thought AMT-061 was close to approval, and 2 nominated AVXS-101 and LentiGlobin. One key informant nominated Valoctocogene roxaparvovec, SPK-9001, and SPK-8011. The fact that key informants were aware of these 6 therapies among the large number of potential pipeline therapies we identified suggests that these 6 are closer to approval than the others. AVXS-101, Valoctocogene Roxaparvovec, SPK-9001, and SPK-8011 are AAV interventions; we identified completed trials with promising evidence for AVXS-10111 and for SPK-9001.12 As noted above AVXS-101 was approved as Zolgensma on May 24, 2019. LentiGlobin is an autologous cell intervention; along with Valoctocogene Roxaparvovec, SPK-801, and AMT-061, it might still be undergoing testing in trials that have yet to complete recruitment.

Adenoviral, AAV, and CRISPR Gene Therapy Eligible interventions included those that used adenovirus, AAV, or CRISPR therapies. In the following sections we describe the indications and their US prevalence for approved and as yet unapproved adenoviral, AAV, and CRISPR technologies.

Adenoviral Therapy Indications Unapproved Our search identified one adenoviral-based therapy that targets brain cancer tumor cells. In 2015, an estimated 166 039 people in the United States were living with brain and other nervous system cancer.13 This therapy has not yet been approved by the FDA.

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AAV Therapy Indications Approved Luxturna (Voretigene neparvovec, an AAV therapy) is the only approved therapy that falls into the scope of this report. Luxturna is used to treat biallelic RPE65 mutation-associated retinal dystrophy, which currently afflicts approximately 1000 to 2000 individuals in the United States.14 RPE65 is a key isomerase for normal enzymatic processes, and mutations in RPE65 can result in blindness.15 Luxturna acts by delivering the expression of a healthy copy of the RPE65 gene directly to the subretinal space. Unapproved Multiple AAV therapies are in trials to treat a broad array of conditions. Ocular disorders • Leber congenital amaurosis is an eye disorder that affects the retina and is associated with vision impairment. Although it is relatively rare, occurring in just 2 to 3 of 100 000 newborns, it is the most common form of inherited blindness in children.16 • Achromatopsia is associated with partial or total lack of color vision as well as photosensitivity and low visual acuity; it occurs in 1 of 30 000 people globally.17 • Choroideremia begins with impaired vision at night and progresses to reduced visual acuity. Its prevalence is estimated to be 1 in 50 000 to 100 000 among people of European descent.18 • X-linked retinitis pigmentosa expresses only in males; the estimated prevalence is 1 to 5 of 10 000 people.19 Heterozygous females are assumed to be unaffected because they can have a healthy copy of the gene to compensate for the mutated copy; however, carrier females may suffer from symptoms.20

Blood disorders • Hemophilia A is a bleeding disorder that affects 1 in 5000 newborn males; it is 4 times more common than hemophilia B. An estimated 20 000 people in the United States have hemophilia.21,22

Metabolic disorders • Sanfilippo syndrome is a rare inherited metabolic disorder that leads to progressive neurocognitive degeneration. The prevalence for all subtypes is estimated to be 1 to 9 in a million people.23

Muscular disorders • Duchenne muscular dystrophy is a genetic disorder that manifests most severely in males (1 in 3500 to 5000 males) and leads to progressive muscle degeneration.24 • Spinal muscular atrophy is a neuromuscular disease that results in muscle weakness and paralysis. The estimated incidence is 1 in 6000 to 10 000 live births.25 • Limb-girdle muscular dystrophy results in atrophy of the limb-girdle muscles; it can cause serious disability or may progress slowly and cause minimal disability.26 Prevalence estimates range from 1 in 14 500 to 1 in 123 000 individuals.27 • Parkinson’s is the most common movement disorder; it is characterized by tremors, rigidity, and bradykinesia.28 It is estimated that almost a million people in the United States will have Parkinson’s in 2020.

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CRISPR Therapy Indications Unapproved While CRISPR has not yet been approved in the United States, many trials are under way and recruiting patients to treat cancer and blood disorders. Cancer • Solid tumor cancers (eg, sarcomas, carcinomas, lymphomas) are defined as abnormal masses that do not contain cysts or liquid areas,29 so they can affect multiple organs (eg, colon). It was estimated that across all types of cancer there were 1 735 350 new cases in 2018.30 • T-cell acute lymphoblastic leukemia is an aggressive type of leukemia31; the estimated incidence in the United States is 0.13 cases per 100 000 people.32 • T-cell lymphoma is a rare malignancy based on a collection of subtypes of non-Hodgkin lymphoma.33 Non-Hodgkin lymphoma accounts for 4% of cancers in the United States; it includes mycosis fungoides, anaplastic large-cell lymphoma, and precursor T-lymphoblastic lymphoma.34 An estimated 74 680 adults and children were diagnosed with non-Hodgkin lymphoma in 2018.35 • Diffuse large B-cell leukemia or lymphoma is an aggressive cancer and the most common type of non-Hodgkin lymphoma in the United States.36 On the basis of 2011-2015 data, the estimated number of new cases in 2011-2015 was 5.5 per 100 000 adults.37 • Gastric cancer is the fourth most common form of cancer. There were an estimated 989 000 new cases globally in 2008.36 • The estimated incidence for nasopharyngeal carcinoma, a relatively rare malignancy, is less than 1 per 100 000 person years.38 • After skin cancer, prostate cancer is the next most common cancer in men; it was estimated that there were 164 690 new cases in 2018.39 • Bladder cancer is the fourth most common cancer in men; it is less common in women.40 It was estimated there were 81 190 new cases in 2018. • About 80% to 85% of lung cancers are non-small-cell lung cancer; it was estimated that there were 234 030 new cases of lung cancer in 2018.41 • Kidney cancer (also known as renal cell carcinoma) is among the 10 most common cancers in men and women.42 It was estimated that there were 65 340 new cases in 2018.43 • Esophageal cancer is a relatively rare malignancy; it was estimated that there were 17 290 new cases in 2018.44

Blood disorders • A CRISPR-based intervention has been designed to treat beta-thalassemia, which is a rare inherited blood disorder. The severity of beta-thalassemia can range from mild to severe, in which patients require regular blood transfusions throughout their lives. The incidence of symptomatic cases is estimated to be approximately 1 in 100 000 people globally.45

Key Question 1a. What different types or modalities of the intervention have been used in clinical practices and clinical research studies? Gene therapy interventions either modify or edit a patient’s gene, or they introduce a protein-producing gene into the patient. In the following sections we describe the adenovirus, AAV, and CRISPR interventions according to whether they act by modifying or editing, or by delivering a gene and its expression to a patient.

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Gene Editing or Replacement The CRISPR-Cas (CRISPR-associated protein) system was recently identified as a powerful tool for therapeutic gene editing. CRISPR-Cas was first discovered in bacteria and archaea46 as an adaptive defense system that uses antisense RNAs as memory signatures of past invasions.47,48 Functional CRISPR-Cas loci comprise identical repeats, intercalated with invader DNA-targeting spacers that encode the Cas components. The CRISPR-associated protein Cas9 is a dual RNA–guided endonuclease that uses a guide sequence within an RNA duplex to form base pairs with DNA target sequences, allowing Cas9 to create a site-specific double-strand break in the DNA.49,50 The CRISPR- Cas9 system requires only a change in the guide RNA sequence, which avoids substantial protein engineering.51 Both ex vivo and in vivo approaches can deliver CRISPR to patients. In vivo, CRISPR machinery is packaged in a vector, such as a virus, and then delivered directly via injection into the target organ. With the ex vivo approach, cells are extracted from patients, modified with CRISPR technology, grown, and then reintroduced into the patient. Genes can be modified in 1 to 2 weeks, then grown within 2 to 3 weeks.52 Gene Delivery and Expression Adenovirus and AAV vector-based therapies both use a nonpathogenic virus to deliver DNA and gene transfer to patients. They act by incorporating the virus’s genome into a patient’s dividing and nondividing cells. The adenovirus genome is a linear double-stranded DNA between 26 kb and 48 kb in size.53 Adenoviruses can deliver up to 30 kb of DNA and produce gene expression in less than 24 hours, but the expression is transient. Another drawback of this therapy is that administration of adenovirus creates an innate immune response that may include production of pro-inflammatory chemokines and cytokines, thrombocytopenia, coagulopathy, and liver damage.54 AAV is a single- stranded DNA genome that is approximately 5 kb long.54 AAVs can deliver smaller DNA segments than adenoviruses, elicit low immune responses, and can potentially provide long-lasting gene expression. Luxturna (Voretigene neparvovec) is an AAV-based gene therapy to treat children and adults with a confirmed biallelic RPE65 mutation-associated retinal dystrophy that leads to vision loss and can cause complete blindness. Both adenovirus and AAV can be used alone or in combination with other modalities; for example, AAV can be used to deliver CRISPR. Both adenoviral and AAV therapies can be injected locally to the site of disease; for example, Luxturna is injected directly into the retina. Other delivery approaches being tested include systemic delivery targeting the liver, intramuscular injection, and central nervous system delivery.55

Key Question 1b. What are the potential/presumed advantages and disadvantages in contrast to current practices? Advantages Many of the potential and presumed advantages of gene therapies are similar across all types of gene therapy. The first and foremost advantage is to cure previously incurable and fatal diseases. For many diseases the only treatment is supportive care, or the treatment has dire side effects or is very costly. In cases in which a disease has a spectrum of severity, our review of the literature found that gene therapy typically targets the most refractory types. On the basis of our review and interviews with key informants, the greatest advantage of gene therapy is reducing suffering by addressing unmet need. A gene therapy would have an advantage over current therapy if it could negate the need for any future treatment. Theoretically, some gene therapies can cure the targeted condition with only one administration, and some conditions can be manageable with current practices and repeated treatments.

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However, high prices for a curative genetic treatment may limit access to the treatment and thus reduce or obviate the therapy’s advantage over current treatment. AAV is one type of gene therapy that is expected to require only one administration, which would be a significant advantage over current treatments that require repeated procedures. For example, hemophilia is currently managed with repeated transfusions, which are a physical burden to patients and costly to patients and health care payer institutions.56,57 Several AAV therapies are being tested that would treat these patients with one administration, eliminating the need for regular transfusions. Multiple adenoviral and CRISPR therapies are in trials to cure certain types of cancer. If these therapies are successful, they will replace chemotherapy, which has the potential to cause chemotoxicity in patients. Disadvantages All types of gene therapies share many unknowns and uncertainty related to the following: (1) their clinical effectiveness versus the risk for adverse outcomes; (2) how they will be implemented in health care systems in which patients are not well informed about what gene therapies are and providers are unfamiliar with requisite procedures; (3) the ethical implications of altering the genome; and (4) how patients and payers will pay for potentially expensive gene therapy. We need more evidence to know whether these unknowns will become disadvantages. Several cases of gene-related death slowed gene therapy progress in the early years of this research. Jesse Gelsinger was the first patient to die in 1999 after receiving gene therapy that triggered a fatal immune response, which resulted in the suspension of adenoviral trials.58 Later in 2002, a French child developed a leukemia-like illness that was thought to have been caused by the gene therapy.59 Other cases include a death due to a fatal infection after a patient had received immunosuppressive therapy in a gene therapy trial.60 Longer-term disadvantages are related to risks that are not yet understood, including future malignancy. All of our key informants were concerned about the risk of adverse health outcomes, including off-target effects, viral spread, genotoxicity, and alterations to the germline. Viral-delivered gene therapy can induce genotoxic events, including inflammation, immune response, random insertion that can disrupt normal genes, activation of proto- oncogenes, and insertional mutagenesis.61 Specific targeting of disease cells with viral cells is a challenge, and viruses can spread and infect healthy cells. Moreover, a virus might insert DNA into the wrong location, causing off-target effects even in reproductive cell lines, thereby changing the germline. These effects are all possible outcomes of adenoviral, AAV, and CRISPR treatment. Price may be a disadvantage of these therapies relative to others. New gene therapies are expensive to patients and payers; for example, Luxturna treatment is priced at $425 000 per eye.62 During our interviews, one key informant expressed concern about the viability of small insurers, small self-insured employers, and even large self-insured employers with limited reinsurance in the face of very high treatment prices. Because these therapies may provide large health gains and reduce future treatment prices, manufacturers can charge high upfront payments to reflect the value of the therapy. Such high short-term fees can limit access if the treatment is unaffordable. While the price of a particular therapy might be based on its perceived value, little clinical evidence exists to establish value for money. The Institute for Clinical and Economic Review white paper “Gene Therapy: Understanding the Science, Assessing the Evidence, and Paying for Value” discusses the challenges gene therapy presents in the United States, as well as potential solutions (eg, annualized payments or risk sharing).63 Preclinical work and drug optimization in mice might not translate to human treatment outcomes. For example, AAV serotypes might be very effective in mice or nonhuman primates but might not be optimized for humans, so they would be less effective for human disease treatment. At

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the same time, industry might be reluctant to repeat human trials if the therapy doesn’t perform, because industry would have to invest additional substantial financial and labor resources. However, if a treatment doesn’t work and scientists don’t understand why, they might be missing an opportunity to improve the therapy. Key informants noted that the FDA requires multiple preclinical studies to reapprove slightly modified gene therapy vectors for new clinical trials. This can make it more difficult for researchers to learn and adapt from failed clinical trials. Also, key informants noted that the FDA’s emphasis on functional outcomes rather than biomarkers in clinical trials with small sample sizes can make it difficult to find statistically significant effects. Functional outcomes have a higher interpatient variation than biomarkers and posttreatment improvement, so such studies require larger patient sample sizes than studies with biomarker outcomes.64 Even among completed human trials, the patient populations can be very small and evidence about the treatment’s effectiveness can be limited. One key informant noted that patients might be reluctant to participate in trials to assess minimal dosage safety because they want the higher dose. For example, with adenovirus and AAV therapies, one treatment will create host immunity to the vector, so patients have only one chance for treatment; most will not want their chance for a cure to be lost because the dose was too low to be effective. Other disadvantages discussed by key informants were related to manufacturing and the need for greater understanding of the science and mechanisms of action to regulate and ensure quality in manufacturing. Gene therapies are regulated as products, but one key informant suggested that perhaps they should be regulated as procedures, which raises the question of who should be permitted to administer gene therapy. Gene therapies could become like stem cell therapies, which have been administered at certain times and places with little regulation of who provides the therapy. For example, in Australia, autologous cellular treatments were once exempt from the regulatory framework that applies to cell and gene treatments; thus, the use of unproven autologous cell interventions grew markedly in Australia.65 Key informants also noted that the current health care system is not structured to deliver gene therapy, in part because providers are not fully aware of new, nonstandard therapies. Moreover, patients seeking care may have to travel far to the few existing gene therapy treatment centers, which can be costly and place heavy burdens on families. Key informants expressed concern about several other potential disadvantages. Several were concerned about delivering interventions to target cells more effectively and balancing minimally effective dosages with adverse reactions. And one key informant noted that much of the evidence for gene therapies includes small study samples, even though the indication is prevalent in large populations (eg, patients older than 65). For example, we found in our second report that many of the gene therapy trials treating cardiovascular disease have a relatively small number of participants compared with the burden of the disease in the population. Thus, lack of generalizability of findings is a concern. Most key informants expressed concern about the state of science education, the average person’s understanding of gene therapy, and how patients can weigh the risks and the information about effectiveness.

Key Question 1c. What are the safety issues or expected adverse events? Some therapies produce physiologic responses that can be dangerous if they are not managed clinically. The primary safety concerns mentioned by key informants involved unintended consequences of editing and introducing genetic material to a person. The concerns included malignancy, off-target effects, viral shedding or spread, and genotoxicity, in which integration of the vector DNA can have genotoxic effects. Regarding gene editing, a lack of specificity with CRISPR can have off-target effects that might result in malignancy.66 Immune responses to AAV pose few safety

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concerns, although preexisting anti-adenoviral antibodies might stimulate stronger innate immune responses after vector administration,67 which could be a serious concern. Regarding Luxturna (Voretigene neparvovec), some potential adverse events are related to the surgery68; for example, injection can introduce infection and inflammation that can cause vision loss. Other possible adverse events related to the treatment and eye response are retinal abnormalities (eg, tears), increased intraocular pressure, air bubbles, and cataract. The most common adverse events are conjunctival hyperemia, cataract, increased intraocular pressure, retinal tear, dellen (thinning of the corneal stroma), macular hole, subretinal deposits, eye inflammation, eye irritation, eye pain, and maculopathy (wrinkling on the surface of the macula).

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Context in Which Gene Therapy Is Used

Key Question 2a. What is the approval process and current approval/certification status for gene therapies? In the United States, the FDA’s Center for Biologics Evaluation and Research reviews and regulates the use of human gene therapies, cellular therapy products, and devices related to gene and cell therapy. Within the National Institutes of Health (NIH), the Office of Biotechnology Activities and the Recombinant DNA Advisory Committee (RAC) review gene therapy protocols. While both the FDA and the RAC consider clinical and preclinical issues, the RAC also serves as an open forum for consideration of ethical, legal, and societal issues.69 Gene therapy sponsors submit an investigational new drug application, a biologics license application, and an investigational device exemption application to the FDA. In the investigational new drug application, manufacturers must describe how they will conduct the study, possible risks that might be involved and what steps they will take to protect patients, and the evidence that supports the study. The FDA has published guidance documents70 to facilitate the approval process. Manufacturers of gene therapy products must comply with FDA requirements for safety, purity, and potency before their therapies can be approved and sold. In August 2018, NIH reported that the RAC will no longer review all gene therapy applications and will instead provide guidance in an advisory role.71,72 The FDA will now review and approve gene therapies and biologic products as it does other treatments and drugs. In cases in which the disease is severe and no alternative treatments exist, the FDA has approved gene therapies on the basis of single-arm phase II trials. Luxturna (Voretigene neparvovec) was approved after one randomized phase III trial reported improved visual acuity 30 days post treatment that persisted 1 year.73,74 Thirteen of the 20 intent-to-treat intervention participants passed the multi-luminance mobility test at the lowest luminance level tested at 1 year, and none of the control participants did. Other evidence came from a small open-label dose escalation phase I trial.75,76 A follow-up trial tested treatment in the contralateral eye and reported persistent improved functional vision after 3 years.77 Ten participants showed improvements in mean mobility and full-field light sensitivity in the injected eye by day 30 that persisted to year 3, but no significant change was seen in the previously injected eyes for mobility or white light full-field sensitivity. Patients who have been treated in both trials will be followed for 15 years.73 It does not appear that longer follow-up was a postmarketing requirement of the FDA, because the applicants included a plan for longer follow-up in their pharmacovigilance plan.78 The FDA’s Regenerative Medicine Advanced Therapy program provides support and assistance for the production of gene therapies that treat serious or life-threatening conditions and address unmet medical needs.79 In December 2016, Congress passed the 21st Century Cures Act, designed to accelerate medical product development in order to treat patients faster and more efficiently.80 In June 2018, the Center for Biologics Evaluation and Research granted the regenerative medicine advanced therapy designation to 24 products.79 The FDA currently has more than 700 active investigational new drug applications for gene therapy81 and recently announced that it plans to provide policy guidance to accelerate approval for the rapid expansion of gene therapy development.82

Key Question 2b. Are any additional accompanying resources or technologies required? Before a patient receives any gene therapy that requires injection delivery, he or she might be given prophylactic antibiotics. In one study, the providers used ultrasound to guide the injection of

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adenoviral therapy into the tumor.83 Immunosuppression therapy (eg, methylprednisone) might be given prior to AAV administration. For the specific AAV treatment of ocular disorders, vitrectomy must precede the subretinal injection of the drug (eg, Luxturna). Other cotreatments include disease- related standard of care; for example, chemotherapy might accompany any gene therapy that targets cancer.

Key Question 2c. What is the current state of adoption in practice and settings? Gene therapy is available through standard care when the intervention is approved. For example, Luxturna is available to patients at 10 designated ocular gene therapy treatment centers, 5 of which are in the Northeastern region of the United States. Two centers are in the South and 2 are on the West Coast, so patients might have to travel many miles for treatment.84 Gene therapy is also available through participation in a clinical trial. However, as the field is rapidly advancing, more treatments will likely be approved for use beyond trials. Key informants noted several challenges facing the adoption of gene therapies and gaps in our understanding of this question. For example, current approaches to payer approval are based on population estimates for risk management and not on personalized medicine. Generally, risks can be spread across populations, but this is less feasible when patients’ specific disease traits (eg, genetics) are used to develop and deliver tailored treatment. Further, even though high upfront prices can be offset by downstream health benefits, current insurance structures might not be able to cover expensive gene therapies. CMS will reimburse hospitals $400 000 for Yescarta and $500 000 for Kymriah for inpatient treatment. The patient is responsible for a $1340 deductible for inpatient treatment per benefit period.85 However, when patients are treated in outpatient settings they could be responsible for 20% ($79 000-$100 000). How other payers will handle payment is less clear. Payers and manufacturers are considering alternative payment scenarios that include staggered or long-term payment agreements or linking payment to patient outcomes.86 In August 2018, the Medicare Evidence Development and Coverage Advisory Committee met to discuss whether and how CMS should incorporate patient-reported outcomes (PROs) into coverage decisions for CAR-T and other gene therapies. While committee members expressed confidence in specific tools to measure PROs, some voiced concern about the difficulty of collecting and interpreting such data.87 In February 2019, CMS proposed to cover FDA-approved CAR T-cell therapy if the treatment is in a CMS-approved registry or clinical study and patients are monitored for at least 2 years posttreatment.88

Key Question 2d. What are the operator factors, such as training and staffing? The manufacturing process is a major operator factor in the success of gene therapy. The clinical production process is challenged by consistent process control and by its ability to achieve the scale needed for commercial use. For example, limited cell line availability and lack of manufacturing capacity hinder the consistent production and characterization of cellular products. In July 2018, the FDA published guidance on the manufacturing information required to support the clinical development of investigational human gene therapies.89 The need for rigorous, high-quality biomanufacturing processes increases the demands on staff training and hiring. In fact, staffing challenges might limit commercial-scale production of gene therapies more than facility availability.90 Key informants also noted that the current health care system is not structured to deliver gene therapy, in part because providers are not fully aware of new nonstandard gene therapies.

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Ongoing Premarket and Postmarket Gene Therapy Studies

Ongoing Trial Characteristics Table 4 describes 19 ongoing studies we identified that are evaluating adenovirus, AAV, or CRISPR interventions. We identified only one CRISPR study that met our criteria for an ongoing trial that might be close to approval in which patient recruitment is complete. This trial is testing a CRISPR intervention to treat metastatic non-small-cell lung cancer.91 All other currently registered CRISPR trials in clinicaltrials.gov are still recruiting participants.

Table 4. Ongoing Adenovirus, AAV, and CRISPR Trials Indication Indication Intervention Outcomes Measured Author Concurrent/Previous Comparator Trial Number Treatments Study Design Months of Follow-up Estimated Completion Adenovirus Cancer Age group: adults Ad.hIFN-β; BG00001 Tumor regression, time University of Pennsylvania, Malignant metastatic or No comparison group to progression, survival 202392 pleural mesothelioma NCT00299962 Treatment enrollment: Trial (multiple groups), none planned N = 18 Treatment requisite: none Completion date: 04/2023 Follow-up: 180 AAV Blood disorders Age group: adults AskBio009 Change from baseline Shire, 203093 Hemophilia B No comparison group in clinical laboratory Weber et al, 201894 Treatment enrollment: evaluations; changes from baseline in FIX NCT01687608 hemophilia B with ≥3 hemorrhages per year activity levels, FIX Single-arm trial, planned protein levels, and N = 30 requiring treatment with exogenous FIX or use of bleeding episode Completion date: 10/2030 FIX prophylaxis due to severity and history of frequent bleeding frequency; immune episodes response to AskBio009; detection of AskBio009 Treatment requisite: none genomes in blood, Follow-up: 204 saliva, urine, stool, and semen Cancer Age group: adults AAV-DC-CTL ELISPOT assays, The First People's Hospital Stage IV gastric cancer No comparison group overall survival 95 of Changzhou, 2026 Treatment enrollment: NCT02496273 none Single-arm trial, planned Treatment requisite: none N = 60 Follow-up: 1 Completion date: 12/2030

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Indication Indication Intervention Outcomes Measured Author Concurrent/Previous Comparator Trial Number Treatments Study Design Months of Follow-up Estimated Completion Muscular conditions Age group: adults rAAVrh74.MHCK7.DYSF.DV Toxicity Hospital, 201696 Dysferlinopathy No comparison group NCT02710500 Treatment enrollment: Single-arm trial, planned none N = 2 Treatment requisite: none Completion date: 04/2019 Follow-up: 24 Muscular conditions Age group: children rAAVrh74.MHCK7.micro- Adverse events, Gross Nationwide Children's Duchenne muscular dystrophin Motor Subtest Scaled Hospital, 201897 dystrophy No comparison group score, 100 Meter NCT03375164 Treatment enrollment: Timed Test, North Star Ambulatory Trial (multiple groups), none Assessment, Timed Up planned N = 12 Treatment requisite: none and Go modified for Completion date: 01/2021 Follow-up: 36 children, ascend and descend 4 steps, handheld dynamometry, microdystrophin gene expression quantification by immunofluorescence, microdystrophin gene expression quantification by western blot Muscular conditions Age group: children and scAAVrh74.tMCK.hSGCA Change in 6-minute Mendell, 201998 adults No comparison group walk test NCT01976091 Limb-girdle muscular Trial (multiple groups), dystrophy type 2D planned N = 6 Treatment enrollment: Completion date: 02/2019 none Treatment requisite: none Follow-up: 24 Neurodegenerative Age group: adults AAV2-GDNF Safety, tolerability, disorders Advanced Parkinson’s No comparison group potential for clinical National Institute of disease responses Neurological Disorders and Treatment enrollment: 99 Stroke, 2025 disability despite optimal NCT01621581 antiparkinsonian medication Trial (multiple groups), therapy planned N = 25 Treatment requisite: none Completion date: 12/2027 Follow-up: 144

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Indication Indication Intervention Outcomes Measured Author Concurrent/Previous Comparator Trial Number Treatments Study Design Months of Follow-up Estimated Completion Neurodegenerative Age group: children AAVrh.10CUCLN2 Weill Cornell LINCL disorders Late infantile neuronal No comparison group Scale, Child Health Weill Medical College of ceroid lipofuscinosis, Batten Questionnaire, Mullen Cornell University, 2020100 disease Scale, MRI (percentage NCT01414985 Treatment enrollment: gray matter volume, percentage ventricular Trial (multiple groups), none volume, and cortical planned N = 8 Treatment requisite: none apparent diffusion Completion date: 12/2022 Follow-up: 18 coefficient) Neurodegenerative Age group: children AAVRh.10CUhCLN2 Weill-Cornell LINCL disorders Batten disease, late No comparison group scale score, disease Weill Medical College of infantile neuronal ceroid progression based on Cornell University, 2020101 lipofuscinosis change in MRI imaging NCT01161576 Treatment enrollment: parameter (percentage gray matter volume), Trial (multiple groups), none disease progression planned N = 25 Treatment requisite: none based on change in Completion date: 08/2032 Follow-up: 18 MRI imaging parameter (percentage ventricular volume), disease progression based on change in MRI imaging parameter cortical apparent diffusion coefficient), Quality of Life Survey, Mullen Scale score Neurodegenerative Age group: adults VY-AADC01 Columbia-Suicide disorders Advanced Parkinson’s No comparison group Severity Rating Scale, Therapeutics Voyager, disease change in Parkinson’s 102 2021 Treatment enrollment: medications, motor Single-arm trial, planned none function, dyskinesia occurrence, mood, N = 16 Treatment requisite: cognitive function, Completion date: 12/2021 patients will continue compulsive behavior, medications for Parkinson’s; sleep quality and patients undergo a disturbance, Non- neurosurgical procedure Motor System Scale, with real-time MRI quality of life monitoring so that therapy is delivered directly into the striatum Follow-up: 36

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Indication Indication Intervention Outcomes Measured Author Concurrent/Previous Comparator Trial Number Treatments Study Design Months of Follow-up Estimated Completion Neurodegenerative Age group: adults VY-AADC01 Parkinson’s symptoms, disorders Parkinson’s disease No comparison group PET scan imaging Voyager Therapeutics, Treatment enrollment: (AADC expression) 103 2019 Levodopa therapy NCT01973543 Treatment requisite: none Trial (multiple groups), Follow-up: 36 planned N = 15 Completion date: 12/2019 Ocular disorders Age group: adults AAV2-hCHM Safety and tolerability Spark Therapeutics, 2019104 Choroideremia, CHM gene No comparison group (assessed by physical NCT02341807 mutations exam, vital signs, laboratory changes Trial (multiple groups), Treatment enrollment: over time, and adverse planned N = 15 none events) Completion date: 01/2019 Treatment requisite: none Follow-up: 24 Ocular disorders Age group: adults GS010; rAAV2/2-ND4 ETDRS visual acuity, GenSight Biologics, 2018105 Leber hereditary optic Placebo improvement from NCT02652767 neuropathy, affected 6 baseline by 15 ETDRS letters or having visual RCT, planned N = 36 months or less acuity >20/200, optic Completion date: 09/2019 Treatment enrollment: none nerve retinal nerve fiber layer (RNFL) Treatment requisite: none thickness and the Follow-up: 24 thickness/volume of the retinal layers of the macula, Humphrey Visual Field 30-2, Pelli- Robson Contrast Sensitivity, Farnsworth-Munsell Color 100 Hue Vision Test, immune responses, blood biodissemination of AAV2 vector DNA, Better-seeing Eye Comparison, Visual Functioning Questionnaire-25, 36- item Short Form Health Survey

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Indication Indication Intervention Outcomes Measured Author Concurrent/Previous Comparator Trial Number Treatments Study Design Months of Follow-up Estimated Completion Ocular disorders Age group: adults GS010; rAAV2/2-ND4 ETDRS visual acuity, GenSight Biologics, 2018106 Leber hereditary optic Placebo improvement from NCT02652780 neuropathy, affected for Baseline by 15 ETDRS letters or having visual RCT, planned N = 36 more than 6 months acuity >20/200, optic Completion date: 01/2019 Treatment enrollment: none nerve RNFL thickness and the Treatment requisite: none thickness/volume of Follow-up: 24 the retinal layers of the macula, Humphrey Visual Field 30-2, Pelli- Robson Contrast Sensitivity, Farnsworth-Munsell Color 100 Hue Vision Test, immune responses, blood biodissemination of AAV2 vector DNA, Better-seeing Eye Comparison, Visual Functioning Questionnaire-25, 36- otem Short Form Health Survey Ocular disorders Age group: adults rAAV2.REP1 Best corrected visual STZ eyetrial, 2018107 Choroideremia No comparison group acuity in treated eye, NCT02671539 Treatment enrollment: absence of vector- related adverse Single-arm trial, planned none reactions, fundus N = 6 Treatment requisite: none autofluorescence Completion date: 03/2021 Follow-up: 24 analysis, central visual field using microperimetry readings, contrast sensitivity, color vision Ocular disorders Age group: children and rAAV2-ND4 Best corrected visual Huazhong University of adults No comparison group acuity, computerized Science and Technology, Leber hereditary optic visual field, visual 2019108 neuropathy evoked potential, NCT03153293 Treatment enrollment: no retinal nerve fiber layer, liver function in Single-arm trial, planned eyesight improvement in plasma, kidney N = 40 Leber's optic hereditary function in plasma Completion date: 06/2019 neuropathy patients or any other treatment within the past year Treatment requisite: none Follow-up: 12

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Indication Indication Intervention Outcomes Measured Author Concurrent/Previous Comparator Trial Number Treatments Study Design Months of Follow-up Estimated Completion Ocular disorders Age group: children and Voretigene neparvovec-rzyl; Bilateral mobility, full- Spark Therapeutics, 203073 adults AAV2-hRPE65v2 field light sensitivity NCT03602820 Inherited retinal dystrophy No comparison group threshold, visual acuity, visual field Single-arm trial, planned Treatment enrollment: testing, visual function N = 41 none questionnaire Completion date: 06/2030 Treatment requisite: none Follow-up: 180 Other: metabolic disorders Age group: children AAVrh.10cuARSA Tolerance, efficacy to Institut National de la Santé Metachromatic No comparison group stop disease Et de la Recherche leukodystrophy progression 109 Médicale, France, 2018 Treatment enrollment: NCT01801709 none Single-arm trial, planned Treatment requisite: none N = 5 Follow-up: 24 Completion date: 04/2019 CRISPR Cancer Age group: adults PD-1 knockout Response rate (per Sichuan University, 201891 Metastatic non-small-cell T cells RECIST v1.1), disease NCT02793856 lung cancer No comparison group control rate (response per RECIST v1.1), Trial (multiple groups), Treatment enrollment: progression-free planned N = 12 none survival, overall Treatment requisite: Completion date: 12/2018 survival, peripheral Peripheral blood blood-circulating tumor lymphocytes were collected DNA, cytokine (IL-10, from each patient and TNF-a, IL-6) changes CRISPR-Cas9 technology in peripheral blood was used to knock out the programmed cell death protein 1 gene to make PD- 1 knockout T cells; patients received a single dose of cyclophosphamide (chemotherapy conditioning) 3 days prior to cell infusion; patients received 6 infusions of knockout cells (2 cycles with 3 infusions each). Follow-up: 24

Abbreviations: AADC, aromatic L-amino acid decarboxylase; ; AAV, adeno-associated virus vector; CHM, choroideremia; CRISPR, clustered regularly interspaced short palindromic repeats; ; CRISPR-Cas, CRISPR-associated protein; DNA, deoxyribonucleic acid; ETDRS, Early Treatment Diabetic Retinopath Study; INCL, late infantile neuronal ceroid lipofuscinosis; MPS, mucopolysaccharidosis; RCT, randomized controlled trial; RECIST, response evaluation criteria in solid tumors; RNA, ribonucleic acid; RNAi, RNA interference; ZFN, zinc finger nuclease.

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Key Question 3a. What are the indication/patient inclusion criteria in ongoing trials? Ocular disorders are the most common indication in the identified studies (32%). Other indications are neurodegenerative disorders (26%),99-103 cancer (16%),91,92,95 muscular conditions (16%),96-98 blood disorders (eg, hemophilia B, 5%),93 and metachromatic leukodystrophy (5%).109 Patient inclusion criteria in ongoing trials differ depending on the indication; for example, the hemophilia B patients had to have had a previous need for factor IX.93 While some of the cancer studies required patients to have failed previous surgery or chemotherapy,110 others required patients to be treatment naïve or did not describe eligibility criteria.92,95 Most of the studies testing treatment for muscular and neurodegenerative conditions, ocular disorders, and metachromatic leukodystrophy described no additional patient inclusion criteria, except for a few that required that patients had showed no improvement despite previous therapy.99,103,108 Patient populations are mostly adult gene therapy candidates (63%)91-93,95,96,99,102-107; however, 21% of the studies are testing interventions specifically in children97,100,101,109 and 16% are testing interventions in both children and adults.73,98,108

Key Question 3b. What are the types of interventions in ongoing trials? The 19 ongoing trials are primarily testing AAV gene therapy approaches (n = 17, 89%).73,93,95-109 However, we also identified an ongoing adenovirus trial (n = 1, 5%)92 and a CRISPR trial (n = 1, 5%).91 The Current Evidence Supporting Gene Therapy section of this report describes a large number of published adenovirus studies. The literature on AAV gene therapy is growing, as documented in the next section. CRISPR specifically has received a lot of attention because of its ability to correct any region of DNA. Expectations are high that this new approach will be approved as a treatment for multiple diseases, including cancer and blood disorders.111 Trials are under way in the United States,112 but it is difficult to estimate when they will report results. We did identify one ongoing CRISPR trial in China91; the trial is registered in the clinicaltrials.gov database and is no longer recruiting. Given the lack of published findings, it is difficult to know how these trials might lead to FDA approval of CRISPR in the near future. In Table 3 we identified 7 distinct CRISPR therapies in the grey literature that are currently being discussed. The trial testing P-1 and TCR gene-knocked mesothelin-directed CAR-T cells is expected to be complete soon (current estimate is June 2019). The others appear to be further from approval, with completion years ranging from 2022 to 2038. However, even though the trial testing CTX001 to treat beta-thalassemia and sickle cell disease113 is not expected to conclude until 2022, several key informants named CTX001 as an intervention that may be approved within the next 5 years.

Key Question 3c. What are the study designs/sizes of ongoing trials? The most common study design in ongoing studies is a trial with multiple study arms in which patients are treated with different doses of the same therapy (47%).91,92,97-101,103,104 Single-arm trials accounted for 42%73,93,95,96,102,107-109 and RCTs for 11%.105,106 On average, the sample size was 21.4 (SD= 16) and ranged from 2 to 60 enrolled participants.

Key Question 3d. What are the comparators in ongoing trials? All the trials with multiple study arms compare groups of patients that receive different dosages of the treatment. The single-arm trials and trials with multiple study arms in which patients are treated with

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different doses of the same therapy have no comparators.73,93,95,96,99,102-104,107-109 The RCTs compare the adenoviral or AAV gene therapy with placebo.105,106 We identified no ongoing nonrandomized trials that included an untreated control group.

Key Question 3e. What are the previous/concurrent treatments in ongoing trials? Most trials required no previous treatment for enrollment (78%),73,91,92,95-98,100-102,104-107,109 although some required patients to have exhausted all previous treatment options (22%).93,99,103,108 Most trials require no concurrent treatment (89%)73,92,93,95-101,103-109; however, patients enrolled in the CRISPR trial receive a single dose of cyclophosphamide (chemotherapy conditioning) 3 days before CRISPR- modified cell infusion.91 In an AAV trial to treat Parkinson’s, patients continue their medications and undergo a neurosurgical procedure with real-time MRI monitoring so the therapy is delivered directly into the striatum.102

Key Question 3f. What is the length of follow-up in ongoing trials? Average length of follow-up is 55.6 months (SD = 66 months) and ranges from 1 to 204 months with variation by indication. The one trial of AAV in hemophilia B has 204 months of follow-up. The cancer trials have the next longest follow-up (mean 68 months), and the trials treating ocular and neurodegenerative disorders followed patients for about 50 months. The group of trials for patients with muscular conditions had, on average, 28 months of follow-up.

Key Question 3g. What outcomes are measured in ongoing trials? Table 4 shows the planned outcomes. Health-related effectiveness outcomes assessed in ongoing trials are mostly indication specific; for example, bleeding episodes and severity rates are being measured for gene therapy intervention in hemophilia patients.93 In studies testing interventions to treat cancer, tumor progression and overall survival are being measured in multiple trials.91,92,95 Change in symptoms (eg, 6-minute walk test, best-corrected visual acuity) are being measured in the muscular condition, neurodegenerative disorder, and ocular disorder trials.73,96-109 Across all trials, safety and toxicity are commonly measured outcomes.

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Current Evidence Supporting Gene Therapy Two tables (Tables A1 and A2) in the Appendix F document in detail each of the 251 completed trials that tested either adenovirus or AAV gene therapies. Documented trials were completed and published, partially published in interim analyses, or had their results documented in a trial registry without scientific publication to date. The evidence tables show study details, study design and comparator, sample size, indication, length of follow-up, indication, previous and concurrent treatment, outcomes measured, reported harms, and the authors’ conclusions.

Characteristics of Published Trials In Table 5 we summarize the evidence according to indication and study design for the adenoviral and AAV interventions. All trials in the table have been completed and have reported at least partial results. More published research studies tested adenoviral therapies (74%) than AAV (26%). We did not identify any completed CRISPR studies.

Table 5. Summary of Published Adenovirus and AAV Studies Indication Study Design by Indication Blood disorders (eg, hemophilia) (4%, n = 9)12,114-121 Single-arm trial (56%, n = 5)12,114,116-118 Trial (multiple groups) (44%, n = 4)115,119-121 Cancer (51%, n = 128)7,122-248 Single-arm trial (41%, n = 53)7,127- 129,133,135,136,140,142,147,148,150,156,158,161,164,174,177,181,183- 185,189,190,192,194-198,200,202-204,206,208,210,211,213,216- 220,222,226,228,233,235,241,242,246,247 Trial (multiple groups) (42%, n = 54)122- 126,131,132,134,137,138,141,143-146,151,157,159,160,162,163,165,167-173,175,178- 180,182,186-188,191,193,199,201,205,209,214,223,227,229,231,234,238-240,245,248 Controlled trial (5%, n = 6)155,166,215,221,225,244 RCT (12%, n = 15)130,139,149,152- 154,176,207,212,224,230,232,236,237,243 Cardiovascular disease (10%, n = 24)249-272 Single-arm trial (8%, n = 2)261,262 Trial (multiple groups) (13%, n = 3)249,266,267 Controlled trial (8%, n = 2)250,254 RCT (71%, n = 17)251-253,255-260,263-265,268-272 Immune deficiency (4%, n = 11)273-283 Trial (multiple groups) (18%, n = 2)274,281 RCT (82%, n = 9)273,275-280,282,283 Inflammatory disorders (2%, n = 4)284-287 Single-arm trial (25%, n = 1)287 RCT (75%, n = 3)284-286 Muscular conditions (1%, n = 3)288-290 Single-arm trial (33%, n = 1)289 Trial (multiple groups) (33%, n = 1)290 RCT (33%, n = 1)288 Neurodegenerative disorders (8%, n = 19)11,291-308 Single-arm trial (42%, n = 8)294,296,298-300,303,304,308 Trial (multiple groups) (21%, n = 4)11,295,305,306 Controlled trial (5%, n = 1)292 RCT (32%, n = 6)291,293,297,301,302,307

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Indication Study Design by Indication Ocular disorders (8%, n = 19)74,75,309-325 Single-arm trial (37%, n = 7)75,311,315-319 Trial (multiple groups) (47%, n = 9)310,312-314,320-323,325 Controlled trial (5%, n = 1)324 RCT (11%, n = 2)74,309 Respiratory conditions (5%, n = 13)326-338 Single-arm trial (31%, n = 4)328,329,332,334 Trial (multiple groups) (54%, n = 7)326,327,330,331,333,336,338 RCT (15%, n = 2)335,337 Other (8%, n = 21)339-359 Single-arm trial (14%, n = 3)357-359 Trial (multiple groups) (24%, n = 5)344,349,353-355 Controlled trial (10%, n = 2)340,352 RCT (53%, n = 11)339,341-343,345-348,350,351,356

Abbreviation: RCT, randomized controlled trial. Note: Controlled trial, intervention and untreated control group; single-arm trial, case series; trial (multiple groups), multiple groups but no untreated control group.

Key Question 4a. What are the indication/patient inclusion criteria in published studies? Across both adenoviral and AAV therapies, the most common indication was cancer (51%), followed by cardiovascular disease (10%), neurodegenerative disease (8%), ocular disorders (8%), respiratory conditions (eg, cystic fibrosis) (5%), blood disorders (eg, hemophilia) (4%), immune deficiency (4%), inflammatory disorders (eg, arthritis) (2%), and muscular conditions (1%). Twenty-one studies (8%) tested interventions for other conditions such as vaccination studies (eg, Ebola, malaria). Others were trials in healthy volunteers, chronic wound healing in diabetic patients, ornithine transcarbamylase deficiency, and venous leg ulcer disease. Most trials had no specific inclusion criteria regarding previous treatment (74%); however, among the studies that did, patients had to have severe disease or have exhausted all treatment options. For example, in one study, patients had to have received a liver transplant to be eligible. Of the studies, 89% were in adult patient populations. The remaining studies tested interventions in children (5%) or in both children and adults (6%).

Key Question 4b. What are the types of interventions in published studies? The identified trials tested a large variety of adenovirus (n = 186, 74%) and AAV (n = 65, 26%) gene therapy approaches. The evidence tables in Appendix F provide details for all included studies.

Key Question 4c. What are the study designs/sizes of published studies? The 2 most common study designs in completed gene therapy trials were trials with multiple groups of patients who were treated with different levels of the same therapy (36%) and single-arm trials (33%). RCTs made up 26% of the completed studies. Only 5% were controlled trials; that is, studies that compared an untreated control group in a nonrandomized treatment allocation design. On average, the sample size was 43 (SD =78), but sample sizes varied widely across indications and trials. The number of enrolled patients in the published gene therapy trials ranged from 3 to 800; a substantial number of the trials had fewer than 10 participants. The trials with large sample sizes were either vaccination trials in healthy volunteers341,343,346-348 or trials with large patient populations, such as cancer or cardiovascular disease.

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Key Question 4d. What are the comparators in published studies? Studies with no comparator were the most common in our sample (67%) given the large number of single-arm trials and trials comparing different doses of the same intervention. Other published trials compared against placebo (19%), usual care (8%), or no treatment (3%).

Key Question 4e. What are the previous/concurrent treatments in published studies? Most studies reported no other treatment concurrently administered with the gene therapy (74%). When concurrent treatments were described they differed by indication; for example, concurrent chemotherapy was required for several cancer gene therapies.135,146,149,174,180,203,204,216,218,237,360,361 Enzyme replacement therapy was concurrently administered for a few muscular condition gene therapies,290,362 while prednisolone11,291,292,306 and antibiotics308 were concurrently administered for neurodegenerative and ocular disorder treatments.

Key Question 4f. What is the length of follow-up in published studies? The length of follow-up averaged 18.1 months (SD =23 months) and ranged from less than 4 weeks to 180 months. The length of follow-up varied widely across trials and by indication. Published ocular disorder trials reported the longest follow-up, with a mean of 28.7 months, whereas the trials treating cancer and neurodegenerative disorders followed patients on average 20 months. The group of trials for patients with cardiovascular disease, immune deficiency, inflammatory disorders, respiratory conditions, muscular conditions, and otherwise classified conditions averaged 16, 9, 4, 8, 14, and 8 months of follow-up, respectively.

Key Question 4g. What outcomes are measured in published studies? The evidence tables in Appendix F document all outcomes measured and the authors’ conclusions for adenovirus and AAV gene therapy studies that have been published to date. All identified trials measured safety and toxicity aspects. Health-related outcomes depended on the trial (some studies concentrated on safety alone) as well as the type of indication. While a quantitative summary of results in the form of a meta-analysis was beyond the scope of this report, we describe reported positive clinical benefits versus suggestive or no evidence of benefits based on the study authors’ summaries of their findings. Trials that tested gene therapy in blood disorders (n = 9) frequently measured bleeding rate, immune responses, and quality of life. Most studies (78%) described a positive clinical health benefit (eg, reduction in factor VIII use), while 2 trials (22%) did not summarize effectiveness results. Trials in cancer patients (n = 128) that reported on health outcomes other than safety measured tumor response, overall survival, progression-free survival, immune response, death, and quality of life. Judging from the authors’ conclusions, a minority of studies (27%) reported definitive positive clinical benefits (eg, increased survival). Most studies (34%) indicated only suggestive evidence that warranted future research (eg, larger studies), and 6% of studies did not report on clinical or suggestive evidence that the therapy was effective in treating cancer. Forty-one trials (32%) did not summarize effectiveness results. Cardiovascular-targeted gene therapy trials (n = 24) measured vital signs, death, amputation, exercise capacity, lipid profiles, and quality of life. A minority of these trials (29%) concluded that evidence exists of clear positive clinical benefit (eg, increased survival). Seven studies (29%) indicated only suggestive evidence that warranted future research, and 17% reported no clinical or suggestive

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evidence that the therapy was effective in reducing cardiovascular disease. Six trials (25%) did not did not summarize effectiveness results. Studies that tested gene therapy in immune deficiency (n = 11) collected clinical indicators of immune response; for example, CD4+ and CD8+ T-cell responses were frequently collected in HIV studies.281,282 Only 18% of studies reported definitive positive clinical benefit (eg, antibody response). Results were reported as suggestive, with study authors supporting future work (45%), and 18% of studies reported no clinical or suggestive evidence that the therapy was effective. Two trials (18%) did not address whether the therapy was effective in treating immune deficiency. Published inflammatory disease trials (n = 4) reported outcomes related to tender joints and hormonal and lipid profiles. Half the studies reported positive clinical benefits (50%) and the other half did not summarize effectiveness results. Studies of gene therapy in patients with muscular disorders (n = 3) measured muscle transduction and muscle response in addition to safety parameters. Two studies (67%) indicated only suggestive effects that supported future studies. One study (33%) did not summarize effectiveness results. Among the published gene therapy trials in patients with neurodegenerative disorders (n = 19), the measured outcomes included neurological deficit, motor ratings, timed walking tests, cognition, muscle response, and time to death. A minority of studies (32%) reported positive clinical benefit (eg, improved motor skills). Most studies (42%) concluded that suggestive evidence warrants future research, and 5% reported no clinical or suggestive evidence that the therapy was effective in treating neurological disease. Five studies (26%) did not summarize effectiveness results. Studies testing gene therapies for ocular disorders (n = 19) reported retinal sensitivity, visual acuity, and function outcomes. Almost half of the studies (42%) indicated positive clinical benefit (eg, improved visual acuity). Five studies (26%) reported only suggestive evidence that warranted future research, and 5% reported no clinical or suggestive evidence that the therapy was effective in treating ocular disorders. Five studies (26%) did not summarize effectiveness results. Gene therapies that targeted respiratory disease (n = 13) reported pulmonary function and lung response. None of the identified studies concluded that gene therapy resulted in a distinct positive clinical benefit. Most studies (54%) reported only suggestive evidence that warranted future research. A third of the trials (31%) reported no clinical or suggestive evidence that the therapy was effective against respiratory disease. Two studies (15%) did not summarize effectiveness results. The studies with otherwise categorized indications (n = 21) measured, for example, immune response, seroconversion, parasite response, and wound response. Most of these trials (67%) reported only suggestive evidence that warranted future research. Two trials (9%) reported no clinical or suggestive evidence that the therapy was effective for the indication. Five studies (24%) did not summarize effectiveness results. The authors’ conclusions for all included trials are documented in the evidence tables in Appendix F, along with more information on the effects of the individual interventions.

Key Question 4h. What are the adverse events in published studies? Adverse events varied by indication, as documented in detail in the evidence tables in Appendix F. The following is a summary of the percentage of adenovirus and AAV interventions that reported severe (grade 3 or higher) treatment-related adverse events by indication. Other studies either reported no serious treatment-related adverse events or did not discuss adverse events. Because many of the studies have no untreated comparator, we cannot use these percentages to assign adverse event rates to the treatments.

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In blood disorder trials, 44% of the studies reported serious adverse events (eg, hospital admission for insulin pump infusion placement and myocarditis121). In cancer trials, 39% of the studies reported serious adverse events; for example, some patients experienced neutropenia,138 lymphopenia,214 raised liver enzymes,214 high fever,153 or cytotoxic response.126 In 2% of the studies, adverse events were not discussed. In cardiovascular disease trials, 50% of the studies reported serious adverse events. Across studies, 31 deaths were reported.253,254,270,272 In immune deficiency trials, 18% of the studies reported serious adverse events; for example, some patients experienced increased creatine kinase,284 acute coronary syndrome,284 or lower respiratory tract infection.282 In inflammatory disease trials, 50% of the studies reported serious adverse events; for example, some patients experienced increased septic arthritis286 and one patient died.284 None of the muscular condition trials reported serious adverse events. In neurodegenerative disease trials, 32% of the studies reported serious adverse events; for example, some patients experienced elevated liver enzymes.11 In the ocular disorders trials, 26% of the studies reported serious adverse events; for example, some patients experienced hemorrhaging in the eye.324 In the respiratory condition trials, 23% of the studies reported serious adverse events; for example, some patients experienced toxicity effects.329 In the other group, 24% of the studies reported serious adverse events. Some patients experienced pulmonary embolism352 or neutropenia,353 and one patient died because of systemic inflammatory response syndrome.357 The evidence tables in Appendix F provide a detailed overview and describe the documented adverse effects for all included trials.

Evidence Map of Ongoing and Completed Gene Therapy Trials Figure 2 is a visualization of the currently available evidence on research supporting adenovirus, AAV, and CRISPR gene therapies. In the figure, each of the 270 included trials, reported in 435 publications, is represented as a bubble in the plot, with the bubble size indicating the number of patients in the study. The studies are plotted according to indication category and by study design. (See Appendix E for separate evidence maps for each intervention.) As Figure 2 shows, the largest amount of evidence is for adenoviral- and AAV-based therapies for treating cancer. These cancer studies are primarily single-arm or multiple-group trials, and they vary in sample size. There are some large RCT trials testing adenoviral or AAV gene therapy in cancer but few controlled trials. Relatively fewer studies have been conducted among patients with muscular conditions or blood, inflammatory, neurodegenerative, ocular, or respiratory disorders, and many of these are based on small sample sizes, probably because the diseases are rare. However, evidence appears to be increasing from RCTs that evaluate the use of adenoviral or AAV therapies in cardiovascular disease, immune deficiency, and otherwise specified diseases.

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Figure 2. Gene Therapy Evidence Map: Adenovirus, AAV, and CRISPR

Abbreviations: AAV, adeno-associated virus; Ad, adenovirus; CRISPR, clustered regularly interspaced short palindromic repeats; RCT, randomized controlled trial. Notes: Evidence map is based on searches to November 2018. Single-arm trial, prospective case series; Trial (multiple groups), trial with multiple arms receiving different doses but no untreated control group; Controlled trial, intervention and untreated control group. The bubbles are staggered in their vertical and horizontal placement for greater visibility.

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Important Issues Raised by Gene Therapy

Key Question 5a. What are the implications of the current level of adoption and future diffusion, given the current level of evidence: efficacy/safety, ethical, disparity, resource allocation, and decision making? In this review we identified 270 complete studies and ongoing trials that evaluate gene therapy treatment using adenoviral, AAV, or CRISPR technologies. Among the ongoing trials, AAV technologies are the most common (n = 17, 89%). We identified only one adenoviral trial (5%) and one CRISPR-based (5%) trial. Among the 251 complete studies, the evidence base differs substantially according to intervention and indication. The most common intervention was adenoviral, perhaps because this vector was one of the first tested in gene therapy. AAV studies currently produce 26% of the evidence from completed studies. While our search identified 7 potential pipeline CRISPR interventions, only one ongoing trial met our inclusion criteria and had completed participant recruitment. This trial has multiple groups but no untreated comparison group and includes 12 metastatic non-small-cell lung cancer adult patients. Results are expected in 2019 (previous estimates indicated December 2018). Most existing gene therapy trials are based on single-arm trials or trials with multiple groups treated with different levels of the same therapy; however, RCTs are increasingly being implemented in cancer, cardiovascular disease, immune deficiency, and otherwise categorized indication trials. Sample sizes remain relatively small across the studies, probably owing to the rarity of the indications addressed by gene therapy. We observed larger sample sizes in the RCTs for the more common diseases, such as cardiovascular conditions, or for vaccine trials. Most studies evaluated safety and indicators of biodistribution, but relatively few studies tested clinical efficacy. This result suggests that, because many of these interventions are still in early development, the primary outcomes are toxicity and maximum tolerable dose. The studies in which AAV or adenoviral gene therapies were used to treat ocular disorders had the largest proportion of study authors concluding positive clinical benefit. Adverse events varied across indication; cardiovascular studies reported the most, including multiple patient deaths. Because many of the studies had no control group, this finding is difficult to assess, especially as the trials were in high-risk patients with critical conditions. Luxturna (Voretigene neparvovec)—currently the only FDA-approved AAV intervention—was approved after only 2 documented trials: an open-label phase I trial and a randomized phase III trial. The sample size of the phase III trial was small (n = 31), but the follow-up was 1 year and the researchers plan to follow patients up to 15 years. Even though the evidence base for adenoviral and AAV gene therapies is relatively large, few studies to date include multiple years of follow-up. Because the durability of treatments that are expected to cure disease remains uncertain, the lack of long-term studies is an obvious and significant gap in this research area. All key informants interviewed for this report mentioned issues they thought were important to patients; they said patients are mostly concerned about the basic components of obtaining and receiving gene therapy, including treatment availability, effectiveness, safety, cost, price, and risks. However, the key informants were concerned about the lack of scientific education and understanding in the public regarding how gene therapies work. They said patients might have to make decisions about care without a thorough understanding of the intervention, including its risks and benefits. A few key informants suggested that, since the true (eg, curative) value of gene therapies is still unknown, other factors should be considered in pricing the therapies; for example, more transparency would be helpful regarding the resources invested in research and development, in addition to the social or individual benefit of patients living longer and healthier lives. Finally, one key informant

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expressed concern about providing insurance for gene therapies in an actuarially sound manner if rare diseases are collectively common but individually rare. This key informant suggested that an insurance-based solution might not exist and that patients might have to pay for the treatment themselves (eg, through a lifelong loan), despite the fact that this option would be politically unfavorable and regressive. (For a discussion of payment options and their implications, see the Institute for Clinical Research and Economics report.63) Ethical Challenges Key informants raised several ethical challenges related to gene therapy. Gene therapy has the potential to alter the germline, which could affect the genome of subsequent generations without their consent. Given the many unknowns about policy implications and unanticipated harmful effects of gene therapy, the American Society of Human Genetics holds the position that performing germline gene editing that culminates in human pregnancy is inappropriate.363 Even with somatic cell gene therapy, ethical issues arise when patients are children and consent is given by parents; for example, parents could seek out gene therapy for nondisease or cosmetics enhancement of their children. While drug pricing presents ethical issues that are not unique to gene therapy, significant ethical concerns for expensive gene therapies may exist. Few groups of patients may be able to afford expensive, lifesaving gene therapies, and the impact of such therapies on payers and insurance premiums could disproportionately burden subpopulations. In addition, some key informants voiced the need for more transparency regarding the balance among pricing, private pharmaceutical company profit, and the public funding of gene therapy research. Key informants were also concerned about the lack of scientific understanding in the general population. Unless we as a society improve science education, patients might not have a strong enough understanding of the potential risks of undertaking certain therapies to give fully informed consent.

Key Question 5b. What are the key issues pertaining to decisional uncertainty? Key informants expressed concerns about the unknown benefits and risks involved in gene therapies. For example, patients might underestimate the risk of a novel gene therapy versus a less efficacious standard therapy because the alternatives are limited. Moreover, patients have to consider the unknown long-term effects of altered genes and the insurance-related and indirect costs (eg, traveling to a gene therapy center, taking leave from work) of receiving new and expensive therapies. Moreover, safety trials treated with the minimum dose may have no clinical benefit, and patients who participate in a safety trial viral-based vector therapy may not be able to participate in a future effectiveness trial because they are immune to the virus. Payers also face uncertainty regarding reimbursement for gene therapy because of the unknowns related to upfront payment, durability, and the impact on later fees. As new therapies are approved, payers will have to decide whether to include them in their plans.

Key Question 5c. What are potential areas of research focus for PCORI and others? To address the gaps in knowledge that might support the approval and use of gene therapies, several areas of research might warrant future work. To help stakeholders understand the effectiveness of treatments versus the risk for adverse outcomes, research could examine in more detail how each intervention type is effective in treating disease, by indication and patient population. Several key informants noted a need for patient registries with long-term follow-up to increase the clinical effectiveness evidence base. Long-term follow-up will be especially important for the CAR-T therapies, in which one treatment is expected to cure. Other research could examine disparities in how

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gene therapy is implemented and the implications for treatment success or adverse events; for example, does gender bias exist in preventing pregnancy during gene therapy treatment? Several key informants suggested the development of Centers of Excellence for gene therapies, where training, manufacturing, and intervention delivery can be closely monitored to provide high standards of care. Research could also examine how Centers of Excellence can ensure that high-quality manufacturing processes and health care delivery standards are followed. Other research could examine how training and education programs of providers and patients might improve provider familiarity with procedures and patient understanding of how the therapy works and the risks of unintended effects. Understanding how people feel about the risks and potential promise of gene therapies and how providers communicate these issues to their patients could shed light on the ethical implications of altering the genome. Qualitative (eg, focus groups) and quantitative (eg, surveys) research could lay the groundwork for a discussion of ethical issues. Key informants suggested that relaxing some of the regulatory processes that determine FDA approval could streamline drug development. For example, research could optimize vector delivery and gene transfer without having to return to the very beginning of the regulatory process. Key informants also suggested relaxing some of the preclinical requirements; for example, in September 2018, NIH and the FDA eliminated the requirement for Recombinant DNA Advisory Committee review and NIH reporting for gene therapy protocols.364 Future research could determine where and how to address regulatory bottlenecks to facilitate drug development and approval, thus accelerating the delivery of high-quality treatments to patients. Future work also could assess how gene therapy trials change over time and whether methodological concerns arise or can be resolved as studies are completed. Given the tremendous uncertainty about how to pay for gene therapies, we recommend future work that explores microsimulation modeling of different payer policy scenarios. Microsimulations would allow researchers to examine the effects of health care system changes on access to various gene therapies across different interventions and patient populations. Also, given the rapid pace at which research is advancing and results are being published, an automated system to curate and track the mounting evidence might be needed to inform key informants and potentially to update model parameters.

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462. Heald AE, Fudman EJ, Anklesaria P, Mease PJ. Single-joint outcome measures: preliminary validation of patient-reported outcomes and physical examination. J Rheumatol. 2010;37(5):1042-1048. PMID:20231202 463. Genethon. Clinical study of AAV1-gamma-sarcoglycan gene therapy for limb girdle muscular dystrophy type 2C. https://ClinicalTrials.gov/show/NCT01344798. Published 2010. Accessed March 1, 2019. 464. Byrne PI, Collins S, Mah CC, et al. Phase I/II trial of diaphragm delivery of recombinant adeno- associated virus acid alpha-glucosidase (rAAaV1-CMV-GAA) gene vector in patients with Pompe disease. Hum Gene Ther Clin Dev. 2014;25(3):134-163. PMID:25238277 465. Corti M, Liberati C, Smith BK, et al. Safety of intradiaphragmatic delivery of adeno-associated virus- mediated alpha-glucosidase (rAAV1-CMV-hGAA) gene therapy in children affected by Pompe disease. Hum Gene Ther Clin Dev. 2017;28(4):208-218. PMID:29160099 466. Nationwide Children's Hospital, Asklepios Biopharmaceutical Inc. Safety study of mini-dystrophin gene to treat Duchenne muscular dystrophy. https://ClinicalTrials.gov/show/NCT00428935. Published 2009. Accessed March 1, 2019. 467. Nationwide Children's Hospital, Parent Project Muscular Dystrophy, The Myositis Association. Follistatin gene transfer to patients with Becker muscular dystrophy and sporadic inclusion body myositis. https://ClinicalTrials.gov/show/NCT01519349. Published 2017. Accessed March 1, 2019. 468. Crystal RG, Sondhi D, Hackett NR, et al. Clinical protocol. Administration of a replication-deficient adeno-associated virus gene transfer vector expressing the human CLN2 cDNA to the brain of children with late infantile neuronal ceroid lipofuscinosis. Hum Gene Ther. 2004;15(11):1131-1154. PMID:15610613 469. Ceregene. Double-blind, multicenter, sham surgery controlled study of CERE-120 in subjects with idiopathic Parkinson's disease. https://ClinicalTrials.gov/show/NCT00400634. Published 2008. Accessed March 1, 2019. 470. Bartus RT, Baumann TL, Siffert J, et al. Safety/feasibility of targeting the substantia nigra with AAV2- neurturin in Parkinson patients. Neurology. 2013;80(18):1698-1701. PMID:23576625 471. Sangamo Therapeutics. Safety and Efficacy of CERE-120 in Subjects With Parkinson's Disease. https://ClinicalTrials.gov/show/NCT00985517. 2018.Accessed March 1, 2019. 472. Marks WJ Jr, Baumann TL, Bartus RT. Long-term safety of patients with Parkinson's disease receiving rAAV2-neurturin (CERE-120) gene transfer. Hum Gene Ther. 2016;27(7):522-527. PMID:26711317 473. Ceregene, Sangamo Therapeutics. CERE-110 in subjects with mild to moderate Alzheimer's disease. https://ClinicalTrials.gov/show/NCT00087789. Published 2010. Accessed March 1, 2019. 474. Sangamo Therapeutics, Alzheimer's Disease Cooperative Study. Randomized, controlled study evaluating CERE-110 in subjects with mild to moderate Alzheimer's disease. https://ClinicalTrials.gov/show/NCT00876863. Published 2019. Accessed March 1, 2019. 475. Neurologix Inc., Weill Medical College of Cornell University, North Shore University Hospital. Safety study of subthalamic nucleus gene therapy for Parkinsonʼs disease. https://ClinicalTrials.gov/show/NCT00195143. 2008. Accessed March 1, 2019. 476. Genzyme a Sanofi Company. A study of AAV-hAADC-2 in subjects with Parkinson's disease. https://ClinicalTrials.gov/show/NCT00229736. Published 2013. Accessed March 1, 2019. 477. Valles F, Fiandaca MS, Eberling JL, et al. Qualitative imaging of adeno-associated virus serotype 2- human aromatic L-amino acid decarboxylase gene therapy in a phase I study for the treatment of Parkinson disease. Neurosurgery. 2010;67(5):1377-1385. PMID:20871425 478. AveXis Inc. Gene transfer clinical trial for spinal muscular atrophy type 1. https://ClinicalTrials.gov/show/NCT02122952. Published 2017. Accessed March 1, 2019. 479. Ceregene, Sangamo Therapeutics. Safety of CERE-120 (AAV2-NTN) in subjects with idiopathic Parkinson's disease. https://ClinicalTrials.gov/show/NCT00252850. Published 2007. Accessed March 1, 2019. 480. Mendell JR, Duchenne Alliance, Milo Therapeutics, Nationwide Children's Hospital. Clinical intramuscular gene transfer of rAAV1.CMV.huFollistatin344 trial to patients with Duchenne muscular dystrophy. https://ClinicalTrials.gov/show/NCT02354781. Published 2017. Accessed March 1, 2019.

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481. Mendell JR, Rodino-Klapac LR, Rosales XQ, et al. Sustained alpha-sarcoglycan gene expression after gene transfer in limb-girdle muscular dystrophy, type 2D. Ann Neurol. 2010;68(5):629-638. PMID:21031578 482. Nationwide Children's Hospital, National Institute of A, Musculoskeletal, Skin D, Muscular Dystrophy A. Gene transfer therapy for treating children and adults with limb girdle muscular dystrophy Type 2D (LGMD2D). https://ClinicalTrials.gov/show/NCT00494195. 2011. Accessed March 1, 2019. 483. Lysogene. Long-term follow-up of Sanfilippo type A patients treated by intracerebral SAF-301 gene therapy. https://ClinicalTrials.gov/show/NCT02053064. 2017. Accessed March 1, 2019. 484. Chen XS, Han GZ, Guo ZP, Lu NZ, Chen J, Wang JB. A comparison of topical application of penciclovir 1% cream with acyclovir 3% cream for treatment of genital herpes: a randomized, double- blind, multicentre trial. Intern J STD AIDS. 2000;11(9):568-573. http://www.embase.com/search/results?subaction=viewrecord&from=export&id=L30711665 485. University of Oxford, Oxford University Hospitals NHST, Moorfields Eye Hospital NHSFT, et al. Gene therapy for blindness caused by choroideremia. https://ClinicalTrials.gov/show/NCT01461213. Published 2017. Accessed March 1, 2019. 486. Xue K, Oldani M, Jolly JK, et al. Correlation of optical coherence tomography and autofluorescence in the outer retina and choroid of patients with choroideremia. Invest Ophthalmol Vis Sci. 2016;57(8):3674-3684. PMID:27403996 487. Simunovic MP, Jolly JK, Xue K, et al. The spectrum of CHM gene mutations in choroideremia and their relationship to clinical phenotype. Invest Ophthalmol Vis Sci. 2016;57(14):6033-6039. PMID:27820636 488. Xue KM, Jolly JK, Barnard AR, et al. Beneficial effects on vision in patients undergoing retinal gene therapy for choroideremia. Nature Med. 2018;24(10):1507 489. Simunovic MP, Xue K, Jolly JK, MacLaren RE. Structural and functional recovery following limited iatrogenic macular detachment for retinal gene therapy. JAMA Ophthalmol. 2017;135(3):234-241. PMID:28152124 490. Simonelli F, Maguire AM, Testa F, et al. Gene therapy for Leber's congenital amaurosis is safe and effective through 1.5 years after vector administration. Mol Ther. 2010;18(3):643-650. PMID:19953081 491. Ashtari M, Cyckowski LL, Monroe JF, et al. The human visual cortex responds to gene therapy- mediated recovery of retinal function. J Clin Invest. 2011;121(6):2160-2168. PMID:21606598 492. Spark T. Safety study in subjects with Leber congenital amaurosis. https://ClinicalTrials.gov/show/NCT00516477. Accessed March 1, 2019. 493. Bennett J, Ashtari M, Wellman J, et al. AAV2 gene therapy readministration in three adults with congenital blindness. Sci Transl Med. 2012;4(120):120. PMID:22323828 494. Spark T. Phase 1 follow-on study of AAV2-hRPE65v2 vector in subjects with Leber congenital amaurosis (LCA) 2. https://ClinicalTrials.gov/show/NCT01208389. Accessed March 1, 2019. 495. Testa F, Maguire AM, Rossi S, et al. Three-year follow-up after unilateral subretinal delivery of adeno- associated virus in patients with Leber congenital amaurosis type 2. Ophthalmology. 2013;120(6):1283- 1291 496. Ashtari M, Nikonova ES, Marshall KA, et al. The role of the human visual cortex in assessment of the long-term durability of retinal gene therapy in follow-on RPE65 clinical trial patients. Ophthalmology. 2017;124(6):873-883. PMID:28237426 497. Byron Lam, University of Miami. Choroideremia gene therapy clinical trial. https://ClinicalTrials.gov/show/NCT02553135. 2018. Accessed March 1, 2019. 498. Genzyme. Safety and tolerability study of AAV2-sFLT01 in patients with neovascular age-related macular degeneration (AMD). https://ClinicalTrials.gov/show/NCT01024998. 2014. Accessed March 1, 2019. 499. Rakoczy EP, Lai CM, Magno AL, et al. Gene therapy with recombinant adeno-associated vectors for neovascular age-related macular degeneration: one-year follow-up of a phase 1 randomised clinical trial. Lancet. 2015;386(10011):2395-2403. PMID:26431823

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500. Constable IJ, Pierce CM, Lai CM, et al. Phase 2a randomized clinical trial: safety and post hoc analysis of subretinal rAAV.sFLT-1 for wet age-related macular degeneration. EBioMedicine. 2016;14:168-175. PMID:27865764 501. Constable IJ, Lai CM, Magno AL, et al. Gene therapy in neovascular age-related macular degeneration: three-year follow-up of a phase 1 randomized dose escalation trial. J Ophthalmol. 2017;177:150-158. PMID:28245970 502. Lions Eye Institute, Perth Western Australia, Adverum biotechnologies Inc. Safety and efficacy study of rAAV.sFlt-1 in patients with exudative age-related macular degeneration. https://ClinicalTrials.gov/show/NCT01494805. 2017. Accessed March 1, 2019. 503. Cideciyan AV, Hauswirth WW, Aleman TS, et al. Human RPE65 gene therapy for Leber congenital amaurosis: persistence of early visual improvements and safety at 1 year. Hum Gene Ther. 2009;20(9):999-1004. PMID:19583479 504. Cideciyan AV, Aguirre GK, Jacobson SG, et al. Pseudo-fovea formation after gene therapy for RPE65- LCA. Invest Ophthalmol Vis Sci. 2014;56(1):526-537. PMID:25537204 505. Ian MM, Alberta Innovates Health Solutions, Canada Foundation for Innovation, et al. An open label clinical trial of retinal gene therapy for choroideremia. https://ClinicalTrials.gov/show/NCT02077361. Published 2017. Accessed March 1, 2019. 506. Bainbridge JW, Smith AJ, Barker SS, et al. Effect of gene therapy on visual function in Leber's congenital amaurosis. N Engl J Med. 2008;358(21):2231-2239. PMID:18441371 507. Ripamonti C, Henning GB, Robbie SJ, et al. Spectral sensitivity measurements reveal partial success in restoring missing rod function with gene therapy. J Vis. 2015;15(15):20. PMID:26605849 508. University College London, Moorfields Eye Hospital NHSFT, Targeted Genetics Corporation. Safety study of RPE65 gene therapy to treat Leber congenital amaurosis. https://ClinicalTrials.gov/show/NCT00643747. Published 2014. Accessed March 1, 2019. 509. Applied Genetic Technologies Corp, Oregon Health Science University, University of Massachusetts Worcester. Phase 1/2 safety and efficacy study of AAV-RPE65 vector to treat Leber congenital amaurosis. https://ClinicalTrials.gov/show/NCT00749957. Published 2014. Accessed March 1, 2019. 510. Cideciyan AV, Aleman TS, Boye SL, et al. Human gene therapy for RPE65 isomerase deficiency activates the retinoid cycle of vision but with slow rod kinetics. Proc Natl Acad Sci U S A. 2008;105(39):15112-15117. PMID:18809924 511. Hadassah Medical Organization. Clinical trial of gene therapy for Leber congenital amaurosis caused by RPE65 mutations. https://ClinicalTrials.gov/show/NCT00821340. Published 2016. Accessed March 1, 2019. 512. Yang S, Yang H, Ma SQ, et al. Evaluation of Leber's hereditary optic neuropathy patients prior to a gene therapy clinical trial. Medicine (Baltimore). 2016;95(40):e5110. PMID:27749593 513. Yang S, Ma SQ, Wan X, et al. Long-term outcomes of gene therapy for the treatment of Leber's hereditary optic neuropathy. EBioMedicine. 2016;10:258-268. http://www.embase.com/search/results?subaction=viewrecord&from=export&id=L613384759 514. Spark Therapeutics, Childrenʼs Hospital of Philadelphia, University of Iowa. Safety and efficacy study in subjects with Leber congenital amaurosis. https://ClinicalTrials.gov/show/NCT00999609. Published 2015. Accessed March 1, 2019. 515. Wagner JA, Nepomuceno IB, Shah N, et al. Maxillary sinusitis as a surrogate model for CF gene therapy clinical trials in patients with antrostomies. J Gene Med. 1999;1(1):13-21. PMID:10738581 516. Moss RB, Rodman D, Spencer LT, et al. Repeated adeno-associated virus serotype 2 aerosol-mediated cystic fibrosis transmembrane regulator gene transfer to the lungs of patients with cystic fibrosis: a multicenter, double-blind, placebo-controlled trial. Chest. 2004;125(2):509-521. PMID:14769732 517. Flotte TR, Trapnell BC, Humphries M, et al. Phase 2 clinical trial of a recombinant adeno-associated viral vector expressing alpha1-antitrypsin: interim results. Hum Gene Ther. 2011;22(10):1239-1247. PMID:21609134 518. Brantly ML, Spencer LT, Humphries M, et al. Phase I trial of intramuscular injection of a recombinant adeno-associated virus serotype 2 alphal-antitrypsin (AAT) vector in AAT-deficient adults. Hum Gene Ther. 2006;17(12):1177-1186. PMID:17115945

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519. University of Massachusetts Worcester, National Heart, Lung, and Blood Institute, Alpha Foundation, University of Florida, National Center for Research Resources.Experimental gene transfer procedure to treat alpha 1-antitrypsin (AAT) deficiency. https://ClinicalTrials.gov/show/NCT00430768. 2015. Accessed March 1, 2019. 520. Mueller C, Chulay JD, Trapnell BC, et al. Human Treg responses allow sustained recombinant adeno- associated virus-mediated transgene expression. J Clin Invest. 2013;123(12):5310-5318. PMID:24231351 521. Applied Genetic Technologies C, National Heart L, Blood I. Safety and efficacy study of rAAV1-CB- hAAT for alpha 1-antitrypsin deficiency. https://ClinicalTrials.gov/show/NCT00749957. 2014. Accessed March 1, 2019. 522. National Institute of Diabetes, Digestive, Kidney Diseases, University of Florida. Phase I randomized study of adeno-associated virus-CFTR vector in patients with Cystic Fibrosis. https://ClinicalTrials.gov/show/NCT00004533. Published 2002. Accessed March 1, 2019.

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Appendix

Appendix A: Report Methodology This project aimed to produce a landscape review of multiple data sources. Our landscape review includes an evidence map based on the published studies and ongoing trials that evaluate the application of gene therapy approaches.

Literature Searches We searched multiple data sources to cover critical aspects of current and future gene therapy interventions. Specifically, our goal was to describe the evidence base for FDA-approved gene therapies currently used to treat or cure conditions. In addition, we describe the evidence for pending or ongoing clinical trials, testing applications that may be poised to gain FDA approval as well as the conditions relevant for future applications. The FDA has approved gene therapies based on the results of single-arm trials when a life-threatening condition has no alternative treatment and its severity6 justifies not performing controlled trials. Thus, we included controlled and uncontrolled trials. We then described the important issues for stakeholders (eg, patients) that the genetic technologies/interventions raise. To identify FDA-approved gene therapies and applications, we referred to the FDA website that lists approved products (https://www.fda.gov/NewsEvents/ProductsApprovals/default.htm). We first reviewed the list of “Approved Cellular and Gene Therapy” products available on the FDA website (July 2018). We continued to monitor the FDA site for 2018 novel therapeutic approvals not otherwise captured on the aforementioned page, searching each drug name. For each therapy, we used the FDA approval letters and accompanying documentation to determine indication, orphan designation status, approval date for each approved indication, whether additional resources are required to utilize the therapy, whether repeat procedures are expected, summary information on adverse events, and anything else that might impact uptake of the therapeutic. If the FDA approval letter did not specify which clinical trials resulted in the approval, we searched the therapy name (generic and branded) on ClinicalTrials.gov and abstracted all associated registered clinical trials. To collect evidence about the current use of approved gene therapies in patients, we searched for empirical published literature across clinical trials in the databases PubMed and EMBASE. We limited searches to English-language studies of human subjects and focused on trials of at least 3 months’ duration. The search strategy included a combination of indexing terms (MeSH terms in MEDLINE/PubMed and EMTREE terms in EMBASE), as well as free-text terms. Another key source of literature was the Web of Science. The Web of Science systematically catalogues conference abstracts, a source of the latest research information that is often not yet captured in the traditional scientific output (ie, journal manuscripts indexed in research databases). The literature searches were conducted by an Evidence-based Practice Center librarian experienced in transparent and comprehensive literature searches. Effective search strategies often require thorough knowledge of indexing terms and scientific database functionality. Searches explicitly addressed the available evidence on the applications in treating human disease and the current and anticipated applications. The review identified remaining evidence gaps that future research studies should address. To capture new advances that are near approval we searched the grey literature online with a selection of the search terms below and subscribed to the newsletter of Gene Therapy Net (http://www.genetherapynet.com/united-states-of-america.html). We identified ongoing trials in

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clinicaltrials.gov (US research). In addition, we checked a database maintained by the Journal of Gene Medicine (http://www.abedia.com/wiley/search.php) and reviewed information in Gene Therapy Net. We limited our searches to US news releases 12 months prior to November 2018 to identify therapies that may be approved in the near future. Gene Therapy Net also provides information on legislative issues. Many nonacademic government and trade publications discuss promising pipeline technologies and products. For example, Pink Sheet of Pharma Intelligence (https://pink.pharmaintelligence.informa.com) publishes news on gene therapy for pharmaceutical professionals on a wide array of topics, including regulation and research/development/strategies. We also searched Lexis-Nexis Academic (https://advance.lexis.com/) for information on developing technologies. We conferred with content experts and screened pertinent online resources, such as Massachusetts Institute of Technology’s NEW Drug Development ParadIGmS (NEWDIGS) site. New Drug Applications (NDA) represent therapies that are closer to approval than the hundreds of Investigational New Drugs (INDs), so while we explored NDAs we did not include IND in our search for near-approval therapies. The newly released FDA suite of 6 draft guidance documents for industry (https://www.fda.gov/biologicsbloodvaccines/guidancecomplianceregulatoryinformation/guidances/cel lularandgenetherapy/default.htm) also provided information for disease-specific clinical and preclinical considerations for gene therapy development. All relevant literature was imported into an EndNote library. As we reviewed the evidence of current and impending gene therapies, we described the prevalence of the indications. We used statistics from published literature and national estimates drawn from the National Cancer Institute Surveillance, Epidemiology, and End-Results (SEER, https://seer.cancer.gov/data/) registry data.

Literature Review Procedure Search outputs were screened by two independent reviewers to avoid missing relevant citations. All citations deemed potentially relevant by at least one reviewer were obtained as full text. Full text publications were screened against the eligibility criteria. We documented the reasons for exclusion in a citation management database. One reviewer abstracted and appraised publications and content and methodological experts checked summaries.

Inclusion Criteria We applied explicit inclusion and exclusion criteria designated a priori following a “PICOTSS” framework, as follows: • Participants: Participants receiving any gene therapy were eligible for inclusion. Studies may include men and women of all ages but need to include human participants. • Interventions: Eligible interventions included those that either replaced a disease-causing gene with a healthy copy of the gene, inactivated a disease-causing gene that was not functioning properly, or introduced a new or modified gene into the body to help treat a disease. • Comparator: Any comparator or no comparator studies were eligible. • Outcomes: Outcomes of interest included disease-related effectiveness/benefit indicators such as complete response (eg, remission), partial response, disease recurrence, mortality, patient- centered outcomes including psychosocial outcomes such as anxiety and worry, and treatment- associated adverse events/harms (eg, cytokine release syndrome). • Timing: Any treatment duration and follow-up of included studies were eligible. • Setting: Any setting was eligible.

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• Study design: Primary clinical research studies were eligible. Publications either self-identified as experimental or described a planned scientific evaluation of gene therapy. Eligible study designs included randomized controlled trials, clinical trials with experimental assignment without randomization, and 1-arm trials (eg, experimental case series). Observational studies were excluded. Conference abstracts were excluded. • Studies published in English since 1989 were eligible. Adenovirus, AAV, and CRISPR Report • Interventions: Eligible interventions included all studies that used adenovirus, AAV, or CRISPR as part of the evaluated gene therapy approach. The literature flow diagram is shown in Appendix C.

Data Extraction We used a standardized form with explicit and pilot-tested categorization rules to extract the data. For each study, we extracted the author and publication year (if applicable), indication/inclusion criteria, type of intervention, study design and study size, comparator, concurrent and prior treatments, length of follow-up, outcomes measures, authors’ conclusions of the effect, and reported adverse events. We used the following categories for indication: • Immune deficiency • Blood disorders (eg, hemophilia) • Ocular disorder • Neurodegenerative disorders • Cancer • Cardiovascular disease • Respiratory conditions (including cystic fibrosis) • Inflammatory disease (including arthritis) • Muscular conditions • Other We consolidated all reports of the same participants into one study entry. This often included trial registry records, progress reports, subgroup analyses, and the main trial publication.

Evidence Map The evidence map is based on a bubble plot. Each “bubble” represents an identified study. The evidence map documents ongoing (open bubbles) and published (filled bubbles) gene therapy trials. The evidence map shows the presence as well as the absence of evidence.

Evidence Tables We also provide the reader with evidence tables to provide a concise overview. We differentiated ongoing and partially (eg, preliminary results have been published) or fully published studies. We limited our search to ongoing studies that have completed recruiting and are more likely near the FDA approval stage than non-RCTs that are still recruiting patients.

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Appendix B: Search Strategies

Published Literature PubMed Run November 2, 2018 Filters activated: Clinical Trial, Clinical Trial, Phase I, Clinical Trial, Phase II, Clinical Trial, Phase III, Clinical Trial, Phase IV, Controlled Clinical Trial, Randomized Controlled Trial, Publication date from 1989/01/01, Humans, English. (((gene-editing[title/abstract] OR “gene editing”[title/abstract]) AND (therap*[title/abstract] OR treatment*[title/abstract])) OR ("gene therapy"[title/abstract] OR "gene therapies"[title/abstract] OR “gene engineering”[title/abstract] OR “gene transfer”[title/abstract]) OR “RNA targeted therapeutic”[tiab] OR “RNA-targeted therapeutics”[tiab] OR ("Gene Editing/ethics"[Mesh] OR "Gene Editing/instrumentation"[Mesh] OR "Gene Editing/legislation and jurisprudence"[Mesh] OR "Gene Editing/standards"[Mesh] OR "Gene Editing/statistics and numerical data"[Mesh] OR "Gene Editing/trends"[Mesh] OR "Gene Editing/utilization"[Mesh] OR "Genetic Therapy/adverse effects"[Mesh] OR "Genetic Therapy/classification"[Mesh] OR "Genetic Therapy/epidemiology"[Mesh] OR "Genetic Therapy/ethics"[Mesh] OR "Genetic Therapy/instrumentation"[Mesh] OR "Genetic Therapy/legislation and jurisprudence"[Mesh] OR "Genetic Therapy/methods"[Mesh] OR "Genetic Therapy/mortality"[Mesh] OR "Genetic Therapy/nursing"[Mesh] OR "Genetic Therapy/psychology"[Mesh] OR "Genetic Therapy/standards"[Mesh] OR "Genetic Therapy/statistics and numerical data"[Mesh] OR "Genetic Therapy/therapeutic use"[Mesh] OR "Genetic Therapy/therapy"[Mesh] OR "Genetic Therapy/trends"[Mesh] OR "Genetic Therapy/utilization"[Mesh] OR "Gene Transfer Techniques/adverse effects"[Mesh] OR "Gene Transfer Techniques/drug effects"[Mesh] OR "Gene Transfer Techniques/immunology"[Mesh] OR "Gene Transfer Techniques/therapy"[Mesh] OR "Gene Transfer, Horizontal/adverse effects"[Mesh] OR "Gene Transfer, Horizontal/drug effects"[Mesh] OR "Gene Transfer, Horizontal/immunology"[Mesh] OR "Gene Transfer, Horizontal/therapeutic use"[Mesh] )) OR CAR-T[title/abstract] OR “CAR T” [title/abstract] OR ("CAR Modified"[title/abstract] AND T[title/abstract]) OR (“chimeric antigen receptor” [title/abstract] AND “T” [title/abstract]) OR CART- 19[title/abstract] OR CRISPR/Cas[title/abstract] OR “clustered regularly interspaced short palindromic repeats”[tiab] OR “zinc-finger nucleases” [title/abstract] OR “zinc finger nucleases” [title/abstract] OR “zinc finger nuclease” [title/abstract] OR “zinc finger nuclease” [title/abstract] OR ZFN[title/abstract] OR TALEN[title/abstract] OR “TAL effector nucleases”[title/abstract] OR “TAL effector nuclease”[title/abstract] OR “Transcription activator-like effector nucleases”[title/abstract] OR “Transcription activator-like effector nuclease”[title/abstract] OR “Transcription activator like effector nucleases”[title/abstract] OR “Transcription activator like effector nuclease”[title/abstract] OR "voretigene neparvovec-rzyl" OR luxturna[title/abstract] OR “axicabtagene ciloleucel” OR “Axi- Cel”[title/abstract] OR “KTE-C19”[title/abstract] OR KTEC19[title/abstract] OR yescarta[title/abstract] OR tisagenlecleucel[title/abstract] OR CTL019[title/abstract] OR CTL- 019[title/abstract] OR kymriah[title/abstract] OR “talimogene laherparepvec”[title/abstract] OR imlygic[title/abstract] OR Provenge[title/abstract] OR “sipuleucel-T” [title/abstract] OR “adeno

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associated virus”[title/abstract] OR “adenoassociated virus”[title/abstract] OR (“adeno-associated virus”[tiab] AND gene*[tiab]) OR (“herpes simplex virus”[tiab] AND gene*[tiab]) OR (HSV[tiab] AND gene*[tiab]) OR tegsedi[tiab] OR inotersen[tiab] OR (("oncolytic virus"[tiab] OR “oncolytic viral”[tiab]) AND (gene*[tiab] OR engineered[tiab] OR modified[tiab])) NOT ("trial protocol"[ti] OR "study protocol"[ti]) Results: 940

(Autologous[tiab] AND cell*[tiab] AND (gene[tiab]OR genes[tiab] OR genetic*[tiab] OR RNA[tiab] OR DNA[tiab])) OR lentiglobin OR bb305 OR 'otl 101' OR 'otl 200' OR gsk269274 OR 'eb 101' OR 'fcx 007' OR 'fcx 013' Results: 315

(RNAi[tiab] OR “RNA interference”[tiab]) OR (“ribonucleic acid”[tiab] AND interfer*[tiab]) OR “interfering ribonucleic acid”[tiab] OR siRNA[tiab] OR miRNA[tiab] OR patisran OR onpattro OR ALN-TTr02 OR ALN-TTRsc02 OR “AMG 890” OR ARO-LPA OR ARO-AAT OR ARO-HBV OR Cemdisiran OR ALN-CC5 OR Fitusiran OR ALN-AT3sc OR ALN-AS1 OR Inclisiran OR ALN- PCSsc OR Lumasiran OR ALN-GO1 OR Revusiran OR ALN-TTRsc) AND (gene*[tiab] AND therap*) NOT (marker* OR biomarker*) Results: 24

(antisense[tiab] AND gene*[tiab]) OR mipomersen OR Kynamro OR eteplirsen OR “exondys 51” OR nusinersen OR spinraza OR tegsedi OR inotersen OR ionis-maptrx OR rg6042 OR ionis-httrx OR volanesorsen Results: 89

Embase Run November 2, 2018 Limits: ([controlled clinical trial]/lim OR [randomized controlled trial]/lim) AND [english]/lim AND [humans]/lim AND [embase]/lim AND [1989-2018]/py (gene-editing):ab,ti OR (“gene editing”):ab,ti AND ((therap*):ab,ti OR (treatment*):ab,ti) OR ("gene therapy"):ab,ti OR ("gene therapies"):ab,ti OR (“gene engineering”):ab,ti OR (“gene transfer”):ab,ti OR (“RNA targeted therapeutic”):ab,ti OR (“RNA-targeted therapeutics”):ab,ti OR (CAR-T):ab,ti OR (“CAR T”):ab,ti OR (("CAR Modified"):ab,ti AND (T):ab,ti) OR ((“chimeric antigen receptor”):ab,ti AND (T):ab,ti) OR (CART-19):ab,ti OR (CRISPR):ab,ti OR (“clustered regularly interspaced short palindromic repeats”):ab,ti OR (“zinc-finger nucleases”):ab,ti OR (“zinc-finger nuclease”):ab,ti OR (“zinc finger nucleases”):ab,ti OR (“zinc finger nuclease”):ab,ti OR (ZFN):ab,ti OR (TALEN):ab,ti OR (“TAL effector nucleases”):ab,ti OR (“TAL effector nuclease”):ab,ti OR (“transcription activator-like effector nucleases”):ab,ti OR (“transcription activator-like effector nuclease”):ab,ti OR (“transcription activator like effector nucleases”):ab,ti OR (“transcription activator like effector nuclease”):ab,ti OR (“voretigene neparvovec-rzyl”):ab,ti OR (luxturna):ab,ti OR (“axicabtagene ciloleucel”):ab,ti OR

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(“Axi-Cel”):ab,ti OR (“KTE-C19”):ab,ti OR (KTEC19):ab,ti OR (yescarta):ab,ti OR (tisagenlecleucel):ab,ti OR (CTL019):ab,ti OR (CTL-019):ab,ti OR (kymriah):ab,ti OR (“talimogene laherparepvec”):ab,ti OR (imlygic):ab,ti OR (Provenge):ab,ti OR (“sipuleucel-T”):ab,ti OR (“adeno associated virus”):ab,ti OR (“adenoassociated virus”):ab,ti OR ((“adeno-associated virus”):ab,ti AND (gene*):ab,ti) OR ((“herpes simplex virus”):ab,ti AND (gene*):ab,ti) OR ((HSV):ab,ti AND (gene*):ab,ti) OR (tegsedi):ab,ti OR (inotersen):ab,ti OR (("oncolytic virus”):ab,ti OR (“oncolytic viral”):ab,ti AND ((gene*):ab,ti OR (engineered):ab,ti OR (modified):ab,ti)) NOT (“trial protocol”):ti OR (“Study protocol):ti EXCLUDE: CONFERENCE ABSTRACT; CONFERENCE REVIEW; EDITORIAL; NOTE; REVIEW; SHORT SURVEY; LETTER; CONFERENCE PAPER Results: 515 – duplicates = 251

(autologous:ab,ti AND cell*:ab,ti AND (gene:ab,ti OR genes:ab,ti OR genetic*:ab,ti OR rna:ab,ti OR dna:ab,ti) OR lentiglobin OR bb305 OR 'otl 101' OR 'otl 200' OR gsk269274 OR 'eb 101' OR 'fcx 007' OR 'fcx 013') Results: 177 - – duplicates with PubMed above = 87

((RNAi):ab,ti OR (“RNA interference”):ab,ti OR (“ribonucleic acid” AND interfer*):ab,ti OR (“interfering ribonucleic acid” OR siRNA OR miRNA):ab,ti OR patisran OR onpattro OR ALN-TTr02 OR ALN-TTRsc02 OR “AMG 890” OR ARO-LPA OR ARO-AAT OR ARO-HBV OR Cemdisiran OR ALN-CC5 OR Fitusiran OR ALN-AT3sc OR ALN-AS1 OR Inclisiran OR ALN-PCSsc OR Lumasiran OR ALN-GO1 OR Revusiran OR ALN-TTRsc AND (gene*):ab,ti AND (therap*):ab,ti NOT (marker* OR biomarker*) Results: 35 – duplicates with PubMed above = 29

((antisense):ab,ti AND (gene*):ab,ti) OR mipomersen OR Kynamro OR eteplirsen OR “exondys 51” OR nusinersen OR spinraza OR tegsedi OR inotersen OR ionis-maptrx OR rg6042 OR ionis-httrx OR volanesorsen) Results: 266 – duplicates with PubMed above = 223

Web of Science Run November 2, 2018 English: Article Indexes=SCI-EXPANDED, SSCI, A&HCI, ESCI Timespan=1989-2018 (TS=(gene-editing) OR TS=(“gene editing”)) AND (TS=(therap*) OR TS=(treatment*)) OR TS=("gene therapy") OR TS=("gene therapies") OR TS=(“gene engineering”) OR TS=(“gene transfer”) OR TS=(“RNA targeted therapeutic”) OR TS=(“RNA-targeted therapeutics”) OR TS=(CAR-T) OR TS=(“CAR T”) OR (TS=("CAR Modified") AND TS=(T)) OR (TS=(“chimeric antigen receptor”) AND TS=(T)) OR TS=(CART-19) OR TS=(CRISPR/Cas) OR TS=(“clustered regularly interspaced short palindromic repeats”) OR TS=(“zinc-finger nucleases”) OR TS=(“zinc-finger nuclease”) OR TS=(ZFN) OR TS=(TALEN) OR TS=(“zinc finger nuclease”) OR TS=(“zinc finger nucleases”) OR TS=(“TAL effector nucleases”) OR TS=(“TAL effector nuclease”)

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OR TS=(“transcription activator-like effector nucleases”) OR TS=(“transcription activator-like effector nuclease”) OR TS=(“transcription activator like nucleases”) OR TS=(“transcription activator like nuclease”) OR (TS=(“voretigene neparvovec-rzyl”) OR TS=(luxturna) OR TS=(“axicabtagene ciloleucel”) OR TS=(“Axi-Cel”) OR TS=(“KTE-C19”) OR TS=(KTEC19) OR TS=(yescarta) OR TS=(tisagenlecleucel) OR TS=(CTL019) OR TS=(CTL-019) OR TS=(kymriah) OR TS=(“talimogene laherparepvec”) OR TS=(imlygic) OR TS=(Provenge) OR TS=(“sipuleucel-T”) OR TS=(“adeno associated virus”) OR TS=(“adenoassociated virus”) OR (TS=(“adeno-associated virus”) AND TS=(gene*)) OR (TS=(“herpes simplex virus”) AND TS=(gene*)) OR (TS=(HSV) AND TS=(gene*)) OR TS=(tegsedi) OR TS=(inotersen) OR (TS=("oncolytic virus" OR “oncolytic viral”) AND TS=(gene* OR engineered OR modified))) AND TS=(“randomized controlled trial”) OR TS=(RCT) OR TS=(“controlled clinical trial”) OR TS=(“clinical trial”) NOT TS=(rat OR rats OR monkey OR monkeys OR primates OR primate OR macaque* OR mouse OR mice OR dog OR canine OR canines OR rabbit OR rabbits) NOT TI=(“trial protocol”) OR TI=(“study protocol”) EXCLUDE: Proceedings Papers; Book Chapters also, in EndNote (all results), pulled out pig/horse articles and some additional “overviews” not called “Reviews” Results: TOTAL: 898 – duplicates = 672 TOTAL: 1863

(TS=(autologous) AND TS=(cell) AND (TS=(gene OR genes OR genetic* OR rna OR dna)) OR TS=(Strimvelis OR Lenti-D OR “elivaldogene tavalentivec” OR Lentiglobin OR BB305 OR OTL-101 OR OTL-200 OR GSK269274 OR EB-101 OR FCX-007 OR FCX-013) AND TS=(“randomized controlled trial”) OR TS=(RCT) OR TS=(“controlled clinical trial”) OR TS=(“clinical trial”) NOT TS=(rat OR rats OR monkey OR monkeys OR primates OR primate OR macaque* OR mouse OR mice OR dog OR canine OR canines OR rabbit OR rabbits) NOT TI=(“trial protocol”) OR TI=(“study protocol”) EXCLUDE: Proceedings Papers; Results: 199 – duplicates with above = 134

(TS=(RNAi) OR TS=(“RNA interference”) OR TS=(“ribonucleic acid” AND interfer*) OR TS=(“interfering ribonucleic acid”) OR TS=(siRNA OR miRNA OR patisran OR onpattro OR ALN- TTr02 OR ALN-TTRsc02 OR “AMG 890” OR ARO-LPA OR ARO-AAT OR ARO-HBV OR Cemdisiran OR ALN-CC5 OR Fitusiran OR ALN-AT3sc OR ALN-AS1 OR Inclisiran OR ALN- PCSsc OR Lumasiran OR ALN-GO1 OR Revusiran OR ALN-TTRsc)) AND (TS=(gene) AND TS=(therap*))

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NOT (TS=(marker* OR biomarker*) AND TS=(“randomized controlled trial”) OR TS=(RCT) OR TS=(“controlled clinical trial”) OR TS=(“clinical trial”) NOT TS=(rat OR rats OR monkey OR monkeys OR primates OR primate OR macaque* OR mouse OR mice OR dog OR canine OR canines OR rabbit OR rabbits) NOT TI=(“trial protocol”) OR TI=(“study protocol”) EXCLUDE: Proceedings Papers; Results: 38 – duplicates with above =36

(TS=(antisense) AND TS=(gene*)) OR TS=(mipomersen OR Kynamro OR eteplirsen OR “exondys 51” OR nusinersen OR spinraza OR tegsedi OR inotersen OR ionis-maptrx OR rg6042 OR ionis-httrx OR volanesorsen) AND TS=(“randomized controlled trial”) OR TS=(RCT) OR TS=(“controlled clinical trial”) OR TS=(“clinical trial”) NOT TS=(rat OR rats OR monkey OR monkeys OR primates OR primate OR macaque* OR mouse OR mice OR dog OR canine OR canines OR rabbit OR rabbits) NOT TI=(“trial protocol”) OR TI=(“study protocol”) EXCLUDE: Proceedings Papers; in EndNote (all results), pulled out pig/horse articles and some additional “overviews” not called “Reviews” Results: 79 – duplicates with above = 61

Grey Literature ClinicalTrials.gov Run November 2, 2018 Searching the “other” field—limiting to active, not recruiting, completed, suspended, terminated, withdrawn studies Modalities "Zinc finger nuclease" OR "Zinc-finger nuclease" OR "Zinc-finger nucleases" OR "Zinc finger nucleases" OR ZFN OR "Transcription Activator-like Effector Nucleases" OR "Transcription Activator-like Effector Nuclease" OR “TAL effector nuclease” OR “TAL effector nucleases” OR TALEN OR CRISPR OR CRISPR/Cas9 OR "Clustered Regularly Interspaced Short Palindromic Repeats" OR CAR-T OR "chimeric antigen receptor" OR (Adenovirus AND "gene therapy") OR (Adenovirus AND "gene transfer") OR Oncolytic Virus OR Luxturna OR "Voretigene neparvovec" OR "Axicabtagene ciloleucel" OR KTE-C19 OR Yescarta OR Tisagenlecleuce OR Kymriah OR "talimogene laherparepvec" OR Imlygic OR Tegsedi OR Inotersen Results: 360 Other Biological Approved Products ("Recombinant protein" AND "gene therapy") OR ("Recombinant protein" AND "gene transfer") OR "Allogenic cord blood hematopoietic progenitor cell therapy" OR "Allogenic umbilical cord blood"

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OR "autologous cellular product" OR "autologous cellularized scaffold product" OR "autologous cellularised scaffold product" OR Allocord OR Azficel OR laviv OR Andexxa OR Clevecord OR Gintuit OR Hemacord OR Ducord OR ("gene therapy" AND RNA) OR ("gene transfer" AND RNA) OR patisiran OR (lentivirus AND "gene therapy") OR (Lentivirus AND "gene transfer") OR (vector AND "gene therapy") OR (vector and "gene transfer") OR “adeno associated virus” OR “adenoassociated virus” OR (“adeno-associated virus” AND gene*) OR (“herpes simplex virus” AND gene*) OR (HSV AND gene*) OR (“oncolytic viral”) Results: 251 – duplicates with modalities =177 TOTAL: 537

((autologous AND cell*) AND (gene OR genes OR genetic* OR rna OR dna)) OR Strimvelis OR Lenti-D OR “elivaldogene tavalentivec” OR Lentiglobin OR BB305 OR OTL-101 OR OTL-200 OR GSK269274 OR EB-101 OR FCX-007 OR FCX-013 Results: 4

RNAi OR “RNA interference” OR (“riboneucleic acid” AND interfer*) OR “interfering ribonucleic acid” OR siRNA OR miRNA OR patisran OR onpattro OR ALN-TTr02 OR ALN-TTRsc02 OR “AMG 890” OR ARO-LPA OR ARO-AAT OR ARO-HBV OR Cemdisiran OR ALN-CC5 OR Fitusiran OR ALN-AT3sc OR ALN-AS1 OR Inclisiran OR ALN-PCSsc OR Lumasiran OR ALN- GO1 OR Revusiran OR ALN-TTRsc AND gene AND therap* NOT marker* OR biomarker* Results: 0

(antisense AND gene*) = 0 (mipomersen OR Kynamro OR eteplirsen OR “exondys 51” OR nusinersen OR spinraza OR tegsedi OR inotersen OR ionis-maptrx OR rg6042 OR ionis-httrx OR volanesorsen) = 45 Results: 45

FDA Records We used the list of “Approved Cellular and Gene Therapy” products available on the FDA website (accessed July 2018) and monitored the FDA site for 2018 novel therapeutic approvals.

LexisNexis and Web Searches We used the search terms “gene therapy” and “nda” in combination with a source restriction to “Pink Sheet: Pharma Intelligence” and date restrictions of (01/01/2017 and 08/30/2018) or (01/01/2017 and 09/15/2018). We abstracted names of any gene therapies that were mentioned. Next, we searched the name of each of the gene therapies for specific information in ClinicalTrials.gov as well as in the Google search engine. In the Google search engine, we used the name of each gene therapy alone and in conjunction with “FDA designation” to determine the modality, indication, determine/confirm the current phase of the therapy, and determine if it had received any specific FDA designation (fast track, breakthrough, orphan drug, rare pediatric disease, or regenerative medicine advanced therapy).

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Gene Therapy Clinical Trials Worldwide We used a database maintained by the Journal of Gene Medicine (http://www.abedia.com/wiley/search.php) to check for less-frequent therapies to ensure all relevant trials had been identified.

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Appendix C: Literature Flow Diagram

Figure notes: AD adenovirus; AAV, adeno-associated virus therapy; CAR-T, chimeric antigen receptor; CRISPR, Clustered Regularly Interspaced Short Palindromic Repeats; RNAi, ribonucleic acid interference; ZFN, zinc finger nucleases

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Appendix D: Stakeholder and Key Informant Interview Information

Interviews We conducted key informant interviews to gather detailed information about gene therapy from a variety of perspectives. The purpose of the interviews was to collect perspectives from patients or patient advocates, clinicians, payers/insurers, public policy-maker representatives, and industry. The goal was to ensure that perspectives of different stakeholders had been considered and were reflected in the research approach. Our application to RAND’s Human Subjects Protection Committee (HSPC: 2018-0560) was approved for exempt status on August 24, 2018.

Recruitment We developed the key informant list by first selecting relevant stakeholder groups. In a second step, the team identified possible representatives of the stakeholder groups. If we could not reach the key informant or he or she declined, we referred to our list to identify an alternate key informant. We also asked anyone who declined if he or she could recommend someone else. Emails were sent to individuals and organizations requesting their participation in phone interviews. These emails included a 1-page brief description of the assessment, written in plain English. We informed the individual that we contacted him or her because of his or her experience (as a patient or advocacy group member, clinician, payer or insurer, public policy representative, or industry member or industry analyst, depending on the interviewee). At this time, we asked if he or she would agree to participate in our assessment and if there were any other experts he or she suggested we contact for his or her relevant experience. We contacted 45 individuals: 15 clinicians, 7 patient and patient advocates, 8 payers/insurers, 5 public policymaker representatives, and 10 industry/industry analysts. Among those we contacted, 2 clinicians, 2 patient advocates, 2 payers/insurers, 2 public policymaker representatives, and 2 industry analysts participated. We also invited industry professionals, although none agreed to participate.

Procedure Semi-structured telephone interviews lasting approximately 1 hour were conducted with each key informant. The interviews were guided by semi-structured interview protocols following a list of set questions but allowing the interviewee to expand or deviate to other topics that he or she thought were important to discuss. We aimed to understand the process by which potential gene therapies progress from the laboratory bench to patient treatment options and to identify positive and negative impacts of the therapies for many stakeholders, including patients/consumers, payers/insurers, purchasers/employers, and public policymakers. The interviewer asked about barriers and facilitators to gene therapy implementation, adjustments to policies and procedures, safety, efficacy, and cost. The interview questions were designed for a wide set of stakeholder perspectives and not all questions were relevant to each interviewee. Thus, interviewees were free to skip questions he or she felt he or she could not answer.

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Informed Consent Protocol Interviewers read the information below before proceeding with the interview.

Research Description The goal of this study is to use an evidence map format and landscape review to describe the current evidence base available for gene therapy applications. The evidence map will include a central figure that visualizes the state of the research. The evidence maps provide a visual overview, communicating the results of multiple dimensions to best document a research area. As part of this work, we will conduct key stakeholder interviews to gather detailed information about gene therapy from a variety of perspectives. Informative research questions need to consider a range of stakeholders. In particular, patient-centered outcomes should not be selected by clinicians and policymakers but require engaging patients in the research process. The key informant interviews will include a range of respondents from patients or patient advocates, clinicians, payers/insurers, public policy maker representatives, industry, and industry analysts. The goal is to ensure that perspectives of different stakeholders have been considered and are reflected in the research approach. This project is supported by the Patient-Centered Outcomes Research Institute (PCORI) and facilitated by the RAND Corporation. Risks and Benefits There will be no risks related to participation in this study. During the completion of the interview, if a question is not applicable to you or you feel uncomfortable answering, please complete it to the best of your ability. The project aims to improve our understanding of approved gene therapies as well as those that may become available for use in the next five years, as a critical means to spread knowledge that will benefit patients, researchers, and practitioners. Duration Your participation will last about an hour. Participation and Withdrawal Participation in this study is entirely voluntary. You have the right not to participate at all, to leave the project at any time, and can decline to be acknowledged as a panelist. Deciding not to participate or choosing to leave the study will not result in any negative consequences. If you decide that you cannot participate, we will ask that you nominate someone to join the panel in your place (if possible). Confidentiality Your name will not be linked to your interview responses in any feedback of results to the group and your name and your responses will not be stored together. Your interview responses will be kept confidential. Study results will be accessible only to the study team. Interview responses and discussion points will be documented in aggregate form across participants. You will be asked for consent to be acknowledged as a key stakeholder in future publications describing the results of the project. Contact Information If you have questions about this research, please email Dr. Andrea Richardson ([email protected]) or Dr. Susanne Hempel ([email protected]). If you have questions about your rights as a research participant or need to report a research- related injury or concern, you can contact RAND’s Human Subjects Protection Committee toll-free at 866 697 5620 or by emailing [email protected]. If possible, when you contact the Committee, please reference Study # 2017-0640. Consent Your consent to participate in this project will be implied by selecting the option, “Yes, I consent to participate in this study (continue to survey)" and by submitting your responses after completion. Interview Guide Introduction Brief Project Background Reason for Reaching Out/Expertise of Stakeholder Representative Questions to Discuss During Phone Conversations: • Are there interventions missing from our preliminary search strategy? • Which clinical effectiveness outcomes should be assessed when evaluating the effects of gene therapy? • Which adverse events should be assessed when evaluating the effects of gene therapy? • Which other outcomes should be assessed? • What is an appropriate timeframe for the assessment? • What information is important to patients that has not been the focus of expert discussions or has been neglected in existing research? • What are the decisional dilemmas for patients? • What considerations will help key stakeholders (will be specified) to decide the value of the intervention?

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• Our evidence map can display only a limited number of dimensions to establish a picture of the existing evidence base. Using the draft example, which dimensions do you think are critical to display (eg, number of patients, number of successful interventions, study design distribution, replication by independent researchers, clinical effectiveness, severity of adverse events, costs, quality of evidence across studies)? • Which new developments are close to approval? Please do not share any proprietary information that you may know. • What are the major challenges you see facing gene therapy advances? • Are there ethical considerations that should be discussed? • What are your major hopes you see facing gene therapy treatments? • What are the major gaps in our understanding that need to be addressed to improve the development of FDA-approved gene therapies? • Is public policy lacking with respect to gene therapy approval and use? • What suggestions do you have to bring more approved gene therapies from bench to bedside?

Interview Data Collection and Analysis Strategy Interviews were conducted by phone with 1 researcher and 1 note taker and, if the interview participant agreed, recorded to ensure information was accurately captured. Results of the interviews were documented in aggregated and deidentified format. We analyzed the content of the interviews by tabulating responses according to the topic addressed.

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Appendix E: Evidence Maps Stratified by Intervention

Appendix Evidence Map 1. Adenovirus Trials

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Appendix Evidence Map 2. AAV Trials

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Appendix Evidence Map 3. CRISPR

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Appendix F: Evidence Tables Published Trials

Appendix Table A1. Published Adenovirus Trials Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Blood disorders Age group: children Recombinant adeno- Development of NAbs None NAbs against AAV2 Li et al, 2012114 Hemophilia A associated virus (43.5%) were observed Single-arm trial Treatments enrollment: No comparison group more frequently and at higher titers compared N: 62 none with both AAV5 (25.8%) Treatment requisite: and AAV8 (22.6%). NAbs none against AAV5 or AAV8 Follow-up: 48 were rarely observed in the absence of coprevalent and higher- titer AAV2 NAbs, suggesting that NAbs to AAV5 and AAV8 were detected following AAV2 exposure due to partial cross-reactivity of AAV2- directed NAbs. Cancer Age group: adults Ad.hIFN-beta Time to progressive Unlikely or not related: Direct Ad.hIFN-beta Chiocca et al, 2008209 Malignant glioma No comparison group disease (size of lesions or deep vein thrombosis injection into the tumor BB-IND 9924 Treatments enrollment: appearance of new (grade 3, n = 2), and the surrounding lesions), progression-free depressed level of normal brain areas after Trial (multiple groups) Tumors had to be either survival, overall survival consciousness (grade 3, surgical removal was N: 11 recurrent or progressive following prior treatment. n = 2), thrombosis feasible and associated (grade 3, n = 1), with apoptosis induction. Treatment requisite: pulmonary embolism surgical removal of tumor (grade 4, n = 1), 4 to 8 days plus convulsions NOS (grade additional dose of gene 3, n = 1), choroidal therapy after initial dose infarction (grade 2, n = Follow-up: 2 to 11 1); likely related: (ongoing as of paper) confusional state (grade 3, n = 1, resolved; grade 4, n = 1, not resolved)

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad.HSVtk Tumor response, overall No grade 3 or greater The results suggest that Sterman et al, 2000168 Malignant pleural No comparison group survival toxicities reported systemic administration Sterman et al, 1998365; mesothelioma of steroids in the context Molnar-Kimber et al, Treatments enrollment: of adenoviral-based gene 1998366; Albelda, none delivery may limit acute 367 clinical toxicity but may 1997 ; Benard et al, Treatment requisite: 368 not inhibit biological 1998 placement of a responses to adenoviral ID NR thoracostomy tube for vectors. Trial (multiple groups) delivery of gene therapy; N: 10 ganciclovir for 2 weeks for all patients; corticosteroid by intravenous administration (n = 5, started ~10 hours before gene therapy) Follow-up: until death Cancer Age group: adults Ad.IFNalpha Safety, toxicity, and Pleural catheter infection The combination of Sterman et al, 2016245 Unresectable malignant No comparison group objective response rate, (n = 2), hypoxia (n = 2), intrapleural Ad.IFN, NCT01119664 pleural mesothelioma progression-free and supraventricular celecoxib, and overall survival; tachycardia (n = 1), chemotherapy proved Trial (multiple groups) Treatments enrollment: chemotherapy biocorrelates on blood esophagitis (n = 1) safe in patients with N: 40 and tumor were MPM. Treatment requisite: measured none Follow-up: 6 Cancer Age group: adults Ad.IFN-alpha2b Antitumor immunologic None These data show Ad.IFN- Sterman et al, 2011226 Malignant pleural No comparison group responses, antiviral alpha2b has potential NCT01212367 mesothelioma immune responses, gene therapeutic benefit in transfer assessment MPM and it generates Single-arm trial Treatments enrollment: none antitumor immune N: 9 responses that may Treatment requisite: induce anatomic and/or none metabolic reductions in Follow-up: none distant tumors.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad.IFN-beta Innate immune Transient hypoxia (n = Intrapleural instillation of Sterman et al, 2007206 Malignant pleural No comparison group responses, humoral 1) elevations in serum Ad.IFN-beta is a Sterman et al, 2006369 mesothelioma (MPM), responses to known MPM transaminases (n = 1) potentially useful tumor antigens, cellular approach for the BB-IND 10603 metastatic pleural effusions (MPE) immune responses generation of antitumor Single-arm trial Treatments enrollment: immune responses in N: 10 none MPM and MPE patients and should be Treatment requisite: investigated further for none overall clinical efficacy. Follow-up: 6 Cancer Age group: adults Ad.IFN-beta Safety and toxicity, gene None This approach was safe Sterman et al, 2010218 Malignant pleural No comparison group transfer assessment, and induced immune ID NR, protocol reviewed mesothelioma, malignant antiviral immune responses and disease by NIH-RAC and FDA pleural effusions responses stability. However, rapid development of Nabs Single-arm trial Treatments enrollment: none prevented effective gene N: 17 transfer after the second Treatment requisite: dose, even with a dose chemotherapy interval as short as 7 Follow-up: 42 days. Cancer Age group: adults Ad.IL-12 Toxicity, biologic None Intratumoral injection of Sangro et al, 2004189 Advanced digestive No comparison group response, antitumor up to 3 1012 viral ID NR, protocol approved tumors activity particles of Ad.IL-12 to by the Spanish Treatments enrollment: patients with advanced government’s Ethical none digestive malignancies is a feasible and well- Committee for Clinical Treatment requisite: tolerated procedure that Investigation, the surgery, chemotherapy, exerts only mild National Biosafety radiotherapy, antitumor effects. Commission, and the transarterial Spanish Agency for the chemoembolization Evaluation of Medicinal Follow-up: 12 Products Single-arm trial N: 21

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad.RSVtk Survival, ongoing 1 moderate neutropenia Given these caveats, with Sterman et al, 1998143 Pleural mesothelioma No comparison group disease, stable disease, event a median follow-up of ID NR Treatments enrollment: tumor progression approximately 12 months, 12 of 21 Trial (multiple groups) none patients have died, with Treatment requisite: Completed no fatal complications none N: 21 attributable to the gene Follow-up: 24 therapy protocol. Autopsies were performed on 9 patients, uniformly demonstrating progressive mesothelioma, often with widespread metastatic disease as the cause of death. Although no definite tumor regressions were noted, 3 of the initial 18 patients remain clinically stable, with no evidence of tumor growth on serial chest radiographs and chest CT scans. Cancer Age group: adults Ad.TK Tumor response and No complications The US-guided Sangro et al, 2010222 Advanced hepatocellular No comparison group toxicity, imaging administration of Ad.TK Penuelas et al, 2005370; carcinoma was feasible in 100% of Clinica Universidad de Treatments enrollment: cases and no Navarra, 2003371 none complications associated with the procedure were NCT00844623 Treatment requisite: reported. Single-arm trial none N: 10 Follow-up: 26 Cancer Age group: adults Ad/PNP Tumor size None This first-in-human Rosenthal et al, 2015242 Advanced solid tumors No comparison group clinical trial found NCT01310179 Treatments enrollment: localized generation of fluoroadenine within Single-arm trial none tumor tissues using E. Treatment requisite: N: 12 coli PNP and fludarabine none is safe and effective. Follow-up: none

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults AD21 Percentage with stable No vector-related The study suggests this a Alvarez et al, 2000163 Recurrent erbB-2- No comparison group disease, percentage with hematological, feasible approach, but Hemminki et al, 2002372 overexpressing ovarian progressive disease coagulation, or needs more research: electrolyte abnormalities “This study suggests that ID NR cancer but 4 grade 3/4 events: adenoviral-mediated Trial (multiple groups) Treatments enrollment: none dyspnea (n = 1), gene therapy using an N: 15 circulatory (n = 1), anti-erbB-2-directed Treatment requisite: Tenckhoff catheter intrabody is feasible in none infections (n = 2) the context of human Follow-up: until death ovarian cancer." Cancer Age group: adults Ad5 [E1-, E2b-]-CEA(6D) Survival None observed An overall survival of Balint et al, 2015150 Late-stage colorectal vaccine 20% (median survival 11 Morse et al, 2013373 cancer No comparison group months) was observed during long-term follow- NCT01147965, IND14325 Treatments enrollment: none up and no long-term Single-arm trial adverse effects were Treatment requisite: Completed reported. none N: 32 Follow-up: 29 Cancer Age group: adults Ad5.SSTR/TK.RGD Toxicity associated with None This study shows the Kim et al, 2012228 Recurrent gynecologic No comparison group intraperitoneal safety, potential efficacy, ID NR, protcol reviewed cancer (epithelial administration of and possible gene by NICH-RAC and FDA ovarian, primary Ad5.SSTR/TK.RGD, transfer imaging capacity Clinical efficacy of Ad5.SSTR/TK.RGD in Single-arm trial peritoneal, fallopian tube, or endometrial) patients with recurrent N: 12 Treatments enrollment: gynecologic cancer. conventional surgical procedures and standard adjuvant therapies Treatment requisite: none Follow-up: none

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: children Ad5/3-D24-GMCSF Tumor size/response No grade 4 or 5 events, Ad5/3-D24-GMCSF Koski et al, 2010221 Refractory advanced No comparison group (complete response, but grade 3 events seems safe in treating Bramante et al, 201483 solid tumors partial response, stable reported: anemia (n = cancer patients, and disease, progressive 1), neutropenia (n = 1), promising signs of ID NR Treatments enrollment: none disease) cholecystitis (n = 1), efficacy were observed. Controlled trial increased alanine Treatment requisite: N: 21 aminotransferase (ALT, n ultrasound to guide = 1), increased aspartate intratumoral injection aminotransferase (AST, n (single dose), 24 hours = 1), hyperbilirubinemia in-hospital monitoring (n = 1), hyponatremia (n post-therapy, and 4- = 1) week outpatient monitoring Follow-up: until death Cancer Age group: adults Ad5/3-Delta24 Clinical efficacy None This study suggests the Kim et al, 2013233 Recurrent ovarian cancer No comparison group feasibility and safety of a ID NR (title states trial) Treatments enrollment: serotype chimeric infectivity-enhanced Single-arm trial none CRAd, Ad5/3-Delta24, as Treatment requisite: N: 10 a potential therapeutic none option for recurrent Follow-up: none ovarian cancer patients. Cancer Age group: adults Ad5-CD/TKrep Recurrence Of the adverse events There were no dose- Freytag et al, 2002142 Locally recurrent prostate No comparison group observed, 94% were mild limiting toxicities, and the Freytag et al, 2003374; cancer or moderate (grade 1/2) maximum tolerated dose Freytag et al, 2007375; Treatments enrollment: in nature. of the Ad5-CD/TKrep Freytag et al, 2007376 none vector was not defined. Together, the results ID NR (self-identified as Treatment requisite: demonstrate that trial in abstract) none intraprostatic Single-arm trial Follow-up: 60 administration of the Completion date: replication-competent 12/2001 Ad5-CD/TKrep virus N: 16 followed by 2 weeks of 5-fluorocytosine and ganciclovir prodrug therapy can be safely applied to humans and is showing signs of biological activity.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad5CMV-p53 Complete response, SAEs: 2 Ad5CMV-p53 combined Cristofanilli et al, 2006204 Locally advanced breast No comparison group partial response, stable with PST is safe, ID NR, reviewed by NIH- cancer disease, progressive biologically active, and RAC Treatments enrollment: disease associated with local immunomodulatory Single-arm trial none effects. The promising Treatment requisite: Completed clinical activity of this chemotherapy, surgery N: 13 combination deserves Follow-up: 41 further investigation in randomized studies. Cancer Age group: adults Ad5CMV-p53 Toxicity, vector None Multiple courses of Fujiwara et al, 2006200 Advanced non-small-cell No comparison group distribution, antibody intratumoral Ad5CMV- ID NR (abstract states lung cancer formation, tumor p53 injection alone or in trial) Treatments enrollment: response combination with intravenous Single-arm trial none administration of cisplatin Treatment requisite: N: 15 were feasible and well none tolerated in advanced Follow-up: none NSCLC patients, and appeared to provide clinical benefit. Cancer Age group: adults Ad5CMV-p53 Safety, pharmacokinetics None Ad5CMV-p53 can be Tolcher et al, 2006136 Advanced cancer No comparison group and pharmacodynamics, safely and repetitively ID NR (but trial stated in Treatments enrollment: detection of Ad5CMV-p53 administered up to 1 title) chemotherapy DNA expression in tumor ten(12) virus particles tissue, skin biopsies, intravenously daily for 3 Single-arm trial Treatment requisite: urine, rectal swabs, oral consecutive days. none N: 17 rinses, detection of Follow-up: 1 antibodies to adenovirus 5, antitumor activity Cancer Age group: adults Ad5CMV-p53; INGN 201; Adverse events, None Observations from this Shimada et al, 2006203 Chemoradiation-resistant ADVEXIN detection of p53 study indicate this Shimada et al, 2008377 advanced esophageal No comparison group transgene, p21 induction treatment results in local and MDM2 induction antitumor effects in ID NR, protocol reviewed squamous cell carcinoma chemoradiation-resistant by NIH-RAC Treatments enrollment: none esophageal squamous Single-arm trial cell carcinoma. N: 10 Treatment requisite: chemoradiation therapy Follow-up: 47

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad5-yCD/utTK(SR39)rep- None None The results demonstrate Barton et al, 20087 Localized prostate cancer hNIS that noninvasive imaging Barton et al, 2011378 Treatments enrollment: No comparison group of adenovirus-mediated gene therapy in humans BB-IND 12786 none is feasible and safe. Single-arm trial Treatment requisite: N: 12 none Follow-up: 12 Cancer Age group: adults AdCAIL-2 Local tumor regression No grade 3 or 4 toxicities Incomplete local tumor Stewart et al, 1999160 Metastatic breast cancer, No comparison group observed regression occurred at Stewart et al, 1997379 melanoma the site of injection in 24% of patients, but no ID NR, protocol reviewed Treatments enrollment: conventional clinical by the Health Protection none responses were seen. Branch of Canada Treatment requisite: Trial (multiple groups) isolation for 72 hours N: 23 after injection to check for viral shedding Follow-up: 1 Cancer Age group: adults AdCAIL-2 Toxicity, systemic None This trial demonstrates Trudel et al, 2003185 Localized prostate cancer No comparison group immune responses, the feasibility and safety ID NR, protocol reviewed Treatments enrollment: tumor response, clinical of intraprostatic by University of Toronto none outcome adenovectormediated IL- 2 gene delivery. Institutional Review Treatment requisite: Board and by the Health none Protection Branch of Follow-up: 18 Canada Single-arm trial N: 12 Cancer Age group: adults Ad-CCL21- DC Survival, safety No serious treatment- Intratumoral vaccination Lee et al, 2017124 Lung cancer No comparison group related adverse events with Ad-CCL21-DC Trial (multiple groups) Treatments enrollment: reported resulted in (1) induction of systemic tumor N: 16 none antigen-specific immune Treatment requisite: responses, (2) enhanced leukapheresis tumor CD8þ T-cell Follow-up: 4 infiltration, and (3) increased tumor PD-L1 expression.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults AdCD40L Safety and toxicity of None Local AdCD40L gene Malmstrom, 2010219 Invasive bladder cancer No comparison group AdCD40L gene therapym, therapy was safe, Uppsala University, Treatments enrollment: AdCD40L vector boosted immune 2009380 none spreading and host activation, and should be antiviral immune further evaluated as a NCT00891748 Treatment requisite: responses, CD40L protein single or an adjuvant none Single-arm trial expression in biopsies, therapy for urothelial N: 8 Follow-up: none CD40L-mediated cytokine malignancies. burst, tumor-infiltrating inflammatory cells, circulating immune cell populations, and antitumor effects Cancer Age group: adults Ad-CD40L Reduction in blood 1 patient experienced Infusions of autologous, Wierda et al, 2010223 Chronic lymphocytic No comparison group lymphocyte counts, grade 3 headache (dose- ISF35-transduced chronic Wierda et al, 2000381; leukemia reduction in limiting toxicity) lymphocytic leukemia Okur et al, 2011382 Treatments enrollment: lymphadenopathy, (CLL) cells were well enhanced/new tolerated, had biological ID NR none expression of target for and clinical activity, and Treatment requisite: Trial (multiple groups) susceptibility for might enhance the confirmation of eligibility N: 9 targeting, number of susceptibility of CLL cells (clinical history, physical infusions, time to with del(17p) to examination, routine progression, time to chemoimmunotherapy. laboratory evaluation), salvage therapy, type of leukapheresis, outpatient salvage therapy, clinical infusion of transduced response cells (test dose followed by remaining dose), scheduled follow-ups (as per eligibility confirmation), option to enroll in long-term study (and additional doses of the therapy) Follow-up: until death

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad-DLMP1-LMP2 Safety, immune response None Ad-DLMP1-LMP2 Chia et al, 2012128 Advanced metastatic No comparison group transduced DCs can be ID NR nasopharyngeal successfully generated and safely administered Single-arm trial carcinoma to patients with N: 16 Treatments enrollment: none advanced NPC. Treatment requisite: none Follow-up: 7 Cancer Age group: adults AdGLIPR1 Changes in pathology Grade 3 toxicities: The intraprostatic Sonpavde et al, 2011227 Localized intermediate or No comparison group (diagnosis, morphology, urinary tract infection (n administration of GLIPR1 ID NR high-risk prostate cancer apoptosis) = 2); no grade 4 tumor suppressor gene toxicities expressed by an Trial (multiple groups) preceding radical prostatectomy adenoviral vector was N: 19 Treatments enrollment: safe in men, with none localized intermediate or high-risk prostate cancer Treatment requisite: oral preceding radical broad spectrum antibiotic prostatectomy. was taken starting the Preliminary evidence of day before the injection biologic antitumor and continued for 4 days activity and systemic after the injection; immune response was intraporstate injection of documented. therapy delivered using transrectal ultrasound for guidance; patients were admitted to hospital for 23 hours Follow-up: 1

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults AdhAQP1 Improvement in parotid No deaths, dose-limiting AdhAQP1 vector delivery Baum et al, 2012229 Radiation-induced No comparison group saliva flow rates toxicities, or serious to a single parotid gland Dental National Institute, salivary hypofunction in (absolute volume, adverse events observed was safe and increased 2014383; Alevizos et al, surviving head and neck proportional change), because of vector parotid flow and relieved 2017384; cancer patients with improvement in visual delivery symptoms in a subset of analog scale assessments patients. Zheng et al, 2010385 Radiation Therapy Oncology Group late of perception of oral NCT00372320 grade 2 to 3 toxicity dryness and amount of Trial (multiple groups) Treatments enrollment: saliva present in their N: 11 none mouth Treatment requisite: glycopyrrolate received intravenously 30 minutes before administration of gene therapy; dilation of Stensen’s duct for cannulation for infusion of gene therapy Follow-up: 12 Cancer Age group: adults AdHSV-TK Clinical efficacy/disease No vector-associated Treatment with AdHSV- Alvarez et al, 2000165 Recurrent ovarian cancer No comparison group status grade 3/4 laboratory TK and ganciclovir in Alvarez and Curiel, Treatments enrollment: abnormalities were combination is feasible in 1997386 Patients had to have observed, but 1 patient the context of human ovarian cancer and ID NR, protocol reviewed failed standard debulking died due to a pulmonary tolerated at the dosages by NIH-RAC and FDA and chemotherapy embolism 3 weeks post- therapy. studied. Trial (multiple groups) treatments. N: 14 Treatment requisite: placement of a Tenchkoff catheter for intraperitoneal administration of gene therapy; 2 days after gene therapy delivery, patients received intravenous ganciclovir for 2 weeks Follow-up: until death

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad-IFNalpha Urine INFa levels A patient was found to A prior trial with a single Navai et al, 2016246 Non-muscle-invasive No comparison group have nonorgan-confined instillation compared ID NR bladder cancer disease with multiple similarly with the current positive lymph nodes at study, suggesting a Single-arm trial Treatments enrollment: radical cystectomy. He second instillation is not N: 7 Patients with non- muscle-invasive bladder succumbed to the necessary to achieve cancer refractory to disease 20.1 months sufficient urinary IFNa Bacillus Calmette–Guerin after study treatment levels. therapy who refused and 16.7 months after radical cystectomy were radical cystectomy. eligible for inclusion Treatment requisite: none Follow-up: 0 Cancer Age group: adults Ad-ISF35 Progressive disease, SAEs (events): 5 Ad-ISF35 IDI was safely Castro et al, 2012138 Chronic lymphocytic No comparison group stable disease, partial neutropenia, 3 delivered in patients with NCT00783874, BB-IND leukemia response, complete hypophosphatemia, 1 CLLs and induced 13046 Treatments enrollment: response AST elevation systemic biologic and clinical responses. These Trial (multiple groups) declined chemoimmunotherapy results provide the Completed rationale for phase II Treatment requisite: N: 15 studies in CLLs, none lymphomas, and CD40- Follow-up: 12 expressing solid tumors. Cancer Age group: adults Ad-mda7; INGN 241 Safety, clinical response Fatigue (n = 1) Intratumoral Cunningham et al, Resectable solid tumors No comparison group administration of INGN 194 2005 Treatments enrollment: 241 is well tolerated, Tong et al, 2005387 surgery, chemotherapy, induces apoptosis in a large percentage of ID NR, protocol reviewed and radiotherapy tumor cells, and by FDA Treatment requisite: demonstrates evidence none Single-arm trial of clinically significant N: 22 Follow-up: 1 activity.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad-OC-hsv-TK Safety, titers of anti- Lymphopenia (n = 1) Data suggest this form of Kubo et al, 2003181 Localized and metastatic No comparison group adenovirus 5 antibody, gene therapy requires Malaeb et al, 2005388 hormone-refractory bioassay of viral titers in further development for urine and serum, the treatment of ID NR, protocol reviewed prostate cancer detection of HSV TK androgen-independent by FDA Treatments enrollment: gene in biopsy prostate cancer Single-arm trial lymph node, retropubic prostatectomy, radiation specimens, circulating metastasis, although N: 11 therapy HSV TK gene histopathological and immunohistochemical Treatment requisite: evidence of apoptosis none was observed in the Follow-up: 12 specimens treated. Cancer Age group: adults Ad-OC-TK Adverse events related to Lymphopenia (n = 3) The phase I/II clinical Shirakawa et al, 2007208 Hormone-refractory No comparison group gene therapy, PSA trial of Ad-OC-TK/VAL Hinata et al, 2006389; metastatic prostate response and gene therapy was Terao et al, 2009390 cancer progression, local completed without any antitumor effect serious adverse events. ID NR, protocol reviewed Treatments enrollment: The efficacy of by Japanese Ministry of androgen deprivation docetaxel-based Health, Labor, and therapy, hormone chemotherapy in Welfare therapy, and chemotherapy hormone-refractory Single-arm trial prostate cancer has been Treatment requisite: N: 6 confirmed. Ad-OC- docetaxel and TK/VAL is the first in vivo estramustine adenoviral gene therapy combination used to treat bone chemotherapy, radical metastasis, and the prostatectomy, radiation expected response to therapy docetaxel-based Follow-up: 34 chemotherapy was preserved, suggesting potential benefit to future combined approaches for the treatment of hormone- refractory metastatic prostate cancer.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Adp53 Safety, clinical activity, Injection site pain (n = Intratumoral injection Nemunaitis et al, 2000161 Advanced non-small-cell No comparison group tumor biopsy 1) with Adp53 in Weill et al, 2000391 lung cancer combination with cisplatin is well tolerated, ID NR, protocol Treatments enrollment: and there is evidence of reviewed by RAC chemotherapy clinical activity. Single-arm trial Treatment requisite: N: 24 none Follow-up: 1 Cancer Age group: adults Adp53 Mean survival time, SAEs: 2 patients For patients with Ning et al, 2011145 Non-small-cell lung No comparison group overall survival, intervention, 3 patients relapsed NSCLC, trans- ID NR. All patients were cancer progressive disease, control (unclear) tracheal injection of required to sign an Treatments enrollment: partial response, stable Adp53 in addition to informed consent form none disease docetaxel did not improve overall survival and the protocol was Treatment requisite: or efficacy. However, approved by the docetaxel direct trans-tracheal institutional ethics Follow-up: 20 committee. injection of Adp53 proved to be a safe Trial (multiple groups) procedure, with tolerable Completed levels of toxicity. N: 42 Cancer Age group: adults Adp53 Tumor size, tumor No dose-limiting toxicity Adp53 combined with Zhang et al, 2005199 Advanced-stage cancer No comparison group, response (complete and adverse events hyperthermia was safe SFDA China project No. with measurable disease other: All patients response, partial noted except transient and effective in patients 2000-54 but no distant metastasis received gene therapy response, stable disease. fever (grade 1 or 2) with advanced cancer, and progressive disease), and p53 gene therapy Trial (multiple groups) Treatments enrollment: and hyperthermia; none subgroups received low-density area has potential to sensitize N: 15 advanced cancer to Treatment requisite: All radiotherapy (n = 5) or hyperthermia treatment. patients received gene chemotherapy (n = 3). therapy and hyperthermia weekly (4- 8 total doses per patient); subgroups received radiotherapy (n = 5) or chemotherapy (n = 3). Follow-up: 2

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad-p53 Disease-specific survival, None The safety and overall Clayman et al, 1999159 Squamous cell carcinoma No comparison group disease-free interval patient tolerance of Ad- Clayman et al, 1998392; of the head and neck p53 has been Nemunaitis et al, 2009393 Treatments enrollment: demonstrated. Of 15 resectable but historically ID NR none noncurable patients in Treatment requisite: Trial (multiple groups) the surgical arm of a surgery Completed phase I study, 4 patients N: 15 Follow-up: 29 (27%) remain free of disease, with a median follow-up time of 18.25 months. Surgical and gene transfer related morbidities were minimal. Cancer Age group: adults Ad-p53 Safety and determination Dyspnea, hypoxia, Ad-p53 can be Keedy et al, 2008211 Bronchioloalveolar cell No comparison group of MTD, P53 transgene pneumonitis/infiltrates administered safely with Eastern Cooperative lung carcinoma expression, monitored repeated dosing. Oncology, 2010394; Treatments enrollment: biodistribution of ad-p53, Eastern Cooperative none clinical response 395 Oncology, 2009 ; Treatment requisite: NCT00003649 none Single-arm trial Follow-up: 16 N: 25 Cancer Age group: adults Ad-p53 Complete response, None Patients in this study had Yen et al, 2000162 Advanced non-small-cell No comparison group partial response, stable evidence of an ID NR lung cancer disease, progressive antitumoral effect of this disease treatment with prolonged Trial (multiple groups) Treatments enrollment: none tumor stability or Completed regression; however, Treatment requisite: N: 10 neither Abs to p53 none protein nor increased Follow-up: 0 lymphocyte proliferative responses to wild-type or mutant p53 peptides have been consistently detected.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Ad-p53; INGN 201 Toxicity Hemiparesis (n = 1) Transduced cells were Lang et al, 2003184 Recurrent glioma No comparison group found only within a short Sidney Kimmel Treatments enrollment: distance of the injection Comprehensive Cancer radiotherapy, site. Although toxicity Center, 2002396 procarbazine, bis- was minimal, widespread distribution of this agent ID NR chloronitrosourea, remains a significant Single-arm trial temozolomide, stereotactic radiosurgery, goal. N: 15 thioguanine Treatment requisite: none Follow-up: 2 Cancer Age group: adults Ad-p53; INGN 201 Complete response, SAEs: 6 patients Intratumoral injection of Swisher et al, 2003179 Nonmetastatic non-small- No comparison group partial response, ptable Ad-p53 (INGN 201) in Swisher et al, 1999397; cell lung cancer disease, progressive combination with Roth et al, 1998398 Treatments enrollment: disease radiation therapy is well tolerated and ID NR none demonstrates evidence Treatment requisite: Trial (multiple groups) of tumor regression at radiation therapy Completed the primary injected N: 19 Follow-up: 47 tumor. Serial biopsies of the tumor suggest BAK gene expression is most closely related to Adp53 (INGN 201) gene transfer.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Adp53; INGN 201; Complete response, SAES: 18 events/patients Multiple dosing of IP Wolf et al, 2004191 Platinum- and paclitaxel- ADVEXIN partial response, mnor Adp53 was well tolerated National Cancer Institute, resistant epithelial No comparison group response, progressive in this group of heavily 2002399 ovarian cancer disease pretreated patients; however, the dosing NCT00003588 Treatments enrollment: schedule and the amount Trial (multiple groups) platinum- or paclitaxel- based therapy cannot be concluded Completed from this study. With a Treatment requisite: N: 17 negative randomized trial none of ovarian cancer in Follow-up: 0 front-line treatment that included an adenovirus p53 plus chemotherapy, further refinement of gene therapy is required before additional trials are undertaken. Cancer Age group: adults Ad-REIC Changes in serum- None In the present Kumon et al, 2016247 High-risk localized No comparison group neutralizing antibody titer neoadjuvant study, we Kumon et al, 2015400 prostate cancer against believe that the proof of concept of simultaneous UMIN000004929 undergoing radical adenovirus, quantitative prostatectomy kinetic analysis of induction of selective Single-arm trial Treatments enrollment: adenoviral DNA in blood killing of cancer cells and N: 18 none by real-time PCR, augmentation of antitumor immunity by Treatment requisite: quantitative kinetic Ad-REIC has been none analysis of adenoviral DNA in urine by real-time successfully established. Follow-up: 46 PCR, cytopathic effects, lymphocytic infilt

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: children Ad-RSVtk Ocular inflammatory None In contrast to studies IIdefonso et al, 2010220 Retinoblastoma No comparison group response, humoral showing significant ID NR Treatments enrollment: immune response immunogenicity of Ad- RSVtk following injection Single-arm trial chemotherapy or radiation therapy into extraocular tumors, N: 7 injection into the eye Treatment requisite: produces only a mild Patients received an local inflammatory intraocular injection of response without the vector followed by evidence of systemic intravenous ganciclovir cellular or humoral for 7 days; patients immune responses to received topical steroids adenovirus. immediately following the procedure. Follow-up: None Cancer Age group: children and Ad-siSSF Clinical efficacy and None This study, with Wang et al, 2018130 adults Usual care: standard adverse events combinatory NCT01956630 Acute leukemia donor lymphocyte associated with Ad-siSSF- modifications in DCs, DC treatment, survival, demonstrates the safety RCT Treatments enrollment: infusion chemotherapy TAA-specific CTL and efficacy of SOCS1- N: 48 responses in vitro, hDCs silenced DCs in treating Treatment requisite: in vitro relapsed acute leukemia. none Follow-up: 23 Cancer Age group: adults AdTG5327 Remission Safety assessed, no No adverse effect related Griscelli et al, 2003180 Unresectable lung cancer No comparison group adverse events reported to positive culture has Molinier-Frenkel et al, (stage IIIB or IV) been observed, thus 2000401; Tursz et al, Treatments enrollment: consistent with the safety 19968; Gahery-Segard et none of this therapy. 402 al, 1997 Treatment requisite: ID NR, protocol reviewed chemotherapy by French regulatory Follow-up: 24 agencies geverning human biomedical research and genetic engeneering Trial (multiple groups) Completed N: 21

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Adv.HSV-tk Safety considerations None This study confirmed the Nasu et al, 2007205 Local recurrence of No comparison group and adverse events, PSA safety profile and ID NR, protcol reviewed prostate cancer after response, detection of possibility of clinical by Japanese government hormone therapy HSV-tK gene in biopsy response at the specimens, anti- surrogate marker level in Trial (multiple groups) Treatments enrollment: none adenovirus neutralizing a clinical trial of HSV-tk N: 8 antibody response, gene therapy for Treatment requisite: systematic hormone-refractory none immunoresponses prostate cancer. Follow-up: 7 Cancer Age group: adults Adv.RSVtk Clinical course during None We conclude the form of Trask et al, 2000140 Malignant brain tumors No comparison group treatment gene therapy used in our ID NR Treatments enrollment: phase I trial is safe when the vector dose does not Single-arm trial none exceed 2 3 1011 VP (1 3 Treatment requisite: N: 13 1010 IU). none Follow-up: 25 Cancer Age group: adults Adv.RSV-tk None Hepatic toxicities were These results indicate Sung et al, 2001170 Metastatic colorectal No comparison group low, with transient grade Adv.RSV-tk can be safely ID NR adenocarcinoma in the 1 elevations in serum administered by aminotransferase levels percutaneous Trial (multiple groups) liver in 3 of 16 patients. Other intratumoral injection in N: 16 Treatments enrollment: none toxicities were also patients with hepatic transient: grade 2 to 3 metastases and can Treatment requisite: fevers in 5 of 16 provide the basis for none patients, grade 3 future clinical trials Follow-up: 12 thrombocytopenia in 1 of involving intratumoral 16 patients, and grade 2 adenoviral vector leucopenia in 3 of 16 injection. patients.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults ADV/HSV-tk Reduction in serum PSA No patient experienced Repeated cycles of in situ Miles et al, 2001171 Prostate cancer with local No comparison group (PSAR), return to initial any toxicity. HSV-tk plus GCV gene Herman et al, 1999403; recurrence after PSA (TR-PSA), PSA- therapy can be Ayala et al, 2005404 radiotherapy doubling time (PSADT) administered to patients where PSA = prostate- with prostate cancer BB-IND 6636 Treatments enrollment: none specific antigen and refractory to Trial (multiple groups) PSADT is considered to radiotherapy with a Treatment requisite: All N: 36 be a surrogate for tumor localized recurrence. patients received at least doubling time Biological responses to one transrectal this experimental therapy ultrasound-guided include increases in injection of gene therapy PSADT, PSAR, and TR- into a cancer positive PSA, and activated CD81 lobe of the prostate; a T cells present in the subgroup received peripheral blood. intravenous ganciclovir infusions (n = 29) starting 24 hours later (every 12 hours for up to 28 doses). Follow-up: until death Cancer Age group: adults ADV/HSV-tk Prostate-specific antigen 2 patients developed Combined RT, short- Teh et al, 2004187 Prostate cancer No comparison group, (PSA) local failure, PSA allergic reaction to course hormonal Teh et al, 2001405 Treatments enrollment: other: gene therapy with distant failure, valacyclovir and therapy, and in situ Fujita et al, 2006406 Patients had to be radiotherapy or with biochemical control, withdrew therapy appeared to Teh et al, 2002407 treatment naïve. hormone therapy and disease-free survival, provide good overall survival locoregional control but ID NR, protocol reviewed Treatment requisite: radiotherapy (2 different inadequate systemic by NIH-RAC and FDA administered broad- groups) control in patients with spectrum antibiotic Trial (multiple groups) positive pelvic lymph beginning day before N: 62 nodes. Longer-term use injections and for 3 more of hormonal therapy in days, gene therapy addition to gene therapy administered by and RT has been intraprostatic injections, adopted for this group of prodrug valacyclovir patients to maximize administered 24 hours both locoregional and after each injection for systemic control. 14 days; for hormone and radiation Follow-up: 24

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults ADV/HSV-tk/GC Quality of life, tumor 11 serious adverse The study is still ongoing. Germano et al, 2003186 Recurrent malignant No comparison group progression, survival events (required “These data show the ID NR gliomas hospitalization) ADV/HSV-tk/GC complex experienced by 6 at the dose used in this Trial (multiple groups) Treatments enrollment: patients: tumor study is safe. Additional N: 11 Patients must have failed conventional external progression (n = 4), of dose escalation is in beam radiation and which 3 required re- progress.” possibly chemotherapy, operation for resection of and external beam recurrent tumor 12 radiation and weeks after gene chemotherapy. therapy; leaked CSF from the surgical incision 2 Treatment requisite: weeks after surgery (n = none 1); perioperative status Follow-up: 12 epilepticus followed by prolonged need for intubation, sepsis, and death 4 weeks after gene therapy (n = 1, only 1 considered to be possibly related to the therapy) Cancer Age group: adults Adv/tk Time to disease No severe adverse We conclude that HSV-tk Sandmair et al, 2000166 Primary or residive lobar Other: transfection with progression, survival effects detected and Adv/tk gene therapy ID NR malignant gliomas control marker gene is safe and well tolerated. On the basis of Controlled trial Treatments enrollment: these results, further N: 14 tumor(s) had to be eligible for surgical trials are justified, removal especially with adenovirus vectors. Treatment requisite: craniotomy and radical tumor resection prior to localized delivery of gene therapy and ganciclovir treatment post surgery (different protocol for controls) Follow-up: until death

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults AdV/TK Safety and tumor None In summary, the results Xu et al, 2009213 Refractory malignant No comparison group response, vector suggest that AdV/TK ID NR tumors dissemination and /GCV combination suicide biodistribution, detection gene therapy is safe in Single-arm trial Treatments enrollment: none of neutralizing antibodies Chinese patients and N: 18 directed to achieved a local response Treatment requisite: with few environmental none adenovirus 5 and herpes simplex virus, Detection effects. Follow-up: none of plasma concentrations of GCV Cancer Age group: adults AdvHSV-tk Death or surgery for None The median survival time Immonen et al, 2004193 Malignant glioma Usual care recurrence, all-cause increased from 37.7 to ID NR Treatments enrollment: mortality, tumor 62.4 weeks. Six patients progression, QoL had increased anti- Trial (multiple groups) none adenovirus antibody Treatment requisite: Completed titers without adverse intravenous ganciclovir N: 36 effects. The treatment Follow-up: 54 was well tolerated. It is concluded that AdvHSV- tk gene therapy and GCV is a potential new treatment for operable primary or recurrent high-grade glioma.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Adv-HSV-tk Overall survival, survival Survival comparable to Two out of 5 patients Hasenburg et al, 2001172 Recurrent ovarian cancer No comparison group time, time to disease patients receiving were free of tumor. Hasenburg et al, 2000408; Treatments enrollment: progression, quality of surgery and Survival was comparable Hasenburg et al, 2002409 Patients must have life chemotherapy to that of patients of other studies with BB-IND 7311 completed initial surgery secondary cytoreductive Trial (multiple groups) and chemotherapy (cisplatin or carboplatin) surgery in combination N: 10 and should have been off with chemotherapy. treatment for at least 6 weeks. Treatment requisite: Topotecan treatment commenced 24 hours after gene therapy injection and continued in 3-week intervals until progression. Follow-up: 60 Cancer Age group: adults Adv-HSV-tk Toxicity, cellular and Cytolysis (n = 1) Neo-adjuvant van der Linden et al, Prostate cancer No comparison group humoral immune adenovirus-mediated 196 2005 Treatments enrollment: responses cytotoxic gene therapy ID NR, protocol reviewed none prior to prostatectomy for prostate cancer is by Dutch Commission for Treatment requisite: feasible and safe in an Genetic Modified Agents pelvic lymph node outpatient setting for and by the Ethics dissection, retro-pubic intraprostatic vector Committee of the radical prostatectomy Erasmus MC doses up to 2 1011 viral Follow-up: 12 particles. Single-arm trial N: 8

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Adv-IL2 None No adverse effects and Intermediate results of Gilly et al, 1999158 Unresectable digestive No comparison group no acute toxicity this phase I to II trial Gilly et al, 1998410 cancer using a single dose (107 and 108 pfu) of ID NR, protocol approved Treatments enrollment: recombinant replication- by French Ministry of pathologically proven deficient adenovirus Health, French Ethical digestive cancer that was expressing the human IL- Committee revealed to be 2 gene in unresectable Single-arm trial unresectable through surgical laparotomy digestive cancers showed N: 6 Treatment requisite: an excellent clinical, none biological, and immune tolerance with no Follow-up: none adverse effects and no acute toxicity. No spreading or shedding of AdV-IL2 was observed. Cancer Age group: adults AdV-tk Progression-free survival, Surgical group grade 3: AdV-tk can be safely Aguilar et al, 2015240 Pancreatic No comparison group, overall survival, clinical abdominal pain (n = 2), combined with pancreatic Advantagene, 2013411 adenocarcinoma other: gene therapy + response (progressive dehydration (n = 1), cancer SOC without disease, partial response, laboratory tests (n = 17); added toxicity. Response NCT00638612 Treatments enrollment: product as adjunct to none surgery or stable disease) surgical group grade 4: and survival compare Trial (multiple groups) laboratory tests (n = 7); favorably to expected Treatment requisite: chemoradiation N: 27 chemoradiation group outcomes and immune surgery or grade 3: dehydration (n activity increased. These chemoradiation = 1), laboratory tests (n results support further Follow-up: 66 = 15); chemoradiation evaluation of GMCI with group grade 4: more modern laboratory tests (n = 6) chemoradiation and surgery as well as PD- 1/PD-L1 inhibitors in pancreatic cancer.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: children AdV-TK Apparent clinical Inflammation AdV-TK followed by Chevez-Barrios et al, Bilateral retinoblastoma No comparison group resolution of vitreous ganciclovir can be 2005198 with vitreous tumor tumor seeds. administered safely to ID NR seeding refractory to children with retinoblastoma. Single-arm trial standard therapies N: 8 Treatments enrollment: All patients had experienced treatment failure in prior therapies including chemo, radio, cryo, and laser. Treatment requisite: none Follow-up: 40 Cancer Age group: adults Adv-tk Classification of 2 reported serious This gene therapy (AdV- Chiocca et al, 2011141 Malignant glioma No comparison group, prognostic groups, adverse events that may tk) plus valacyclovir can Advantagene, 2010412 Treatments enrollment: other: resection in disease progression, have been related to be safely delivered in appearance of new lesion gene therapy: patient 5 combination with surgery NCT00751270 none combination with or neurologic (fever, hyponatremia, and accelerated radiation Treatment requisite: valacyclovir and radiation Trial (multiple groups) deterioration, nausea/vomiting, in newly diagnosed delivery of gene therapy (all patients received N: 13 corticosteroid dose stable confusion, grade 2 AST malignant gliomas. by injection into tumor radiotherapy along with or increasing, quality-of- and ALT and creatinine bed after surgical gene therapy) life assessment increases) and patient 4 resection of tumor; all (progressive extra-axial patients received oral fluid had to be collected valacyclovir 1 to 3 days from under the surgical post–gene therapy wound closure) delivery for 14 days followed by temozolomide as standard of care; standard radiation therapy began Follow-up: until death

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Adv-TK Progression-free survival, Stated no severe adverse The study demonstrated Ji et al, 2016243 Recurrent high-grade Usual care: conventional overall survival, objective events were observed; a notable improvement Huazhong University of glioma (HGG) therapy: surgery, response evaluation, reported grade 3 events: of PFS-6, PFS, and OS in Science and Technology, Treatments enrollment: chemotherapy, or clinical benefit of anemia (n = 1 and n = 2 the ADV-TK-treated 2011413 chemotherapy palliative care treatment in treatment and control group, and the efficacy groups, respectively) and and safety appear to be NCT00870181 completion ≥ 4 weeks to start of trial neutropenia (n = 1 in comparable to other RCT both groups) reported treatments used Treatment requisite: N: 53 for recurrent HGG. ADV- none TK gene therapy is Follow-up: 8 therefore a valuable therapeutic option for recurrent HGG. Cancer Age group: adults Adv-TK Recurrence-free survival, Well tolerated and no HCC patients with no Li et al, 2007207 Advanced hepatocellular Usual care: liver overall survival significant toxicity vascular invasion could ID NR carcinoma transplant evident; mild catarrhal be selected for LT symptoms (10/23), slight followed by adjuvant RCT Treatments enrollment: liver transplant fever with no chills, light ADV-TK therapy, N: 45 headache regardless of intrahepatic Treatment requisite: huge or diffuse tumor. none Follow-up: 26

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults AdV-tk Clinical response, serum Grade 3: lymphopenia (n In vivo transrectal Rojas-Martinez et al, Prostate cancer No comparison group PSA = 1) ultrasound guided 235 2013 Treatments enrollment: instillation of an ID NR excluded if had received adenoviral vector into 4 sites in the prostate was Single-arm trial previous treatment for prostate cancer practical as an outpatient N: 10 procedure, was well Treatment requisite: oral tolerated, and led to ciproflaxin and distribution throughout phosphate enema on the intraprostatic tumor morning of injection; mass. AdV-tk intraprostatic injections demonstrated no guided by transrectal significant acute or late ultrasonography; toxicities. gancyclovir or valacyclvir after injection (prodrug); radical prostatectomy after injection (9 weeks later on av) Follow-up: median: 136+ ongoing Cancer Age group: adults AdV-tk Survival None GMCI can be safely Wheeler et al, 2016244 Newly diagnosed No treatment combined with SOC in Advantagene, 2015414 malignant glioma newly diagnosed malignant gliomas. NCT00589875 Treatments enrollment: Controlled trial standard of care (SOC) treatment N: 48 Treatment requisite: none Follow-up: 88 Cancer Age group: adults Adv-TK Recurrence-free survival No evident adverse Double-dose ADV- Zhu et al, 2018154 Advanced hepatocellular Usual care: liver (RFS) and overall survival effects TK/GCV gene therapy Huazhong University of carcinoma transplant (OS), combined with LT was Science and Technology, Treatments enrollment: safety of double-dose safe and improved RFS 2011415 none ADV-TK/GCV adjuvant and OS in advanced HCC patients without vascular NCT02202564 Treatment requisite: therapy invasion during the 5- none RCT year follow-up period. N: 86 Follow-up: 60

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: children and Adv-VST Production of ADV-VST, None Adoptive transfer of Qian et al, 2017129 adults No comparison group ADV-VST infusion rapidly isolated ADV-VST NCT02851576 Hematological tolerance, Immune is an effective reconstitution, ADV therapeutic option for Single-arm trial malignancies response achieving in vivo N: 14 Treatments enrollment: ilogeneic hematopoietic expansion of specific T stem cell transplantation cells and clearance of viral load, even as a Treatment requisite: preemptive treatment. none Follow-up: 9 Cancer Age group: adults Aglatimagene Tumor response, Grade 3 cytotoxic Gene-mediated cytotoxic Aggarwal et al, 2018126 Malignant pleural besadenovec progression-free survival, response (n = 1) immunotherapy was safe Trial (multiple groups) effusion No comparison group overall survival, safety and well tolerated with chemotherapy in patients N: 19 Treatments enrollment: none with malignant pleural effusion and showed Treatment requisite: encouraging response. celecoxib Follow-up: 44 Cancer Age group: adults CG7870 Treatment-related Fatigue, fever PSA decreases Small et al, 2006202 Hormone-refractory No comparison group toxicity; PSA response; demonstrated on this ID NR, protocol reviewed metastatic prostate neutralizing antibodies; trial are the first reported by RAC and FDA cancer CG7870 in blood, saliva, evidence of PSA and urine; cytokines modulation after Single-arm trial Treatments enrollment: none blood levels intravenous N: 23 administration of an Treatment requisite: oncolytic Ad. none Follow-up: 6

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults CI-1042 Complete response, SAEs (events/patients): 1 In this phase II study, Hamid et al, 2003134 Metastatic colorectal No comparison group partial response, severe lethargy; 1 intravenous CI-1042 was Trial (multiple groups) cancer progressive disease, increase in partial administered safely to stable disease thromboplastin time patients with advanced Completed Treatments enrollment: none colorectal cancer. N: 18 Toxicity was Treatment requisite: manageable, consisting none primarily of flu-like Follow-up: 17 symptoms. Stable disease was experienced by 7 patients for 11 to 18 weeks Cancer Age group: adults CTL102 Immune responses, No dose-limiting toxicity Direct intratumoral Palmer et al, 2004188 Hepatic metastatic No comparison group, immunohistochemical observed, including at delivery of CTL102 to ID NR colorectal cancer or other: intratumoral analysis of resected the maximum dose of patients with primary and tumors for transgene virus particles secondary liver cancer is Trial (multiple groups) hepatocellular carcinoma injection followed by (1 to 5 cm) laparotomy and hepatic expression feasible and well N: 18 Treatments enrollment: resection 3 to 8 days tolerated with minimal none later side effects. Treatment requisite: Gene therapy was delivered by ultrasound- guided intratumoral injection followed by laparotomy and hepatic resection 3 to 8 days later. Follow-up: Until death Cancer Age group: adults CTL102 Change in observed time All adverse events With further clinical Patel et al, 2009214 Localized prostate cancer No comparison group to PSA progression (TTP) attributable to treatment development, it is ID NR Treatments enrollment: were transient and no believed VDEPT can additional treatment was become a new Trial (multiple groups) none required. Lymphopenia: therapeutic option. Treatment requisite: N: 39 grade 3 (n = 8), grade 4 prophylactic ciprofloxacin (n = 1); raised hepatic before injection and for 3 enzymes: grade 3 (n = days after 1); fever: grade 3 (n = Follow-up: 15 1); amnesia: grade 3 (n = 1)

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults dl1520 Tumor response by No serious adverse Further improvement in Habib et al, 2001167 Primary and secondary No comparison group computed tomography events recombinant vector ID NR liver tumors scan or tumor markers design is necessary to (complete response, achieve better clinical Trial (multiple groups) Treatments enrollment: none partial response, stable response. N: 16 disease, progressive Treatment requisite: disease) Phase I patients received multiple doses by 1 of 3 methods: intratumoral administration of gene therapy with computed tomography guidance, intraarterial administration through the hepatic artery, or intravenous administration via a vein in the arm. Follow-up: Until death Cancer Age group: adults dl1520; ONYX-015 Physical examination, Fever, rigour, Although only a few Habib et al, 2002176 Hepatocellular carcinoma Placebo liver function test, anti- hypotension monitored; patients were treated in ID NR Treatments enrollment: adenovirus type 5 vector was well this study, it appears antibody titer, analysis tolerated, although more effective vectors RCT none of adenovirus presence antitumor activity was are needed to achieve a Treatment requisite: Completion date: in serum samples using modest useful clinical impact. none 08/1999 polymerase chain N: 10 Follow-up: none reaction

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults dl1520; ONYX-015 Complete response, SAEs: 45 events No dose-limiting toxicity, Reid et al, 2001174 Colorectal carcinoma No comparison group partial response, stable maximally tolerated ID NR metastatic to the liver disease, progressive dose, or treatment- disease emergent clinical Single-arm trial Treatments enrollment: none hepatotoxicity were Completed identified following Treatment requisite: N: 11 dl1520 infusion. Mild to chemotherapy moderate fever, rigors, Follow-up: 2 and fatigue were the most common adverse events. An objective response was demonstrated in combination with chemotherapy in a patient who was refractory to both 5-FU and dl1520 as single agents. Therefore, hepatic artery infusion of the attenuated adenovirus dl1520 was well tolerated at doses resulting in infection, replication, and chemotherapy-associated antitumoral activity. Cancer Age group: adults E10A Tumor response, None reported Direct intratumoral Li et al, 2008137 Advanced tumors No comparison group progressive disease, injection of up to 1012 ID NR, China’s State Treatments enrollment: stable disease vp of E10A to patients is Food and Drug none well tolerated, and further studies are Administration regulatory Treatment requisite: necessary to define and authorities none increase clinical efficacy. Trial (multiple groups) Follow-up: 1 Completed N: 15

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults E10A Minor response, stable None Weekly intratumoral Lin et al, 2007144 Solid tumors No comparison group disease, progressive injection of up to 1 × ID NR, protocol approved Treatments enrollment: disease 1012 VP of E10A to by Chinese State FDA none patients with solid tumor is a feasible and well- and the institutional Treatment requisite: tolerated procedure that ethics committees none exerts mild antitumor Trial (multiple groups) Follow-up: 1 effects. A small and Completed sustained elevation of N: 15 endogenous endostatin in blood possibly has antitumor activity. Cancer Age group: adults E10A Objective response rate, SAEs: 24 intervention, 20 The overall disease Ye et al, 2014149 Advanced head and neck Usual care objective disease control placebo control rate significantly NCT00634595 carcinoma rate, overall response increased from 80.6% in rate, overall disease the control group to RCT Treatments enrollment: none control rate, overall 92.6% in the test group Completed survival, progression-free (P = 0.034). Except for Treatment requisite: N: 136 survival fever, no adverse events chemotherapy were associated with the Follow-up: 40.3 E10A treatment. In sum- mary, E10A plus chemotherapy is a safe and effective therapeutic approach in patients with advanced head and neck squamous cell carcinoma or nasopharyngeal carcinoma. Cancer Age group: adults H101 Complete regression, SAEs (events): 1 fever, 7 Intratumor injection of Lu et al, 2004135 Advanced cancerous No comparison group partial regression, minor leucopenia, 1 liver the genetically ID NR, protocol tumors response, progressive dysfunction, 1 alopecia engineered adenovirus approved by the State Treatments enrollment: disease, stable disease H101 exhibits potential Food and Drug surgery and/or antitumor activity to Administration of China chemotherapy and/or refractory malignant tumors in combination Single-arm trial radiotherapy with chemotherapy. Low Treatment requisite: Completed toxicity and good chemotherapy N: 46 tolerance of patients to Follow-up: 0 H101 were observed.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults INGN 201 (Ad5CMV-p53) Overall patient status, None Intraoperative INGN 201 Yoo et al, 2009216 Carcinoma of the oral No comparison group tumor status, adverse gene therapy is NCT00017173 cavity, oropharynx, effects, accrual rate, technically feasible, but it percentage of patients has many logistical Single-arm trial hypopharynx, and larynx successfully receiving problems when N: 13 Treatments enrollment: none required doses of INGN performed in a multi- 201 institutional setting. Treatment requisite: chemoradiotherapy Follow-up: 3 Cancer Age group: adults Ad5CMV-p53 Toxicity, detection of p53 Safe Intravesical instillation of Pagliaro et al, 2003183 Locally advanced bladder No comparison group transgene, Ad5CMV-p53 is safe, National Cancer Institute, cancer immunohistochemistry feasible, and biologically 2002416 Treatments enrollment: and TUNEL active when administered in multiple doses to NCT00003167 none patients with bladder Treatment requisite: Single-arm trial cancer. N: 13 none Follow-up: 44 Cancer Age group: adults OAMCGT Expanded Prostate Grade 3/4 AEs: 14 Combining OAMCGT with Freytag et al, 2014238 Prostate cancer Other: intensity- Cancer Index Composite, OAMCGT plus IMRT, 10 IMRT does not Lu, 2011417; Henry Ford Treatments enrollment: modulated radiation EuroQol EQ-5D IMRT only exacerbate the most Health, 2013418 none therapy common side effects of prostate radiation ID NR Treatment requisite: therapy and suggests a intensity-modulated Trial (multiple groups) clinically meaningful radiation therapy Completed reduction in positive N: 44 Follow-up: 24 biopsy results at 2 years in men with intermediate-risk prostate cancer. Cancer Age group: adults ONYX-015 Toxicity assessment None This trial has shown the Chiocca et al, 2004192 Recurring malignant No comparison group relative safety of ID NR (title states trial) glioma injection of ONYX-015 into the brain Single-arm trial Treatments enrollment: none surrounding a resected N: 24 malignant glioma. Treatment requisite: none Follow-up: 19

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults ONYX-015 Toxicity, None Intratumoral Galanis et al, 2005195 Advanced sarcoma No comparison group Immunohistochemistry administration of ONYX- ID NR, protocol reviewed Treatments enrollment: for p53 and mdm2 015 in combination with by Mayo Clinic none expression, plasma MAP chemotherapy is neutralizing antibody well tolerated with no Institutional Review Treatment requisite: titers, detection of ONYX- significant toxicity due to Board none 015 viral genome in ONYX-015 being Single-arm trial Follow-up: 11 plasma encountered. N: 6 Cancer Age group: adults ONYX-015 Complete response, SAEs (patients): 2 sepsis, This study indicates Hecht et al, 2003132 Unresectable pancreatic No comparison group partial response, 2 duodenal perforation, 2 ONYX-015 injection via Mulvihill et al, 2001419 carcinoma progressive disease, cystic fluid collection, 2 EUS into pancreatic stable disease, survival hyperlipasemia, 1 carcinomas by the Trial (multiple groups) Treatments enrollment: none hyperamylasemia, 1 transgastic route with Completed elevated liver tests, 1 prophylactic antibiotics is Treatment requisite: N: 21 fever, 6 feasible and generally ciprofloxacin, leukopenia/neutropeni, 1 well tolerated either chemotherapy lymphopenia, 1 anemia, alone or in combination Follow-up: 36 1 increased partial with gemcitabine. thromboplastin time, 1 Transgastric EUS-guided gemcitabine lung toxicity injection is a new and practical method of delivering biological agents to pancreatic tumors.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults ONYX-015 Tumor size, necrosis Grade 3 mucositis In patients with recurrent Lamont et al, 2000164 Recurrent squamous cell No comparison group induction of injected (n = 2) squamous cell carcinoma ID NR carcinoma of the head tumor (complete of the head and neck, regression, partial ONYX-015 adenovirus Single-arm trial and neck regression, minor plus chemotherapy N: 14 Treatments enrollment: none regression, stable (cisplatin and 5- disease, progressive fluorouracil) provided Treatment requisite: disease) antitumor activity and intratumoral injection of local tumor control. the gene therapy with local anesthesia and/or intravenous sedation as needed for each injection (daily for 5 consecutive days, repeated every 3 weeks until tumor progression or adverse event); treatment cycles included cispla Follow-up: until death

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults ONYX-015 Toxicities, response to None Intralesional treatment Makower et al, 2003133 Hepatobiliary tumors No comparison group treatment, analysis of with ONYX-015 in Wadler, 2003420 Treatments enrollment: p53 status, correlation of patients with p53 status with hepatobiliary tumors is ID NR, protocol reviewed none response, antiadenoviral safe and well tolerated by NIH Treatment requisite: The antibody studies, and some patients had first 8 patients treated Single-arm trial analysis of viral shedding evidence of an anticancer received prophylactic N: 20 in body fluids effect. antibiotic coverage with oral erythromycin (1 g every 6 hours) and kanamycin (1 g every 6 h), beginning on the day before ONYX- 015 administration and continuing for 24 hours after treatment.Also, patients received i.v. ticarcillin/clavulanate (3.1 g every 4 h) and gentamycin (80 mg every 8 h). Patients allergic to penicillin received vancomycin (1 g every 12 h), aztreonam (2 g every 8 h), and metronidazole (500 mg every 6 h). Antibiotics began on the day before ONYX-015 administration and continued for 1 week after treatment. Follow-up: 0

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults ONYX-015 Complete regression, None The findings suggest that Nemunaitis et al, 2000131 Advanced head and neck No comparison group partial regression, minor belagenpumaucel-L Ganly et al, 2000421, cancer response, stable disease, improved survival for Khuri et al, 2000422 Treatments enrollment: progressive disease patients who were randomized within 12 ID NR (title describes none weeks of completion of study as trial) Treatment requisite: chemotherapy and in none Trial (multiple groups) those who had received Completed Follow-up: 4 prior radiation. Further N: 37 studies of belagenpumatucel-L in NSCLC are warranted. Cancer Age group: adults ONYX-015 (d11520) Clinical response, survival Grade 3 events: Patients generally Vasey et al, 2002175 Recurrent or refractory No comparison group time pyrexia/sweats (n = 3), tolerated the treatment ID NR epithelial ovarian cancer headaches (n = 2), but there was no clear myalgias (n = 1), malaise clinical benefit: “There Trial (multiple groups) Treatments enrollment: (n = 1), nausea (n = 3), was no clear-cut N: 16 had to have received platinum-containing vomiting (n = 2), evidence of clinical or chemotherapy with diarrhea/ileitis (n = 1), radiologic response in relapse within the abdominal pain (n = 1) any patient.” previous 6 months Treatment requisite: laparoscopy or laparotomy for intraperitoneal catheter placement, baseline measurements of biomarkers and tumor Follow-up: unclear

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults pVAX/L523S + Ad/L523S Relapse-free survival, No patients developed Results suggest a high Nemunaitis et al, 2006201 Early stage (IB, IIA, IIB) No comparison group overall survival grade 3 or 4 toxicity that level of safety. but ID NR non-small-cell lung was characterized as evidence of L523S- possibly, probably, or directed immune Trial (multiple groups) cancer definitely related to the activation was limited, N: 13 Treatments enrollment: none gene therapy or the suggesting a need for procedure. modification of dose, Treatment requisite: schedule, and site of intramuscular need and vaccination with further syringe injections at clinical testing. weeks 0, 2, 4, and 8 Follow-up: until death Cancer Age group: adults rAd/hup53 Tumor response None The treatment was well Kuball et al, 2002177 Advanced stage cancer No comparison group tolerated. To date, no ID NR Treatments enrollment: evidence for objective tumor responses was Single-arm trial none observed. An Treatment requisite: Completed amplification of humoral none N: 6 and cellular anti- Follow-up: 3 adenoviral immune responses was demonstrated in all patients following rAd/hup53 vaccination. Cancer Age group: adults rAd/p53; SCH 58500 Tumor sampling, toxicity, Fever (100%), CT scans are not a valid Buller et al, 2002178 Recurrent ovarian cancer Usual care: patient monitoring— hypotension (89%), measure of response to ID NR Treatments enrollment: chemotherapy ELISA Laboratory, data- variety of abdominal i.p. SCH 58500 due to serum, ascites sampling complaints (79%), extensive adenoviral- Trial (multiple groups) none for pharmacokinetic hypertension (68%), induced inflammatory Treatment requisite: N: 41 studies, complete blood nausea (63%), changes. Intraperitoneal none counts, fibrinogen, fibrin tachycardia (58%), SCH 58500 is safe, well Follow-up: 3 split products, PT, PTT, vomiting (58%), fatigue tolerated, and combined serum C3, C4, CH50, (53%) with platinum-based electrolytes including chemotherapy can be magnes associated with a significant reduction of serum CA125 in heavily pretreated patients with recurrent ovarian, primary peritoneal, or fallopian tube cancer.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults rAd2p53 Toxic effects None This recombinant virus– Lu et al, 2011224 Esophageal carcinoma Usual care RT combination is ID NR Treatments enrollment: significantly more beneficial in palliation RCT none than RT alone, with Treatment requisite: N: 45 minor side affects. none Follow-up: None Cancer Age group: adults rAd-IFNalpha Tumor response No significant treatment- Intravesical rAd-IFN Dinney et al, 2013231 Non-muscle-invasive No comparison group (complete response), related adverse events alphaSyn3 therapy was ID NR (but trial stated in bladder cancer duration of response, were observed. One well tolerated with no title) Treatments enrollment: progression-free survival patient was hospitalized dose-limiting toxicity for adverse events encountered. Efficient Trial (multiple groups) Patients were required to deemed not related to gene transfer and N: 17 have proven disease recurrence after 2 or the study therapy. expression was more cycles of Bacillus confirmed by dose- Calmette-Guerin (BCG), dependent urinary with or without interferon-alpha (IFNα) recombinant interferon- concentrations. alpha (IFNα) protein, and Intravesical rAd- at least 3 months since IFNα/Syn3 demonstrated last treatment. promising clinical activity in non-muscle-invasive Treatment requisite: bladder cancer (NMIBC) Cystoscopy, biopsies, recurring after BCG. urinary cytology, and bladder capacity evaluation were performed 2 to 3 weeks before therapy. Patients were hospitalized for 48 hours post-therapy and monitored daily for 5 days for adverse events. Follow-up: 24

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults rAd-IFNalpha/Syn3 High-grade disease Nine patients (22%) rAd-IFN alpha/Syn3 was Shore et al, 2017248 High-grade, bacillus No comparison group recurrence reported a total of 19 well tolerated. It NCT01687244 calmette-guerin- grade 3 AEs: 12 in the demonstrated promising low-dose arm (coronary efficacy for patients with Trial (multiple groups) refractory or relapsed non-muscle-invasive artery occlusion, HG NMIBC after BCG N: 43 bladder cancer diarrhea, sepsis, therapy who were unable Treatments enrollment: arthralgia, renal or unwilling to undergo Patients were unable or neoplasm, transitional radical cystectomy. unwilling to undergo cell carcinoma, carotid radical cystectomy. artery occlusion, syncope, renal failure, Treatment requisite: nephroureterectomy, none COPD, and hypotension) Follow-up: 12 and 7 in the high-dose arm (abdominal pain, back pain, fracture, syncope, dysuria [n = 2]), and acute renal failure. Cancer Age group: adults rAd-p53 Efficacy None The rAd-p53 gene Guan et al, 2011225 Advanced hepatocellular Usual care therapy in combination ID NR carcinoma with TACE is a safe and effective treatment Controlled trial Treatments enrollment: none modality for advanced N: 82 HCC. Treatment requisite: none Follow-up: 12 Cancer Age group: adults rAd-p53 Survival rate None Sequential therapy of Chen et al, 2014236 Advanced hepatocellular Usual care p53 gene transcatheter ID NR carcinoma arterial infusion was safe and could prolong the RCT Treatments enrollment: survival of the patients. N: 48 none Treatment requisite: none Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults rAd-p53 Progression-free survival, Observed no serious The wound surface Deng et al, 2017152 Non-small-cell lung Usual care: radical overall survival adverse effect related to injection of rAd-p53 NCT01574729 cancer surgery alone, other: treatment thus far. showed efficacious effects in preventing RCT Treatments enrollment: radical surgery alone plus none local injecton of vector recurrence or metastasis N: 163 and improving PFS and Treatment requisite: All OS after a radical surgery patients had radical in patients with NSCLC. survery. Follow-up: minimum 36, median 43 (as of report) Cancer Age group: adults rAd-p53 Overall antitumor efficacy None The results show the Guan et al, 2009215 Advanced non-small-cell Usual care: therapy of combination of rAd-p53 ID NR lung cancer bronchial arterial infusion and BAI was well tolerated in patients with Controlled trial Treatments enrollment: (BAI) none NSCLC and may have N: 12 improved the quality of Treatment requisite: life and delayed the none disease progression. Follow-up: 12 Cancer Age group: adults rAd-p53 Infection efficiency of Harms not reported Intraepithelial injections Li et al, 2009217 Oral leukoplakia No comparison group rAd-GFP, cell of recombinant human Zhang et al, 2009423 Treatments enrollment: morphology, growth adenovirus-p53 were inhibition ratio, apoptosis safe, feasible, and ID NR (described as trial) none and cell cycle, messenger biologically active for Treatment requisite: Single-arm trial RNA and protein patients with dysplastic none N: 22 expression of p53, p21, oral leukoplakia. Follow-up: 24 bcl-2

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults rAd-p53 Complete response, None Intra-arterial infusion of Li et al, 2014237 Stage III or IV oral Placebo partial response, stable combined rAd-p53 and ChiCTR-TRC-09000392 carcinoma disease, progressive chemotherapy disease, survival significantly increased RCT Treatments enrollment: none the survival rate of Completed patients with stage III Treatment requisite: N: 99 but not stage IV oral chemotherapy cancer, compared with Follow-up: 86 intra-arterial chemotherapy. Intra- arterial infusion of combined rAd-p53 and chemotherapy may represent a promising alternative treatment for oral squamous cell carcinoma. Cancer Age group: adults rAd-p53 Local recurrent rate, 3- SAEs (oral membrane The 3-year overall- Liu et al, 2013232 Oral cancer No treatment year disease-free burn and pain): 0 survival (OS) rate and 3- Liu et al, 2012424 Treatments enrollment: survival, 3-year overall intervention, 2 placebo year disease-free survival survival (DFS) rate of EG are ID NR none 100% and 93%, Treatment requisite: RCT respectively. The 3-year radiotherapy (after gene Completed OS and DFS rates of CG therapy) N: 107 are 94% and 68%, Follow-up: 36 respectively. Mild or medium fever and flu-like symptoms were more frequently observed in EG and were considered to be associated with application of rAd-p53. Post-tumorectomy wound surface injection of rAd-p53 combined with radiotherapy is a safe and effective regimen for the patients with TGa or GCa.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults rAd-p53 Overall survival rate, None In patients with NPC, Pan et al, 2009212 Nasopharyngeal Placebo, usual care: tumor control rAd-p53 was safe and ID NR, but trial— carcinoma radiotherapy biologically active. The including protocols and Treatments enrollment: results indicated rAd-p53 informed consent none improves radiotherapeutic tumor process—approved by Treatment requisite: control and survival rate the Chinese State Food none in patients with NPC. and Drug Administration Follow-up: 61 and Beijing Hospital Ethics Committee on September 12, 2001 RCT N: 82 Cancer Age group: adults rAd-p53 Survival, complete SAEs: late radiorectitis: 4 The 5-year progress-free Su et al, 2016139 Locally advanced cervical Other: radiotherapy response, partial PRT, 3 RT; late survival rate of the PRT RCT cancer alone response, stable disease, radiocystitis: 1 PRT, 0 group was 17.1% higher progressive disease RT; fever: 18 both than that of the RT Completed Treatments enrollment: none groups group (HR = 0.485; 95% N: 104 CI, 0.234-1.006; p = Treatment requisite: 0.047). rAd-p53 radiotherapy administration did not Follow-up: 97 increase the adverse events caused by radiotherapy, except for transient fever after rAd- p53 administration. rAd- p53 was safe and biologically active in improving radiotherapeutic survival rates in patients with cervical cancer.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults rAd-p53 Complete response, SAEs (events): 5 Taken together, Wang et al, 2016151 Renal cell carcinoma Other: lenvatinib partial response, stable hypertension, 4 lenvatinib and rAd-p53 CH21010110 Treatments enrollment: disease, progressive proteinuria, 1 diarrhea, 2 are well tolerated at the disease, progression-free rash, 1 fatigue, 2 pain in indicated dose. Gene and Trial (multiple groups) none survival injection site target therapy for RCC Treatment requisite: Completed could be further lenvatinib N: 382 enhanced by synergistic Follow-up: 42 effects of lenvatinib and rAd-p53. Cancer Age group: adults rAd-p53 Disease control rate, No significant difference rAd-p53 combined with Xiao et al, 2018155 Unresectable advanced Usual care: recombinant progressive disease rate, in all adverse events, radiotherapy and ID NR. This study was soft-tissue sarcomas adenovirus-p53 (rAd- progression-free survival, except for transient hyperthermia can approved by the Treatments enrollment: p53) gene therapy overall survival fever, which occurred in effectively improve the Institutional Review advanced unresectable combined with 89% of patients therapeutic efficacy and Board of the authors’ STS radiotherapy and survival outcomes in patients with advanced hospital. Written Treatment requisite: hyperthermia for unresectable STS, informed consent was none advanced STS providing a new obtained from all Follow-up: 6 patients. therapeutic strategy. Controlled trial Completion date: 11/2014 N: 71

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Replication-defective Tumor progression Not discussed After 28 days, 4 of the 6 Kauczor et al, 1999157 Incurable ((stage IIIb or adenoviral expression (stable disease or patients showed stable ID NR IV) non-small-cell lung vector encoding wild- progressive disease) disease at the treated type p53 tumor site, whereas Trial (multiple groups) cancer No comparison group other tumor sites N: 15 Treatments enrollment: none progressed. Computed tomography–guided Treatment requisite: injections are a sufficient spiral computed and easy-to-perform tomography (CT) procedure for performed prior to intratumoral gene therapy start; dose levels therapy delivery. 1 to 3 were delivered bronchoscopically and dose levels 3 to 4 were deilvered percutaneously under CT guidance; spiral CT was performed again at 28 days post- treatment Follow-up: 1 Cancer Age group: adults Replication-defective None No clinically significant Wild-type p53 gene Schuler et al, 1998156 Advanced non-small-cell adenoviral expression toxicity observed therapy by intratumoral Boulay et al, 2000425 lung cancer vector encoding wild- injection of a replication- type p53 defective adenoviral ID NR, protocol approved Treatments enrollment: expression vector is safe, by German and Swiss none No comparison group feasible, and biologically national reugulatory Treatment requisite: effective in patients with offices none advanced non-small-cell Single-arm trial Follow-up: 1 lung cancer. N: 15 Cancer Age group: adults rhAd5 Progression-free survival, None This study suggests Dong et al, 2014239 Unresectable Usual care overall survival rhAd5 is a safe, effective Control group was from hepatocellular carcinoma gene therapy that patients who received Treatments enrollment: prolongs the PFS and OS TACE along the same none time of patients with unresectable HCC. period Treatment requisite: Trial (multiple groups) none N: 299 Follow-up: 3

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults rhAd-p53 Reduction in size of Bone marrow Intratumoral injection of Xiao et al, 2017153 Locally advanced cervical Usual care PVB group cancer foci, total efficacy suppression (n = 1), rhAd‑p53 combined with ID NR cancer (LACC) (cisplatin + vincristine + rate (complete response, elevated neoadjuvant partial response, stable aminotransferase levels chemotherapy may have RCT Treatments enrollment: bleomycin), other: The treatment group is the disease, progressive (n = 2), high fever (n = advantages over N: 40 Patient had to have treatment for naïve gene therapy + PVB disease) 5) conventional cancer (ie, no prior group (cisplatin + chemotherapy for its chemotherapy and/or vincristine + bleomycin). high efficacy, safety, and radiotherapy). synergism in the therapy for LACC. Treatment requisite: All patients had chemotherapy (the gene therapy group was also adminsitered vector). Follow-up: Not discussed Cancer Age group: adults SCH 58500 Immune responses to None IHA administration of a Atencio et al, 2006197 Colorectal cancer No comparison group SCH 58500, tissue- replication defective ID NR metastatic to the liver specific detection of SCH adenovirus is a feasible 58500 DNA and mode of delivery, Single-arm trial Treatments enrollment: none transgene p53 mRNA, allowing for exogenous N: 29 distribution of SCH 58500 transfer of the p53 gene Treatment requisite: in liver, Ad transduction into target tissues, with none and pretreatment total or evidence of functional Follow-up: none serum neutralizing Ad p53. Limited and antibody levels, Ad transient inflammatory transduction and CAR responses to the drug expression occurred, but preexisting immunity to Ad did not preclude SCH 58500 delivery. Cancer Age group: adults SCH 58500; rAd/p53 None None Intratumoral adenoviral Schuler et al, 2001169 Advanced non-small-cell No comparison group p53 gene therapy ID NR, protocol reviewed lung cancer appears to provide no by national regulatory Treatments enrollment: additional benefit in offices of each none patients receiving an effective first-line participating center Treatment requisite: chemotherapy for Trial (multiple groups) none advanced NSCLC. N: 25 Follow-up: none

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults SCH 58500; rAd/p53 Toxicity and survival No observed dose- Intravesical instillation of Kuball et al, 2002173 Bladder cancer No comparison group limiting toxicity SCH58500 combined with ID NR Treatments enrollment: a transduction-enhancing agent is safe, feasible, Trial (multiple groups) none and biologically active in Treatment requisite: N: 12 patients with bladder cystoscopy or catheter cancer. placement Follow-up: 12 Cancer Age group: adults Sitimagene ceradenovec Time to death or re- SAEs: 111 intervention, The findings suggest use Westphal et al, 2013234 High-grade glioma Usual care: 2004- intervention 81 placebo of sitimagene Trial (multiple groups) Treatments enrollment: 000464-28 ceradenovec and ganciclovir after resection Completed none can increase time to Treatment requisite: N: 251 death or re-intervention none in patients with newly Follow-up: 43 diagnosed supratentorial glioblastoma multiforme, although the intervention did not improve overall survival. Locally delivered gene therapy for glioblastoma should be further developed, especially for patients who are unlikely to respond to standard chemotherapy.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults Telomelysin Adverse events, clinical None Telomelysin was well Nemunaitis et al, 2010127 Solid tumors No comparison group response, viral tolerated. Evidence of ID NR Treatments enrollment: pharmacokinetics antitumor activity was analysis, viral e1A, hexon suggested. Single-arm trial XRT, carboplatin, expression in treated N: 16 docetaxel, anastrozole, interferon, dacarbazine tumors, neutralizing lenalidomide, vinblastine, antibody response, cisplatin, IL‐2, interferon, serum cytokines, perifosine, cisplatin, peripheral blood doxorubicin, ifosfamide, lymphocyte gemcitabine, perifosine, immunophenotyping, irinotecan, topotecan, htert mrnA pemetrexed, CT 2103, paclitaxel, carbo Treatment requisite: none Follow-up: 0 Cancer Age group: adults TG1024 Serum IL-2 levels None The results show Dummer et al, 2008210 Advanced solid tumors No comparison group intratumoral injections of ID NR, reviewed by Swiss and melanoma TG1024 are safe and well and Belgium federal Treatments enrollment: tolerated. agencies none Single-arm trial Treatment requisite: N: 35 none Follow-up: none Cancer Age group: adults TG1041 Size of tumor, time to No patients died because It is possible that a Khorana et al, 2003182 Malignant melanoma No comparison group disease progression, of therapy administration different dose schedule ID NR Treatments enrollment: achievement of stable and no may enhance clinical disease, survival rate efficacy of the study Trial (multiple groups) Patients had cancer not grade 4 toxicities were observed. Two grade 3 drug. N: 11 treatable with current therapy. toxicities were observed Treatment requisite: in one patient: high fever none and deep venous thrombosis (both Follow-up: until death required hospitalization).

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults TG1042 Complete response, SAEs: 1 event The study showed Dreno et al, 2014148 B-cell lymphomas No comparison group partial response, stable treatment with TG1042 NCT00394693 Treatments enrollment: disease, duration of was associated with a response, dermatology clinical benefit in most of Single-arm trial none life quality index, time to the patients with Treatment requisite: Completed disease progression relapsing CBCL, including none N: 13 tumor regression, a Follow-up: 53 clinically meaningful duration of response, and a good treatment tolerance. Cancer Age group: adults TG1042 Clinical outcome, safety, None The study shows Dummer et al, 2004190 Advanced primary No comparison group immune response at intralesional injections of ID NR, protocol reviewed cutaneous T-cell injection site, systemic TG1042 are both safe by Swiss Agency for lymphomas, multilesional immune response and well tolerated. Therapeutic Products cutaneous B-cell Single-arm trial lymphomas N: 9 Treatments enrollment: topical steroids, PUVA, IFN-, acitretin, bexarotene, imiquimod, radiotherapy, methotrexate, antibiotics, rituximab Treatment requisite: none Follow-up: 32

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults TG1042 Tumor response Grade 3 adverse events: Translational research on Khammari et al, 2015241 Metastatic melanoma No comparison group (complete regression, flu-like symptoms (n = predictive markers did NCT00720031 Treatments enrollment: partial response, stable 1), asthenia (n = 3), not significantly differ disease, disease nausea/vomiting (n = 1), between responder and Single-arm trial Patients had to have progression), objective axillary pain (n = 1) nonresponder patients. N: 18 received at least one treatment with a response, disease control However, specifically chemotherapy or rate, overall survival, regarding injected immunotherapy. lesion size and number lesions, the clinical response correlated with Treatment requisite: CD3-/CD56+ NK cells blood draw 3 to 6 to that could be activated generate T-cell clone(s), by TG1042. Further treatment with deticene larger studies of this while T-cell clones were combined being produced, TG1042 immunotherapy are (adenovirus expressing needed to confirm these interferon-alpha) and findings. Intralesional Interleukin-2 were TG1042 combined with infused after each T-cell antigen-selected TILs clone infusion should be discussed. Follow-up: median: 30 Cancer Age group: adults TNFerade Overall survival, disease- SAEs: 3 patients Preoperative TNFerade, Chang et al, 2012146 Advanced resectable No comparison group free survival (unclear) in combination with GenVec, 2003426 esophageal cancer chemoradiotherapy is biologically active and NCT00051480 Treatments enrollment: none safe and is associated Trial (multiple groups) with long survival. This Treatment requisite: Completed regimen warrants chemoradiation, N: 24 additional studies. esophagectomy Follow-up: 60 Cancer Age group: adults TNFerade Death, progression-free SAEs (grades 3-4): 116 Standard of care plus Herman et al, 2013230 Locally advanced No treatment survival, tumor response intervention, 50 placebo TNFerade is safe but not NCT00051467 pancreatic cancer rates, surgical effective for prolonging downstaging rates survival in patients with RCT Treatments enrollment: LAPC. Completed none N: 304 Treatment requisite: standard of care Follow-up: 50.5

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cancer Age group: adults TNFerade Tumor response, safety No serious adverse This is the first human Senzer et al, 2004123 Solid tumors No comparison group events reported study with TNFerade and Trial (multiple groups) Treatments enrollment: radiation. The combined treatment was well N: 36 exhausted all prior treatment options tolerated in patients with predominantly prior Treatment requisite: treatment-refractory solid external beam radiation tumors. Follow-up: 3 Cancer Age group: adults Ad.hB7-1 Toxicity, survival, tumor No treatment-related The B7-1 modified Antonia et al, 2002125 Renal cell carcinoma No comparison group response serious adverse events autologous tumor cell Trial (multiple groups) Treatments enrollment: reported vaccine is safe and can BB-MF7553 none be combined with systemic IL-2 with N: 15 Treatment requisite: acceptable toxicity. interleukin 2, intradermal Immunological and injection of unmodified clinical responses were irradiated autologous observed in some tumor cells patients. Follow-up: 10 Cardiovascular disease Age group: adults Ad2/HIF-1alpha/VP16 Claudication onset time, None Gene therapy with Creager et al, 2011265 Intermittent claudication No treatment ankle-brachial index, intramuscular NCT00117650 Treatments enrollment: quality of life administration of Ad2/HIF-1 was not an RCT none effective treatment for Treatment requisite: N: 289 patients with intermittent none claudication. Follow-up: none

Cardiovascular disease Age group: adults Ad2/HIF-1alpha/VP16 Deaths and amputations None HIF-1 therapy in patients Rajagopalan et al, Critical limb ischemia Placebo with critical limb ischemia 260 2007 Treatments enrollment: was well tolerated, ID NR none supporting further, larger, randomized RCT Treatment requisite: efficacy trials. N: 28 none Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cardiovascular disease Age group: adults Ad5.hAC6 LVEF (echocardiography, SAE: 18 intervention, 5 AC6 gene transfer safely Hammond et al, 2016270 Heart failure Placebo before and during control; HF admission: 4 increased LV function Hammond, 2015427 Treatments enrollment: dobutamine infusion), LV intervention, 4 control; beyond standard heart death: 1 intervention, 1 failure therapy, NCT00787059 implanted cardiac peak +dP/dt control attainable with one-time RCT defibrillator (ICD) and at and peak –dP/dt before least 1 major coronary and during dobutamine administration. Larger N: 56 artery or graft with less infusion, exercise trials are warranted. than 50% proximal capacity assessed by obstruction exercise treadmill testing, Treatment requisite: symptoms, ICD events, intracoronary hemodynamic values nitroprusside (50 μg/min) obtained at catheteriz Follow-up: 3 Cardiovascular disease Age group: adults Ad5FGF-4 Reversible perfusion SAEs: 44 intervention, 22 Intracoronary injection of Grines et al, 2003256 Stable angina Placebo defect size, perfusion placebo Ad5FGF-4 showed an Cardium Therapeutics, Treatments enrollment: defect size encouraging trend for 2016428; Kaski and antianginal drugs improved myocardial perfusion; however, Consuegra-Sanchez, Treatment requisite: 429 further studies of 2013 none therapeutic angiogenesis ID NR. The study was Follow-up: 12 with Ad5FGF-4 will be approved by the Food necessary. and Drug Administration, Recombinant DNA Advisory Committee, appropriate institutional review boards and biosafety committees, and was conducted in accordance with the International Conference on Harmonization and Good Clinical Practice Guidelines. All patients provided written informed consent. RCT Completed N: 53

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cardiovascular disease Age group: adults Ad5FGF-4 Rest pain, use of SAEs: 14 total Injections of Ad5FGF-4 Matya et al, 2005257 Unreconstructable critical Placebo analgesics, leg were generally well ID NR limb ischemia ulcer/gangrene, ABI and tolerated and considered TBI, DSA, MRI, Tc-Scint safe. Transfection RCT Treatments enrollment: none efficacy at these Completed concentrations may have Treatment requisite: N: 13 been limited or local. The none small sample size did not Follow-up: 6 allow any firm conclusions regarding clinical efficacy, but a trend toward more and slightly larger blood vessels was observed in the angiograms. It is concluded that intramuscular injection of Ad5FGF-4 into CLI patients seemed safe, but transfection efficacy was limited at the assessed doses. Conclusions regarding clinical efficacy are impossible to draw from this small patient cohort. Cardiovascular disease Age group: adults Ad5-FGF4 None None Results show evidence of Grines et al, 2002251 Stable angina pectoris Placebo favorable anti-ischemic ID NR Treatments enrollment: effects with Ad5-FGF4 compared with placebo, RCT none and it appears to be Treatment requisite: N: 79 safe. Angiogenic gene none transfer with Ad5-FGF4 Follow-up: 3 shows promise as a new therapeutic approach to the treatment of angina pectoris.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cardiovascular disease Age group: adults Ad-5PDGF-B Ulcer wound area, ulcer SAEs (events): 3 In the 12 patients who Mulder et al, 2009249 Nonhealing diabetic foot No comparison group closure osteomyelitis, 1 septic completed the study, ID NR ulcers arthritis, 1 acute angle ulcer closure was glaucoma, 1 proteinuria, observed by month 3 in Trial (multiple groups) Treatments enrollment: none 1 hypersomnolence, 1 10 patients, 7 of whom Completed increased ammonia, 1 received a single Treatment requisite: N: 15 increased transaminases, application of GAM501. none 1 myocardial In conclusion, GAM501 Follow-up: 6 infarction, 1 cardiogenic did not appear to have shock, 2 gastrointestinal any toxicity at doses that disorders showed biological activity. GAM501 holds promise as a potentially effective treatment for nonhealing diabetic foot ulcers. Cardiovascular disease Age group: adults Ad5PDGF-B (GAM501) Acetate wound size, None Both GAM501 and FCG Blume et al, 2011264 Diabetic neuropathic foot Usual care wound healing rate appeared to be safe and NCT00493051 ulcers well tolerated, and alternate dosing RCT Treatments enrollment: none schedules hold promise Completed to improve overall Treatment requisite: N: 124 complete wound closure formulated collagen gel in adequately powered Follow-up: 3 trials. Cardiovascular disease Age group: adults Ad-HGF Left ventricular end- None Percutaneous endocardial Meng et al, 2018250 Postinfarction heart No treatment diastolic dimension, left administration of Ad-HGF NCT01925352 failure ventricular ejection is safe and potentially fraction, summed rest efficient in improving Controlled trial Treatments enrollment: none score of single photon LVEF and lowering LVDd N: 30 emission computed of patients with Treatment requisite: tomography postinfarct failure. none Follow-up: 6

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cardiovascular disease Age group: adults Ad-HGF Changes of vital None These preliminary clinical Yuan et al, 2008262 Coronary heart disease No comparison group symptoms, blood and data indicate direct ID NR (described as trial Treatments enrollment: urea routine analyses, intramyocardial in abstract) coronary artery bypass myocardial profusion administration of Ad-HGF was well tolerated and Single-arm trial surgery could improve myocardial Treatment requisite: N: 18 perfusion with a dose– none effect relationship, Follow-up: 1 encouraging larger and randomized efficacy trials. Cardiovascular disease Age group: adults Ad-hHGF Supraventricular or None, safe The study demonstrates Yang et al, 2009261 Severe and diffused No comparison group ventricular arrhythmias, it is feasible to safely use Yang et al, 2009430 triple-vessel coronary derum HGF levels, an adenovirus gene- hepatocyte growth transfer vector to deliver ID NR (title states trial) artery disease the human hepatocyte Single-arm trial Treatments enrollment: none growth factor gene to N: 18 individuals with clinically Treatment requisite: significant coronary none artery disease by direct Follow-up: 14 intracoronary injection. Cardiovascular disease Age group: adults AdVEGF121 Exercise treadmill test SAEs: 1 intervention, 0 Percutaneous, catheter- Fuchs et al, 2006259 Refractory advanced Placebo (ETT), Canadian placebo based AdVEGF121 ID NR coronary artery disease Cardiovascular Society intramyocardial injection Angina Classification, is a practical, feasible, RCT Treatments enrollment: none Seattle Angina and potentially safe Completed Questionnaire approach for Treatment requisite: N: 10 intramyocardial gene none transfer. A larger Follow-up: 12 randomized, phase II efficacy study is warranted.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cardiovascular disease Age group: adults AdVEGF121 Change in peak walking SAEs: 6 intervention, 5 A single unilateral Rajagopalan et al, Peripheral arterial Placebo time, claudication onset placebo intramuscular 2003255 disease time, postexercise ankle- administration of Rajagopalan et al, Treatments enrollment: brachial index intreated AdVEGF121 was not 2003431 stable medication limb, physical component associated with improved summary scale of SF-36, exercise performance or ID NR regimen health survey and quality of life in this Treatment requisite: RCT walking impairment study. This study does none Completed questionnaire not support local delivery N: 105 Follow-up: 6.5 of single-dose VEGF121 as a treatment strategy in patients with unilateral PAD. Cardiovascular disease Age group: adults AdVEGF121 Survival, angina score Incidences of malignancy Adenovirus-mediated Rosengart et al, 2013267; Coronary artery disease No comparison group and retinopathy were no VEGF direct myocardial Rosengart et al, 1999432; Treatments enrollment: greater than expected for administration is safe, Rosengart et al, 1999433 none the age-matched general and future larger trials to population. assess efficacy are NCT01174095 Treatment requisite: warranted. Trial (multiple groups) none N: 31 Follow-up: 132 Cardiovascular disease Age group: adults AdVEGF121 Total exercise time SAEs: 11 intervention, 11 Overall there was no Stewart et al, 2006258 Nonrevascularizable Usual care (ETT), onset of 1 mm placebo significant difference in ID NR ischemic heart disease additional ST-segment adverse events between depression while the 2 groups, despite the RCT Treatments enrollment: 2 exercising fact that procedure- Completed of the 3 long-acting nitrates, calcium-channel related events were seen N: 67 blockers, and beta- only in the thoracotomy blockers. group. Therefore, administration of Treatment requisite: AdVEGF121 by direct none intramyocardial injections Follow-up: 6.5 resulted in objective improvement in exercise- induced ischemia in patients with refractory ischemic heart disease.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cardiovascular disease Age group: adults AdVEGF-DDeltaNDeltaC Myocardial perfusion, Death (control n = 0, AdVEGF-DeltaNDeltaC Hartikainen et al, Refractory angina (AdVEGF-D) angina pectoris treated n = 3), ACS or gene therapy was safe, 272 2017 ; Treatments enrollment: Placebo symptoms (stress and at MI (control n = 1, feasible, and well Kuopio University, severe refractory angina rest), quality of life treated n = 3), stroke tolerated. Myocardial 434 (control n = 0, treated n perfusion increased at 1 2015 Treatment requisite: = 2) year in the treated areas NCT01002430, EudraCT none with impaired MPR at 003295-22 Follow-up: 12 baseline. Plasma Lp(a) RCT may be a potential N: 30 biomarker to identify patients that may have the greatest benefit with this therapy. Cardiovascular disease Age group: adults BIOBYPASS Improvements in None Direct intramyocardial Kastrup et al, 2011263 Refractory advanced (AdGVVEGF121.10NH) exercise capacity, time to injection of BIOBYBASS ID NR coronary artery disease Placebo ischemic threshold, (AdGVVEFG121.10NH) myocardial perfusion was safe but did not RCT Treatments enrollment: none improve exercise N: 17 capacity, time to Treatment requisite: ischemic threshold, or intramyocardial injection myocardial perfusion of therapy or placebo. compared with placebo Follow-up: 12 injection in refractory myocardial ischemia patients. Cardiovascular disease Age group: adults CI-1023; Resting ankle-brachial 1 death, 1 bladder CI-1023 appears to be Mohler et al, 2003254 Critical limb ischemia Ad(GV)VEGF121.10 indexes neoplasm, 1 increased well tolerated and safe ID NR Treatments enrollment: Placebo swelling for single-dose administration in patients Controlled trial none with critical limb ischemia Treatment requisite: N: 15 due to peripheral arterial none disease. Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cardiovascular disease Age group: children and VEGF-Ad, VEGF-P/L DSA analysis of SAEs: 5 intervention (2 For the primary endpoint, Makinen et al, 2002252 adults Placebo vascularity, restenosis groups), 1 placebo follow-up DSA revealed ID NR Lower-limb ischemia rate, Rutherford class, increased vascularity in ABI the VEGF-treated groups RCT Treatments enrollment: none distally to the gene Completed transfer site (VEGF-Ad P Treatment requisite: N: 54 = 0.03, VEGF-P/L P = none 0.02) and in the VEGF-Ad Follow-up: 36 group in the region of the clinically most severe ischemia (P = 0.01). As for the secondary endpoints, mean Rutherford class and ABI showed statistically significant improvements in the VEGF-Ad and VEGF-P/L groups, but similar improvements were also seen in the control patients. Catheter-mediated VEGF gene therapy is safe and well tolerated. Angiography demonstrated that VEGF gene transfer increased vascularity after PTA in both VEGF-Ad-, and VEGF-P/L-treated groups.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Cardiovascular disease Age group: adults VEGF-Adv, VEGF-P/L Minimal lumen diameter 1 death, 1 cancer, 1 Gene transfer with VEGF- Hedman et al, 2003253 Catheter-based local Placebo and percentage diameter sarcoidosal granulomas, Adv or VEGF-P/L during Hedman et al, 2009435 intracoronary vascular stenosis, myocardial 1 “no flow" PTCA and stenting shows perfusion, exercise phenomenon and intracoronary gene ID NR, protocol approved endothelial growth factor tolerance, incidence of cardiac arrest during transfer can be by the Finnish Board for gene transfer new cardiac events, control angiography, a performed safely (no Gene Technology and the Treatments enrollment: emergency or elective aspiratory pneumonia major gene transfer– Finnish Agency for none angiography or and pseudomembranotic related adverse effects Medicinal Products Treatment requisite: revascularization, colitis, 1 stroke, fever, were detected), no none RCT functional class, working increase in anti-Adv differences in clinical N: 103 Follow-up: 6 ability, need for peroral antibodies, transient restenosis rate or nitrates decrease in the platelet minimal lumen diameter count, increase in LDH, were present after the serum IL-6 level increase 6-month follow-up, and a significant increase was detected in myocardial perfusion in the VEGF-Adv–treated patients. Immune deficiency Age group: adults Ad26.ENVA.01 and Ad35- Immunogenicity of No vaccine-associated Baseline vector immunity Baden et al, 2016279 Prophylactic HIV-1 ENV vaccines, evaluation of serious adverse events did not markedly affect NCT01215149 vaccine Placebo anti-tumor immunity reported responses, and a induced by vaccines significant immune RCT Treatments enrollment: none response by both N: 218 vaccines was observed in Treatment requisite: all populations. Patients had to be willing to undergo HIV testing and accept the HIV test results, undergo risk- reduction counselling, and commit to low-risk behavior for the duration of the trial. Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Immune deficiency Age group: adults Ad26-EnvA Safety and tolerability, Safe This Ad26 vectored Baden et al, 2013275 HIV Placebo immunogenicity vaccine was generally NCT00618605 Treatments enrollment: safe and immunogenic at all doses tested. RCT none Reactogenicity was Treatment requisite: N: 60 minimal with doses of 5 none × 1010 vp or less. Ad26 Follow-up:12 is a promising new vaccine vector for HIV-1. Immune deficiency Age group: adults Ad35-GRIN Safety and tolerability None Coadministration of an Omosa-Manyonyi et al, HIV Other: 4 regimens: adjuvanted protein and 278 2015 Treatments enrollment: heterologous prime- an adenovirus vector NCT01264445 none boost with 2 doses of showed an acceptable safety and reactogenicity RCT Treatment requisite: F4/AS01E or F4/AS01B followed by Ad35-GRIN; profile and resulted in N: 146 none Ad35-GRIN followed by 2 strong, multifunctional, Follow-up: 16 doses of F4/AS01B; or 3 and complementary HIV- coadministrations of specific immune Ad35-GRIN and responses. F4/AS01B Immune deficiency Age group: adults Ad5HVR48.ENVA.01 Safety and tolerability, None Ad5HVR48.ENVA.01 was Baden et al, 2014276 HIV Placebo Ad5 nAb responses, Ad48 safe, well tolerated, and NCT00695877 Treatments enrollment: nAb responses, HIV-1 immunogenic at all doses Env-specific antibody tested. RCT none responses, HIV-1 EnvA- Treatment requisite: N: 48 specific cellular immune none responses Follow-up: 12 Immune deficiency Age group: adults AERAS-402/AD35.TB-S CD4+ T-cell response, SAES (events/patients): AERAS-402/AD35.TB-S Churchyard et al, 2015282 HIV Placebo CD8+ T-cell response, 1 lower-respiratory tract was well tolerated, safe, NCT01017536 Treatments enrollment: antibody response infection; 1 joint injury and induced predominantly RCT Bacillus Calmette-Guérin vaccination polyfunctional CD4+ and Completed CD8+ T-cell responses to Treatment requisite: N: 26 the vaccine. none Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Immune deficiency Age group: adults rAd5 Vaccine safety, None The higher frequency of Koblin et al, 2011274 HIV No comparison group immunogenicity local reactions after ID NCT00384787 Treatments enrollment: and SC administration and the lack of sufficient Trial (multiple groups) none evidence to show there Treatment requisite: N: 90 were any differences in none immunogenicity by route Follow-up: 12 of administration do not support changing route of administration for the rAd5 boost. Immune deficiency Age group: adults rAd5 CD4+ T-cell response, None Heterologous prime- Koup et al, 2010281 HIV No comparison group CD8+ T-cell response, boost using vector-based NCT00102089 and Treatments enrollment: antibody response gene delivery of vaccine NCT00108654 none antigens is a potent immunization strategy for Trial (multiple groups) Treatment requisite: 4 to inducing antibody and T- 6 plasmid DNA Completed cell responses. N: 14 Follow-up: 6 Immune deficiency Age group: adults rAd5 Analytic treatment None Therapeutic vaccination Li et al, 2011273 HIV Placebo interruption, plasma viral with an rAd5-HIV gag Schooley, 2010436 Treatments enrollment: load vaccine was associated with lower ATI week 16 NCT00080106 pre-antiretroviral therapy PVL even after Treatment requisite: ART RCT controlling for viral and if needed Completed host genetic factors. N: 110 Follow-up: 16 Immune deficiency Age group: adults rcAd26.MOS1.HIV-Env Vaccine safety, vaccine- None The highly attenuated Stephenson et al, 2018280 HIV-1 Placebo specific immune rcAd26.MOS1.HIV-Env NCT02366013 Treatments enrollment: responses, viral shedding vaccine was well tolerated up to 10^11 VP RCT none in healthy, HIV-1- Treatment requisite: N: 33 uninfected adults, though none the single dose was Follow-up: 8 poorly immunogenic, suggesting the replicative capacity of the vector was too attenuated.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Immune deficiency Age group: adults rAd5 Change in total HIV DNA 2 increase in creatine Antiretroviral therapy Achenbach et al, 2015277 HIV Other: antiretroviral in the peripheral blood, kinase, 1 acute coronary intensification followed NCT00976404 Treatments enrollment: therapy intensification adverse events syndrome, 1 acute by DNA prime and rAd5 coronary syndrome boost vaccine did not RCT none plus injections of HIV significantly increase HIV Treatment requisite: oral DNA prime vaccine N: 28 expression or reduce the raltegravir 400 mg and latent HIV reservoir. oral maraviroc 150 mg, 300 mg, or 600 mg Follow-up: 2 Neurodegenerative Age group: adults Ad5-VEGF Change in neurological No serious undesirable The life span of patients disorders Amyotrophic lateral Placebo deficit, change in vital events observed under conditions of Zavalishin et al, 2008297 sclerosis lung capacity, change in hypoxia increased after ID NR Treatments enrollment: muscle force of 36 treatment with the test muscles, improvement in drug, which was RCT none survival probably related to Treatment requisite: N: 10 improved resistance of none motoneurons. The Follow-up: 24 presence of virus- neutralizing antibodies decreases the effectiveness of adenoviral vectors, which necessitates a differential approach to the selection of patients and continuous monitoring of gene therapy.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Ocular disorders Age group: adults AdPEDF.11 Safety, tolerability, visual Eye pain (n = 1) These data suggest the Campochiaro et al, Age-related macular No comparison group acuity possibility of 2006314 degeneration antiangiogenic activity ID NR Treatments enrollment: that may last for several months after a single Trial (multiple groups) none intravitreous injection of Treatment requisite: N: 28 doses greater than 108 topical anesthetic PU of AdPEDF.11. This injection under the study provides evidence conjunctiva that adenoviral vector- Follow-up: 12 mediated ocular gene transfer is a viable approach for the treatment of ocular disorders and that further studies investigating the efficacy of AdPEDF.11 in patients with neovascular AMD should be performed. Other: normal patients Age group: adults Ad(GV)CD.10 Vector persistence, skin Not reported Intradermal Harvey et al, 1999355 Healthy patients No comparison group induration, systemic anti- administration of an E12 BB-IND 6950 Treatments enrollment: adenovirus immunity E32 Ad vector to normal against Ad5 patients induces Trial (multiple groups) none mild/moderate local Treatment requisite: N: 6 cellular responses, even Each patient received in Ad-immunized one intradermal individuals, and these administration of the observations provide a vector in each of 2 sites baseline to determine if + skin biopsy. human anti-Ad vector Follow-up: 1 host responses can be circumvented by using other means.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Other: healthy adults Age group: adults Ad35.CS.01 Solicited local reactions, While the vaccine was The Ad35.CS.01 vaccine Creech et al, 2013342 Malaria (vaccine) Placebo overview of serious generally well tolerated, was most immunogenic NCT00371189 Treatments enrollment: adverse events and adverse events were at the highest dosages laboratory adverse more frequent in the (1010 and 1011 vp/mL). RCT none events, unsolicited highest-dose groups Treatment requisite: N: 72 adverse events, (1010 and 1011 vp/mL). none immunogenicity (humoral More robust humoral Follow-up: 1 response) responses were also noted at the highest doses, with 73% developing a positive ELISA response after the 3-dose series of 1011 vp/mL. Other: healthy adults Age group: adults Ad4-H5-Vtn Seroconversion by None Oral Ad4 vector priming Gurwith et al, 2013341 Influenza Placebo haemagglutination- might enhance the NCT01006798 Treatments enrollment: inhibition efficacy of poorly immunogenic vaccines RCT none such as H5N1. N: 166 Treatment requisite: none Follow-up: 6 Other: ebola vaccine Age group: adults Ad-5 Ebola glycoprotein- Safe The adenovirus 5- Li et al, 2017348 Ebola Placebo specific ELISA antibody vectored ebola vaccine of Zhu et al, 2015437 Treatments enrollment: responses 28 days post 1·6 × 1011 viral particles boost and the was highly immunogenic NCT02326194 and none occurrences of adverse and safe. NCT02533791 Treatment requisite: reactions post boost RCT none N: 120 Follow-up: 5 Other: healthy adults Age group: adults Ad-5 Adverse reactions, Safe The recombinant Zhu et al, 2017347 Ebola Placebo serious adverse events, adenovirus type 5 vector- PACTR201509001259869 Treatments enrollment: HIV infection rate based ebola vaccine was safe and highly RCT none immunogenic in healthy Treatment requisite: N: 500 Sierra Leonean adults, none and 8·0 × 1010 viral Follow-up: 6 particles was the optimal dose.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Other: partial ornithine Age group: adults Ad-OTC Toxicity Myalgia and flulike The low levels of gene transcarbamylase Ornithine No comparison group symptoms, fever, transfer detected in this deficiency transcarbamylase anemia, trial suggest that at the Raper et al, 2002357 deficiency thrombocytopenia, doses tested, significant Raper et al, 2003438 Treatments enrollment: 1 case of fatal systemic metabolic correction did not occur. However, ID NR, protocol reviewed none inflammatory response substantial subject-to- by NIH-RAC and FDA Treatment requisite: syndrome subject variation in host none Single-arm trial responses is a limitation. N: 18 Follow-up: 18 Other: healthy/infectious Age group: adults ChAd3-EBO-Z CD4 T-cell response, CD8 SAEs (events): deployed: ChAd3-EBO-Z was safe disease vaccine Health volunteers Placebo T-cell response, antibody 2 headache, 1 fever, 3 and well tolerated, 346 De Santis et al, 2016 Treatments enrollment: response other; nondeployed: 0 although mild to NCT02289027 none moderate systemic adverse events were RCT Treatment requisite: common. A single dose none Completed was immunogenic in N: 120 Follow-up: 6 almost all vaccine recipients. Antibody responses were still significantly present at 6 months. There was no significant difference between doses for safety and immunogenicity outcomes. This acceptable safety profile provides a reliable basis to proceed with phase II and phase III efficacy trials in Africa. Other: uncertain Age group: adults ChAd3-EBO-Z Safety, measured with No AEs 1×1011 pu single-dose Tapia et al, 2016345 Ebola Placebo occurrence of adverse ChAd3-EBO-Z could NCT02231866 and Treatments enrollment: events suffice for phase III NCT02267109 none efficacy trials of ring- vaccination containment RCT Treatment requisite: needing short-term, none N: 91 high-level protection to Follow-up: 0 interrupt transmission.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Other: malaria vaccine Age group: children ChAd63 Safety No AEs The vaccines were well Mensah et al, 2017350 Malaria (vaccine) Usual care tolerated in all age NCT02083887 Treatments enrollment: groups, with no vaccine- related serious AEs. RCT none N: 65 Treatment requisite: none Follow-up: 0 Other: health/infectious Age group: adults ChAd63 Parasite concentration in No SAE reported in text The results demonstrate disease vaccine Healthy volunteers Other: infected controls blood, malaria diagnosis (only in figures) immune interference 340 Sheehy et al, 2012 Treatments enrollment: whereby responses Controlled trial none against merozoite surface protein 1 are Completed Treatment requisite: MVA dominant over apical MSP1 N: 44 membrane antigen 1 and Follow-up: 2 ME-TRAP. The results show induction of strong cellular immunity against MSP1 and AMA1 is safe but does not impact on parasite growth rates in the blood. The data call into question the utility of T cell–inducing blood- stage malaria vaccines and suggest the focus should remain on high- titer antibody induction against susceptible antigen targets. Other: malaria vaccine Age group: adults ChAd63 Adverse events, None reported The results demonstrate Venkatraman et al, Malaria No comparison group Immunogenicity Matrix-M can be safely 349 2017 Treatments enrollment: used in combination with NCT01669512 none ChAd63-MVA ME-TRAP heterologous prime- Trial (multiple groups) Treatment requisite: boost immunization none N: 23 without any reduction in Follow-up: 4 cellular or humoral immunogenicity.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Other: none Age group: adults ChAd63-KH None None ChAd63-KH was safe, Osman et al, 2017354 Healthy volunteers No comparison group and whole-blood ISRCTN 07766359 Treatments enrollment: transcriptomic profiling indicated ChAd63-KH Trial (multiple groups) none induced innate immune Treatment requisite: Completed responses characterized none N: 20 by an interferon Follow-up: 3 signature and the presence of activated dendritic cells. Broad and quantitatively robust CD8 + T-cell responses were induced by vaccination in 100% of vaccinated patients. Other: influenza Age group: adults ChAdOx1 Safety No treatment-related ChAdOx1 NP + M1 Antrobus et al, 2014344 Influenza No comparison group serious adverse events appears to be safe and Coughlan et al, 2012439 Treatments enrollment: reported immunogenic and may be a promising vaccine Trial (multiple groups) none vector that could be used Treatment requisite: N: 15 to deliver vaccine none antigens where strong Follow-up: 4 cellular immune responses are required for protection. Other: loosened hip Age group: adults CTL 102 Safety, tolerability, Two patients in the high- These data show this prosthesis Loosened hip prosthesis No comparison group walking distance, patient prodrug dose group had gene therapy approach 358 de Poorter et al, 2008 Treatments enrollment: satisfaction grade 2 rises in AST and targeted at the interface de Poorter et al, 2008440 arthrogram of the hip ALT levels and 2 had membrane around a diarrhea. loosened hip prosthesis is ID NR, protocol approved with aspiration of joint a feasible treatment by the Central Committee fluid to exclude bacterial option for elderly on Research Involving infection patients for whom Human Subjects (CCMO Treatment requisite: hip surgical intervention is and Ministry of Housing, synovial fluid aspiration, not appropriate. Spatial Planning, and the joint rinsed to remove Environment (VROM, The neutralizing antibody Netherlands) Follow-up: 6 Single-arm trial N: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Other: venous leg ulcer Age group: adults H5.020CMV.PDGF-beta Wound response, routine The injections were well The study demonstrated disease Venous leg ulcer disease No comparison group blood testing, H5 tolerated and patients that H5.020CMV.PDGF-B 359 Margolis et al, 2009 Treatments enrollment: adenovirus detection, were able to use a was able to transfect Margolis et al, 2004441 none PDGF antibody, detection compression bandage cells in the wound. of adenoviral DNA within during the 28 days of the ID NR, protocol reviewed Treatment requisite: wound bed, assessment initial study period. by FDA none of wound Single-arm trial Follow-up: 6 neovascularization, N: 15 assessment of tissue inflammation, assessment of progenitor cell recruitment Other: healthy/infectious Age group: adults MRKAd5 HIV infection None The increased HR of HIV- disease vaccine Healthy volunteers Placebo 1 acquisition, irrespective 343 Gray et al, 2014 Treatments enrollment: of number of doses Gray et al, 2011442 none received, warrants further investigation to RCT Treatment requisite: understand the biological none Completed mechanism. Further use N: 800 Follow-up: 78 of the Ad5 vector for HIV vaccines is not warranted. Other: infectious disease Age group: adults NMRC-M3V-D/AdPfCA Development of SAEs (events/patients): 1 The DNA/Ad regimen Chuang et al, 2013352 Malaria (DNA plasmids + Ad parasitemia, antibody pulmonary embolism provided the highest NCT00870987 Treatments enrollment: vector) response sterile immunity achieved against malaria following Controlled trial none Other: infected with immunization with a Treatment requisite: malaria Completed gene-based subunit none N: 26 vaccine (27%). Follow-up: 1 Protection was associated with cell- mediated immunity to AMA1, with CSP probably contributing. Substituting a low seroprevalence vector for Ad5 and supplementing CSP/AMA1 with additional antigens may improve protection.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Other: adults Age group: adults rAd5 Vaccine safety, ebola GP- None The vaccine was well Ledgerwood et al, Ebola Placebo specific antibody tolerated by patients in 339 2010 Treatments enrollment: responses, ebola GP- both dose groups NCT00374309 none specific T-cell responses, regardless of preexisting adenovirus 5-specific Ad5 antibody status. RCT Treatment requisite: antibody and T-cell none N: 31 responses Follow-up: 1 Other: healthy volunteers Age group: adults VRC-HIVADV038-00-VP None SAEs (events/patients): Nonserious AEs were Walsh et al, 2016353 Healthy volunteers (rAd5-EnvA), VRC- 1 severe neutropenia, 1 reported by patients NCT01095224 Treatments enrollment: HIVADV052-00-VP (rAd5- upper UTI; among the during the study duration EnvB), and VRC- AEs, the vast majority and coded for their Trial (multiple groups) none HIVADV027-00-VP (93.7%) were judged to relationship to the study Treatment requisite: Completed (rAd35-EnvA) be not related to the product (related or not none N: 180 No comparison group study products; most AEs related). Follow-up: 3 judged to be related to vaccination were local injection site reactions; 2 SAEs occurred during the study Other: norovirus Age group: adults VXA-G1.1-NN Antibody responses to No grade 3 or 4 solicited This vaccine was well Kim et al, 2018351 Norovirus Placebo norovirus VP1, memory adverse events tolerated and generated NCT02868073 Treatments enrollment: and antibody-secreting substantial immune cell responses, activation responses, and these RCT none properties of B cells results represent a major Treatment requisite: N: 66 step forward for none development of a safe Follow-up: 12 and novel norovirus vaccine. Respiratory conditions Age group: adults Ad CFTR Toxicity, lung response, No treatment-related No significant changes in Bellon et al, 1997327 Cystic fibrosis No comparison group hematological, adverse events reported immunological and Trial (multiple groups) Treatments enrollment: biochemical, inflammatory parameters inflammatory, were observed. N: 6 Confirmation of adenoviral Ela-like DNA immunological, sequences were in nasal virological parameters, or bronchial airway. safety Treatment requisite: none Follow-up: 15

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Respiratory conditions Age group: children and Ad(GV)CFTR.10 Not discussed No significant adverse This impressive level of Harvey et al, 1999331 adults No comparison group effects clearly expression is linked to ID NR Cystic fibrosis attributable to therapy the challenging fact that expression is limited in Trial (multiple groups) Treatments enrollment: none time. Although this can N: 14 be initially overcome by Treatment requisite: repetitive administration, baseline biomarker unknown mechanisms assessment, biomarkers eventually limit this collected at various time strategy, and further points repetitive administration Follow-up: 8 does not lead to repetitive expression. Respiratory conditions Age group: children and Ad2/CFTR Pulmonary function No serious adverse Vector administration Joseph et al, 2001333 adults No comparison group testing, reduction in reactions was tolerated but Perricone et al, 2001443 Cystic fibrosis bronchiectasis on CT transfer of cystic fibrosis scan (48 hours post- transmembrane ID NR Treatments enrollment: none treatment compared with conductance regulator Trial (multiple groups) baseline) (CFTR) cDNA was Treatment requisite: N: 36 inefficient and transgene bronchoscopic delivery (n expression was transient = 20) or aerosol delivery for the doses and (n = 16) of gene therapy method of Follow-up: 12 administration, as evidenced by no consistent change in pulmonary function.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Respiratory conditions Age group: adults Ad2/CFTR-2 Measurement of serum No adverse events The vector partially Zabner et al, 1996330 Cystic fibrosis No comparison group chemistries and blood observed that could be corrected the defect in Welsh et al, 1995444 Treatments enrollment: counts, pulmonary attributed to vector airway epithelial Cl- function tests, sputum transport in some ID NR none cultures patients, with variability Treatment requisite: Trial (multiple groups) observed between vector applied to nasal N: 6 patients and less epithelium with a correction with modified Foley catheter subsequent placed beneath the administration. Future inferior turbinate; work must focus on measurements (with vectors with increased associated perfusions) efficiency and with the were done with a rubber ability to evade host catheter telescopically defenses. introduced into the nostril Follow-up: 1 Respiratory conditions Age group: adults Ad5-CB-CFTR None Toxic effects (n = 2) In patients with cystic Knowles et al, 1995329 Cystic fibrosis No comparison group fibrosis, adenoviral ID NR, approved by NIH- Treatments enrollment: vector–mediated transfer RAC and FDA none of the CFTR gene did not correct functional defects Single-arm trial Treatment requisite: in nasal epithelium, and none N: 12 local inflammatory Follow-up: 1 responses limited the dose of adenovirus that could be administered to overcome the inefficiency of gene transfer.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Comparator Design Treatments N Months of Follow-up Respiratory conditions Age group: adults AdCFTR Electric potential No adverse effects were An adenovirus vector to Crystal et al, 1994328 Cystic fibrosis No comparison group difference between nasal observed with AdCFTR transfer and express the Hay et al, 1995445 Treatments enrollment: epithelium surface and administration to the CFTR cDNA in the subcutaneous tissues nasal epithelium. One respiratory epithelium of ID NR none patient needed treatment individuals with cystic Treatment requisite: All Single-arm trial after AdCFTR fibrosis is feasible, and patients underwent N: 4 administration to the correction of the cystic baseline (1 week), bronchial epithelium (2-4 fibrosis phenotype in the vehicle (1 week), and weeks for complete airway epithelium may treatment. patients were recovery). be achieved with this isolated for 2 days prior strategy. to treatment, and vehicle/treatment was administered to the nasal epithelium by syringe and catheter on day 1 and on days 2 to 3, vehicle/treatment was administered by bronchoscope to the bronchial epithelium

Follow-up: 12 Respiratory conditions Age group: adults AdGVCD.10 Systemic and mucosal No serious adverse These results suggest Harvey et al, 2001326 Cystic fibrosis No comparison group humoral and cellular events reported that, while adaptive anti- Trial (multiple groups) Treatments enrollment: immune responses, Ad immune responses safety likely play some role in N: 6 fiber-optic bronchoscopy, atropin, meperidine, the disappearance of the midazolam, lidocaine vector DNA following vector administration to Treatment requisite: the human lung, other none mechanisms may also be Follow-up: 2 involved in the response of humans to Ad gene transfer vectors.

Abbreviations: AAV, adeno-associated virus therapy; AEs, adverse events; ID, trial identification number; mL, milliliter; NR, not reported; RCT, randomized controlled trial; SAEs, serious adverse events.

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Appendix Table A2. Published AAV Trials Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Blood disorders Age group: adults AAV-1-LPL(S447X) Transient elevation of None Four of 8 patients had Mingozzi et al, 2009118 Lipoprotein lipase No comparison group CPK following vector detectable T-cell ID NR, protcol reviewed deficiency administration, responses to capsid by NIH Treatments enrollment: activation of capsid T- with dose-dependent cell responses, MHC kinetics of appearance. Single-arm trial none class I and II epitopes Patients with detectable Treatment requisite: N: 8 T-cell responses to none capsid also had higher Follow-up: 31 anti-AAV-1 IgG3 antibody titer. No subject developed B- or T-cell responses to the LPL transgene product. These findings suggest that T-cell responses directed to the AAV-1 capsid are dose- dependent. Blood disorders Age group: adults AAV2-FIX FIX expression Not reported These results Jiang et al, 2006117 Severe hemophilia B No comparison group demonstrate, for the ID NR protcol reviewed Treatments enrollment: first time, multiyear FIX by NIH none expression by AAV2 vector in humans and Single-arm trial Treatment requisite: suggest improved none N: 8 muscle delivery Follow-up: 10 provides effective treatment for protein deficiencies or muscle- specific diseases.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Blood disorders Age group: adults AAV5-hFVIII-SQ Factor VIII use, number Progression of chronic AAV5-hFVIII-SQ was Rangarajan et al, Hemophilia A No comparison group of bleeding episodes arthropathy in 1 patient associated with 121 2017 Treatments enrollment: persistent normalization BioMarin, 2022446 Eight patients had been of factor VIII activity level over 1 year in 6 of NCT02576795 receiving regular factor 7 patients who received Trial (multiple groups) VIII prophylaxis before the study. One patient a high dose, with N: 9 used on-demand factor stabilization of VIII for bleeding hemostasis and a episodes. profound reduction in factor VIII use in all 7 Treatment requisite: patients. glucocorticoids if the alanine aminotransferase level increased from the individual participant’s baseline level Follow-up: 12 Blood disorders Age group: adults AMT-060 Treatment-related None A single infusion of Miesbach et al, 2018116 Hemophilia B No comparison group adverse events, NAbs to AMT-060 had a positive UniQure Biopharma, Treatments enrollment: AAV5, total safety profile and 2015447 none immunoglobulin M resulted in stable and (IgM) and IgG clinically important NCT02396342; EudraCT Treatment requisite: antibodies against increases in FIX activity, 2013-005579-42 none AAV5, AAV5 capsid- a marked reduction in Single-arm trial Follow-up: 12 specific T cells, spontaneous bleeds and N: 10 antibodies to FIX FIX concentrate use, (including inhibitors), without detectable shedding of AMT-060 cellular immune vector, inflammatory responses against markers capsids.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Blood disorders Age group: adults DTX101; AAVrh10FIX EuroQoL-5D-5 level SAEs: 1 patient None Ultragenyx Hemophilia B No comparison group questionnaire, (musculoskeletal and Pharmaceutical Inc, Treatments enrollment: haemophilia-specific connective tissue 115 2017 none quality of life disorders) questionnaire NCT02618915 Treatment requisite: Trial (multiple groups) none 11/2018 Follow-up: 12 N: 6 Blood disorders Age group: adults rAAV2/5-PBGD Immune responses, Four severe adverse RAAV2/5-PBGD D’Avola et al, 2016120 Acute intermittent No comparison group viral shedding, events observed in 2 administration is safe, Digna Biotech, 2014448 porphyria aminolevulinic acid and patients: pilonidal cyst but AIP metabolic porphobilinogen levels, surgery, hospital correction was not NCT02076763, Treatments enrollment: requirement of AIP- admission for insulin- achieved at the doses NCT02082860 none specific therapies and pump infusion tested in the study. (experimental phase) Treatment requisite: hospitalizations, placement, myocarditis, However, the treatment none Trial (multiple groups) depression, anxiety, upper abdominal pain had a positive impact in N: 8 Follow-up: 12 quality of life, liver clinical outcomes in transduction most patients. Blood disorders Age group: adults scAAV2/8-LP1-hFIXco Increase in steady-state Unlikely related: The infusion of a single Nathwani et al, 2014119 Severe hemophilia B No comparison group transgenic factor IX staphylococcus septic dose of AAV8 vector Nathwani et al, 2011449 Treatments enrollment: activity, reduction or arthritis infection (grade resulted in long-term elimination of use of 4, n = 1); therapeutic factor IX NCT00979238 none factor IX concentrate, supraventricular expression associated Treatment requisite: Trial (multiple groups) reduction in the number tachycardia (grade 4, n with clinical none N: 10 of bleeding episodes = 1); improvement. Follow-up: median = 38 related to treatment: increase in alanine aminotransferase (grade 3, n = 1)

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Blood disorders Age group: adults SPK-9001 Change in annualized No serious adverse The study found George et al, 201712 Hemophilia B No comparison group bleeding rate, factor IX events observed sustained therapeutic Pfizer, 2019450 Treatments enrollment: concentrate expression of factor IX consumption, number coagulant activity after NCT02484092 Patients were required of infusions gene transfer in 10 Single-arm trial to have ≥50 prior exposure days to any patients with N: 10 factor IX (FIX) protein hemophilia who products in their received the same lifetime. vector dose. Transgene derived factor IX Treatment requisite: coagulant activity Patients received a enabled the termination 100% correction dose of baseline prophylaxis of their FIX product and the near immediately before elimination of bleeding transfusion of gene and factor use. therapy. Follow-up: 12 (invited to enroll in long-term follow-up protocol after 12 months) Cancer Age group: adults rAAV-DC Complete remission, None These preliminary data Di, 2012147 Cancer No comparison group partial remission, minor suggest that the rAAV- ID NR Treatments enrollment: remission, stable DC immunotherapy is disease, progressive well tolerated and Single-arm trial none disease, progression- showed no severe Treatment requisite: Completed free survival adverse reactions in none N: 27 cancer patients. Follow-up: 9

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Cancer Age group: adults TNFerade Tumor response, safety Grade 4 wound TNFerade and radiation Mundt et al, 2004122 Soft tissue sarcoma No comparison group infection and therapy were well Trial (multiple groups) Treatments enrollment: gangrenous necrosis (n tolerated in the = 2) treatment of patients N: 14 none with soft-tissue Treatment requisite: sarcoma of the external beam extremity. The number radiotherapy of objective responses Follow-up: 2 observed warrants additional studies of this approach in a larger controlled prospective trial. Cardiovascular disease Age group: adults AAV1/SERCA2a LV end-systolic No safety issues noted At 6 months, LVESV Hulot et al, 2017271 Systolic heart failure Placebo volumes, LV end- was increased in both None Treatments enrollment: diastolic volume, groups compared with ejection fraction, baseline: median NCT01966887 Patients had undergone echocardiography, peak (interquartile range) in RCT visually complete resection of papillary VO2 max, NT-proBNP AAV1/SERCA2a vs N: 9 lesions. Patients could placebo: 13 (13;14) mL not have received vs 3.5 (−36;36) mL, P intravesical therapy = 0.74, with a mean within 3 months before difference between beginning study groups of 11.4 mL in treatment, with the favor of placebo. exception of cytotoxic agents when administered as a single instillat Treatment requisite: none Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Cardiovascular disease Age group: adults AAV1/SERCA2a Time to recurrent All-cause hospital Despite promising Greenberg et al, 2016269 Cardiac disease Placebo events (hospital admission: placebo: results from previous Greenberg et al, 2014451 Treatments enrollment: admission because of 240; intervention: 172 studies, AAV1/SERCA2a heart (100); heart failure- at the dose tested did NCT01643330 receiving optimum failure or ambulatory related hospital not improve the clinical RCT tolerated stable medical treatment for at least treatment for worsening admission: placebo: course of patients with Completed 30 days heart failure), time to 121; intervention: 99; heart failure and N: 250 ambulatory treatment reduced ejection Treatment requisite: first terminal event (all- for worsening heart fraction. Although the none cause death, heart transplantation, failure: placebo: 7; study did not find Follow-up: 12 implantation of a intervention: 8; nonfatal evidence of improved durable mechanical myocardial infarctions: outcomes at the dose of circulatory support devi) placebo: 5; AAV1/SERCA2a studied, intervention: 3; Non- the findings should fatal strokes: Placebo: stimulate further 3; Intervention: 5; research. Heart transplantation Placebo: 4; Intervention: 7; Durable MCSD implant; Placebo: 8; Intervention: 7; Deaths: Placebo: 20; Intervention: 25 Cardiovascular disease Age group: adults AAV1/SERCA2a Mortality, None After a single Zsebo et al, 2014268 Advanced heart failure Placebo hospitalization, intracoronary infusion Jessup et al, 2011452; Treatments enrollment: symptomatic efficacy of AAV1/SERCA2a in Jaski et al, 2009453; none patients with advanced 454 heart failure, positive Hjjar et al, 2008 ; Treatment requisite: 455 signals of Celladon, 2010 ; none Celladon, 2015456 cardiovascular events Follow-up: 24 persist for years. NCT00454818 RCT N: 39

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Cardiovascular disease Age group: adults Alipogene tiparvovec Reduction in fasting High fever (39.9 ºC) This is the first Gaudet et al, 2013266 Lipoprotein lipase (AAV1-LPLS447X) median plasma postinjection (n = 1); demonstration of a Stroes et al, 2008457 deficiency No comparison group triglyceride long-term follow-up had single gene therapy 6 serious adverse intervention leading to NCT01109498, CT-AMT- Treatments enrollment: events, including a persistent transgene 011–01 none patient who had a expression and Treatment requisite: Trial (multiple groups) cardiac arrest leading to sustained clinical controlled low-fat diet N: 14 death but none of the benefit in a systemic as part of serious AEs were metabolic disorder. preobservational study related to alipogene (low-fat diet continued tiparvovec during study) administration Follow-up: 24 Immune deficiency Age group: adults tgAAC09 Change in 5 serious adverse Two doses of tgAAC09 Vardas et al, 2010283 HIV-1 Placebo immunogenicity events, all considered were well tolerated at Mehendale et al, Treatments enrollment: unrelated to the the dosage levels given. 2010458; International none therapy (1 in the Fewer than half the placebo group): malaria recipients of the highest AIDS Vaccine Initiative, Treatment requisite: 459 (n = 1), preeclampsia vaccine dosage, 3 1012 2007 contraception (4 (n = 2), inguinal hernia DRP, had T-cell NCT00482027 (also months), HIV testing, (n = 1), anemia due to responses to HIV. NCT00888446) counseling (pre-HIV a bleeding ulcer (n = and post-HIV test, HIV RCT 1), death due to viral risk reduction) N: 91 encephalitis (n = 1), Follow-up: 18 occurred in the low dose group)

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Inflammatory disorders Age group: adults AAV1-LPL(S447X)) Effect on fasting plasma None Intramuscular Carpentier et al, 2012287 Lipoprotein lipase No comparison group metabolites, hormones, administration of Ferreira et al, 2014460 deficiency (LPLD) and lipids; postprandial alipogene tiparvovec NCT00891306 Treatments enrollment: glucose, insulin, and C- resulted in a significant peptide and total improvement of Single-arm trial none plasma and chylomicron postprandial Treatment requisite: N: 5 TG levels; postprandial chylomicron metabolism none glucose, insulin, and C- in LPLD patients, Follow-up: 3 peptide and total without inducing large plasma and chylomicron postprandial NEFA TG levels; postprandial spillover. plasma NEFA and glycerol levels and appearance rates Inflammatory disorders Age group: adults rAAV2-TNFR:Fc Improvement in tender 4 SAEs requiring It is feasible to proceed Mease et al, 2009285 Inflammatory arthritis Placebo joint count (68 joints), hospitalization in the with larger trials to Targeted Genetics, Treatments enrollment: improvement in swollen same participant: further test the safety 2005461 Patients with joint count (66 joints), labyrinthitis, myocardial and efficacy of local improvement in infarction, and 2 TNFR:Fc gene transfer NCT00617032 rheumatoid arthritis had patient’s self episodes of congestive as a therapeutic RCT to have an adequate trial of at least 1 assessment of pain, heart failure; thought to modality for patients N: 15 DMARD prior to improvement in patient be unrelated to with inflammatory screening and couldn't and physician global treatment arthritis. be taking a TNF-α assessment of disease antagonist. activity, improvement in patient’s Treatment requisite: none assessment Follow-up: 6

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Inflammatory disorders Age group: adults rAAV2-TNFR:Fc Decrease in target joint Ten patients in the Patient-reported Mease et al, 2010286 Inflammatory arthritis Placebo swelling, decrease in gene therapy outcome measures of ID NR with peripheral joint target joint tenderness, experienced 12 serious clinical response improvement in target adverse events. showed greater RCT involvement joint global visual Unrelated (n = 1 for improvement in treated N: 127 Treatments enrollment: rheumatoid arthritis: analog scale, each): degenerative patients than in placebo failure or inadequate improvement in joint joint disease requiring L patients. response to at least 1 function visual analog knee replacement, disease-modifying scale, improvement in infected incision after antirheumatic drug target joint pain repair of traumatic ankle fracture, syncope Treatment requisite: from coronary artery none disease. Unlikely related Follow-up: Unclear— (n = 1 for each): likely at least 7 months abdominal pain from but no median value is constipation, cellulitis given and ulcer leg calf, acute pyelonephritis, myocardial infarction, massive pulmonary emboli (fatal), pneumonia, disseminated histoplasmosis + retroperitoneal hemorrhage (fatal). Probably related (n = 1 for each): septic arthritis of injected joint.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Inflammatory disorders Age group: adults tgAAC94 Serious adverse events, 1 reported death The case study Targeted Genetics Inflammatory arthritis Placebo, usual care severe or very severe reinforces the 284 Corporation, 2008 Treatments enrollment: adverse events, importance of Frank et al, 200960 none study drug–related considering infectious 462 complications. Heald et al, 2010 Treatment requisite: adverse events NCT00126724 none RCT Follow-up: 4 10/2008 N: 120 Muscular conditions Age group: children and AAV1.des.h(gamma) Tolerability of None The protein expression Herson et al, 2012289 adults SGC AAV1.des.h can be induced in Genthon, 2010463 Limb girdle muscular No comparison group patients with limb girdle muscular dystrophy NCT01344798 dystrophy type 2C type 2C by adeno- Single-arm trial Treatments enrollment: none associated virus N: 9 serotype 1 gene Treatment requisite: transfer with no serious none adverse effects. Follow-up: 6 Muscular conditions Age group: children AAV1-CMV-GAA Maximal inspiratory 1 slight increase in Combined AAV1-CMV- Smith et al, 2017290 Infantile-onset Pompe No comparison group pressure, respiratory antiacid alpha- GAA and exercise Smith et al, 2013362 disease responses to inspiratory glucosidase, 1 right training conferred Byrne et al, 2014464 Treatments enrollment: threshold loads (load lung contusion with benefits to dynamic Corti et al, 2017465 had chronic respiratory compensation), physical apical pneumothorax motor function of the evidence of diaphragm (resolved), 1 pleural diaphragm. Children NCT00976352 insufficiency despite activity (descent on effusion (determined to with a higher baseline Trial (multiple groups) long-term use of enzyme-replacement MRI, EMG activity) be unrelated) neuromuscular function N: 9 therapy may have greater potential for functional Treatment requisite: gains. enzyme-replacement therapy (ERT) infusions every 1 to 2 weeks for >1 year and remained on ERT throughout study Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Muscular conditions Age group: children AAV2.5 Muscle transduction Save and well tolerated This trial established Bowles et al, 2012288 Duchenne muscular Placebo that the rationally Nationwide Children's dystrophy designed AAV2.5 vector Hospital, 2009466 Treatments enrollment: was safe and well tolerated. NCT00428935 none RCT Treatment requisite: N: 6 methylprednisolone Follow-up: 24 Neurodegenerative Age group: adults AAV1.CMV.FS344 Walk distance No significant adverse The results are disorders Becker muscular No comparison group improvement (6-minute events observed in encouraging for Mendell et al, 2015306 dystrophy walk test), either arm treatment of Nationwide Children’s Treatments enrollment: improvement in muscle dystrophin-deficient Hospital, 2017467 none strength muscle diseases. NCT01519349 Treatment requisite: Trial (multiple groups) treatment with N: 6 prednisone for 1 month pre (and 1 month post) viral gene therapy injection Follow-up: 12 Neurodegenerative Age group: children AAV2CUhCLN2 Safety of administration None The data support the disorders Late infantile neuronal No comparison group of AAV2CUhCLN2, anti- hypothesis that AAV- Worgall et al, 2008296 ceroid lipofuscinosis AAV antibodies, disease mediated CNS gene ID NR, protocol Treatments enrollment: progression assessed by transfer can slow the reviewed by NIH-RAC none CNS imaging, safety rate of progression of the CNS manifestations Single-arm trial Treatment requisite: of the disease and none N: 10 support the concept Follow-up: none that additional studies using AAV-mediated CNS gene transfer to treat the CNS manifestations of LINCL are warranted.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Neurodegenerative Age group: children AAV2(CU)hCLN2 Target site infusion None The neurosurgical disorders Late infantile neuronal No comparison group considerations and Souweidane et al, ceroid lipofuscinosis results of this study are 2010300 disease presented to offer Crystal et al, 2004468 Treatments enrollment: guidance and a basis for the design of future ID NR, protocol none gene therapy or other reviewed by NIH-RAC Treatment requisite: clinical trials in children none Single-arm trial that Follow-up: 1 N: 10 utilize direct therapeutic delivery. Neurodegenerative Age group: adults AAV2-GAD UPDRS motor ratings, SAE: 1 intervention; 0 The efficacy and safety disorders Advanced Parkinson’s Placebo clinical global placebo of bilateral infusion of LeWitt et al, 2011302 disease impression of severity, AAV2-GAD in the NCT00643890 Treatments enrollment: consistent medication subthalamic nucleus effects, on–off support its further RCT levodopa fluctuations, freezing development for Treatment requisite: Completed gait Parkinson’s disease and none N: 45 show the promise for Follow-up: 6 gene therapy for neurological disorders. Neurodegenerative Age group: children AAV2-hAADC Improvement in weight, Orofacial dyskinesia (n Gene therapy targeting disorders Aromatic L-amino acid No comparison group improvement in motor = 2) primary AADC Hwu et al, 2012304 decarboxylase (AADC) functions, improvement deficiency is well ID NR deficiency in cognitive functions, tolerated and leads to improvement in improved motor Single-arm trial Treatments enrollment: none oculogyric crises function. N: 4 symptoms, Treatment requisite: improvement in stereotactic surgical symptoms (ptosis, delivery of gene dyskinesia, head therapy and control, sitting, hospitalization for 1 standing) week after therapy delivery Follow-up: max 24+, but still ongoing

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Neurodegenerative Age group: adults AAV2-hAADC FMT uptake, clinical None This study showed disorders Parkinson's disease No comparison group safety, efficacy AAV2-hAADC treatment Mittermeyer et al, Treatments enrollment: was safe and that AADC 303 2012 none gene expression was maintained at least 4 ID NR Treatment requisite: years after Single-arm trial none administration of N: 10 Follow-up: 48 therapy. Neurodegenerative Age group: adults AAV2-neurturin UPDRS part 3 score in SAEs (patients): 13 Intraputaminal AAV2- disorders Parkinson’s disease Placebo the off state, UPDRS intervention, 4 placebo neurturin is not superior 301 Marks et al, 2010 Treatments enrollment: score in the off state, to sham surgery when Ceregene, 2008469 levodopa UPDRS score in the on assessed using the state, home diary UPDRS motor score at NCT00400634 Treatment requisite: assessment of motor 12 months. However, none RCT state, timed walking the possibility of a Completed Follow-up: 18 test, Purdue pegboard benefit with additional N: 58 test—most-affected targeting of the hand, Purdue pegboard substantia nigra and test—least-affected longer-term follow-up hand, Dyskinesia rating should be investigated in further studies. Neurodegenerative Age group: adults AAV2-neurturin UPDRS motor SAEs: 5 patients There was no disorders Parkinson disease Placebo component, total intervention; 8 patients significant difference 307 Olanow et al, 2015 Treatments enrollment: UPDRS, subject- placebo; 2 patients had between groups in the stable doses of completed home diary cerebral hemorrhages primary endpoint and in assessment of motor with transient most secondary Bartus et al, 2013470; antiparkinsonian drugs state, Clinical Global symptoms. No clinically endpoints. Therapeutics Sangamo for at least 6 weeks Impression, Patient meaningful adverse Therapeutics, 2018471; Treatment requisite: Global Impression, events were attributed Marks et al, 2016472 none Unified Dyskinesia to AAV2-neurturin. NCT00985517 Follow-up: 15 Rating Scale, nonmotor RCT symptom questionnaire, Completed and Parkinson’s N: 51 disease

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Neurodegenerative Age group: adults AAV2-NGF Cognitive function All adverse events were This trial provides disorders Alzheimer’s disease No comparison group (Alzheimer’s Disease thought to be unelated important evidence that 305 Rafii et al, 2014 Treatments enrollment: Assessment Scale– or unlikely to be related bilateral stereotactic Ceregene, 2010473 none Cognitive, Alzheimer’s to the study drug. administration of AAV2- Disease Cooperative NGF is feasible, well NCT00087789 Treatment requisite: Serious adverse events: Study Clinical Global tolerated, and able to stereotactic abdominal pain (n = 1), Trial (multiple groups) Impression of Change , produce long-term, neurosurgical cholecystitis (n = 1), N: 10 digit span backward, biologically active NGF administration of gene pneumonia (n = 1), judgment of line expression. therapy under general pelvic fracture (n = 1), orientation, category anesthesia failure to thrive fluency), stabilization or resulting in death (n = Follow-up: 24, invited improvement in glucose 1) to join long-term follow- metabolism (PET scans) up study after while taking the Snodgrass Picture Naming Activation Task Neurodegenerative Age group: adults AAV2-NGF ADAS-Cog 11, clinical SAEs: 22 intervention, This multicenter disorders Alzheimer’s disease Placebo dementia rating—sum 16 placebo randomized clinical trial 293 Rafii et al, 2018 Treatments enrollment: of boxes, Modified demonstrated the Sangamo and cholinesterase Clinical Global feasibility of sham Alzheimer’s Disease inhibitors, memantine Impression of Change, surgery—controlled 474 MMSE, neuropsychiatric stereotactic gene Cooperative, 2019 Treatment requisite: inventory, Alzheimer's delivery studies in NCT00876863 none disease cooperative patients with AD. AAV2- RCT Follow-up: 24 study—activities of daily NGF delivery was well- Completed living tolerated but did not N: 49 affect clinical outcomes or selected AD biomarkers. Pathological confirmation of accurate gene targeting is needed.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Neurodegenerative Age group: adults AAV-GAD Clinical improvement in None AAV-GAD gene therapy disorders Parkinson's disease No comparison group motor ratings of the subthalamic 294 Kaplitt et al, 2007 Treatments enrollment: nucleus is safe and well Neurologix Inc, 2008475 none tolerated by patients with advanced NCT00195143 Treatment requisite: Parkinson’s disease, surgery Single-arm trial suggesting in vivo gene N: 12 Follow-up: 12 therapy in the adult brain might be safe for various neurodegenerative diseases. Neurodegenerative Age group: adults AAV-hAADC Unified Parkinson’s No direct adverse This study provides disorders Parkinson disease No comparison group disease rating scale, events were noted, but class IV evidence that 298 Christine et al, 2009 Treatments enrollment: imaging results 2 patients sustained bilateral intrastriatal Genzyme, 2013476 none vascular complications infusion of adeno- 477 distant from the associated viral type 2 Valles et al, 2010 Treatment requisite: infusion site. vector containing the NCT00229736 none human AADC gene Single-arm trial Follow-up: none improves mean scores N: 10 on the Unified Parkinson’s disease rating scale by approximately 30% in the on and off states, but the surgical procedure may be associated with an increased risk of intracranial hemorrhage and self-limited headache

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Neurodegenerative Age group: adults AAV-hAADC-2 UPDRS, motor state None The findings provide disorders Parkinson's disease No comparison group diaries, MMSE, short class IV evidence Muramatsu et al, Treatments enrollment: form of the GDS, regarding the safety 299 2010 at least 5 years of laboratory tests and efficacy of AADC ID NR levodopa therapy gene therapy and warrant further Single-arm trial Treatment requisite: evaluation in a none Completed randomized controlled, N: 6 Follow-up: 12 phase II setting. Neurodegenerative Age group: children AVXS-101 Time until death or the Patient 1 in cohort 1 In patients with SMA1, disorders Spinal muscular atrophy No comparison group need for permanent had elevations in serum a single intravenous Mendell et al, 201711 type 1 ventilatory assistance, aminotransferase levels infusion of adeno- AveXis, 2017478 Treatments enrollment: Children’s Hospital of (31 times the upper associated viral vector Philadelphia infant test limit of the reference containing DNA coding NCT02122952 Patients had genetically of neuromuscular range for alanine for SMN resulted in Trial (multiple groups) confirmed diagnosis of SMA1. Patients with the disorders aminotransferase longer survival, superior N: 15 c.859G→C disease [ALT]). One patient in achievement of motor modifier in exon 7 of cohort 2 required milestones, and better SMN2 were excluded. additional prednisolone motor function than in to attenuate elevated historical cohorts. Treatment requisite: serum ALT and AST prednisolone level. Follow-up: 30

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Neurodegenerative Age group: adults CERE-120 Change in Unified No serious adverse The initial data support disorders Parkinson's disease No comparison group Parkinson’s Disease events occurred in any the safety, tolerability, 295 Marks et al, 2008 Treatments enrollment: Rating Scale 22 motor patient. and potential efficacy; Ceregene, 2007479 Patients had to be on score (part III) (25% however, these results reduction is considered must be viewed as NCT00252850 stable doses of clinically meaningful); preliminary until data Trial (multiple groups) antiparkinsonian drugs in the month preceding change in time spent in from blinded, controlled N: 12 the trial. the on state without clinical trials are dyskinesia; change in available. Treatment requisite: complete UPDRS, The therapy was change in patient- administered bilaterally completed diary into the putamen in assessments of motor four needle tracks on function, change in each side. timed motor tests (e.g. Follow-up: 12 timed walking test, Purdue Pegboard), change in dyskinesia rating scale, clinical global impression of severity, change in clinical global impression of improvement scales, and improvement in quality of life measures

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Neurodegenerative Age group: adults rAAV1.CMV.huFS344 Improvements in 6- No observed therapy- The findings show disorders Sporadic inclusion body No treatment minute walk test, related serious adverse promise for follistatin Mendell at al, 2017292 myositis exercise tolerance, events (fall, n = 2; gene therapy for mild to Mendell et al, 2017480 Treatments enrollment: activities of living biking accident, n = 1) moderately affected, ambulatory sporadic NCT02354781, IND none inclusion body myositis 14845 Treatment requisite: patients. Controlled trial pretreatment muscle N: 14 biopsy, prednisone treatment for 1 month prior to and after vector administration Follow-up: 2 to 24 (range) Neurodegenerative Age group: children rAAV1.tMCK.hSGCA Improvement in gene None observed We observed sustained disorders Muscular dystrophy Placebo expression, gene expression but Mendell et al, 2009291 (limb-girdle muscular improvement in muscle studies over longer time Mendell et al, 2010481 dystrophy type 2D) size periods without Nationwide Children's Treatments enrollment: immunotherapy will be Hospital, 2011482 Patients had to be required for design of vascular delivery gene NCT00494195 treatment-free for 3 therapy trials. RCT months prior to enrollment. N: 6 Treatment requisite: methylprednisolone at 4, 24, and 48 hours post gene therapy injection Follow-up: 6

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Neurodegenerative Age group: children rAAV2/5_NAGLU Brain cells showed All adverse events were Intracerebral rAVV2/5 disorders Mucopolysaccharidosis No comparison group sustained enzyme mild, chiefly diarrhea or was well tolerated and Tardieu et al, 2017308 type IIIB syndrome production that led to anesthesia related. induced sustained Lysogene, 2017483 (Sanfilippo type B improved cognitive enzyme production in outcomes in all patients the brain. The initial SRCTN19853672 syndrome) compared with the specific anti-NAGLU Single-arm trial Treatments enrollment: none natural history of immune response that N: 4 mucopolysaccharidosis later subsided Treatment requisite: type III syndromes, suggested acquired Immunosuppression with the outcomes immunological was started 14 days being best in the tolerance. The best before surgery with 0·2 youngest patient. We results being obtained mg/kg oral tacrolimus saw an initial specific in the youngest patient per day and 1200 immune response to implies a potential mg/m² oral the therapeutic enzyme window of opportunity. mycophenolate mofetil that later subsided, Longer follow-up is per day. A 4-hour which indicated needed to further pharmacokinetic profile acquired immunological assess safety outcomes was determined when tolerance for three and persistence of steady state was patients. The treatment improved cognitive achieved and 7 days had a good safety development. after. profile within the Follow-up: 30 follow-up period in all patients. Ocular disorders Age group: adults AAV.REP1 Retinal sensitivity, Complications in 2 The initial results of this MacLaren et al, 2014318 Choroideremia No comparison group vision change patients retinal gene therapy Chen et al, 2000484 Treatments enrollment: trial are consistent with University of Oxford, none improved rod and cone 485 function that overcome 2017 Treatment requisite: 486 any negative effects of Xue et al, 2016 surgery 487 retinal detachment. Simunovic et al, 2016 Follow-up: 6 Xue et al, 2018488 Simunovic et al, 2017489 NCT01461213 Single-arm trial N: 6

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Ocular disorders Age group: adults AAV2.hRPE65v2 Visual acuity No serious adverse Although normal vision Maguire et al, 200875 Leber's congenital No comparison group events related to the was not achieved, this Maguire et al, 200976 amaurosis type 2 vector study provides the basis Simonelli et al, 2010490 Treatments enrollment: for further gene therapy Ashtari et al, 2011491 none studies in patients with Leber's congenital Spark Therapeutic, Treatment requisite: 492 amaurosis. 2024 none 493 Bennett et al, 2012 Follow-up: 20 Spark Therapeutics, 2026494 Bennett et al, 201677 Testa et al, 2013495 Ashtari et al, 2017496 NCT00516477, NCT01208389 (follow- up) Single-arm trial N: 3 Ocular disorders Age group: adults AAV2-REP1 Best-corrected visual No serious adverse AAV2-REP1 can Fischer et al, 2018309 Choroideremia No treatment acuity events reported maintain and, in some study eye Treatments enrollment: cases, improve visual acuity in choroideremia. RCT confirmed REP gene mutation Longer-term follow-up N: 6 is necessary to establish Treatment requisite: whether these benefits none are maintained. Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Ocular disorders Age group: adults AAV2-REP1 (10(11) Early treatment diabetic No serious adverse Sustained improvement Lam et al, 2018310 Advanced No comparison group retinopathy study events reported or maintenance of best- Byron Lam and choroideremia letters best-corrected correct visual acuity can University of Miami, Treatments enrollment: visual acuity in treated be achieved in patients 2018497 none versus untreated eye, with choroideremia with intraocular pressure, high-dose AAV2-REP1, NCT02553135 Treatment requisite: change from baseline in and the therapy can be oral corticosteroid Trial (multiple groups) microperimetry, change delivered with (prednisone) for a total N: 6 from baseline in fundus intraoperative OCT with of 21 days, starting 2 autofluorescence, a good safety profile. days before surgical change from base procedure; surgical procedure: 3-port pars plana vitrectomy performed under general anesthesia and an automated injection system for the vector was used to deliver treatment to the subretinal space Follow-up: 24

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Ocular disorders Age group: adults AAV2-sFLT01 Best-corrected visual 1 death of a 91-year- Intravitreous injection Heier et al, 2017325 Advanced neovascular No comparison group acuity old patient 1 year after of AAV2-sFLT01 Genzyme, 2014498 age-related macular study completion and 2 seemed to be safe and degeneration years after vector well tolerated at all NCT01024998 Treatments enrollment: injection. The patient doses. Trial (multiple groups) Patients had best chose not to enter the N: 19 corrected visual acuity extended follow-up of 20/100 or worse in study, and information the study eye and regarding the cause of 20/400 or better in death was fellow eye; showed unobtainable. intraretinal or subretinal fluid in macula; and had adequate pupillary dilation. Treatment requisite: broad-spectrum topical antibiotic solution in the study eye for 3 days before the injection Follow-up: 12 Ocular disorders Age group: adults AAV8-RS1 Visual acuity, The investigational This study showed the Cukras et al, 2018312 X-linked retinoschisis No comparison group retinal sensitivity on product was generally feasibility of this NCT02317887 Treatments enrollment: MP1 testing, well tolerated in all but approach for delivery of 1 individual. an AAV8 vector into the Trial (multiple groups) none ocular CT, eye. As found in other Treatment requisite: electroretinogram N: 9 trials, a dose-dependent none increase in ocular Follow-up: 6 inflammation that resolved with topical and systemic steroids was also observed. Cavity closure in one participant provided the first signal of possible efficacy.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Ocular disorders Age group: adults rAAV.sFlt-1 Phase I: reduced Subconjunctival rAAV.sFlt-1 gene Constable et al, 2016324 Wet age-related No treatment, other: all number of subsequent hemorrhage; subretinal therapy may prove to Rakoczy et al, 2015499; macular degeneration patients received 0.5 RBZ retreatments hemorrhage; mild cell be a potential adjunct Constable et al, Treatments enrollment: mg ranibizumab (RBZ) during the first year, debris in the vitreous; 2 or alternative to 2016500; Constable et Most patients had at baseline and week 4; improvement and phakic patients required conventional intravitreal al, 2017501; previous anti-VEGF patients were maintenance of center- cataract surgery injection for patients Perth Western Australia therapy and no randomized to no point thickness, vision with wet AMD by Lions Eye Institute, washout period was additional treatment, improvement, BCVA providing extended 2017502 required. high dose, or low dose change from baseline delivery of sFlt-1. NCT01494805 Treatment requisite: Phase II: data not yet available Controlled trial none N: 40 Follow-up: 36 Ocular disorders Age group: adults rAAV2-RPE65 Systemic and ocular None Gene therapy for Leber Jacobson et al, 2012317 Leber congenital No comparison group safety congenital amaurosis Cideciyan et al, 2009503; amaurosis caused by RPE65 Cideciyan et al, 2014504 Treatments enrollment: mutations is sufficiently safe and substantially NCT00481546 none efficacious in the Treatment requisite: Single-arm trial extrafoveal retina. N: 15 none Follow-up: 36

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Ocular disorders Age group: adults rAAV2.REP1 Safety, best-corrected 1 localized intraretinal One serious adverse Dimopoulos et al, Choroideremia No comparison group visual acuity, fundus immune response event was experienced 311 2018 Treatments enrollment: autofluorescence, in 6 patients treated Ian et al, 2017505 none spectral-domain optical with a subfoveal coherence tomography, injection of AAV2.REP1. NCT02077361 Treatment requisite: microperimetry, quality The area of remaining pretreatment with oral Single-arm trial of life functional RPE in the prednisone N: 6 treated eye and Follow-up: 24 untreated eye declined at the same rate during a 2-year period. Fundus autofluorescence area is a remarkably predictive biomarker and objective outcome measure for future studies of ocular gene therapy in CHM patients. Ocular disorders Age group: adults rAAV2/2 RPE65 Visual acuity, contrast None Gene therapy with Bainbridge et al, Leber's congenital No comparison group sensitivity, color vision, rAAV2/2 RPE65 vector 2015319 amaurosis spectral sensitivities improved retinal Bainbridge et al, Treatments enrollment: sensitivity, albeit 2008506; Ripamonti et none modestly and 507 temporarily. al, 2015 ; Treatment requisite: University College none 508 London, 2014 Follow-up: 36 NCT00643747 Single-arm trial N: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Ocular disorders Age group: children and rAAV2/4.RPE65.RPE65 Improvement in Observed no local or The therapy is well Le Meur et al, 2018313 adults No comparison group perception of detail, systemic adverse tolerated but the NCT01496040 Leber congenital fixation, color vision; effects related to efficacy varies reduction in treatment: depending on the Trial (multiple groups) amaurosis photophobia, visual thyroidectomy (n = 1), patient. N: 9 Treatments enrollment: none fatigue supraventricular tachycardia (n = 1) Treatment requisite: 3- were deemed serious way 20G vitrectomy to adverse events not inject vector; related to treatment prednisolone daily treatment for 1 week prior to injection and continued for 1 month post injection; dexamethasone- tobramycin eye drops for 1 month post injection; atropine eye drops for 7 days post injection Follow-up: maximum 42 Ocular disorders Age group: children and rAAV2-CB-hRPE65 Improvements in visual Serious adverse event Treatment with rAAV2- Weleber et al, 2016323 adults No comparison group acuity, optical not considered to be CB-hRPE65 was not Applied Genetic Leber congenital coherence tomography, due to therapy: associated with serious Technologies, 2014509 amaurosis, severe early perimetry (static, noncardiac chest pain adverse events, and kinetic), and hypertension (n = improvement in 1 or NCT00749957 childhood-onset retinal degeneration electroretinography, 1) more measures of Trial (multiple groups) Treatments enrollment: quality of life visual function was N: 12 none observed in 9 of 12 patients. The greatest Treatment requisite: improvements in visual vitrectomy (for acuity were observed in subretinal injection of younger patients with therapy), postoperative better baseline visual treatment (local acuity. corticosteroids, antibiotics) Follow-up: 24

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Ocular disorders Age group: adults rAAV2-CBSB-hRPE65 Ocular safety, visual None No positive result was Hauswirth et al, 2008315 Leber congenital No comparison group function, central retinal evident. Cideciyan et al, 2008510 amaurosis structure ID NR Treatments enrollment: Single-arm trial none N: 3 Treatment requisite: none Follow-up: 3 Ocular disorders Age group: children and rAAV2-hRPE65 Visual function, ocular None The investigated Banin et al, 2010316 adults No comparison group safety process of genetically Hadassah Medical, Leber congenital analyzing affected 2016511 amaurosis isolated populations as a screen for gene-based NCT00821340 Treatments enrollment: none therapy suggests a new Single-arm trial paradigm for disease Treatment requisite: N: 208 diagnosis and none treatment. Follow-up: none

Ocular disorders Age group: adults rAAV2-ND4 Visual acuity, vision None Findings support the Wan et al, 2016322 Leber’s hereditary optic No comparison group field, VEP, OCT, blood feasible use of gene Yang et al, 2016512 neuropathy and urine, liver, kidney, therapy for Leber’s Yang et al, 2016513 Treatments enrollment: immune function hereditary optic neuropathy. NCT01267422 none Trial (multiple groups) Treatment requisite: N: 9 none Follow-up: 9

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Ocular disorders Age group: children and rAAV2-VMD2-hMERTK Improvements in visual None listed as serious Gene therapy for Ghazi et al, 2016321 adults No comparison group function: visual acuity, MERTK-related RP using NCT01482195 Retinitis pigmentosa retinal function, central careful subretinal macular and foveal injection of rAAV2- Trial (multiple groups) Treatments enrollment: none thickness VMD2-hMERTK is not N: 6 associated with major Treatment requisite: side effects and may vitrectomy and result in clinical subretinal injection of improvement in a therapy, subset of patients. subconjunctival antibiotics and steroids Follow-up: 180 Ocular disorders Age group: adults scAAV2(Y444,500,730F) Loss of visual acuity to No observed serious No serious safety Feuer et al, 2016320 Leber hereditary optic -P1ND4v2 no light perception ocular or systemic problems were NCT02161380 neuropathy No comparison group adverse events observed in the first 5 patients enrolled in this Trial (multiple groups) Treatments enrollment: none phase I trial of virus- N: 5 based gene transfer in Treatment requisite: this mitochondrial intravitreal injection disorder. Additional Follow-up: 36 study follow-up of these and additional patients planned for the next 4 years is needed to confirm these preliminary observations. Ocular disorders Age group: children and Voretigene neparvovec; Multiluminance mobility No product-related Treatment with Russell et al, 201774 adults AAV2-hRPE65v2 test score (bilateral), serious adverse events Voretigene neparvovec Spark et al, 2015514 Inherited retinal Placebo multiluminance mobility or deleterious immune gene replacement test score (first injected responses occurred. improved functional NCT00999609 dystrophy eye), full-field light vision in RPE65- RCT Treatments enrollment: none sensitivity threshold (2 mediated inherited Completed eye average), best- retinal dystrophy that Treatment requisite: N: 31 corrected visual acuity was previously none (2 eye average) medically untreatable. Follow-up: 12

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Other: prophylactic Age group: adults tgAAC09 HIV test No vaccine-related All responses were to AIDS vaccine candidate Healthy volunteers Placebo serious adverse events gag epitopes. tgAAC09 Mehendale et al, Treatments enrollment: reported appears to be safe, well 356 2008 none tolerated, and modestly immunogenic. ID NR, approved by Treatment requisite: ethical, scientific, and none regulatory authorities in Follow-up: 6 Europe and India RCT N: 80 Respiratory conditions Age group: adults AAV-CFTR None Little or no AAV-CFTR Wagner et al, 1999332 Cystic fibrosis No comparison group inflammatory or administration to the Wagner et al, 1999515 Treatments enrollment: immune responses maxillary sinus results observed in successful, dose- ID NR, reviwed by FDA, none dependent gene NIH-RAC Treatment requisite: transfer to the maxillary none Single-arm trial sinus and alterations in N: 10 Follow-up: none sinus TEPD suggestive of a functional effect, with little or no cytopathic or host immune response.

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Respiratory conditions Age group: children and AAV-CFTR (tgAAVCF) Absolute FEV1, change SAEs (patients): 13 Repeated doses of Moss et al, 2007337 adults Placebo in FEV1, FEV1 intervention, 11 placebo aerosolized tgAAVCF Moss et al, 2004516 Cystic fibrosis percentage predicted, were safe and well forced vital capacity, tolerated but did not ID NR, protocol Treatments enrollment: mid-maximal expiratory result in significant reviewed by NIH-RAC none flow rate (FEF25-75) improvement in lung and FDA Treatment requisite: function over time. none RCT Because gene transfer Completed Follow-up: 7 is the simplest, most N: 109 basic way to correct the underlying genetic defect that leads to disease in CF, further research is warranted to develop an effective gene transfer agent for the treatment of CF. Respiratory conditions Age group: adults rAAV1-AAT Hematology or clinical No SAEs were reported These findings suggest Brantly et al, 2009338 Alpha-1 antitrypsin No comparison group chemistry parameters during the study. immune responses to Flotte et al, 2011517; deficiency Potentially related AEs AAV capsid that develop Brantly et al, 2006518 Treatments enrollment: included mild erythema after intramuscular University of none near the injection site. injection of a serotype 1 There was no indication rAAV vector expressing Massachusetts Treatment requisite: 519 of muscle toxicity or AAT do not completely Worcester, 2015 ; AAT protein 520 inflammation. eliminate transduced Mueller et al, 2013 ; augmentation cells in this context. Applied Genetic Follow-up: none Technologies, 2011521 NCT00430768 Trial (multiple groups) N: 9

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Indication Indication Intervention Outcomes Measured Reported Harms Authors’ Conclusions Citations Concurrent/Prior Design Treatments Comparator N Months of Follow-up Respiratory conditions Age group: children and rAAV2-CFTR (tgAAVCF) Safety outcomes, vector All patients experienced The current study Flotte et al, 2003336 adults No comparison group viremia, shedding, at least 1 AE, most of provides clear evidence National Institute of Cystic fibrosis immune response, DNA which were clearly of the safety of upper- Diabetes, 2002522 Treatments enrollment: transfer outcomes unrelated to vector and lower-respiratory administration. None tract delivery of NCT00004533 none resulted in death or tgAAVCF within a dose Treatment requisite: Trial (multiple groups) withdrawal range resulting in none N: 25 measurable DNA Follow-up: 24 transfer, which supports its continued development as a clinical agent for CFTR gene transfer in the context of CF lung disease. Respiratory conditions Age group: adults tgAAVCF None 242 adverse events This phase I trial shows Aitken et al, 2001334 Cystic fibrosis No comparison group total, 6 serious and 3 aerosolized tgAAVCF is ID NR, protocol Treatments enrollment: possibly related to the safe and widely reviewed by NIH-RAC none study drug delivered to the proximal airways of CF and FDA Treatment requisite: patients by Single-arm trial none nebulization. N: 12 Follow-up: 3 Respiratory conditions Age group: children and tgAAVCF Recurrent sinusitis, The tgAAVCF This phase II trial Wagner et al, 2002335 adults Placebo imaging, histopathology administration was well confirms the safety of ID NR Cystic fibrosis with tolerated, without tgAAVCF but provides adverse respiratory little support of its RCT antrostomies events, and there was efficacy in the within- Completed Treatments enrollment: none no evidence of patient controlled sinus N: 23 enhanced inflammation study. Treatment requisite: in sinus histopathology none or alterations in serum- Follow-up: 3 neutralizing antibody titer to AAV capsid protein.

Abbreviations: AAV, adeno-associated virus therapy; AEs, adverse events; ID, trial identification number; mL, milliliter; NR, not reported; RCT, randomized controlled trial; SAEs, serious adverse events.

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