Abstract B85: Combination therapy using the programmed death receptor-1 and the programmed death-ligand 1 inhibitors and Fluorouracil in human colorectal carcinoma in a humanized patient-derived orthotopic mouse model
Authors: Zhang Xin , Maresh Grace , Hellmers Linh , Yip Henry , Baste Lara McKean , Green Heather , Margolin David , Li Li 
Published: March 2020

Background: One of the recently understood mechanisms associated with the progression of cancer is the immune checkpoint pathway. Many tumors can stimulate the expression of immune checkpoint molecules, resulting in a phenotype of exhausted T cells that cannot restrain tumor progression. One such inhibitory ligand and receptor pair in solid tumors are the programmed death-ligand 1 (PD-1) and programmed death receptor-1 (PD-L1). They prevent the killing of cancer cells by cytotoxic T-lymphocytes. PD-1 receptor is expressed by activated T cells among other cells, while PD-L1 is overexpressed on many tumor types including colorectal carcinoma (CRC). We have developed a humanized patient-derived orthotopic xenograft (hPDOX) model for CRC. Here, we investigate the potential efficacy of combination of immune checkpoint inhibitors (ICIs) and fluorouracil (5FU) in this CRC hPDOX model.

Methods: Humanized mice were established by IP injection of donor peripheral blood mononuclear cells (PBMCs) into recombinase activating gene 2 (Rag2) and common cytokine receptor gamma chain gene (IL2Rγ) double knockout (Rag2-/-/IL2RÎ³-/-) Rag2 mice. Luciferase-tagged patient tumor CoCaPt302 cells were injected intrarectally into Rag2 mice. Groups of mice (n=7-10) received ICIs, anti-PD-1 and anti-PD-L1 antibodies (nivolumab, 200 μg/mouse and atezolizumab, 200 μg/mouse), and 5FU (200 mg/kg) once a week for 4 weeks, IV injection alone or in combination. Tumor growth was measured weekly by bioluminescent imaging (BLI). At necropsy, tumor weights were measured. Human HLA-ABC+CD45+ hematopoietic cells and CD4+ or CD8+ T cells were detected in peritoneal lavage, blood, and spleen by flow cytometry. The presence of human immune cells (humanization) and tumor-infiltrating lymphocytes was confirmed by immunohistochemistry staining.

Results: Humanization was evidenced by mouse blood containing more than 45% HLA-ABC+CD45+ human cells when tested by flow cytometry. Tumor weight showed significant reduction in group treated with combination of ICIs and 5FU compared with untreated controls (p=.0038). Tumors were also significantly smaller than ICIs (p=.0123) or 5FU (p=.0427) monotherapy groups. In addition, the lung and liver metastasis detected by ex vivo BLI were reduced from 50% of mice in control group to 33% and 11%, respectively, and exhibited better antitumor response as well as less development of metastases. CD3+ T cells, especially CD4+, CD8+ T cells in blood and spleen, were reduced in the ICIs only and combination treatment groups, but not in 5FU group compared to the controls.

Conclusion: Our study provides preclinical evidence of establishment of humanized Rag2 mice to be used as a hPDOX model for CRC; and treatment efficacy including inhibiting tumor growth and distant organ metastasis was significantly enhanced through ICI and 5FU combination therapy. The effect of the combination therapy using CRC patient-derived specimens and PBMCs from MHC matched donors or autologous PBMCs will be further investigated.

Citation Format: Xin Zhang, Grace Maresh, Linh Hellmers, Henry Yip, Lara McKean Baste, Heather Green, David Margolin, Li Li. Combination therapy using the programmed death receptor-1 and the programmed death-ligand 1 inhibitors and Fluorouracil in human colorectal carcinoma in a humanized patient-derived orthotopic mouse model [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B85.