Background The level of expression of a mutation-associated neoantigen (MANA) is a critical determinant of effective T cell priming.1 Thus, interventions that increase the expression of a protein containing a MANA could increase responses to immune checkpoint inhibitors (ICI), especially for tumors with relatively low mutation burden like microsatellite stable (MSS) colorectal cancer (CRC).1 We have shown that radiation therapy (RT) upregulates the expression of genes containing MANA in a preclinical model, resulting in enhanced presentation of the derived MANA.2 We also found expansion of CD8 T cells against a MANA upregulated in expression by RT in a lung cancer patient with complete response to RT+ipilimumab.3 Here we set out to test if MANA upregulation is a common response of human tumors to RT.
Methods Patient-derived tumor organoids (PDO) from colorectal (CRC) (n=3) (figure 1a) and lung (n=4) cancer were characterized for radiation response4 (figure 1b). PDOs were then irradiated with doses of 5-8 Gy daily for 3 days (RT) or left untreated (UT) and RNAseq performed 24h post-RT. Differentially expressed genes were identified using DESeq2 differential expression analysis. Ingenuity Pathway Analysis (IPA) was used to identify pathways modulated by RT.
Results In the CRC PDO, 2034 and 362 genes were found to be concordantly upregulated and downregulated, respectively, by RT (figure 1c). Upregulated genes were associated with p38 MAPK signaling and innate immune signaling pathways (figure 1d). This RT-modulated gene set was used to interrogate predicted MANA dataset from CRC patients . We found that ≥1 MANA-encoding gene was predicted to be significantly upregulated by RT in 263/266 patients from the TCGA COADREAD MSS cohort (figure 2a). The median number/patient of upregulated MANA was 11 (range 0-79), while for downregulated MANA it was 0 (range 0-15, figure 2b). In the lung PDO, 439 and 117 genes were found to be concordantly upregulated and downregulated, respectively, by RT. Upregulated genes were largely associated with DNA damage response pathways. We found that ≥1 MANA-encoding gene was predicted to be significantly upregulated by RT in 29/35 patients from a lung cancer dataset.5 The median number/patient of upregulated MANA was 3 (range 0-25), while for downregulated MANA it was 0 (range 0-14).
Conclusions Our data support the novel concept that RT can uncloak neoantigens in tumors like MSS CRC where low neoantigen expression may preclude effective T cell priming and contribute to ICI resistance.
Acknowledgements The authors are grateful for the financial support of The NIH (R01CA198533, SD) and from the AACR (21-40-12-VANN, SV).
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Lhuillier C, Rudqvist NP, Yamazaki T, et al. Radiotherapy-exposed CD8+ and CD4+ neoantigens enhance tumor control. J Clin Invest. 2021;131(5):E138740.
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Martin ML, Adileh M, Hsu KS et al. Organoids reveal that inherent radiosensitivity of small and large intestinal stem cells determines organ sensitivity. Cancer Res. 2020;80(5):1219–1227.
Rizvi NA, Hellmann MD, Snyder A et al. Cancer immunology. Mutational landscape determines sensitivity to PD-1 blockade in non-small cell lung cancer. Science. 2015;348(6230):124–8.
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