Reigniting Recruitment

a Publication of the HIV Vaccine Trials Network

Volume 4, issue 1 | july, 2012

REIGNITING RECRUITMENT

IN THIS ISSUE photo by Sid Niazi

ARTICLES 02 Probing the Diversity of Vaccine Elicited HIV-1 Antibodies: Informed by the 18 Social and Behavioral Science in Clinical Trials of Biomedical HIV Prevention RV144 Correlates Analysis Interventions 03 Assays to Probe the Humoral Response 22 HVTN Annual Network Award Winners 06 Turning up the Heat in Miami 24 HVTN Protocols 07 Orlando HVTN 505 Study Site: From Challenges to Accomplishments CALENDAR [back cover] 10 Assessing Mucosal Immune Responses in HIV Vaccine Trials 13 Highlights from Recent HVTN Publications Probing the Diversity of Vaccine Elicited HIV-1 Antibodies: Informed by the RV144 Correlates Analysis Georgia D. Tomaras

RV144, a trial conducted by the U.S. Military HIV Research IgG antibody responses (ie, ADCC and nAbs) might corre- Program and the Thai Ministry of Health, was the first HIV-1 late with decreased risk of infection if present with low levels vaccine study in which a modest efficacy (31.2%) of HIV‑1 of Env IgA (higher levels of Env IgA seem to counteract the vaccination was shown. The study regimen consisted of a beneficial effects of IgG). Thus, the balance of antibody types, canarypox prime expressing Gag, Pro, and gp120 (ALVAC- and the interactions among them -- particularly in those with HIV), and a gp120 boost (AIDSVAX® B/E). These results different Fc receptor binding properties -- merits further study were published in the New England Journal of Medicine in to determine if specific Env antibody isotypes influence the 2009.1 level of vaccine efficacy.

A concerted effort by an international consortium of investiga- The results from the RV144 correlates analysis have signifi- tors, led by Dr. Barton F. Haynes of Duke University, set out cantly informed our understanding of how to analytically to determine if any immune variables correlated with infection probe the HIV-1 vaccine elicited antibody responses, and risk in the RV144 trial. In the initial phase of the studies, there generated several leading hypotheses that are critical to test was an open call to measure vaccine-elicited immune responses in future HIV-1 vaccine candidates. Measurements of the re- on RV144 samples. In the next phase, a selected set of immune sponse rates and magnitudes of V1V2 IgG antibodies and Env variables, chosen from the pilot study results, were formally IgA binding antibodies elicited by different vaccine regimens tested as primary immune variables for the correlates analysis (different vector, adjuvant, and Env immunogen formulations) of RV144. are important for understanding the broad applicability of these findings -- ie, whether these correlates can be confirmed The RV144 correlates analysis yielded 2 immune correlates with other regimens. of risk: V1V2 IgG antibodies correlated with decreased risk Durability of HIV-1 Env Immune Responses of infection, while HIV-1 Env IgA antibodies correlated with increased risk of infection in the vaccine arm (higher One of the most overarching lessons from RV144, indepen- Env IgA levels seemed to have abrogated the vaccine effect, dent of the immune correlates results, comes from the observa- but the risk of infection in this group was not higher than 2 tion that there was decreased vaccine efficacy over time. This in the placebo group). The combined results of the original waning of vaccine efficacy corresponds to decreasing vaccine- and correlate studies galvanized the HIV-1 vaccine field, and specific immune responses (anti-Env antibody titers). Thus, the have influenced the direction of subsequent protocols testing measurement of the durability of immune responses (antibody HIV-1 vaccine candidates. Notably, these correlates of HIV-1 half-life) is a critical component for evaluating improvements infection risk from RV144 are not necessarily surrogates of in HIV-1 vaccine strategies going forward. protection nor mechanistic correlates. However, toward the goal of identifying the types of immune responses an effica- Antibody Responses to Circulating vs. Vaccine Strain cious vaccine should elicit, the RV144 study has provided key HIV-1 Env hypotheses to test in further HIV-1 vaccine trials. The 2 correlates of risk identified in RV144, along with cor- Key Hypotheses Generated by RV144 responding secondary analyses, highlight an important feature for assessing vaccine efficacy. The measurements that correlated The RV144 correlates analysis evaluated 6 primary variables: most strongly with infection risk were not those against the the binding of plasma IgA antibodies to Env, the avidity of vaccine strain Env in the boost, but rather antibodies against IgG antibodies for Env, antibody-dependent cellular cyto- cross-clade responses: The strongest statistical correlate was toxicity (ADCC), HIV-1 neutralizing antibodies (nAbs), the the association of risk with IgA responses to the C1 region in binding of IgG antibodies to variable regions 1 and 2 (V1V2) gp120 of circulating strain in Thailand (CRF01AE). Moreover, of the gp120 Env, and the level of Env-specific CD4+ T cells.2 IgG responses to the V2 region of CRF01AE Env significant- In addition to the 2 identified immune correlates of risk ly correlated with decreased risk of infection among the differ- (V1V2 IgG and HIV-1 Env IgA), analysis of the 6 primary ent clades of V2 responses tested in the peptide microarray variables using interaction models suggested that functional assay (Gottardo, Montefiori et al., unpublished).2 This

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2 JUNE 2012 VOLUME 4:1 | HVTNEWS Assays to Probe the Humoral Response Georgia D. Tomaras

There are several categories of assays used to examine the diverse humoral response to vaccination: 1) validated assays for HIV-1 vaccine endpoints, 2) qualified and/or standardized assays and 3) assays for research and development. Validated assays are currently used in the first line assessment of HIV-1 vaccine trials. Qualified and/or standardized assays are utilized to generate hypotheses and further characterize vaccine immunogenicity. Assays for research and development are more exploratory in nature and are not suitable as a vaccine study endpoint, but can provide substantial insight into the com- plexity and potential functionality of vaccine elicited immune responses.

Validated Assays for HIV-1 Vaccine V= Variable Region C=Constant Region H=Heavy Chain L=Light Chain

Endpoints Structure of the antibody: The variable regions (VH, VL) contain the an- Neutralizing Binding Antibodies tigen binding sites; the Fc region mediates the effector activity (eg, neutralization); the constant regions present in the heavy chain define the isotype Currently validated assays include the measurement of of antibody (IgA, IgD, IgE, IgG, and IgM) and subclass (eg, IgG1-4). neutralizing antibodies categorized as Tier 1 or Tier 2 neutralization using the TZM-bl assay. Validated assays to clinical trials include the measurement of both monomeric assess binding antibody responses (IgG and IgA) by ELISA IgA (mIgA) and dimeric IgA (dIgA) that will be extended to the and multiplex bead technology are in place and can measure analysis of human mucosal samples. Moreover, the analysis magnitude and breadth of vaccine elicited responses. The goal of IgA responses and IgG subclasses, and the respective ratios of ongoing validation efforts include improving upon existing between different types of antibodies, may provide insights assays (eg, validation of A3R5 neutralization assays) and to into the role of different adjuvants/priming on the quality of the select additional binding and functional assays most relevant humoral responses. for vaccine evaluation. Antibody Epitope Mapping (Linear and Qualified and Standardized Conformational) A potentially important characteristic of antibodies is their Assays breadth of binding to multiple HIV-1 clades (Group M, Clade IgG Subclasses, Isotypes, and Mucosal A, B, C, AE, etc.). Recent assay developments include the measurement of diverse HIV-1 Envs resulting in a single score Antibodies for magnitude and breadth. The linear epitope specificities are Antibody isotypes and IgG subclasses (IgG1-IgG4) are measured using either binding antibody assays with specific currently being measured in response to vaccination. The epitopes (eg, V2, C1, alanine scanning mutants) or through an rationale for measuring antibody isotypes and subclasses is HIV-1 multiclade peptide microarray that can fully measure that they provide insights into potential functional responses. multiclade gp160 epitope specificity. Conformational epitopes IgG1 and IgG3 are considered to be the most functional of (eg, CD4BS, CD4i, C1, and N-glycan dependent epitopes) can the subclasses in that they have been associated with HIV-1 be determined through differential binding to wild type pro- neutralization, complement fixation, FcR binding and antibody teins/scaffolds with and without mutation(s) that alter confor- dependent cellular cytotoxicity / cell mediated virus inhibition mational binding sites, or through competitive binding assays (ADCC/ADCVI). lgA antibodies at mucosal sites have been with monoclonal antibodies of known specificities (eg, A32 correlated with protection in exposed, uninfected subjects and monoclonal antibodies [mAb] blocking). studies in NHPs have indicated a potentially protective role for mucosal antibody responses as well as antibody effector functions and IgA memory B cells. Current assays in human

Continued on page 4... www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 3 Antibody Affinity/Avidity Assays through the generation of mAbs using recombinant DNA Measurements of the strength of specific antibody-antigen technology with memory B-cell cultures and antigen specific interactions may correspond with antibody affinity matura- cell sorts. These mAbs provide a stable source of vaccine tion and specific functional antibody responses that ultimately elicited antibodies to test for antiviral function in complex are linked to preventing HIV-1 acquisition or decreasing assay systems (eg, aggregation, mucus inhibition, inhibition viral load. The HVTN labs are utilizing two types of assays of transcytosis). Furthermore, the analysis of antibody gene for measuring antibody avidity: surface plasmon resonance sequences can determine whether the vaccine elicited anti- (SPR) and antibody avidity index assay. Measurement of body repertoire includes characteristics previously associated avidity by SPR allows characterization of antibody on-rate with broadly neutralizing antibodies (eg, specific VHVL chain and off-rate to specific epitope and protein immunogens. gene usage, high mutation frequency, and CDR3 length within Avidity index assays allow for bridging and comparison the variable domains). These strategies along with detailed between NHP and human clinical trials. Moreover, there is epitope mapping (eg, CD4BS, sensitivity to N-linked glyco- precedence for utilizing avidity measurements for evaluating sylation) can inform whether current vaccines are stimulating vaccines (eg, pneumococcal and hepatitis B vaccines). epitope specific antibodies that potentially may be driven toward induction of broadly neutralizing or virus inhibitory antibodies. With this knowledge in hand, vaccine strategies B Cell ELISpot may build upon current vaccine approaches by further boost- Long-lived plasma B cells are responsible for continued ing with novel immunogen designs to achieve higher levels of production of antibodies; whereas memory B cells are re- affinity maturation or specific immune focusing. sponsible for producing an anamnestic response after antigen re-exposure. B cell ELISpots are in place for evaluation of human clinical trials and can be used together with antibody Functional Mucosal Antibody Assays assays (using matched envelope reagents) to characterize the Vaccine elicited antibodies that can block HIV-1 acquisition at vaccine elicited humoral repertoire. mucosal surfaces might be among the most efficacious types of antibodies. In addition to traditional HIV-1 neutralization and Fc receptor mediated virus inhibition; antibodies may Inhibitory Antibody (Non-neutralizing) Assays also aggregate virions, inhibit movement through cervical Standardized assays used in human clinical trials to assess mucus, inhibit transcytosis, mediate intra-epithelial neutraliza- antibody function include ADCC and ADCVI. Alternative neu- tion, block HIV-1 Env gp120 interaction with α4β7 integrin on tralization assays with primary cells, such as peripheral blood CD4+ target cells, and inhibit macrophage infection. A variety mononuclear cells (PBMCs) and monocytes/macrophages of assays are in development with the aim to mimic key steps may also provide information on vaccine elicited antibody during mucosal transmission. Proper assessment in these function. Further examination of Fc receptor function of assays currently may depend on mAbs or purified antibodies vaccine-elicited antibodies is needed, including the roles of from genital fluids and plasma. Further studies using these different antibody isotypes and subclasses, antibody glycosyl- assays to assess vaccine elicited responses at mucosal sites ation, and antibody interaction with a range of effector cells. are needed to determine their role in protection from HIV-1 Additionally, defining the epitope specificity and antibody acquisition. A question remaining from the RV144 corre- isotypes and subclasses that mediate Fc receptor mediated lates analysis, particularly in light of the finding related to virus inhibition, neutralization, and virion capture will enable a monomeric plasma IgA, is whether vaccine elicited immune better understanding of antibody function. responses were present at mucosal surfaces, and whether these responses may have contributed to the observed Assays for Research and efficacy. Future studies are planned for assessing vaccine Development elicited mucosal immunity with RV144 vaccine products, as well as other HIV-1 vaccine candidates. B Cell Technology Phenotypic analysis of vaccine elicited antigen specific B cells NOTE: A fully referenced version can be found at can provide insights for vaccine immunogenicity. Significant http://hvtnews.wordpress.com/. advances have been achieved by probing the B-cell response

4 JUNE 2012 VOLUME 4:1 | HVTNEWS Probing the diversity of Vaccine Elicited HIV-1 Antibodies: Informed by the RV144 Correlates Analysis

assays

ses

ses protocol # BAMA ELISA (binndAinbg Ab) ADCC Ab avidityEpitope anAably repertoBir-ece alln EaLlyISBp coet ll phenotyping

DNA Prime / Viral Vector Boost

076 secondary analysis supports the finding of the primary V2 IgG variable and goes a step further 077 to show increased significance when the clade is 082 matched to the circulating strain in the vaccine population. The outcomes of these 2 CRF01AE 087 measurements provide substantial rationale to 092 include antibody responses to the circulating strains in the vaccine population as key variables 094 to assess going forward in HIV-1 vaccine evalu- 205 ation.

505 Ultimately, these specific hypotheses should be Viral Vector Prime / Boost examined in future HIV-1 vaccine trials in the broader context of examining multiple im- 078 mune parameters. As noted by Plotkin,there is 3 083 a natural redundancy of the immune system. Thus, vaccination may protect through multiple 085 mechanisms that are potentially reliant on host 090 and/or pathogen specific factors. Historically, numerous efficacious vaccines correlate with 091 / IAVI B003 / IPCAVD 004 specific antibody responses, but are not neces- Various Primes / Protein Boost sarily a mechanistic correlate -- that is, the antibody response is not the protective mecha- 073/073E / SAAVI 102 nism, but correlates well with vaccine efficacy (a 4 086 / SAAVI 103 nonmechanistic correlate). This is an important distinction as it is not yet known if the cor- 096 relates identified for RV144 are mechanistic or 097 non-mechanistic correlates. protein only Diversity of HIV-1 Vaccine Elicited 088 Immune Responses

Mucosal specimens Periphery (blood) A striking feature of HIV-1 vaccines, more Abbreviations and Acronyms: Ab = Antibody; ADCC = antibody dependent cellular cytotoxic- generally, is that there is significant heterogeneity; BAMA = Binding Abs multiplex array; ELISA = Enzyme-linked immunosorbent assay; nAb = ity in vaccine responses among individuals. It Neutralizing antibody is precisely this heterogeneity that allows for an immune correlates analysis to be sufficiently powered, since a dynamic range of immune Table1. Vetted Antibody Analyses in HVTN Open / Planned Protocols responses is needed to clearly define a correlate. Measurement of the full repertoire of the

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www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 5 Jim Maynard

Vic Sorrell sings like an angel and he’s found a way to make it this study is in the fight against HIV, reenergized them to do work for him in Nashville. Vic had a great idea to create excite- the work, and sent them home with fresh practical tools to get ment and draw attention to HVTN 505. He did it with song! the job done. One such fresh tool was…singing! Vic launched a city-wide Karaoke competition that used other people’s love of singing to raise money for a local AIDS service In addition to recruitment tools workshops, the study’s co- agency, while educating his community about the trial and col- principal investigators, Drs. Scott Hammer and Magda So- lectingOrlando contacts from HVTNdozens of interested 505 volunteers. Study He took S ite:bieszczyk, shared their scientific expertise to help recruiters feel a fresh idea and turned up the heat on recruitment in Nashville more knowledgeable in talking to potential volunteers. HVTN andF nowrom it was Challengestime to spread that fire in to Miami Accomplishments and beyond! social scientist, Dr. Michele Andrasik, spoke about her research on barriers to recruitment – those things that make people RecruitmentEdwin DeJesus staff from acrossand theKeith United Barsky States joined less likely to take part in a trial – and led a workshop to help HVTN’s Core Community Engagement (CE) Project Man- recruiters do their jobs in a more culturally competent way. It agers in Miami in January to share their very best ideas and was an action packed weekend, but would it pay off in increased brainstorm new ones, all for the sake of increasing enrollment enrollments? in HVTN 505. This trial, the only HIV vaccine efficacy trial in the world, has been very difficult to enroll. “We got back from the recruiters’ retreat and whoosh, new enrollments just took off!” said Jim Carey, Community En- Recruitment staff at 21 gagement Coordinator at the University of Illinois – Chicago sites in 18 U.S. cities (UIC) trials site. “We needed some fresh ideas and there is has been working to no reason to reinvent the wheel. The other sites are a great find men and trans- resource. But it was more than that, the recruiters came back gender women who reenergized. This week we enrolled 3 new participants!” have sex with men to take part in this trial, The Chicago site’s experience is not unique. Sites across the and it hasn’t been easy. Network are launching new programs and events and increas- The original goal of ing their rate of enrollment. HVTN 505 achieved a high water 1350 volunteers was mark the week of April 16, when for the first time, 26 volun- expanded to 2200 after teers were enrolled in one week. The goal of 2200 participants is a protocol amendment, still a challenging one and we need to keep the volume turned making an already up on recruitment, but we are well on our way to reaching it. challenging job that much more difficult. How are things working out for Vic? He is still singing in With a new enrollment Nashville and just recently completed the second annual target, recruiters felt Karaoke competition. And the fire did spread! Recruiters James they were struggling Mapson and Jim Higginbotham at the University of Alabama- for new ways to engage Birmingham site are in the final rounds of their first annual possible volunteers. Karaoke competition, following Vic’s model with great success. We hope that soon it will be the Network’s turn to sing when Vic Sorrell’s karaoke recruitment event Experienced recruiters we reach our goal of 2200 participants in this important inspired other recruiters at the Miami from New York, Bos- trial. Recruiters’ Retreat. ton, Atlanta, Nashville, San Francisco, Roches- Jim Maynard is Associate Director, Community Engagement Unit, ter, and Seattle worked with the Core CE team to put together HIV Vaccine Trials Network a program that reminded workshop participants how important

6 JUNE 2012 VOLUME 4:1 | HVTNEWS Orlando HVTN 505 Study Site: From Challenges to Accomplishments Edwin DeJesus and Keith Barsky

In October 2010, the Orlando Im- a study within the HIV-negative popula- Orlando is a medium-sized metropolitan munology Center (OIC) became an tion. It is also possible for an institution, area, but widely spread out, and with expansion site for the HIV Vaccine having no affiliation with a university no centralized gay neighborhood. There Trials Network (HVTN). Although new hospital or a governmental health pro- are only a few gay bars, one bathhouse, to the HVTN and the NIH’s Division gram, to successfully participate in these and no after-hours clubs. All of these of AIDS (DAIDS), we had over 20 years types of clinical trials. establishments are far from each other, of experience in recruiting patients into and are not easily accessible via public privately-sponsored clinical transportation. trials for various pharmaceu- tical companies and NIH- Despite the lack of gay-night- sponsored programs. However, life entertainment, Orlando we were faced with the reality has been recognized by several that many of the established organizations as one of the HVTN programs, services, gayest cities in the U.S.. It and strategies had not yet been has the largest gay employee tested or implemented at an association in the U.S. (Disney independent private site. Some and Universal Studios Gay of those elements required and Lesbian Association), the modifications to meet the largest U.S. LGBT softball needs of our clinic and our league, as well as one of the community. largest State Universities, Orlando site recruiting event. From right to left: Keith Barsky, Dino Martino, UCF, which has a very strong Our hope was that the skills Tomas Devia, Anthony Egreja, Juan Rodriguez, Saul Leon and recently expanded LGBT that we had acquired over service initiative (ucf.edu/ the years would have prepared us for To prepare for the challenges of recruit- lgbtq). We knew that the at-risk popula- being an HVTN study site. We quickly ing for HVTN 505, we prioritized tion was present in Orlando, but that we discovered that there are tremendous several issues: needed to reach out to them by methods difficulties associated with recruiting other than just visiting clubs or posting HIV-negative volunteers into a govern- The need to know and understand flyers in the bars. It was clear to us from ment-conducted clinical trial, and asking our local gay community the beginning that the most effective ap- them to volunteer to take an experimen- proach was going to require a repetition tal HIV vaccine. At the starting point The need for a thorough under of “the message” by many different types we had no recruitment team, no Com- standing of the science and rationale of vehicles. We wanted to create a sense munity Advisory Board, and only limited of the study of pride in our community by the work associations with the local gay com- being done locally as part of a bigger The need to form strong alliances munity organizations. The hill ahead of picture, a “National Vaccine Study”! with key community organizations us was steep, but our drive, enthusiasm, and desire to be part of something big The need to relate to potential We believe that a well-trained staff is the in our community gave us the impetus volunteers and learn to “speak their key to patient recruitment and reten- we needed to eventually conquer those language” tion. Members of the study team were hurdles. Today, we are happy to see that all required to learn the science behind our original hopes have become a reality, In every major city, the men who have this vaccine study’s development (which and that it is possible to use similar skill sex with men (MSM) community has included a range of general informa- sets learned from enrolling other (HIV- found different patterns of communication about viruses and vaccines to very infected) clinical trials and apply them to tion, evolution/growth, and networking. specific information about HIV), as well Continued on page 8...

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 7 Orlando HVTN 505 Study Site: From Challenges to Accomplishments

as the rationale for the study procedures, so they are able to effectively answer questions from potential volunteers. We also organized several meetings to educate our ancillary staff (including receptionists and any other office staff who may peripherally come in contact with a study volunteer) about the proper way to speak and interact with our volunteers, making them feel important from their very first interaction with OIC. One can never underestimate the importance of a first impression!

Prior to HVTN, there was very little collaboration between the city organizations that serve people with HIV infection and the at-risk population in Orlando. We initiated programs that helped us to develop strong affiliations with key community gay organizations. We learned that in dealing with many Reaching out to the community. Partnerships formed between the Orlando Immunology Center and of these groups, in order to expand community resources and services. *Infectious Disease the relationships, we needed to give something in order to gain something back. One of these organizations is antibody-positive for HIV, we are often role in the dissemination of HVTN The Center, which provides programs able to intervene early and refer them 505 news. We created the “V-Force” our and services for the GLBT community, for continuity of care. This also links us Facebook identity for daily postings and including free HIV testing. OIC has to the many seronegative partners (of notifications. To communicate more made an agreement with The Center to those seropositive patients), for whom effectively with potential study partici- see, free of charge and on the same day we are also able to provide further test- pants we needed to learn their language, of testing, any person who tests posi- ing, counseling, or even possible study which for many is “spoken” through tive at their site. Our initial intent was participation. texting and e-mailing, as opposed to to make sure that individuals newly traditional face-to-face interactions. diagnosed with HIV infections had the We fostered similar relationships with Their schedules of communication can opportunity to discuss prognosis, HIV other key gay organizations in Central be also very unpredictable. Thus, we pathophysiology, and/or treatment op- Florida, helping us to disperse the HIV made sure to extend our phone/text/e- tions with a health care provider, imme- preventive message and to increase the mail availability beyond the regular diately after diagnosis. Engagement in interest of participating in the HVTN business hours (including weekends), HIV care begins at testing, and recent 505 study. These collaborations, which to ensure quick responses for interested studies highlight the importance of the initially started as HVTN 505 promo- candidates. HIV counseling, testing, and referral tions, have now transformed into even experience as it relates to subsequent more important relationships aimed The success with enrollment and linkage to medical care.1-3 at implementing other programs in retention at our site is the result of the our community. The ties that we have collaborative effort that begins with our The value of the services we provide to formed with these organizations will recruiter, is held together by our study The Center extended far beyond our last long after the HVTN 505 study is coordinators, and is sealed by the direct initial expectations. The Center started completed, and could potentially benefit interactions of the principal investigator to also incorporate (as part of their rou- future DAIDS programs. and sub-investigators, who frequently tine HIV screening program) the option get involved at different times during of referring seronegative at-risk po- Social media, the main vehicle in which the process. This team effort is equally tential participants to the HVTN 505 most young people now communicate seen in the community. Potential par- study. For those individuals who test with each other, has played a significant ticipants are more likely to identify this

8 JUNE 2012 VOLUME 4:1 | HVTNEWS Orlando HVTN 505 Study Site: From Challenges to Accomplishments

study as part of an exceptional organi- Edwin DeJesus is Principal Investigator who have never entered HIV medical care. zation when they are able to see many and Keith Barsky is Study Recruiter/Clin- AIDS Educ Prev 2011;23(3 Suppl):117-127. people, of varying backgrounds and ical Research Coordinator at the Orlando 2. Gardner EM, McLees MP, Steiner levels, delivering the same message. HVTN 505 study site. JF, Del RC, Burman WJ. The spectrum of engagement in HIV care and its rel- The path that OIC and HVTN have evance to test-and-treat strategies for paved for the gay community in Orlan- prevention of HIV infection. Clin Infect Dis do will continue to grow and the strong 2011;52(6):793-800. relationships formed will ultimately lead Reference List to greater awareness about HIV and 3. Marks G, Gardner LI, Craw J, Crepaz 1. Garland PM, Valverde EE, Fagan J, N. Entry and retention in medical care STDs, in general. et al. HIV counseling, testing and referral among HIV-diagnosed persons: a meta- experiences of persons diagnosed with HIV analysis. AIDS 2010;24(17):2665-2678.

"Probing the Diversity of Vaccine Elicited HIV-1 Antibodies", continued from page 5

epitope specificities (linear and conformational) of different virus-inhibitory antibodies that can recognize circulating virus antibody isotypes/subclasses both systemically and at mucosal strains and prevent HIV-1 acquisition. sites are needed for vaccine trials to understand the breadth and diversity of responses. In RV144, numerous immune In response to the correlates analysis results, the HVTN in- parameters were untested in the correlates analysis due both to corporated additional antibody assays into currently open pro- inherent assay limitations, sample limitations (lack of mucosal tocols, and into future protocols currently in development. We samples) and the need to keep the correlates analysis focused are analyzing not only blood samples but also mucosal speci- on a limited number of endpoints to achieve statistical power. mens (see Table 1). The addition of mucosal antibody analyses On the other hand, ultimately the heterogeneity of the HIV-1 may expand the knowledge gained from the RV144 correlates vaccine response could become a roadblock to overcome for analysis, and highlight what differences exist between immune improving upon the 31.2% efficacy obtained in the RV144 responses in the mucosa and those in circulation. study. Thus, understanding the genetic, environmental, and immunological determinants of heterogeneous HIV-1 vaccine Georgia Tomaras is Director, HVTN Binding Antibody Labora- elicited immune responses could potentially become critical for tory Program, and Associate Director of Research, Duke Human refining immunogen designs that can achieve more uniformity Vaccine Institute. of specific immune responses among vaccinees.

The HVTN has a wide breadth of humoral, cellular, and innate immunity assays. Furthermore, we are testing and analyzing a wide variety of regimens. Our ability to look across platforms Reference List (regimens, products, delivery mechanisms) allows us to spot patterns that single trials may not expose. 1. Rerks-Ngarm S, Pitisuttithum P, Nitayaphan S, et al. Vaccina- tion with ALVAC and AIDSVAX to Prevent HIV-1 Infection in Thailand. N Engl J Med. 2009. The analysis of the RV144 vaccine and planned follow-up studies will continue to inform us, both technically and 2. Haynes BF, Gilbert PB, McElrath MJ, et al. Immune-cor- conceptually, how best to probe the full repertoire of vac- relates analysis of an HIV-1 vaccine efficacy trial. N Engl J Med. cine elicited HIV-1 antibodies. Going forward, the 2 RV144 2012;366(14):1275-1286. correlates of risk (V1V2 IgG and Env IgA), in terms of both 3. Plotkin SA. Correlates of protection induced by vaccination. response rate and magnitude, should be evaluated in HIV-1 Clin Vaccine Immunol. 2010;17(7):1055-1065. vaccine strategies (different vectors, adjuvants, and boosts), in the context of profiling epitope specificities and functional 4. Plotkin SA, Gilbert PB. Nomenclature for immune correlates of attributes of vaccine elicited antibody responses. Such analyses protection after vaccination. Clin Infect Dis. 2012;54(11):1615-1617. will enable a better understanding of how to elicit durable

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 9 Assessing Mucosal Immune Responses in HIV Vaccine Trials Tracey Day and Florian Hladik

HIV infection most frequently occurs via sexual transmis- sampling is more invasive than blood draws, and requires more sion of the virus through the mucosal surfaces of the genitals time and training for clinical personnel who take such samples. or rectum. The gut mucosa also serves as a site where the virus Sampling procedures also yield a lower amount of fluid or cells inflicts significant damage on the immune system during the compared to blood draws, and processing mucosal samples typi- first few weeks of infection. A primary focus of HIV preventive cally proves more challenging. vaccine research is therefore the induction of protective immune responses in these crucial early battlefields. Despite these challenges, the HIV Vaccine Trials Network (HVTN) has strived to enable mucosal immunity assessments Mucosal surfaces serve as a barrier between the body and in HIV vaccine trials. The large number of HVTN trials col- the environment. From harmless food matter to commensal lecting mucosal specimens reflects this priority (see Table 1). bacteria to pathogens, the mucosal surfaces of the respiratory, The HVTN Laboratory Program has developed procedures to gastrointestinal, and genitourinary tracts are constantly bom- process and analyze mucosal specimens, and actively collaborates barded with foreign particles. The mucosal immune system has with leading mucosal immunologists to further improve these the difficult task of differentiating between objects that should methods. These efforts are increasing the HVTN’s capacity to be ignored and those that represent a threat. To accomplish evaluate vaccine-specific mucosal responses to candidate preven- this balancing act, distinct anatomical and physiological fea- tive HIV vaccines. tures, which govern mucosal compartments, evolved as part of the immune system. Like systemic immunity, mucosal immunity employs all major lymphocyte subtypes -- B cells, CD4+, and CD8+ T cells, γδ T cells and natural killer cells -- as well as their antigen-presenting cellular counterparts, dendritic cells and macrophages.

Some specialized properties of the mucosal immune system have been identified -- the prevalence of different immunoglobulin subtypes, for example. However, due to technical difficulties in investigating human mucosal immune responses, the mechanisms of mucosal immunity in protection from pathogens are not as clearly understood as those of systemic immunity. While systemic immunity can be measured from peripheral blood, such responses do not necessarily reflect the responses in mucosal compartments. In a recent nonhuman primate vaccination study, Bomsel et al.1 found that protection from an intra- vaginal virus challenge was associated with anti-viral antibody responses in the vaginal mucosa that were not detected in blood. These results provide evidence that antiviral mucosal immune responses can be important in preventing HIV acquisition, and that these responses may be missed in peripheral blood analyses.2

Figure 1: Section of a colon biopsy that has been fluorescently stained to visualize Although HIV vaccine researchers have been interested in as- + + saying vaccine- and HIV-specific mucosal responses for many the CCR5 cells (shown in red) and the CD3 T cells (shown in white). Colors overlapping on the same cell indicate T cells that have the CCR5 HIV-receptor. years, a number of challenges have slowed progress in incor- The nuclei of all cells are also stained (blue) so that we can see the tissue structure porating such measurements into clinical research. Mucosal and how many total cells there are in the biopsy.

10 JUNE 2012 VOLUME 4:1 | HVTNEWS Assessing Mucosal Immune Responses in HIV Vaccine Trials

Current assessments aim to evaluate a broad range of mucosal Tissue biopsies provide samples of mucosal immune system immune responses and answer some key questions for HIV cells and can yield detailed information on cellular responses. vaccine development. Do vaccines delivered intramuscularly Currently, cellular responses are evaluated from foreskin, rectal, elicit detectable mucosal responses? Which mucosal immune re- colon, and other gastrointestinal tract biopsies (see Table 1). A sponses may be associated with protection from HIV infection? cervical cytobrush may be used to collect cells from the cervix, a Which mucosal specimens and assays are most relevant to the vulnerable site for HIV infection in women. detection of responses? The answers will be vital for clarifying what mucosal immune responses are capable of blocking HIV T-cell, B-cell, and innate immune cell phenotypes can be ana- infection, and for identifying vaccines that elicit such responses. lyzed from cellular samples by evaluating cell surface markers via flow cytometry, and by measuring gene expression using tran- Measuring Mucosal Immune Responses scriptional profiling. HIV-specific T- and B-cell functionality may be assessed from fresh cellular samples through intracellular To measure mucosal immune responses, a variety of mucosal cytokine staining and B-cell enzyme-linked immunosorbent specimens are collected from some or all trial participants. spot (ELISpot) assays. The organization of immune cells within

MUCOSAL SPECIMENS

obrush

a TRIAL al secretions al biopsy on biopsy saliv semen rect cervical secrcetrivoincasl cytrect col small intestfinoer ebsiokpinsy*

hvtn 076

hvtn 077

hvtn 078

hvtn 086/SAAVI 103

hvtn 088

hvtn 096

hvtn 097

hvtn 205

hvtn 505

hvtn 914

* Terminal ileum

Table 1: HVTN Opened / Planned Protocols that Include Vetted Mucosal Analyses.

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 11 Assessing Mucosal Immune Responses in HIV Vaccine Trials

the tissues can also be examined from tissue biopsies using Cutting Edge Technologies Optimize Output immunohistochemistry. Figure 1, for example, shows a colon from Mucosal Analyses biopsy sample in which T cells are identified by staining with Several U.S. and African MIG laboratories are using new tech- CD3 antibodies (in white). By simultaneously staining with a nologies to maximize the information obtained from mucosal CCR5 antibody (in red), the numbers of T cells expressing the samples collected in HVTN trials. For example, newer multi- CCR5 HIV co-receptor are visualized, as well as their locations laser flow cytometers and, recently, single-cell mass spectrometry in the tissue. cytometers that simultaneously detect more than 30 labeled antigens enable researchers to gain much more information Mucosal surface secretions can provide important information from the same number of cells.4 as well. HIV-specific mucosal antibodies and mucosal cytokine response profiles, for example, may be obtained from vaginal and Other new methods include bead-based immunosorbent rectal secretions, saliva, and semen. A variety of approaches exist approaches to measure small quantities of proteins in muco- to collect vaginal and rectal secretions including adsorbent wicks sal samples, and the use of computerized image analysis of and sponges. In the case of vaginal secretions, cups that collect antibody-stained biopsy sections (immunohistochemistry). small amounts of fluid from volunteers can also be used during Computers can measure 10- to 100-fold more cells, offering far clinic visits. more sensitivity than would manual analysis. Figure 1 shows a sample from one such analysis, in which CCR5 expression on T Peripheral blood samples that are collected during HVTN cells was digitally quantitated within colon biopsy tissue sections clinical trials are typically cryopreserved and shipped to the using this method. central laboratory for analysis. Performing immunogenicity analyses in a centralized lab ensures optimal data quality and Single-cell, subnanoliter well arrays, under development by allows data from multiple trials to be more readily compared. Chris Love at the Massachusetts Institute of Technology and Adopting this approach for mucosal specimens, however, is his collaborators, incorporates 80,000 to 250,000 wells onto challenging because the small number of immune cells pres- microscopy slides, with each well holding up to 125 pL of re- ent in mucosal samples, in conjunction with tissue-derived agents along with one cell.5 Those cells are labeled with specific proteases, make these samples particularly vulnerable during antibodies for phenotyping via multicolor cytometry. Because cryopreservation and subsequent thawing. In addition, the large the cells remain alive, they can be tested further for other func- variety of specimen types requires the development of custom- tions, such as whether they secrete particular cytokines. This ized cryopreservation methods to optimize cell viability for each combined phenotypic and immunofunctional analysis offers specimen type. Maximizing cell viability during the freezing and great advantages over both flow cytometry and in situ analysis. thawing process is critical for some immunogenicity assays, such Flow cytometers query each cell only once before it is discarded, as intracellular cytokine staining to detect HIV-specific T-cell and imaging of processed and sectioned tissue is limited to non- responses. This key assay requires that a certain number of cells functional assays. The single-cell platform permits quantitative remain viable during assaying, which currently limits its applica- analysis of antigen-specific T-cell responses among precisely tion to fresh mucosal samples. phenotyped T-cell subtypes, making it ideal for assessing mucosal cell immune responses to vaccines. To develop improved cryopreservation methods and assays for assessing mucosal immune responses in conjunction with HIV In acknowledgement that mucosal analyses are indispensable for vaccine trials, the HVTN Laboratory Program initiated a col- defining the immunological impact of HIV vaccines and micro- laboration with leading mucosal immunologists, the Mucosal 3 bicides, innovative leaps are being made to optimize their use in Immunology Group (MIG). With support from the National clinical trials. Recent findings of highly cross-reactive antibody Institute of Allergy and Infectious Diseases (NIAID), MIG is responses in HIV-infected individuals, and the uncovering of tasked with developing, optimizing, and standardizing mucosal systemic correlates of risk among those who received the RV144 specimen preserving and processing techniques. MIG also aims vaccine, put us closer to knowing what a vaccine should do to to develop standardized assays to measure and characterize protect against HIV. From here, we still require an iterative mucosal immune responses. A major benefit that standardized process to identify a vaccine that will induce the necessary im- methods provide is that they permit comparison of results across mune responses. It is truly an exciting time for those working to many studies, providing an opportunity for novel insights. prevent HIV infections.

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12 JUNE 2012 VOLUME 4:1 | HVTNEWS Highlights from Recent HVTN Publications Tracey Day

From primary protocol results to auxiliary research findings induce bnAbs when used together with a DNA prime. To to commentaries and review articles, HVTN researchers and enhance the exposure of the coreceptor binding site, a dele- collaborators have published a large number of articles over the tion was introduced in the V2 loop of Env. Although some past year. Here we present a tabular summary of our publica- bnAbs target regions of the V2 loop, its deletion was found to tions from February 2011 – February 2012 and highlight render the virus more susceptible to neutralization in previous several in more detail below. studies and thus warranted testing in human trials. In this trial, the trimeric recombinant protein was administered in MF59 Heterologous vs. Homologous Boosting adjuvant, following priming with gag and env DNA. The DNA (HVTN 068) vaccine component consisted of plasmid DNA adsorbed onto polylactide coglycolide microspheres, which have been shown Vaccine efficacy can be increased by administering multiple to improve immunogenicity of DNA vaccines in animal mod- vaccinations. This strategy, known as a “prime/boost” regimen, els. Remarkably, 100% of vaccinees developed nAb responses is used for many licensed vaccines and by several candidate against the homologous virus isolate, SF162. Neutralization HIV-1 vaccines. De Rosa et al. hypothesized that heterologous against heterlogous tier 2 virus isolates, which are typically vaccination could improve the quality of T-cell responses for more difficult to neutralize, however, was weak. Env-specific recombinant adenoviral serotype 5 (rAd5) vaccines. To test this CD4+ T-cell responses were also detected in the majority of hypothesis, immune responses in Ad5-seronegative adults were vacinees after boosting. The magnitude and potency of neutral- compared for a homologous prime boost regimen (rAd5 prime/ izing antibody responses in this trial were encouraging, but the rAd5 boost) and a heterologous prime boost regimen (DNA challenge of eliciting bnAbs remains.2 prime/rAd5 boost). The heterologous DNA prime/rAd5 boost regimen induced higher levels of CD4+ T cells and altered the Multiclade Prime/Boost Regimen cytokine and memory marker proﬁles of the T-cell responses. (HVTN 204) In contrast, homologous boosting did not increase T-cell responses, but enhanced Env-speciﬁc antibody responses. This The large genetic diversity of circulating HIV-1 strains makes suggests that to optimize both T-cell and antibody responses, development of a universal HIV vaccine challenging. To induce vaccine regimens may need to include both heterologous and responses against a variety of virus subtypes, the Vaccine homologous vaccine modalities. This study also evaluated Research Center (VRC) of the National Institute for Allergy the detailed kinetics of prime/boost responses. By assessing and Infectious Diseases developed a DNA prime rAd5 boost responses at many time points, researchers observed that al- vaccine regimen containing inserts from HIV-1 clades A, B, though limited responses were detected immediately following and C. Churchyard et al. report on the results of evaluating this DNA priming, the presence of a DNA prime influenced post- regimen in a phase 2a clinical trial conducted in healthy adults boost responses. The authors therefore propose that evaluations from South Africa, the United States, Latin America, and the of DNA vaccine immunogenicity should include assessment Caribbean. Similar to other studies involving this regimen, after boosting.1 the vaccines were well tolerated and appeared safe. The results from intracellular cytokine staining revealed that this regimen Neutralizing Exposure induced concurrent, polyfunctional CD4+ and CD8+ T-cell (HVTN 049) responses, as well as central and effector memory CD8+ T-cell responses with antiviral activity. High frequencies of durable Vaccines that elicit broadly cross-neutralizing antibodies Env-specific binding antibody responses were also gener- (bnAbs) against HIV-1 may be effective in preventing infec- ated; however, little virus neutralization activity was detected tion. However, immunogens capable of inducing bnAbs have from these antibody responses. The promising results of this not yet been identified. Alternative froms of HIV envelope and other clinical and preclinical studies contributed to the glycoproteins (Env) that expose conserved regions are be- advancement of this candidate to clinical efficacy testing in the ing investigated as a means to generate bnAbs. Spearman et phase 2b trial, HVTN 505.3 al. examined whether a modified trimeric Env vaccine could

Tables on following page, text continued on page 21...

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 13 Tabular Summary of Recent HVTN Publications

Primary Trial Publications:

Protocol First Author Aim* Result* Citation

HVTN 060, HVTN This first in humans study evaluated a DNA plasmid vaccine expressing HIV-1 Gag-p37 with or without a plasmid cytokine Kalams, S. The vaccine and adjuvants were well tolerated. The vaccine's immunogenicity was not improved by the addition of the adjuvants. PLoS One. 063 adjuvant (IL-12- or IL-15). The adjuvant addition was an attempt to augment the immunogenicity of the DNA vaccine. 2012;7(1):e29231

This study investigated IL-2 as an adjuvant by comparing HIV-1 DNA vaccine (VRC-HIVDNA009-00-VP) alone and with a Plasmid IL-2/Ig “increased immune responses when administered 2 days after the DNA vaccine, compared with simultaneous administration." HVTN 044 Baden, L. J Infect Dis. plasmid coding for a fusion protein of IL-2 and the Fc protein of immunoglobulin G (IL-2/Ig; VRC-ADJDNA004-IL2-VP). This study was the first to demonstrate that cytokine adjuvants can improve the immunogenicity of a DNA vaccine in humans. 2011;204(10):1541-1549

"Homologous boosting greatly enhanced Env-specific Ab responses, but did not increase T-cell responses. Despite limited postprime The study compared immune responses and kinetics of the responses of "homologous priming and boosting with a rAd5 HVTN 068 DeRosa, S. immune responses, DNA priming significantly enhanced the magnitude of postboost Env-specific Ab and CD4+ T-cell responses and J Immunol. vaccine (VRC-HIVADV014-00-VP) to heterologous DNA prime (VRC-HIVDNA009-00-VP) and rAd5 boost vaccination." 2011;187(6):3391-3401 influenced the cytokine and memory marker profiles of the boosted T-cell responses."

A comparison of a DNA vaccine prime (VRC-HIVDNA009-00-VP) given intramuscularly and a rAd5 vaccine boost (VRC- The vaccine's immunogenicty was not affected by giving the rAd5 boost via different administration routes. In addition, there was a higher HVTN 069 Koblin, B. PLoS One. HIVADV014-00-VP) given one of 3 routes: intramuscularly, intradermally, or subcutaneously. frequency of local reactions after intradermal or subcutaneous administration. 2011;6(9):e24517

A phase II trial that evaluated the safety and immunogenicity of a multiclade DNA prime (VRC-HIVDNA-016-00-VP) and a The regimen was well-tolerated and induced IFN- + T cells in 70.8% of participants. T-cell responses were predominantly balanced CD4+/ HVTN 204 Churchyard, G. γ PLoS One. multiclade rAd5 boost (VRC-HIVADV014-00-VP) vaccine in healthy adults from various geographical locations. CD8+ polyfunctional responses. High frequencies (83.7%–94.6%) of multi-clade anti-Env binding antibodies were also observed. 2011;6(8):e21225

This trial was stopped early because the vaccine was not efficacious in another study (Step study). The available data suggested no HVTN 503/ protection from acqusition or reduction of viral load setpoint for vaccine recipients. The vaccine induced IFN- -secreting T-cell responses Gray, G. Assessed safety and efficacy of the MRKAd5 HIV-1 subtype B vaccine in South Africa, a subtype C region. γ Lancet Infect Dis. Phambili gag/pol/nef to clade B (89%) and C peptides (77%). A trend for a lower viral-load setpoint and a slower decline in CD4 count was observed in 2011;11(7):507-515 vaccinated women.

The first human trial of a vaccine regimen consisting of a DNA prime (pCMVgagB, pSINenvB; PLG) and a trimeric, V2- This regimen generated Env-specific CD4+ T cell but not CD8+ T-cell responses and high titers of NAbs against the homologous SF162 HVTN 049 Spearman, P. J Infect Dis. deleted Env protein boost (gp140SF162DV2; MF59) aimed to elicit T-cell responses and bNAbs. virus. The Ab responses however, had limited breadth and did not neutralize heterologous, tier 2 clade B virus strains. 2011;203(8):1165-1173

A phase I trial that Compared the safety and immunogenicity of a recombinant MVA vaccine (GeoVax MVA/HIV62) The MVA62 vaccine was well tolerated and appeared safe. Different patterns of immune responses were induced when MVA62 was HVTN 065 Goepfert, P. J Infect Dis. administered alone or with a DNA vaccine prime (GeoVax pGA2/JS7 DNA). administered alone (higher Ab responses) or together with the JS7 DNA vaccine (better T-cell responses). 2011;203(5):610-619

This first in humans study examined the safety and immunogenicity of homologous and heterologous prime-boost Both the homologous and heterologous prime-boost regimens were well tolerated. Balanced HIV-specific CD4+ and CD8+ T-cell responses 2011;29(10):1948- HVTN 055 Keefer, M. Vaccine. regimens with different replication-defective poxvirus vectors (MVA-HIV and FPV-HIV) containing identical HIV inserts. were more reliably elicited by the heterologous prime boost strategies. 1958

Review, Commentary, and Methods Publications:

Protocol First Author Aim* summary* Citation

Describes strategic accomplishments made by the HIV Vaccine Trials Network. These acomplishments improve the process of designing 2012; Review Kublin, J. HIV Vaccine Trials Network: activities and achievements of the first decade and beyond. Clinical Investigation. and implementing HIV vaccine clinical trials, as well as analyzing their results. 2(3): 245-254

Articulates the scientific value of principal stratification estimands for randomized, placebo-controlled, preventive HIV vaccine efficacy Commentary Gilbert, P. Commentary on "Principal stratification - a goal or a tool?" by Judea Pearl. Int J Biostat. trials. 2011;7(1):Article 36

Describes how immune correlates may be determined from vaccine efficacy trials and from the RV144 trial in particular. Discusses how AIDS Res Hum Review Rolland, M. Evaluating immune correlates in HIV type 1 vaccine efficacy trials: what RV144 may provide. clinical trial designs may be augmented to improve the assessment of correlates. Retroviruses. 2012;28(4):400-404

Genetic studies have identified polymorphisms in genes associated with the innate immune response to HIV infection, including TLR4, Review Sobieszczyk, M. Host genetic polymorphisms associated with innate immune factors and HIV-1. Curr Opin HIV AIDS. TLR9, IRF-3, TRIM5α and the ABOBEC3 gene family, that contribute to protection from disease progression. 2011;6(5):427-434 The assay, based on the hydrolysis of a fluorogenic peptide substrate by granzyme B, provides "rapid quantification of HIV-1 or SIV- Methods Pollara, J. High-throughput quantitative analysis of HIV-1 and SIV-specific ADCC-mediating antibody responses. Cytometry A. specific ADCC-mediating antibodies that develop in response to vaccination, or in the natural course of infection." 2011;79(8):603-612

The use of standardized software, such as LabKey Server's neutralizing antibody (nAb) tool, for analyzing, sharing, and archiving assay 2011 May Methods Piehler, B LabKey Server nAb: a tool for analyzing, visualizing and sharing results from neutralizing antibody assays. BMC Immunol. results improves "the reproducibility, reliability, and comparability of data obtained across many labs." 27;12:33

Proposes to characterize a marker’s "principal surrogate value based on the distribution of risk difference between interventions" and Methods Huang, Y Comparing biomarkers as principal surrogate endpoints. Biometrics. develops "a semiparametric estimated-likelihood method to estimate the joint surrogate value of multiple biomarkers." 2011;67(4):1442-1451

Discusses the rationale behind adaptive trial designs, in which multiple candidates are initially tested and only those showing efficacy Sci Transl Med. Review Corey, L. HIV-1 vaccines and adaptive trial designs. signals are carried through the extended study. This allows a more rational faster paced strategy for vaccine development and improves 2011;3(79):79ps13 the ability to identify immune correlates.

Statistical interpretation of the RV144 HIV vaccine efficacy trial in Thailand: a case study for statistical issues in efficacy Other Gilbert, P. Discusses the interpretation of statistical results from RV144 generated using frequentist statistics and Bayesian framework analyses. J Infect Dis. trials. 2011;203(7):969-975

2011;15 Suppl Other Landers, S Community perspectives on developing a sexual health agenda for gay and bisexual men. Discusses the history and next steps for advocacy for the health of gay and bisexual men of color. AIDS Behav. 1:S101-106

14 JUNE 2012 VOLUME 4:1 | HVTNEWS Article highlighted in HVTNews article on page 13. Primary Trial Publications: * Text adapted or quoted from original articles.

Protocol First Author Aim* Result* Citation

Review, Commentary, and Methods Publications:

Protocol First Author Aim* summary* Citation

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 15 Tabular Summary of Recent HVTN Publications (cont.)

Auxiliary Study Publications:

Protocol First Author Aim* Result* Citation

Higher frequencies of adenovirus-specific CD4+ T-cell responses prior to vaccination were associated with decreased levels of HIV-specific Analyzed adenovirus-specific T cells in recipients of the MRKAd5 HIV-1 vaccine and evaluated HVTN 071, HVTN 502 Frahm, N. gag/pol/nef CD4+ T-cell responses, and decreased breadth of CD8+ T-cell responses. These effects were observed to be independent of pre-existing J Clin Invest. their impact on vaccine-induced HIV insert-specific T-cell responses. 2012;122(1):359–367 adenovirus-specific nAb titers.

HVTN 502/ Merck 023/ The study determined nAb titers for alternative adenovirus serotypes (Ad5, Ad26, Ad35, and Ad48) in 2011;29(32):5203- Barouch, D. "Ad26, Ad35, and Ad48 nAb titers were substantially lower than Ad5 nAb titers in multiple large international human populations." Vaccine. Step Study individuals from the Americas, sub-Saharan Africa, and Southeast Asia. 5209

“Investigated the epitopes and HLA molecules utilized to mount HIV-specific CD8+ T-cell responses in healthy "Certain HLA alleles (B27, B57, B35, and B14) are preferentially utilized to mount HIV-specific responses, whereas other alleles (A02 and J Acquir Immune Defic HVTN 050, HVTN 054 Friedrich, D. HIV-seronegative individuals who participated in clinical trials” of MRKAd5 HIV-1 gag and VRC-HIVADV014- Syndr. 2011;58(3):248-252 B07) are rarely utilized (P < 0.001)." 00-VP rAd5 vaccines.

HVTN 502/ Merck 023/ Examined the motivators and deterrents to HIV vaccine trial participation among women at 'high risk' for HIV The majority of women interviewed were positive about participating in HIV vaccine trials but “were ultimately deterred by concerns about 2011;29(36):6130- Voytek, C. Vaccine. Step Study acquisition. research procedures, potential consequences of participation, and/or their ability to attend study appointments.” 6135

Evaluated "whether the T-cell epitope response specificities elicited by vaccination were within conserved HVTN 502/ Merck 023/ or variable regions of the HIV proteins and whether the vaccine elicited the type of T-cell coverage likely to The study found that "the Merck vaccine tended to elicit responses to fewer highly conserved epitopes and more less- Li, F. gag/pol/nef PLoS One. Step Study be effective after encountering the diversity of HIV-1 circulating in the regions where the vaccine trial was conserved epitopes than would be predicted from the vaccine sequences alone." 2011;6(6):e20479 conducted".

HVTN 502/ Merck 023/ Evaluated a novel method for predicting HLA alleles using unphased flanking single-nucleotide The data support the application of this approach for predicting HLA alleles with genome-wide association study-derived single-nucleotide 2011 Apr Zhang, X. BMC Genet. Step Study polymorphism genotypes in ethnically diverse populations. polymorphism data. The predictive models and the program used are available online. 25;12:39

Devised a novel assay to measure 30 secreted factors from ex vivo-stimulated PBMC to examine "the "The cytokine response profile of Gag-specific T cells mirrored the Ad5-specific response present in all subjects before vaccination, and HVTN 050 Pine, S. relationship between pre-existing Ad5 immunity and T-cell cytokine response profiles in healthy, HIV- included a number of Th1- and Th2-associated cytokines not routinely assessed in current vaccine trials, such as IP-10, IL-10, IL-13, and PLoS One. 2011;6(4):e18526 uninfected recipients of MRKAd5 HIV-1 gag vaccine." GM-CSF."

HVTN 502/ Merck 023/ Investigated the epidemiology of herpes simplex virus type 2 (HSV-2) infection "and its impact on HIV Pre-existing HSV-2 infection was a significant risk factor for HIV acquisition in MSM participating in the Step study. However, past HSV-2 Barnabas, R. J Acquir Immune Defic Step Study incidence, viral load, and time to antiretroviral therapy initiation among Step study participants." infection was not associated with increased HIV viral load or early disease progression. Syndr. 2011;57(3):238-244

HVTN 502/ Merck 023/ Investigated the mechanisms associated with variability in HIV-specific T-cell responses to the MRKAd5 HIV- No genetic variant was found to reach significance; however polymorphisms in MHC alleles HLA-B*2705, B*5101, and B*5701 were Fellay, J. J Infect Dis. Step Study 1 gag/pol/nef vaccine using whole-genome genotyping and HLA class I typing. associated with higher Gag responses, whereas HLA-B*0801 and B*4501 were associated with lower responses. 2011;203(6):773-779

No significant differences between vaccine and placebo recipients were found with regard to HIV RNA levels, CD4+ T-cell counts, time to HVTN 502/ Merck 023/ Evaluated the impact of the MRKAd5 HIV-1 vaccine on disease progression in participants who Fitzgerald, D. gag/pol/nef initiation of anti-retroviral therapy, and AIDS-free survival. The data hinted that vaccinees with protective HLA types (B*27, B*57, B*58) J Infect Dis. Step Study became HIV-1 infected during a 2 year follow-up period. 2011;203(6):765-772 had lower HIV RNA levels than placebo recipients with those HLA types.

HVTN 502/ Merck 023/ Performed a sieve analysis of breakthrough virus sequences from infected Step study participants to The study found that HIV sequences from infected vaccinees had greater distances to the vaccine sequence than those for infected placebo 2011;17(3):366- Rolland, M. Nat Med. Step Study determine whether the MRKAd5 HIV-1 gag/pol/nef vaccine put immune pressure on the infecting viruses. recipients. The first evidence of selective pressure from vaccine-induced T-cell responses on HIV-1 infection in humans. 371

Compared HIV-specific CD8+ T-cell cytotoxic capacity (granzyme B target cell activity and HIV-infected CD4+ HVTN 071, HVTN 502/ Merck Vaccine recipients with "HLA class I alleles B*27, B*57, or B*58, which have been associated with immune control over HIV replication in Migueles, S. T-cell elimination) for uninfected MRKAd5 HIV-1 vaccine recipients, HIV-negative individuals, PLoS Pathog. 023/ Step Study, HVTN 905-02 gag/pol/nef chronic infection," had higher T-cell cytotoxic capacity, suggesting these alleles allow better cytotoxic priming in infection and vaccination. 2011;7(2):e1002002 and chronically infected patients.

16 JUNE 2012 VOLUME 4:1 | HVTNEWS Article highlighted in HVTNews article on page 13. * Text adapted or quoted from original articles. Auxiliary Study Publications:

Protocol First Author Aim* Result* Citation

Abbreviations: Ab = antibody, Ad5 = adenovirus type 5, ADCC = antibody dependent cell-mediated cytotoxicity, bNAb = broadly neutralizing antibody, CTL = cytotoxic T lymphocyte, FPV = fowlpox virus, MSM = men who have sex with men, MVA = modified vaccinia Ankara, nAb = neutralizing antibody, PBMC = peripheral blood mononuclear cell, PLG = polylactide coglycolide microspheres, rAd5 = recombinant adenovirus type 5, VE = vaccine efficacy,

Continued on page 21...

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 17 Social and Behavioral Science in Clinical Trials of Biomedical HIV Prevention Interventions Beryl Koblin, Michele Peake Andrasik, and Judy Austin

The HIV prevention toolbox has been a mixed bag in recent cious interventions. years, with the success of several randomized clinical trials of biomedical interventions in reducing HIV infections, and Documentation of HIV exposure, through self-report of the failure of others to show efficacy. A trial of a recombinant risk behaviors, is used to determine if exposure to the virus is canarypox vector vaccine prime with a recombinant glycopro- equivalent in the different arms of the study at baseline and tein 120 subunit vaccine boost found a modest efficacy (31%) during followup, and if treatment assignment is associated among a general population sample in Thailand (RV144),1 but with any changes in exposure. As mentioned above, differential the Step study found that a replication-defective adenovirus exposure may obscure efficacy. Collection of risk behavior data type 5 (Ad5) vector vaccine with subtype B HIV-1 gag/pol/nef can be used to determine if the level of HIV exposure could act inserts did not protect against HIV infection, and appeared to as an effect modifier to biomedical intervention efficacy. increase the risk of HIV acquisition among MSM who were uncircumcised or had neutralizing antibodies to Ad5 at enroll- Randomization and blinding are in place in clinical trials to ment.2 The Partners PrEP study, which looked at daily oral tenofovir or tenofovir with emtricitabine, provided a 67% and 75% reduction in HIV infection rates, Both individual [behavioral] and contextual [social] respectively, among serodiscordant hetero- 3 factors may influence participation and retention sexual couples. Conversely, a study of daily oral use of tenofovir and emtricitabine rates, adherence, and risk of HIV exposure, as well among heterosexual women (FEM-PREP) did not find any reduction in HIV infec- as dissemination and uptake of highly efficacious tion (HR= 0.94; 95% CI, 0.59-1.52). interventions. The range of efficacies observed with different biomedical prevention interventions could be explained by a number of factors. The biomedical eliminate imbalances in exposure by treatment arms.7 However, intervention itself may not be of sufficient strength or appro- randomization does not preclude a systematic or differential priately targeted to have an impact on HIV. Host factors, such shift in risk occurring during follow-up. Unblinding of treat- as genetics or gender, may influence efficacy.4 Insufficient dos- ment assignment during the study, or perceptions of treatment ing levels, as a result of low adherence, has been the proposed assignment while blinded, may contribute to changes in risk mechanism for the range of efficacies reported in pre-exposure behaviors, leading to changes in exposure to HIV.8 prophylaxis (PrEP) studies.5 Finally, if HIV exposure is not equivalent in the different treatment arms in a clinical trial, the Recent studies within the HVTN illustrate the critical role of ability to detect an efficacious intervention may be undermined. social and behavioral science work in the conduct and interpretation of recent HIV vaccine trials. Behavioral science provides the backbone for investigating several of these factors at the individual level. Social science The HVTN has utilized behavioral investigations in the Step provides the structure to investigate context at the dyadic, study, to determine if the increased risk of HIV acquisition situational and community levels in which individuals oper- among subgroups of vaccinees, compared to placebo recipients, ate.6 Both individual and contextual factors may influence could be explained by differences in HIV exposure. The analysis participation and retention rates, adherence, and risk of HIV found few differences in sexual risk behaviors by treatment exposure, as well as dissemination and uptake of highly effica- arm, and vaccine recipients remained at higher risk of HIV in-

18 JUNE 2012 VOLUME 4:1 | HVTNEWS Social and Behavioral Science in Clinical Trials of Biomedical HIV Prevention Interventions

fection compared to placebo recipients, even when controlling to the community, focus on including participant testimonials for sexual risk behaviors and other potential confounders (HR showcasing participants’ experience in the study, and collect =2.8; 95% CI, 1.7-6.8). Thus, increased risk of HIV infection survey data on experiences of current trial participants. In among subgroups of MSM who received the vaccine was not addition, the HVTN has developed 2 cultural responsive- explained by an increase in HIV exposure in these groups of ness curricula for use at clinical research sites (CRS) and with MSM, ruling out behavioral differences and lending support Core staff. A 5 hour transgender specific cultural responsive- to a biological mechanism.9 ness workshop was provided at the Spring 2012 HVTN Full Group Meeting. A more general full day cultural respon- Participant recruitment, as any recruiter can attest, is an siveness workshop was pilot tested and evaluated with CRS art and, importantly, a science. Recent social science work in staff at 2 sites. Evaluation results were utilized to improve the HVTN provides important insights into recruitment of the curriculum and the workshop was then provided to all MSM and transgender women into HVTN 505. Data from HVTN Core staff. The evaluations and feedback from these surveys (n=1835) and 6 focus groups (n=62) with MSM in 6 workshops will lead to additional improvements in the overall cities (Boston, Chicago, Denver, Houston, Los Angeles and curriculum and plans to make the training available to all sites New York) indicated that more than 70% of eligible MSM are underway. would consider participating in an HIV vaccine study. Lack of knowledge and information about HIV vaccine trials emerged The HVTN has made a significant commitment to inclusion as the primary barrier to participation. Almost 40% of survey of social and behavioral science into its work. In support of respondents indicated having never heard about a vaccine trial, and another 20% indicated hearing about it as a result of taking the survey. Among participants who The HVTN has made a significant commitment to were unsure of their interest in participating (n=254), 63% indicated that interest inclusion of social and behavioral science into its in participating would increase if they had more information. Additional barriers to work. participation included: concerns about side effects including vaccine induced seropositivity (VISP) (77%); concerns about privacy (54%); the mission and goals of the HVTN, the newly formed Social concerns about others perceiving them as “risky” (55%); and and Behavioral Working Group (SBWG) examines social and acquiring HIV from the vaccine (43%, with 31% endorsing behavioral science questions that have an impact on the design, not sure). Participation facilitators included: safety (92%); implementation and interpretation of vaccine trials. These helping to end the epidemic (90%); and potential protection questions include behavioral risk assessment, adherence mea- from HIV (60%). The increased dissemination of community- sures for protocols that would involve the use of PrEP (such level information about the progress that has been made in as HVTN 095-MTN 022), and assessment of understanding HIV vaccine research is sorely needed, as 64% of respondents of the informed consent. The SBWG is identifying social and endorsed believing that there was very little chance of finding behavioral science research priorities within the Network, to a vaccine that works. Two focus groups with African Ameri- facilitate the integration of behavioral and social science work can gay male leaders (N= 15), 4 focus groups with male-to- into existing and future HVTN efforts. Another goal is to female transgender women (N=42) and 6 focus groups with assist investigators in incorporating social and behavioral sci- HVTN Clinical Research Site (CRS) staff (N= 74) were also ence into their research agenda. These efforts will be defined as conducted. Data from these groups highlighted the need for specific objectives related to the overall protocols, employing improved cultural responsiveness throughout the Network and well-established and appropriate behavioral and social science the identification of innovative approaches to strengthen com- methodologies for the questions at hand. munity and research institution partnerships. The current research priorities of the SBWG include: Based on these results, the HVTN is planning to enhance measuring and describing PrEP use among HVTN 505 VISP and other adverse event educational materials available participants,

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 19 Social and Behavioral Science in Clinical Trials of Biomedical HIV Prevention Interventions

examining biomarkers of sexual exposure to HIV through Reference List collaboration between the SBWG and laboratory scientists, 1. Rerks-Ngarm S, Pitisuttithum P, Nitayaphan S, et al. Vaccina- tion with ALVAC and AIDSVAX to Prevent HIV-1 Infection in developing methods and measures for optimal community Thailand. N Engl J Med. 2009. engagement, 2. Buchbinder SP, Mehrotra DV, Duerr A, et al. Efficacy assess- evaluating recruitment and retention strategies, ment of a cell-mediated immunity HIV-1 vaccine (the Step Study): a double-blind, randomised, placebo-controlled, test-of-concept trial. developing new approaches to informed consent assess- Lancet. 2008;372(9653):1881-1893. ment of understanding, 3. Baeten JM, Donnell D, Ndase P, et al. ARV PrEP for HIV-1 improving assessment of understanding, prevention among heterosexual men and women. 19th Conference on evaluating risk factors for HIV infection in different Retroviruses and Opportunistic Infections, Seattle, WA, March 5-8 populations (including substance users), Section 8, Paper #29. 2012. providing data for efficacy trial design, and 4. Stanberry LR, Spruance SL, Cunningham AL, et al. Glyco- protein-D-adjuvant vaccine to prevent genital herpes. N Engl J Med. collaborating closely with the NIH's Division of AIDS' 2002;347(21):1652-1661. and the Office of HIV/AIDS Network Coordination's efforts to develop core measures of risk behaviors. 5. Grant RM, Lama JR, Anderson PL, et al. Preexposure chemo- prophylaxis for HIV prevention in men who have sex with men. N An overarching principal is to incorporate social and Engl J Med. 2010;363(27):2587-2599. behavioral science early in study design and idea generation, to 6. Frye V, Latka MH, Koblin B, et al. The urban environment and ensure appropriate integration of the necessary expertise. As sexual risk behavior among men who have sex with men. J Urban PrEP use becomes more common, the issues of acceptability, Health. 2006;83(2):308-324. adherence, and measurement of HIV exposure through different routes (eg, vaginal vs. anal sex) will become even 7. Schulz KF, Grimes DA. Generation of allocation sequences in more complicated. The upcoming HVTN 095-MTN 022 trial, randomised trials: chance, not choice. Lancet. 2002;359(9305):515- 519. testing HIV vaccines in combination with oral and topical antiretrovirals, will begin to explore some of these issues. Thus, 8. Bartholow BN, Buchbinder S, Celum C, et al. HIV sexual risk biomedical and social-behavioral sciences, often perceived behavior over 36 months of follow-up in the world's first HIV vac- as opposites, are, in essence, complementary and interactive cine efficacy trial. J Acquir Immune Defic Syndr. 2005;39(1):90-101. within a greater whole. 9. Koblin BA, Mayer KH, Noonan E, et al. Sexual risk behaviors, circumcision status and pre-existing immunity to adenovirus type 5 Beryl Koblin is Principal Investigator, Laboratory of Infectious among men who have sex with men participating in a randomized Disease Prevention, New York Blood Center; Michele Peake HIV-1 vaccine efficacy trial: Step Study. J Acquir Immune Defic Syndr. Andrasik is the HVTN Social Scientist; Judy Austin is a doctoral 2012. student in Epidemiology at the Mailman School of Public Health, Columbia University.

20 JUNE 2012 VOLUME 4:1 | HVTNEWS Highlights from Recent HVTN Publications

"Highlights from Recent HVTN Publications", continued from page 13

Hint on Helpful HLAs greater responses to the vaccine-matched clade B antigens, (Step Study) most developed clade C responses as well. Although premature cessation of the trial limited what conclusions could be drawn, T-cell responses play an important role in controlling virus unlike the Step study, there was no increase in risk of HIV-1 replication during natural HIV-1 infection. Vaccines that infection observed in Ad5-seropositive or uncircumcised men. elicit strong T-cell responses, therefore, may lower viral loads Post-hoc analyses indicated a trend toward lower viral load set- and slow disease progression. The Merck adenovirus type points and slower CD4+ T-cell decline for female vaccinees, as 5 trivalent HIV-1 vaccine (MRKAd5/HIV-1 gag/pol/nef), compared to placebo recipients. These data suggest that future which elicits T-cell responses, was evaluated in an efficacy trial, efficacy trials could stratify by sex to examine potential differ- known as the Step study. In 2007, the study was stopped when ences in vaccine efficacy for females and males.5 the first interim analysis indicated that the vaccine would not achieve efficacy for preventing HIV-1 acquisition or reduc- Panning for Vaccine-Induced Pressure ing plasma HIV-1 RNA levels. Fitzgerald et al. evaluated the (Step Study) effect of the vaccine on disease progression in participants who became HIV-1 infected during a 2 year follow-up period. The MRKAd5/HIV-1 vaccine evaluated in the Step study Among 87 male study participants who became HIV-1 in- induced HIV-specific T-cell responses in more than 75% of fected no significant differences between vaccine and placebo vaccinated subjects, yet it did not prevent infection or reduce recipients were found with regard to several disease progres- + viral load set point. Rolland et al. investigated whether the sion measures (HIV-1 RNA levels, CD4 T-cell counts, time vaccine-elicited T-cell responses affected virus strains of the to initiation of anti-retroviral therapy, and AIDS-free survival). + breakthrough infections. A “sieve analysis” approach was used Given the magnitude of CD8 T-cell responses elicited by the to test whether immunization had the effect of sifting out cer- vaccine these results point to a need for additional research to tain virus strains. HIV-1 sequences isolated from 68 infected elucidate why the vaccine failed to control viral replication. In- vaccine and placebo recipients were compared with the vaccine terestingly, a subanalysis of the data hint that the vaccine may insert sequence. Viruses infecting vaccine recipients were more have reduced HIV-1 RNA levels in participants with certain likely to encode epitopes that differed from those present in HLA types (HLA class I alleles B*27, B*57, and B*58) known the vaccine. The most significant site distinguishing vaccine to be associated with lower viral load setpoints and delayed and placebo recipients was Gag amino acid 84. Differences disease progression. These observations suggest that reduction + between sequences of viruses infecting vaccine versus placebo of viral loads may be possible if the right CD8 T-cell respons- recipients were confined to Gag, Pol, and Nef, the portions of es are induced. Further research is needed to identify what 4 the virus that were included in the vaccine. These data support these responses are and how to induce them via vaccination. the notion that vaccine-induced T-cell pressure led to a high rate of mismatch among vaccine recipient founder sequences. Clade Crisscross The data represent the first demonstration of a vaccine exerting (Phambili) immune pressure on HIV-1 and provide impetus for designing vaccines to elicit cytotoxic T lymphocyte responses. The Cross-clade immunity has been shown to develop in indi- researchers propose that new vaccines should aim to trigger viduals infected with HIV-1 and in uninfected individuals mutations in vulnerable regions of HIV-1, potentially ‘corner- vaccinated with a HIV-1 clade-specific vaccine. A test of ing’ the virus into adopting attenuated forms.6 concept study was initiated to assess efficacy of the MRKAd5/ HIV-1 clade B vaccine in South Africa, where there is a high Troubling T-cells prevalence of HIV-1 and the major circulating strain is clade (Step Study/HVTN 071) C. The trial was stopped after the same vaccine was found to be ineffective in the Step study. Gray et al. report that, like HIV-1 insert-containing adenovirus serotype 5 (Ad5) vector the Step study, the MRKAd5/HIV-1 vaccine did not prevent vaccines induce strong HIV-specific T-cell responses. How- HIV-1 infection or lower viral load setpoint among South ever, pre-existing Ad5 immunity can dampen the immune African participants. This was despite having induced a high responses induced by these vaccines. Research has focused frequency and magnitude of T-cell responses as measured by more on the role of Ad5-specific antibodies in reducing Ad5 γ interferon- ELISpot. Although vaccine recipients developed vaccine immunogenicity. Using samples from clinical stud-

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www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 21 HVTN Annual Network Award Winners On May 30th 2012 in Washington DC, the HVTN honored individuals who perform outstanding work at our sites

Keith Barsky (Center), HVTN Service Award winner. HVTN service Award HVTN Mentoring Award

Award Recipient: Award Recipient: Keith Barsky Magdalena Sobieszczyk, MD, MPH Recruiter, Orlando Co-Principal Investigator, New York

The HVTN Service Award recognizes individuals who The HVTN Mentoring Award recognizes an individual have creatively responded to a particular challenge arising who has demonstrated extraordinary commitment as a in their CRS work, and by which value has been added to mentor by promoting research, training/education, and the CRS / HVTN enterprise. professional development of those they have mentored. Accolades for Keith Accolades for Magda

“Keith is the only recruiter at OIC [Orlando Immunology “What I really appreciate about her is that she is completely Center] for the HVTN, which means he has spoken to almost non-intimidating and welcoming, which allows me to ap- every person that has been screened or enrolled at our site. proach her with any question or concern that I have with my His rate of converting a study contact into a screen is among work and research. I think this is a particularly important the highest of all the HVTN centers; all thanks to his excel- quality for people mentoring early researchers, who may find lent communication skills.” the whole process and experience very intimidating. Having someone like Dr. Sobieszczyk as a mentor and guide has “The path that Keith has paved between our center and the really helped me grow as a researcher, as well as know how gay community in Orlando has resulted in strong relation- to be a good mentor myself for those I will work with in the ships which I suspect will ultimately help provide more future.” awareness about HIV and STD prevention in Orlando.” “Magda makes training and mentorship a priority by “Prior to the HVTN, there was very little collaboration providing incredible support for her staff and seeking out between city organizations that serve the HIV-infected educational and training opportunities. She challenges us to and at-risk populations in Orlando. Keith was able to form become analytical and thoughtful researchers. Regardless of strong ties with the HIV community and gay organiza- her own workload, she is always available as a compassion- tions.” ate listener and to offer her knowledge and expertise.”

“She sets a marvelous example. Her kindness and generosity equal her intelligence and hard work!”

22 JUNE 2012 VOLUME 4:1 | HVTNEWS Kyle Rybczyk (Center), HVTN Citizenship Award winner. Octavio Valente, Jr. HVTN Citizenship Award Volunteer Service Award

Award Recipient: Award Recipient: Kyle Rybczyk, FNP-BC Eduviges Cuello Clinic Coordinator, Nashville Community Advisory Board (CAB) member, Dominican Republic The HVTN Citizenship Award is for individuals Octavio Valente, Jr. was a dedicated CAB member from deserving special recognition for good citizenship with Rio de Janeiro, Brazil who passed away on March 21, regard to consistent, reliable, and quality performance of 2006. Octavio served many roles in the HVTN and CRS duties and activities. in the HIV/AIDS community through his advocacy, Accolades for Kyle dedication and spirited energy.

“In my entire career, I have not had the pleasure of In his honor, this service award is given to a CAB working on a team led by someone more vision-oriented member who has demonstrated exemplary leadership and and committed to excellence than Kyle Rybczyk. I am dedication in the HVTN. beyond impressed by the diligence he consistently displays in ensuring a quality experience for each and every volunteer Accolades for Eduviges who walks through our doors. Furthermore, our site auditor, “Eduviges was the First HIV Vaccine volunteer in the Do- on several occasions, has remarked of the near unparalleled minican Republic. Motivated by that experience, she joined excellence that Kyle inspires with regards to documentation the CAB of the Instituto Dermatológico y Cirugía de Piel and accurate clinical process at our site. Kyle is an (IDCP) in Santo Domingo. She has advocated on the impor- outstanding citizen within the HVTN and also within our tance of community participation in prevention research at community here in Nashville. He encourages those around several different international forums, and has been given a him to identify the best within themselves and to strive for national award in the Dominican Republic for her contri- consistently making that best of themselves a reality. I am bution to the advancement of public health.” honored to nominate Kyle for this award.” “Eduivges has served on the HVTN Global CAB, its Ethics and Scientific Working Groups, HANC Partners, and the HVTN 907/906 protocol team. All of this is a testament to her commitment to the defense and protection of the rights of volunteers participating in vaccine studies, contributing significantly to shine the light on the role of activists in the fight against the HIV/AIDS.”

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 23 HVTN Protocols -- Enrolling or in Active Follow Up

Start HVTN# Products and Inserts N Sites Description Date pGA2/JS7 NDA MVA/HIV62; Atlanta; Birmingham; Boston; Iquitos; Lima; Nashville; New York; A phase 2a clinical trial to evaluate the safety and tolerability of a prime/boost regimen of the GeoVax DNA vaccine and modified vaccinia Ankara (MVA) vaccine, or of Jan-09 205 env, pol, gag, 120 tat, rev, vpu, PR, RT Rochester; San Francisco; Seattle MVA vaccine alone, in healthy, HIV-1 uninfected, vaccinia-naïve individuals. Immune responses will be tested in rectal and genital secretions as well as in blood.

DNA/rAd5 Atlanta; Birmingham; Boston; Nashville; New York; Rochester; San A phase 1b clinical trial to evaluate the safety and immunogenicity of recombinant adenoviral serotype 35 (rAd35) and serotype 5 (rAd5) HIV-1 vaccines when given as a Feb-09 077 192 DNA/rAd35; env Francisco; Seattle heterologous prime/boost regimen or as boosts to a recombinant DNA vaccine in healthy, Ad5-naïve and Ad5-exposed, low risk, HIV-1 uninfected adult participants.

A longitudinal study of women at risk for HIV-1 infection, to inform HIV vaccine trial participation. The primary objectives are 1) To determine the feasibility of recruiting Apr-09 907 No Product 800 Port-au-Prince; San Juan; Santo Domingo and retaining women at risk of HIV infection, with a focus on female sex workers with demographic, behavioral, or other social factors associated with high prevalence of HIV; and 2) To determine whether the HIV incidence rate in the longitudinal cohort is at least 1% per year.

A phase 2b, randomized, placebo-controlled, test-of-concept trial to evaluate the effect of the VRC DNA/rAd5 vaccine regimen on the rate of HIV-1 acquisition, and on May-09 505 DNA/Ad5; 2200 All U.S. Sites env, nef, pol, gag HIV-1 viral load setpoint, compared to placebo participants who are diagnosed with HIV-1 infection at or after Day 196 post-enrollment through Month 24 visit.

rAd5 vaccine A phase 1b clinical trial to evaluate the safety and immunogenicity of heterologous prime/boost vaccine regimens (NYVAC-B/rAd5 vs. rAd5/NYVAC-B) in healthy, HIV-1 Aug-09 078 80 Lausanne NYVAC-HIV-B; env , gag, pol, nef uninfected, Ad5 nAb seronegative adult participants.

A phase 1b clinical trial is to examine the role of genetic background in governing the immune responses following vaccination with HIV antigens delivered via a DNA- Jan-10 082 DNA/Ad5; 8 Boston; Rochester; San Francisco; Seattle env, gag, pol, nef prime Ad5-vectored boost regimen.

A phase 1 clinical trial to assess the safety, tolerability, and ability to increase breadth and magnitude of immune responses of prime/boost vaccine regimens in healthy, Atlanta; Birmingham; Boston; Nashville; New York; Rochester; San Sep-10 083 Ad5/Ad35; 180 HIV-1 uninfected adults. The trial uses heterologous inserts ( from different HIV-1 clades) and heterologous vectors (recombinant adenovirus vectors of different env Francisco env serotypes).

A randomized, double blind phase 1b trial designed to examine the influence of antigenic competition on the immunogenicity of HIV-1 Gag/Pol. With this study, we want Jan-11 084 Ad5; 100 Boston; Chicago; Iquitos; Lausanne; Lima; Sao Paulo; New York gag, env, pol to determine whether the breadth and magnitude of T-cell responses to Gag and/or Pol is altered by co-immunization with Env.

A cohort study designed to evaluate the feasibility of measuring immune responses and activation levels in the foreskin and rectosigmoid mucosa in HIV-negative, Apr-11 914 No Product 30 Lima uncircumcised men who have sex with men and who are at risk for HIV acquisition.

091 / IAVI B003 A collaborative effort with IAVI, a phase 1 placebo-controlled, double-blind, randomized trial to evaluate the safety and tolerability of the Ad26.ENVA.01 and Ad35-ENV May-11 Ad26 and Ad35; 80 Cape Town; Klerksdorp; Soweto / IPCAVD 004 env vaccines, administered in heterologous and homologous prime-boost regimens.

A follow-on study to SASHA, an adolescent HIV vaccine preparedness study in South Africa, this is designed to document the incidence of HIV, other sexually May-11 913 No Product 196 Cape Town; Rustenburg transmitted infections, pregnancy, and circumcision in adolescents at clinical research sites in South Africa, extending the study period of observation from 9 to 18 months.

A phase 1b clinical trial to evaluate mucosal immune responses following intramuscular injections of an HIV-1 DNA plasmid vaccine prime followed by an HIV-1 May-11 076 DNA/Ad5; 45 Seattle gag, pol, nef, env adenoviral vector boost in healthy, adenovirus type 5 seronegative HIV-1 uninfected adults.

The primary objectives of this phase 1 open-label trial are 2-fold: 1) To evaluate the safety and tolerability of and 2) To evaluate neutralizing antibody responses to HIV-1 Jul-11 088 Novartis Sub C pg140 with MF59; 40 Birmingham; Nashville; Rochester; Seattle env Sub C gp140 vaccine with MF59 in healthy, HIV-1 uninfected adults, primed or unprimed with HIV-1 subtype B envelope subunit vaccines with MF59.

SAAVI DNA-C2/MVA-C; A phase 1 placebo-controlled study extension to HVTN 073 / SAAVI 102 to evaluate the safety and tolerability of IM administration of Novartis Sub C gp140 vaccine with HVTN 073E / gag, RT, tat, nef Aug-11 48 Boston; Cape Town; Soweto MF59 adjuvant, given to HIV uninfected healthy adult participants in South Africa and the United States who have previously received SAAVI DNA-C2 vaccine followed SAAVI 102 env, gp140 by SAAVI MVA-C vaccine.

A phase 1 placebo-controlled clinical trial to evaluate the safety and immunogenicity of vesicular stomatitis virus serotype Indiana (VSVIN) HIV gag vaccine given to Oct-11 090 VSV; gag 60 Atlanta; Nashville; Philadelphia; San Francisco healthy, HIV-1 uninfected adult participants. The primary objective of this dose-escalation trial is to establish the maximum safe and tolerated dose of a recombinant VSV HIV gag vaccine being given for the first time in humans. SAAVI DNA/MVA 086 / A phase 1 placebo-controlled clinical trial to evaluate the safety and immunogenicity of SAAVI DNA-C2, SAAVI MVA-C and Novartis subtype C gp140 with MF59 Nov-11 Novartis gp140/MF59; 184 Cape Town; Klerksdorp; Soweto SAAVI 103 gag,RT, tat, nef, adjuvant in various vaccination schedules, in HIV-uninfected healthy vaccinia-naïve adult participants in South Africa. env, gp140 Atlanta; Birmingham; Boston; Cape Town; Chicago; Iquitos; This protocol assesses the persistence of HIV vaccine-induced seropositivity in participants who received vaccines in DAIDS-funded preventive HIV vaccine trials. The Nov-11 910 No Product n/a Klerksdorp; Lausanne; Lima; New York; Nashville; Philadelphia; Port primary purpose is to describe how long vaccine-induced antibodies to HIV, detectable on commercially available HIV antibody test kits, persist in participants who have au Prince; Rochester; San Francisco; Sao Paulo; Seattle; Soweto received preventive HIV vaccines.

Boston; Chicago; Nashville; New York; Philadelphia; Rochester; San A phase 1b randomized double-blind clinical trial to examine whether polytopic administration of VRC rAd5 A/B/C vaccine enhances HIV-specific cellular Dec-11 085 rAd5 A/B/C; 90 gag-pol/env gag-pol/env gag, pol, env Francisco immune responses in humans.

GeoVax The primary objective of this trial is to evaluate the safety and immunogenicity of a heterologous prime-boost vaccine regimen of GEO-D03 DNA and MVA/HIV62B Apr-12 094 DNA GM-CSF 48 Birmingham; Boston; Rochester; San Francisco vaccines in healthy, HIV-1 uninfected vaccinia naïve adult participants. MVA; pr, rt, env, tat, rev, vpu

Profectus DNA A phase 1 trial to evaluate the safety and tolerability of a prime-boost regimen of HIV-MAG vaccine given with and without plasmid human IL-12, delivered with May-12 087 100 Nashville; New York; Philadelphia; Rochester IL-12 EP; gag, pol, nef, tat, vif, env electroporation, followed by an rVSVINN4CT1gag1 (VSV HIV gag) boost in healthy HIV-uninfected adult volunteers.

24 JUNE 2012 VOLUME 4:1 | HVTNEWS HVTN Protocols -- Enrolling or in Active Follow Up

An ancillary study to HVTN 205, the aim of this protocol is to characterize the innate immune response to candidate HIV vaccines. The objectives of the study are 1) pGA2/JS7 NDA MVA/HIV62; Jul-09 908 env, pol, gag, 47 Birmingham; San Francisco; Seattle To define the early systemic innate immune response to candidate HIV vaccines; 2) To determine which innate immune responses correlate with the ensuing adaptive tat, rev, vpu, PR, RT immune responses. rAd5 vaccine A phase 1b clinical trial to evaluate the safety and immunogenicity of heterologous prime/boost vaccine regimens (NYVAC-B/rAd5 vs. rAd5/NYVAC-B) in healthy, HIV-1 Aug-09 078 80 Lausanne NYVAC-HIV-B; env , gag, pol, nef uninfected, Ad5 nAb seronegative adult participants.

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 25 HVTN Protocols -- Planned Protocols described below are under external review but not yet open to enrollment at the time of this writing.

Trial Products and In- N Sites Description serts

A phase 1 clinical trial to evaluate safety and to compare the immunogenicity of 3 DNA-C, NYVAC-HIV-C; HVTN 092 gag, pol, 204 U.S., Switzerland DNA vaccine prime schedules, followed by a NYVAC vaccine boost in healthy, HIV- nef, env 1 uninfected adult participants

HVTN 095/MTN 022 is a phase 1 trial designed to evaluate the safety and HVTN 095/ DNA-C, NYVAC-HIV-C, Tenofovir immunogenicity of DNA/NYVAC prime boost vaccination with or without oral 108 U.S., South Africa MTN 022 Gel, Truvada®; gag, pol, nef, env emtricitabine/tenofovir or vaginal tenofovir 1% gel in healthy, HIV-1–uninfected adult female participants

A phase 1 double blind placebo controlled clinical trial to evaluate the safety and to HVTN AIDSVAX B/E, DNA-C, NYVAC- compare the priming ability of NYVAC alone versus NYVAC + AIDSVAX® B/E, and 96 Switzerland 096 HIV-C; gag, pol, nef, env DNA alone versus DNA + AIDSVAX® B/E when followed by NYVAC + AIDSVAX® B/E boosts in healthy, HIV-1 uninfected adult participants

This is a phase 1b randomized double blind placebo controlled clinical trial to HVTN AIDSVAX B/E, ALVAC vCP1521; evaluate the safety and immunogenicity of the vaccine regimen ALVAC-HIV 100 South Africa 097 gag, PR, pol, env, gp120 followed by AIDSVAX® B/E in healthy, HIV-1 uninfected adult participants in South Africa

"Assessing Mucosal Immune Responses in HIV Vaccine Trials", continued from page 12

Acknowledgements References We thank Cecilia Morgan, Shelly Karuna, Eva Chung, Danielle 1. Bomsel M, Tudor D, Drillet AS, et al. Immunization with Harden and the Network Evaluation Committee for help in as- HIV-1 gp41 subunit virosomes induces mucosal antibodies protect- sembling Table 1. We thank Kimberly Smythe for providing Figure ing nonhuman primates against vaginal SHIV challenges. Immunity. 1 and John Hural and Cecilia Morgan for helpful discussions. 2011;34(2):269-280. Portions of this article were adapted from an article by Florian Hladik that appeared in the May 2012 volume 7, number 5 issue of 2. McElrath MJ. Standing guard at the mucosa. Immunity. Microbe, the news magazine of the American Society for Microbi- 2011;34(2):146-148. ology. 3. Mucosal Immunology Group. http://public.hivmucosalgroup.org. 2012. Last accessed: 6-7-2012 Florian Hladik is Research Associate Professor in the Department of Obstetrics and Gynecology at the University of Washington and Affiliate 4. Bendall SC, Simonds EF, Qiu P, et al. Single-cell mass cytometry of Investigator in the Vaccine and Infectious Disease Division at the Fred differential immune and drug responses across a human hematopoietic Hutchinson Cancer Research Center in Seattle. continuum. Science. 2011;332(6030):687-696. Tracey Day is Senior Science Writer at the HVTN. 5. Varadarajan N, Julg B, Yamanaka YJ, et al. A high-throughput single-cell analysis of human CD8+ T cell functions reveals discordance for cytokine secretion and cytolysis. J Clin Invest. 2011;121(11):4322- 4331.

26 JUNE 2012 VOLUME 4:1 | HVTNEWS Highlights from Recent HVTN Publications

"Highlights from Recent HVTN Publications", continued from page 21

ies that evaluated the MRKAd5/HIV-1 vaccine, Frahm et al. Reference List analyzed Ad5-specific T cells and their impact on vaccine- induced HIV-specific T-cell responses. Higher frequencies of 1. De Rosa SC, Thomas EP, Bui J, et al. HIV-DNA priming alters Ad5-specific CD4+ T-cell responses prior to vaccination were T cell responses to HIV-adenovirus vaccine even when responses to + DNA are undetectable. J Immunol. 2011;187(6):3391-3401. associated with decreased magnitude of HIV-specific CD4 + T-cell responses and decreased breadth of HIV-specific CD8 2. Spearman P, Lally MA, Elizaga M, et al. A trimeric, V2-deleted T-cell responses. This inverse correlation between pre-existing HIV-1 envelope glycoprotein vaccine elicits potent neutralizing Ad5-specific T cells and insert-specific T cells was indepen- antibodies but limited breadth of neutralization in human volunteers. dent of adenovirus-specific neutralizing antibody titers. Some J Infect Dis. 2011;203(8):1165-1173. pre-existing Ad-specific T cells recognized epitopes that are 3. Churchyard GJ, Morgan C, Adams E, et al. A phase IIA conserved across adenovirus strains. These data indicate that randomized clinical trial of a multiclade HIV-1 DNA prime fol- pre-existing adenovirus-specific T-cell responses can affect im- 7 lowed by a multiclade rAd5 HIV-1 vaccine boost in healthy adults munogenicity of adenovirus-based vaccines. Frahm et al. (HVTN204). PLoS ONE. 2011;6(8):e21225.

Tracey Day is Senior Science Writer at the HVTN. 4. Fitzgerald DW, Janes H, Robertson M, et al. An Ad5-Vectored HIV-1 Vaccine Elicits Cell-mediated Immunity but does not Affect Disease Progression in HIV-1-infected Male Subjects: Results From a Randomized Placebo-Controlled Trial (The Step Study). J Infect Dis. 2011;203(6):765-772.

5. Gray GE, Allen M, Moodie Z, et al. Safety and efficacy of the HVTN 503/Phambili Study of a clade-B-based HIV-1 vaccine in South Africa: a double-blind, randomised, placebo-controlled test-of- concept phase 2b study. Lancet Infect Dis. 2011.

6. Rolland M, Tovanabutra S, Decamp AC, et al. Genetic impact of vaccination on breakthrough HIV-1 sequences from the STEP trial. Nat Med. 2011;17(3):366-371.

7. Frahm N, Decamp AC, Friedrich DP, et al. Human adenovirus- specific T cells modulate HIV-specific T cell responses to an Ad5- vectored HIV-1 vaccine. J Clin Invest. 2012;122(1):359-367.

www.hvtn.org HVTNEWS | VOLUME 4:1 JUNE 2012 27 Managing Editor: cecilia morgan EDITOR: adi ferrara Calendar senior SCIENCE WRITER: tracey day PRODUCTION MANAGER: courtney liebi design & layout: lisa donohue XIX international aids conference FOR MORE INFORMATION, VISIT: hvtn.org july 22-27, 2012 Walter E. Washington Convention Center (WCC) COMMENTS/QUESTIONS? Washington, D.C. [email protected] Tel: 206 667-6300 aids vaccine 2012 Fax: 206 667-6366 september 9-12, 2012 HVTN/FHCRC, 1100 Fairview Ave North, LE-500 Boston Convention and Exhibition Center PO Box 19024 Seattle, Washington 98109-1024 Boston, MA

Thank you to: Jim Kublin, Danna Flood, Carrie Sopher, Danielle Harden, Sid Niazi, hvtn conference Vic Sorrell, Eva Chung october 29-31, 2012 Grand Hyatt Seattle Seattle, WA For current and archived editions of HVTNews, please visit us here - hvtn.org/science/hvtnews.html keystone symposia: immunological mechanisms of vaccination The HIV Vaccine Trials Network is an international collaboration of december 13-18, 2012 scientists and educators searching for an effective and safe HIV Fairmont Château Laurier vaccine. Support for the HVTN comes from the National Institute of Ottawa, Ontario, Canada Allergy and Infectious Diseases (NIAID) of the U.S. National Institutes of Health, an agency of the U.S. Department of Health and Human Services. The Network and NIAID have a close, cooperative working relationship, with shared attention to intellectual and scientific issues.

Mission of the HVTN To enhance the discovery and drive the development of a safe and globally effective vaccine to prevent HIV.

We do this through well-designed clinical research trials that objectively and ethically address the critical questions of the field. on the cover: Our objective clinical trial platform lets us evaluate safety, immunogenicity Vic Sorrell, Nashville (Vanderbilt site) HVTN and efficacy of candidate vaccines, as well as design clinical trials that will recruiter and singer extraordinaire, combines his provide clues on ways to enhance the effectiveness of new vaccines. talents to increase enrollment in HVTN 505. In We promote the use of new innovations, which may help us get closer to his spare time he continues to work in the music a safe and effective vaccine more quickly. industry.