An Ex Vivo Organotypic Culture Platform for Functional Interrogation of Human Appendiceal Cancer Reveals a Prominent and Heterogenous Immunological Landscape

Abstract

Purpose: Epithelial neoplasms of the appendix are difficult to study preclinically given their low incidence, frequent mucinous histology, and absence of a comparable organ in mice for disease modeling. While surgery is an effective treatment for localized disease, metastatic disease has a poor prognosis as existing therapeutics borrowed from colorectal cancer have limited efficacy. Recent studies reveal that appendiceal cancer has a genomic landscape distinct from colorectal cancer and thus preclinical models to study this disease are a significant unmet need. Experimental Design: We adopted an ex-vivo slice model that permits the study of cellular interactions within the tumor microenvironment. Mucinous carcinomatosis peritonei specimens obtained at surgical resection were cut using a vibratome to make 150 mm slices cultured in media. Results: Slice cultures were viable and maintained their cellular composition regarding the proportion of epithelial, immune and fibroblasts over 7 days. Within donor specimens, we identified a prominent and diverse immune landscape and calciumimaging confirmed that immune cells were functional for 7 days. Given the diverse immune landscape, we treated slices with TAK981, an inhibitor of SUMOylation with known immunomodulatory functions, in early phase clinical trials. In 5 of 6 donor samples, TAK981 treated slices cultures had reduced viability, and regulatory T cells (TREGs). These data were consistent with TAK981 activity in purified TREGs using an in-vitro murine model. Conclusions: This study demonstrates an approach to study appendiceal cancer therapeutics and pathobiology in a preclinical setting. These methods may be broadly applicable to the study of other malignancies.

Distinct cell states define the developmental trajectories of mucinous appendiceal neoplasms towards pseudomyxoma metastases

“…While these cells lacked features suggestive of anti-tumor activity, their cytotoxic potential could be harnessed further for strategies that aim to convert bystander cells into infiltrating cells that can contribute to anti-cancer activity (52). This is further supported by recent studies that demonstrated an ex vivo T cell activation in PMP tumors (55)(56)(57).…”

“…Importantly, these cells retained cytotoxic gene expression suggesting they could be targeted for therapeutic purposes. Recent studies have demonstrated that PMP tumors can respond ex vivo to immune checkpoint blockade or neoantigen vaccination (55)(56)(57). Further functional characterization of this lymphocyte infiltrate will help clarify the role of these cells in anti-cancer immunity.…”

A new immuno-oncology target – SUMOylation

“…Ubc9-deficient Tregs in mice lost Foxp3 expression upon T cell receptor (TCR) stimulation [ 6 ]. In primary patient tumor tissues, pharmacological SUMOylation inhibition reduced Tregs [ 7 ] ( Figure 2 ).…”

Positioning SUMO as an immunological facilitator of oncolytic viruses for high-grade glioma

“…As such, the reliance of Treg expansion and function on SUMO serves as an appealing target. Ding et al reported that the knockout of UBC9 (the SUMO E2 ligase) in a Treg population resulted in impaired proliferation, activation, and suppressive functionality ( Weitz et al, 2022 ). Similarly, Lam et al (2023) treated T-cell populations from patients with chronic lymphocytic leukemia with TAK-981 and reported decreased Treg differentiation.…”

“…As such, the implication of SUMO in the pathogenesis of resistant cancers has positioned SUMOtherapeutics as potential anti-cancer agents and immunotherapeutic adjuvants ( Seeler and Dejean, 2017 ). The first SUMOtherapeutic, TAK-981 (Takeda Pharmaceuticals, Tokyo, Japan), has been reported to induce cell-cycle arrest ( Hanel et al, 2022 ; Kim et al, 2023 ), deplete Treg populations ( Weitz et al, 2022 ) and spur immune activation ( Khattar et al, 2019 ; Lightcap et al, 2021 ; Kumar et al, 2022 ), giving rise to increasing efforts to apply it to a wide set of cancers ( Langston et al, 2021 ) ( Figure 1 ). TAK-981 is presently being investigated for advanced non-small cell lung cancer, cervical cancer, microsatellite-stable colorectal cancer, refractory or relapsed diffuse large B-cell lymphoma, and follicular lymphoma (NCT03648372).…”