Abstract
A recurring historic finding in cancer drug development is encouraging antitumor effects observed in tumor-bearing mice that fail to translate into the clinic. An intriguing exception to this pattern is immune checkpoint therapy, as the sustained tumor regressions observed in subsets of cancer patients are rare in mice. Reasoning that this may be due in part to relatively low mutational loads of mouse tumors, we mutagenized transplantable mouse tumor cell lines EMT-6/P, B16F1, RENCA, CT26, and MC38 in vitro with methylnitro-nitrosoguanidine (MNNG) or ethylmethane sulfonate (EMS) and tested their responsiveness to PD-L1 blockade. Exome sequencing confirmed an increase in somatic mutations by mutagen treatment, an effect mimicked in EMT-6 variants chronically exposed in vivo to cisplatin or cyclophosphamide. Certain mutagenized variants of B16F1, EMT-6/P, CT26, and MC38 (but not RENCA) were more immunogenic than their parents, yet anti-PD-L1 sensitization developed only in some EMT-6/P and B16F1 variants. Treatment response patterns corresponded with changes in immune cell infiltration and especially increases in CD8+ T cells. Chronically cisplatin-exposed EMT-6 variants were also more responsive to anti-PD-L1 therapy. Although tumor PD-L1 expression was upregulated in in vivo chemotherapy-exposed variants, PD-L1 expression levels were not consistently associated with anti-PD-L1 treatment activity across mutagenized or chemotherapy-exposed variants. In summary, mutagenized and more immunogenic mouse tumors were not universally sensitized to PD-L1 blockade. Chemically mutagenized variants may be useful to evaluate the impact of immunologically "hot" or "cold" tumors with a high mutational load, to which certain chemotherapy agents may contribute, on immunotherapy outcomes. Mol Cancer Ther; 17(4); 869-82. ©2018 AACR.
Highlights
Immune checkpoint inhibitors are a relatively new and successful therapeutic modality causing potent responses and prolonged survival times in patient subsets of certain malignancies
Generation and growth of mutagenized mouse tumor variants We first selected EMT-6/P breast cancer, RENCA kidney cancer, and B16F1 melanoma cells for mutagenesis treatment, which are examples of spontaneous (EMT-6 was originally derived from a hyperplastic nodule) and transplantable cell lines that form progressively growing poorly immunogenic tumors [17, 22]
Mutagenized variants remained morphologically similar to parental cells with the exception being MNNG-exposed B16F1, which in addition to some size and shape changes, lost their black pigment phenotype (Supplementary Fig. S2F–S2H)
Summary
Immune checkpoint inhibitors are a relatively new and successful therapeutic modality causing potent responses and prolonged survival times in patient subsets of certain malignancies. Two of the most important immune checkpoints are CTLA-4 that restricts initiation of T-cell responses in lymph nodes, and programmed death receptor 1 (PD-1)/PD-L1 that downregulates the activation and proliferation of antigen-specific T cells in tumors and peripheral tissues [1]. Therapeutic antibodies against CTLA-4 and PD-1/PD-L1 relieve immunosuppression and unmask antitumor effector T-cell responses. It is emerging that cancer types that are most responsive to checkpoint inhibitors often have a high mutational load [2, 3]. Note: Supplementary data for this article are available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/).
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