Abstract
There is a pressing need for novel immunotherapeutic targets in colorectal cancer (CRC). Cytotoxic T cell infiltration is well established as a key prognostic indicator in CRC, and it is known that these tumor infiltrating lymphocytes (TILs) target and kill tumor cells. However, the specific antigens that drive these CD8+ T cell responses have not been well characterized. Recently, phosphopeptides have emerged as strong candidates for tumor-specific antigens, as dysregulated signaling in cancer leads to increased and aberrant protein phosphorylation. Here, we identify 120 HLA-I phosphopeptides from primary CRC tumors, CRC liver metastases and CRC cell lines using mass spectrometry and assess the tumor-resident immunity against these posttranslationally modified tumor antigens. Several CRC tumor-specific phosphopeptides were presented by multiple patients’ tumors in our cohort (21% to 40%), and many have previously been identified on other malignancies (58% of HLA-A*02 CRC phosphopeptides). These shared antigens derived from mitogenic signaling pathways, including p53, Wnt and MAPK, and are therefore markers of malignancy. The identification of public tumor antigens will allow for the development of broadly applicable targeted therapeutics. Through analysis of TIL cytokine responses to these phosphopeptides, we have established that they are already playing a key role in tumor-resident immunity. Multifunctional CD8+ TILs from primary and metastatic tumors recognized the HLA-I phosphopeptides presented by their originating tumor. Furthermore, TILs taken from other CRC patients’ tumors targeted two of these phosphopeptides. In another cohort of CRC patients, the same HLA-I phosphopeptides induced higher peripheral T cell responses than they did in healthy donors, suggesting that these immune responses are specifically activated in CRC patients. Collectively, these results establish HLA-I phosphopeptides as targets of the tumor-resident immunity in CRC, and highlight their potential as candidates for future immunotherapeutic strategies.
Highlights
Tumor infiltration by effector T cells has been confirmed by several large studies to be significantly associated with good prognosis in colorectal cancer (CRC), even in metastatic disease [1,2,3,4,5]
We evaluated the responses of tumor infiltrating lymphocytes (TILs) taken from the same tumors that were used for phosphopeptide identification, and broadened this to determine if the immunogenic phosphopeptides may represent public T cell targets in CRC
Two liver metastases and three CRC cell lines were used to identify a total of 198 phosphopeptides, of which 120 were tumor-associated in our cohort: 74 tumor-specific, 12 tumor-associated, and 34 CRC cell line-associated phosphopeptides (Supplementary Tables 1–5)
Summary
Tumor infiltration by effector T cells has been confirmed by several large studies to be significantly associated with good prognosis in CRC, even in metastatic disease [1,2,3,4,5]. Cytotoxic T cells recognize antigenic peptides presented by HLA-I complexes on the surface of cancer cells. These T cells release cytotoxic factors, which kill the transformed cells and can control tumor growth. The remaining 14% of tumors that may respond to ICB therapies, have a defect in DNA mismatch repair, leading to microsatellite instability (MSI) and a high tumor mutational burden (TMB) [8, 9]. It has been proposed that the T cells infiltrating MSI CRC are targeting the mutational neoantigens that arise from the high TMB in these tumors [12, 13]. Whilst some TILs may target mutational neoantigens in MSS CRC, it seems unlikely that T cell targeting of these limited antigens could fully explain the control of tumor growth in CRC; other classes of tumor antigens are almost certainly implicated [6]
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