Fast-Track proposals will NOT be accepted. Direct Phase II proposal will be accepted Number of anticipated awards: 3 - 5 Budget (total costs, per award): Phase I: up to $400,000 for up to 12 months Phase II: up to $2,000,000 for up to 2 years PROPOSALS THAT EXCEED THE BUDGET OR PROJECT DURATION LISTED ABOVE MAY NOT BE FUNDED. Summary Gene therapy has come of age over the past few years. One of the most promising anticancer approaches in the clinic is chimeric antigen receptor (CAR)-T cell therapy. However, the pioneering first-generation products now on the market for B-cell malignancies, that target a single cancer antigen, have major limitations. First, all normal B cells expressing CD19 are eliminated by the therapy meaning that normal B cell functions are lost. Second, patients may lose expression of the CD19 CAR-T target antigen, rendering the malignant tumor cells invisible to the immune system tasked with its Page 71 destruction. Third, the therapies can trigger toxicities that are hard to predict and control, such as cytokine release syndrome. By combining computer science logic with biology, scientists have developed synthetic gene circuit technologies to redirect genetic events within cells to enable the resulting therapies to sense and adapt to their environment, or be controlled to avoid the safety and efficacy pitfalls that limited first-generation products. For example, new CAR-T approaches involve the delivery to T cells of gene circuits based on Boolean logic that can produce tumor cell killing only when two (or more) cancer antigens are expressed on cancer cells but not on normal cells, preserving normal B cell function. These synthetic gene circuits are assembled of DNA encoding RNA or protein that enable individual cells to respond and interact with each other to perform a function at the desired locations (e.g. within the tumor vs. whole body), targets (e.g. cancer cells vs. healthy neighboring cells), amount (e.g. therapeutic vs. toxic doses) and duration (e.g. shut down before significant side effect occurs). Key components include sensors that detect user-defined inputs, processors that make decisions in response to the inputs, and actuators that produce the desired output activities (payloads). Synthetic gene circuits can be delivered into cells ex vivo as in the CAR-T case, or in vivo using any well-established gene transfer vectors. These gene circuit therapies can be programmed to distinguish cancer cells from normal cells and to activate therapeutic payload expression from inside tumors. Project Goals The goal of the topic is to stimulate the development of gene circuit therapies for cancer. Engineering of immune cells and/or cancer cells is encouraged, while other cell types are not excluded. The recent pioneering work in synthetic biology has shown the potential of overcoming current challenges in gene therapy by creating sophisticated gene circuits to distinguish between malignant and healthy cells and to efficiently kill the former without harming the latter. Unlike conventional small molecules or biologics, including most of the current gene therapies, gene circuit therapies can potentially sense multiple disease signals, integrate this information to make a decision to trigger sophisticated or combinatory therapeutic mechanisms. Alternatively, gene circuit therapies can also be controlled exogenously, therefore allowing precise control over timing, dose and location of the therapies. The activities that fall within the scope of this solicitation include the development of the gene circuits designed and created using synthetic biology approaches into cancer therapies through engineering immune cells ex vivo, or by delivering directly into cancer cells in patients using viral or non-viral gene transfer approaches/vectors, including engineering of bacteria to specifically target cancer. The approach should also allow precise control over timing, dose, and location of the therapies. Examples of appropriate activities include to demonstrate that the gene circuit can be expressed in cancer cells in vitro and in vivo, with increased efficacy and decreased toxicity compared to currently available similar therapies or to standard of care. A system that does not have the potential to allow precise control of the therapeutics over timing, dose, and location as needed will not be responsive. Methodologies to create gene circuits without delivery will not be responsive. Animal studies establishing proof-of-concept efficacy in well-validated in vitro and in vivo models should be completed in Phase I. In Phase II the contractor is expected to perform a large-scale in vivo efficacy study, as well as other studies required for FDA IND submission. Phase I Activities and Deliverables: Establishing proof-of-concept efficacy and/or toxicity: • Demonstrate in vitro sustained and controllable transgene expression with efficacy in appropriate cell lines and/or 3D models • Demonstrate in vivo sustained and controllable transgene expression with efficacy in appropriate small animal models • Conduct gene circuit optimization (as appropriate). • Perform (optional) animal toxicology and pharmacology studies as appropriate. • Demonstrate (optional) increased efficacy and/or decreased toxicity as compared with standard-of-care for the cancer indication in appropriate animal model(s). Phase II Activities and Deliverables: The offerors are encouraged, but not required, to meet FDA before the submission of a Phase II proposal. A detailed experimental plan necessary for filing an IND is expected in the Phase II proposal: • Conduct properly powered efficacy studies, demonstrating benefits with statistical significance. • Complete IND-enabling experiments and assessments according to the plan developed. The plan should be reevaluated and refined as appropriate. • Develop and execute an appropriate regulatory strategy. If warranted, provide sufficient data to file an IND or an exploratory IND for the candidate therapeutic agent. Page 72 • Demonstrate the ability to produce a sufficient amount of clinical grade material suitable for an early clinical trial.