Nonclinical Toxicology, Biodistribution, and Pharmacokinetics of UB-VV111, an In Vivo Anti-CD19 CAR T Cell Therapy

Introduction: Chimeric antigen receptor (CAR) T cell therapy has demonstrated transformative outcomes in hematologic malignancy; however, many challenges remain, such as manufacturing complexity, high cost, requirements for lymphodepletion, and poor product persistence. To address this, Umoja is developing the VivoVecTM platform, an off-the-shelf surface-engineered lentiviral vector (LVV) drug product designed to generate CAR T cells in vivo without lymphodepletion by selectively binding, activating, and transducing T cells. UB-VV111 is a VivoVec LVV that encodes a payload transgene comprised of an anti-CD19 CAR targeting B cells and the rapamycin activated cytokine receptor (RACR) system to promote expansion and persistence of transduced T cells in vivo. Here, we present a comprehensive nonclinical data package, including a GLP toxicology study, that demonstrates a low risk of insertional oncogenesis, off-target transduction, or CAR T cell-related toxicity and supports evaluation of UB-VV111 in clinical trials. Methods: LVV RNA genomes were detected by RT-qPCR. Pharmacokinetics of a surrogate LVV in blood was assessed after intranodal (IN) or intravenous (IV) dosing to canines or NHPs. Vector shedding to feces was assessed after dosing UB-VV111 IV or intraperitoneal (IP) in non-humanized NSG mice. Vector copy number (VCN) and insertion site analysis was assessed in human PBMCs from 3 donors treated with a range of UB-VV111 MOIs. UB-VV111 transduction was assessed in human primary cells isolated from tissues that express LDL-R (liver, adrenal gland, lung, kidney, colon), a ligand potentially engaged by the Cocal fusion glycoprotein expressed on VivoVec particles. An 8-week biodistribution study was performed in canines dosed IN or IV with a GFP- expressing LVV. A 13-week GLP study in CD34-humanized NSG mice was performed to evaluate the toxicology and biodistribution of UB-VV111 with rapamycin in a model with human T and B cells. UB-VV111 was administered IP (surrogate for IN) or IV at up to 5.1 ×107 transducing units (a safety factor of 14x to the highest proposed clinical dose scaled by body weight). Results: Vector RNA genomes were not detected in blood at any time following IN dosing or in feces at any time following IV or IP dosing. Vector RNA genomes were rapidly depleted from circulation following IV dosing, with a peak concentration 5 minutes post injection and < 5% of vector RNA genomes remaining in circulation by 2 hours. The risk of insertional oncogenesis with UB-VV111 is low and no greater than approved lentiviral-based ex-vivo CAR T cell therapies. The VCN per CAR T cell was low, with an average VCN of 1.77 copies per CAR T cell. The insertion site distribution resembled that of a typical LVV, with a low tendency to integrate at recurrent sites and a high prevalence in introns when inserted in the vicinity of genes. UB-VV111 nonclinical data demonstrated a low risk of off-target transduction. UB-VV111 did not transduce human primary cells isolated from non-lymphoid tissues at the maximum MOI achievable in vivo with current proposed clinical doses. In canines, VivoVec was well tolerated and did not result in quantifiable transduction in any non-lymphoid tissues at any timepoint following IN or IV ROA. UB-VV111 with rapamycin was well tolerated in the CD34-humanized mouse GLP toxicity study, with no UB-VV111-related mortalities or effects on body weights, clinical observations, hematology, clinical chemistry, human cytokine expression, or gross pathology findings. UB-VV111 treatment resulted in generation of CAR T cells and dose-dependent B-cell depletion. UB-VV111 vector genome integration was dose dependent, decreased over time, and was highest in liver and spleen. Across tissues, multiplex RNA ISH analysis classified transduced cells as immune cells, predominantly murine macrophages or human T cells. Transduction of human macrophages in spleen and liver was exceedingly rare in CD34-humanized mice, indicating macrophage transduction is a feature specific to the murine model. Conclusion:UB-VV111 demonstrated a favorable safety and biodistribution profile, including a low risk of acute toxicity due to UB-VV111 and a low risk of toxicity from VivoVec mediated on-target (T cell) or off-target (non-T cell) transduction. Taken together, these nonclinical studies support the evaluation of UB-VV111 and rapamycin at the doses proposed in the planned Phase 1 clinical study.