Engineering synthetic immune cells with modular sentinel and therapeutic functions for T1D

PROJECT SUMMARY Our long-term goal is to engineer therapeutic immune cells that can report on and treat early-stage type 1 diabetes (T1D). Ideally, these cells should home to islets, detect and integrate multi-factor signatures of early- stage disease and, in response, induce localized immunosuppression to block destruction of beta cells. Although this seems like an extraordinarily difficult multi-level challenge, the remarkable progress in engineering immune cells to recognize and kill cancer has generated a broad set of new technologies and approaches that could be brought to bear on cell-based therapies for T1D. Here we propose to repurpose, redirect, and extend cell engineering approaches to construct synthetic immune cells (CD4+ T cells) capable of sensing and treating early-stage autoimmune disorders like T1D. Given the multiple challenges in achieving this goal, we propose to take a modular approach – we have broken up what an ideal anti-T1D cell therapy would have to achieve into three distinct subtasks. Our focus will be on independently engineering and validating cell circuit modules that can achieve these subtasks, which can then be linked together in multiple combinations to develop options for a full therapy. These modular objectives are outlined in our specific aims: Aim 1. ISLET SENSING/TARGETING | Engineer synthetic immune cell sentinels that recognize and establish residence/activity in the pancreas/islets. Sensors of pancreatic/islet specific antigens; islet restricted activation using synNotch receptors; and islet trafficking via synthetic adhesion proteins. Aim 2. AUTOIMMUNE DISEASE SENSING | Engineer synthetic immune cells that sense and report on local immune perturbations associated with T1D onset. Engineer sensors that detect presence of autoreactive T cells and elevated local levels of specific inflammatory cytokines Aim 3. IMMUNO-SUPPRESSIVE OUTPUT: Engineer therapeutic cells that protect islets by inducing local immunosuppressive outputs in response to disease sensing. Engineer output responses encompassing induced local production of suppressive cytokines (IL10, TGFb), inflammatory cytokine sinks (CD25), and other beta-cell protective factors. We will test multiple configurations of linking the disease sensing circuits from Aims 1 and 2 to the suppressive outputs from Aim 3. To develop innovative cell engineering platforms for treating T1D, we propose to focus on human pluripotent stem cell (hPSC)-derived islets as a highly flexible system in which to evaluate the immunoprotective function. hPSC-cells can be readily generated and genetically modified to add convenient model antigens for sensing or killing, allowing for the rapid design-build-test iterative cycles for the circuit modules described above (i.e., making each aim non-dependent on the others). hPSC islets can also be used to assess immunoprotective responses against a variety of islet-targeted cytotoxic T cells, both in vitro and in vivo (implantation in mouse kidney capsule), making this platform ideal for proof-of-principle evaluation and cell circuit optimization.