Abstract PO-110: Targeting cathepsin B in the pancreatic stellate cells stimulates CD8+ T cell dependent anti-tumor immune response
Authors: Garg Bharti , Jain Tejeshwar , Vaish Utpreksha , Dudeja Vikas 
Published: November 2021

Background: Cathepsin B (CTSB) is a lysosomal cysteine protease which has been described to have a role in pancreatic cancer development. However, its significance in the tumor microenvironment has never been investigated. Pancreatic stellate cells (PSCs) are one of the main sources of Cancer Associated Fibroblasts (CAFs) in the pancreatic cancer microenvironment. Herein, we describe the effects of targeting CTSB in the pancreatic stellate cells (PSCs) in pancreatic cancer (PDAC). Methods: To specifically study the role of CTSB in PSCs, PSCs were isolated from C57BL/6J (WT) or CTSB -/- mice and were orthotopically injected with KPC cancer cells into the tail of pancreas of 6-week old female WT mice. Tumors were analyzed at endpoint using flow cytometry. To further study the dependence of effects of CTSB in PSCs on adaptive immune cells, WT or CTSB -/- PSCs were co-injected with KPC cells into Rag 1-/- mice or CD8+ T cell depleted mice (anti-CD8 mAb, 250ug/dose every 3 days). In vitro, WT or CTSB-/- PSCs were co-cultured with KPC across a transwell. CD8+ T cells, isolated from KPC tumor bearing mice, were exposed to conditioned media from this co-culture ex-vivo and their cytotoxicity was measured using calcein release assay. In another experiment, WT mice injected with KPC cancer cells were treated with vehicle control or the CTSB inhibitor, CA-074 methyl ester (10 mg/kg), through daily intraperitoneal injections. Results: Co-injections of WT PSC with KPC cells formed larger tumors as compared to KPC cancer cells alone, however, co-injection with CTSB-/- PSCs abrogated this tumor promoting effect. Flow cytometric analysis revealed increased infiltration of Ifn-γ secreting CD8+ T cells in the KPC+CB-/- PSC group as compared to KPC+WT PSC group. Tumor decreasing effect of CTSB-/- PSCs was lost when co-injections were performed in Rag 1-/- mice or CD8+ T cell depleted mice, indicating that this effect relies on a functional adaptive immune response, specifically CD8+ T cells. In vitro, co-culture of CTSB-/- PSCs with KPC led to reduced expression of inflammatory CAF (iCAF) markers like IL-6 and IL-11 when compared to co-culture of WT PSCs with KPC. CD8+ T cells exposed to conditioned media from CTSB-/- PSC co-culture showed increased cytotoxicity against KPC cells ex-vivo, as compared to those exposed to conditioned media from WT PSC co-culture. Finally, treatment with CA-074 was able to decrease orthotopic PDAC tumor growth with increased CD8+ T cell infiltration evident on flow cytometry. Conclusion: Targeting CTSB in the PSCs stimulates anti-tumor adaptive immune response against pancreatic cancer by inhibiting conversion of PSCs into the tumor-promoting iCAF phenotype. CTSB might emerge as a novel target for CAF-directed therapy in pancreatic cancer.

Citation Format: Bharti Garg, Tejeshwar Jain, Utpreksha Vaish, Vikas Dudeja. Targeting cathepsin B in the pancreatic stellate cells stimulates CD8+ T cell dependent anti-tumor immune response [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-110.