Project A-Gene A case study-based approach to integrating QbD principles in Gene Therapy CMC programs Presented by Contributors CHAPTER LEADS / CONTRIBUTING MEMBERS Greg MacMichael Hone Megan Furcolo Patrick Jeffrey Hung CMC Bioservices Pfizer Pfizer Vigene Biosciences Saroj Ramdas Baldus Phoebe Parker Joann Alexandra Beumer-Sassi Amicus Pfizer Pfizer Voisin Consulting Life Sciences Mo Heidaran Banbula Agnieszka Dawn Henke Luis Maranga Parexcel Pfizer Standards Coordinating Body Voyager Therapeutics Paul McCormac Lavoie Janelle (SCB) Shamik Sharma Pfizer Pfizer James Warren Voyager Therapeutics Iryna Sanders McEnroe Janet Ultragenyx Nripen Singh Pfizer Pfizer Jessie Sun Voyager Therapeutics Aili Cheng Micklewright Althea Ultragenyx Pfizer Pfizer Jim Richardson Cirelli David Alphonse Ignatius Arun United State Pharmacopia Pfizer Pfizer (USP) REVIEWERS Note: All chapters were reviewed, but individual reviewers did not review every chapter. Reviewers volunteered to provide editorial review for select chapters according to their expertise and interest. Tayler Renshaw Nadine Sandhöfer Jean Stanton Mike Winkler Audentes Therapeutics Cevec Johnson & Johnson (J&J) REGENEXBIO Malou Gemeniano Tim Farries Kelvin Lee Justin Horvath Audentes Therapeutics ERA Consulting National Institute for REGENEXBIO Innovation in Manufacturing William Werner Raj Puri Rob McCombie Biopharmaceuticals (NIIMBL) Audentes Therapeutics (Regulatory chapter only) Sangamo Therapeutics FDA Hardeep Samra David Gray Robert Shaw Neurogene Audentes Therapeutics Tim Miller SCB Forge Biologics Tina Parikh John Grunkemeier Sven Kili Neurogene Audentes Therapeutics Michele Meyers Sven Kili Consulting GSK David Litwack Dominic DeMuro Khandan Baradaran Prevail Therapeutics Audentes Therapeutics Steven Weisser Ultragenyx GSK Michael Boychyn Isabella Palazzolo Jeffrey Hung REGENEXBIO Biogen Michael Mercaldi Vigene Homology Medicines Mike Byrne Stuart Beattie Erica Giordano REGENEXBIO Biogen Maria Lobikin Voisin Consulting Life Sciences Homology Medicines Tristan Marshall Clare Blue Emmanuelle Sabbah- REGENEXBIO Biogen James McGivney IV Petrover Homology Medicines Don Startt Voisin Consulting Life Sciences Nicole Faust REGENEXBIO Cevec Eric Faulkner Homology Medicines PRODUCTION Medical/Technical Writers Project Management Editor Graphic Design (select figures) Darshana Gupte Michael Lehmicke Marie Daghlian Michaela Flatly Cecelia Wall Adam Roose Overall Graphic Design and Layout Amr Eissa Carol Collier Julia Grant ii Project A-Gene Introduction The effort that has ultimately resulted in the document below began with a short, yet complex question posed to an audience of cell and gene therapy (CGTx) CMC experts. The Alliance for Regenerative Medicine (ARM) convened its first of a now annual series of CMC Summits in December of 2017, inviting individuals from across the industry to join in a conversation on manufac- turing. As part of this discussion, the question of “What is the biggest challenge to cell and gene CMC? How can ARM help to address it?” was raised to the audience and, across the numerous responses and specific examples given, there emerged a common complaint related to the heterogeneity of the process. After exploring this further, there was general agreement that many of the issues driving the challenges to streamlined, cost effective manufacture of CGTx products derive from a lack of standardized methodologies and training around CMC programs. It was further suggested that this phenomenon was not new, and that we should look to the past for inspiration on how to address this challenge for the future. Therefore, it was decided to embark on a mission to recreate ‘A-Mab’ for the CGTx industry. In previous years, the broader life sciences industry has encountered systemic barriers to the continued development of promising technologies. In both the mono- clonal antibody industry and the vaccine development world, the whirl of energy around scientific discovery was stalled by the realities of manufacturing. While a small team of well-trained experts can produce high quality batches of drug product for use in process development and early clinical trials, it is inevitable that this process will need to be dramatically increased in scale, and the process transferred to other parties for commercial production. To help lower the barrier to this nec- essary tech transfer, and to better prepare new entrants to the industry, the leading developers of monoclonal antibodies and vaccines have produced a consolidated set iii of recommendations for implementation of Quality by Design (QbD). A-Mab and A-Vax, respective to each industry, have played a significant role in elevating best practices within their industries, and have been effective in continued workforce development efforts. Borrowing from this model, the members of the Alliance for Regenerative Medicine have worked to produce a similar document for use by the burgeoning gene therapy sector. In order to make A-Gene an effective resource, and reflecting the ongoing inno- vation in the sector, we sought to: 1) draw from as wide a set of expertise as possible, 2) specify our area of focus to direct gene therapy, and 3) focus on AAV as the case study. This effort to catalogue expertise in gene therapy development occurred in parallel to approval of the first cohort of human gene therapies, which have shown the value and clinical relevance of such programs. As the field continues to develop it has been recognized that the future of cost effective gene therapy relies on implementation of common practices, development of specialized technologies, and above all else, standardization of methods. Given the wide scope of innovation underway in each of these areas of focus, ARM and the A-Gene team sought to bring in as much thought leadership as possible to ensure that what was recorded was truly a representation of best practices in the industry. Reflecting on this, A-Gene is a truly collaborative effort that has been crafted with contributions from more than 50 industry experts from more than 20 leading therapeutics developers. To further maximize the utility of A-Gene, the drafting team decided to focus on in vivo gene therapy. Current FDA language around Gene Therapy encompasses in vivo gene therapy, gene editing/manipulation, and gene modified cell therapy. Each application carries a wide array of specific manufacturing steps and consid- erations, and so in order to draft a cohesive and informative narrative, the A-Gene team decided to focus on in vivo gene therapy. Ex vivo gene-modified cell therapies will be the subject of a future case study. Finally, the A-Gene team deliberated on what the underlying case study would be for this document. As each chapter is meant to operate as both an independent re- source, as well as part of a comprehensive narrative, we felt it was necessary to focus on one specific use case to facilitate the utility of A-Gene. Lentiviral and AAV vectors are the two most frequently used viral vector platforms used in gene therapies, and the approach to producing these two vectors are similar. While lentiviral vectors are predominantly used for ex vivo cell modification for gene modified cell therapies, AAV is the major vector type for in vivo applications. Given our previous decision iv to draw from the widest set of expertise possible on direct gene therapies and the focus on in vivo gene therapy approaches, it naturally made sense to focus our case study on human rAAV therapeutic development. This is not to imply either a relative value in rAAV vs other viral platforms for direct gene therapy, but is, in general, recognition of the number of companies pursuing rAAV applications. We deliberated on inclusion of parallel tracks throughout the document, comparing and contrasting rAAV to lentivirus, but in the end decided this would be too cumbersome. Beyond this, there is a great deal of overlap in thought process and methodology between the two approaches, and so in lieu of a running comparison we elected to spend some time in the introduction addressing differences in CMC methods between the two viral platforms. A-Gene is not intended to represent a standard to be rigidly applied. It is a hy- pothetical case study representing an archetype of an AAV vector for gene therapy. Therefore, it is a snapshot in time of current best principles in a rapidly evolving field. The data cited in the document are non-proprietary, and are intended to be for illustrative purposes only. Where appropriate the authors have borrowed formatting and structure from the A-Mab case study. While we have attempted to be as comprehensive as possible, and have subjected the document to rigorous review, it is not a “recipe book” for AAV manufacture. Some aspects of process development (e.g., facility design), were deliberately omitted for the sake of brevity. Importantly, A-Gene is not an example of a mock regulatory submission, nor should it be interpreted as regulatory advice, or cited as regulatory guidance. As a final point, we wish to thank those who contributed to this effort. The Alliance for Regenerative Medicine is grateful to the innumerable thought leaders, subject matter experts, and researchers who have helped to make this project a reality. We also wish to extend our deep appreciation to the National Institute for Innovation in Manufacturing Biopharmaceuticals for their support and contribu- tions to this effort, and for working with ARM to make this project a reality. We look forward to continuing our work with