Toward Precision Radiotherapy for Use with Immune Checkpoint Blockers

Abstract

The first evidence that radiotherapy enhances the efficacy of immune checkpoint blockers (ICB) was obtained a dozen years ago in a mouse model of metastatic carcinoma refractory to anti-CTLA-4 treatment. At the time, ICBs had just entered clinical testing, an endeavor that culminated in 2011 with the approval of the first anti-CTLA-4 antibody for use in metastatic melanoma patients (ipilimumab). Thereafter, some patients progressing on ipilimumab showed systemic responses only upon receiving radiation to one lesion, confirming clinically the proimmunogenic effects of radiation. Preclinical data demonstrate that multiple immunomodulators synergize with radiotherapy to cause the regression of irradiated tumors and, less often, nonirradiated metastases. However, the impact of dose and fractionation on the immunostimulatory potential of radiotherapy has not been thoroughly investigated. This issue is extremely relevant given the growing number of clinical trials testing the ability of radiotherapy to increase the efficacy of ICBs. Recent data demonstrate that the recruitment of dendritic cells to neoplastic lesions (and hence the priming of tumor-specific CD8⁺ T cells) is highly dependent on radiotherapy dose and fractionation through a mechanism that involves the accumulation of double-stranded DNA in the cytoplasm of cancer cells and consequent type I IFN release. The molecular links between the cellular response to radiotherapy and type I IFN secretion are just being uncovered. Here, we discuss the rationale for an optimized use of radiotherapy as well as candidate biomarkers that may predict clinical responses to radiotherapy combined with ICBs. Clin Cancer Res; 24(2); 259-65. ©2017 AACR.

Figure 1. Cancer cell secretion of interferon type I is an essential signal for a radiation-induced in situ anticancer vaccine

(1) Radiotherapy used at the optimal dose and fractionation leads to accumulation of cytosolic double stranded DNA in the cancer cells, which stimulates the production of interferon type I (IFN-I) via cGAS/STING pathway, and the transactivation of interferon stimulated genes (ISGs), including chemokines such as CXCL10. (2) IFN-I binds to its receptor on the cancer cells, promoting further secretion of IFN-I and expression of several ISGs. (3) IFN-I binds to its receptor on BATF3-depenedent dendritic cells (BATF3-DCs) promoting their recruitment to the tumor and their activation. (4) Once in the tumor BATF3-DCs take up tumor associated antigen (TAA) and tumor-derived DNA, which further stimulates production of IFN-I via cGAS/STING. (5) Optimally activated BATF3-DCs then migrate to tumor-draining lymph nodes (TDLN) where they can cross-prime CD8⁺ T cells. (6) Once activated, tumor-specific CD8⁺ T cells differentiate into cytotoxic T cells (CTLs) that express CXCR3. Their homing to the irradiated tumor is facilitated by CXCL10 and other IFN-induced chemokines. CTLs eliminate the residual cancer cells remaining after radiotherapy, leading to durable tumor regression. (7) The CTLs are also capable to home to distant metastatic sites and reject non-irradiated metastases (abscopal effect).

Figure 2. Proposed strategy for rational selection of the radiation dose to be used in combination with ICBs

A biopsy is performed to test the tumor site selected for irradiation for expression of cGAS and STING by immunostaining. If expression is weak or negative the tumor is analyzed for methylation of the promoter of the genes encoding cGAS, STING and IFNbeta. If sufficient tissue is available, comprehensive genomic and epigenomic analysis can provide additional information about specific mutations and mutation load, and RNASeq can be used to confirm cGAS and STING expression. Another portion of the fresh tumor is used to establish the radiation dose threshold for Trex1 induction versus the optimal induction of IFN-I and ISGs. This can be accomplished in two ways: (1) If sufficient material is available tissue fragments are irradiated in ex vivo cultures. (2) Alternatively, the tissue is used to prepare seeds (0.1×0.3×0.3 cm) for implantation in a subcutaneous pocket on the flank areas of NOD.Cg-Prkdcscid B2mtm1Unc Il2rgtm1Wjl/SzJ (NSG) mice to generate Patient-derived tumor xenografts (PDTX). Once tumors reach 5 mm in average diameter they are irradiated. Obtained information is used to choose the radiation dose and fractionation for treatment of the patient in combination with ICBs. In the event that IFN-I is not induced by any radiation dose due to hypermethylation of the promoter of cGAS- and or STING-encoding genes, radiation is preceded by administration of a demethylating agent.