Intratumoral mPH-762, a self-delivering RNAi therapeutic (INTASYL?) targeting mouse PD-1, generates systemic tumor-specific memory CD8 T cells, providing a mechanism for abscopal efficacy toward untreated tumors in a murine hepatocarcinoma model

Abstract Intratumoral (IT) administration of immune checkpoint inhibition (ICI) holds potential to enhance local activity while decreasing systemic toxicity, however, high molecular weight antibodies are not well-suited for this application. PH-762, a self-delivering RNAi compound targeting PD-1 built on proprietary INTASYL™ technology, rapidly enters tissues without need for further formulation or delivery vehicles. In preclinical studies, PH-762 and its murine-targeted analog (mPH-762) provide robust PD-1 silencing, anti-tumor efficacy and associated immunophenotypic changes in the tumor microenvironment (TME). PH-762 is currently under clinical investigation in a Phase 1b study as neoadjuvant therapy for advanced resectable melanoma. We have previously shown that IT mPH-762 provides abscopal efficacy toward untreated distal tumors in a bilateral Hepa1-6 model of murine hepatocellular carcinoma. Here, we identify potential mechanisms underlying this abscopal efficacy in follow-up studies in vivo and ex vivo. Equivalent inoculums of Hepa1-6 cells were subcutaneously implanted into bilateral flanks of C57BL/6J mice (N = 12/group). When tumors reached a mean volume of 150 mm3, vehicle (PBS) or mPH-762 (0.5 mg/dose or 2 mg/dose) were administered to one of the two tumors (the directly treated tumor), on Days 1, 4, 7, 10 and 13. Tumor volumes and body weights were recorded longitudinally. Directly treated (DT) and untreated distal tumors (UT), as well as mesenteric lymph nodes (mLNs) and spleens (SP) were isolated from a satellite group (N = 6) on Day 14. On-target immunomodulatory changes were assessed in DT and UT TME by immunostaining/flow cytometry. Additionally, systemic tumor specific T cells were expanded from bulk mLN and SP by 21-day culture with irradiated Hepa1-6 cells. Tumor specific memory reactivity was assessed by challenge with either intact Hepa1-6 cells; or intact CT26 BALB/c colon cancer cells, PMA/ionomycin, or PBS controls: and assessed by intracellular IFN-γ, TNF-α and surface CD107a staining. IT mPH-762 again provided an abscopal decrease in mean tumor volume of UT tumors compared to IT PBS. On-target immunomodulatory changes to TME appeared primarily confined to the DT only at this timepoint. However, IT mPH-762 increased both the frequency and reactivity of Hepa1-6-specific CD8 T cells expanded from mLN and SP compared to IT PBS. These data indicate that IT mPH-762 generates a systemic immune response of durable tumor-specific reactive memory CD8 T cells in systemic lymphoid organs, providing a mechanism for the abscopal efficacy of mPH-762. These data further support the clinical development of IT PH-762. Citation Format: Benjamin G. Cuiffo, Andrew Boone, Dingxue Yan, Melissa Maxwell, Brianna Rivest, James Cardia, Simon P. Fricker. Intratumoral mPH-762, a self-delivering RNAi therapeutic (INTASYL™) targeting mouse PD-1, generates systemic tumor-specific memory CD8 T cells, providing a mechanism for abscopal efficacy toward untreated tumors in a murine hepatocarcinoma model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1846.