Thanks to the improved awareness of cancer screening, CRC is now detected at an early stage, resulting in a better rate of survival. Surgery without chemotherapy, which was deemed as the curative treatment, was carried out on the majority of patients with stage I/II colon cancer and in some cases of stage I/II rectal cancer. Indeed, it enabled prevention from unnecessary side effects of chemotherapy. Nevertheless, more than 20% of patients with stage I/II CRC who underwent surgical resection still suffered recurrence. Quite a few multigene prognostic signatures have been developed for CRC, but none of them graduated to widespread application due to the uncertainty of prognostic accuracy. Accordingly, an effective prognostic model composed of multiple biomarkers to distinguish early-stage patients with a high risk of recurrence is crucial and necessary for elective adjuvant chemotherapy or other targeted treatments.
Emerging studies have revealed that autophagy functions diversely in the development, maintenance, and progression of tumors. While autophagy may prevent cellular cancerous transformation in normal tissue, it acts as a survival mechanism in established tumors, especially under stress conditions and in response to chemotherapy. Several autophagy inhibitors and activators have been brought up as improved chemotherapeutic options for cancer treatment, but without sufficient clinically significant results, especially in CRC. Accordingly, further investigation on the biological mechanism of autophagy in the tumor microenvironment deserves attention, and more targets associated with autophagy awaits to be found.
In our study, we developed a prognostic model comprised of 10 ATGs for stage I/II CRC. This ATG signature, which classified patients into high and low autophagy risk groups, demonstrated a significant difference in 5-year DFS. The C-index of the ATG signature exhibited a good clinical predicting fitness superior to Oncotype DX colon. Validation results suggested that the ATG signature could successfully predict DFS for stage I/II CRC patients after treatment. This novel model enabled us to identify CRC patients with high autophagy risk stood for increased risk of tumor recurrence. Surprisingly, despite the original intention for DFS prediction of early-stage CRC patients, the ATG signature also showed a significant effect in the prediction for all stages. Therefore, this model could be applied to predict tumor recurrence in all CRC patients independent of tumor stage. Upon the prognostic prediction, clinicians could thus make an informed decision regarding supplementary treatment regimens. For example, adjuvant chemotherapy might improve the outcome of CRC patients with high autophagy risk, for there was no significant difference in DFS between the two autophagy risk groups after adjuvant chemotherapy in this study. It might reduce the impact of autophagy on tumor relapse through massive, indiscriminate cell killing, and immunosuppression.
As we looked over the genes within the ATG signature, some of them have been found correlated to CRC, but mostly bear context-dependent biological functions in cancers, similar to autophagy. For example, the cytosolic histone deacetylase 6 (HDAC6) served as a tumor suppressor in hepatocellular carcinogenesis, while another study revealed HDAC6 inhibitor significantly suppressed the proliferation and viability, and induced apoptosis in CRC cells, where autophagy activation was observed. Elevated Sirtuin 2 (SIRT2) was found to be associated with poor prognosis in CRC patients via its participation in tumor angiogenesis. Meanwhile, in a separate study SIRT2 was found to be downregulated in colon cancer biopsies compared to adjacent noncancerous tissues, and overexpression of SIRT2 inhibited the proliferation and metastatic progression of SW480 cells. In terms of autophagy-related functions, a previous investigation reported that in response to oxidative stress or serum starvation, SIRT2 dissociated as acetylated FOXO1, which later bound to autophagy protein 7 (ATG7) and induced autophagy in tumors. As these inconsistencies make it difficult to clarify the role of autophagy in CRC, we further investigated the biological functions of the ATG signature, expecting to find some clues in the autophagy-related functions in tumors.
GSEA revealed that the ATG signature included genes that were robustly involved in multiple immune/ inflammatory pathways including IL6/JAK/STAT3, IL2/STAT5, IFN-α, IFN-γ, TNF-α/NF-κB, and the inflammatory response presented a particular relation to CRC proliferation or prognosis as previous studies revealed[21-25]. As our findings suggested that a high autophagy risk score correlated with the down-regulation of these immune/ inflammatory pathways, we speculated that autophagy might play a role in CRC tumorigenesis and tumor proliferation via an anti-immune/ anti-inflammatory response. Moreover, increased infiltration of Tregs and decreased infiltration of M1 macrophages observed in the high autophagy risk group during immune infiltration analysis seemingly catered to the anti-immune/ anti-inflammatory response. No significant difference in cytotoxic T lymphocyte was found in the immune infiltration analysis. Tregs are known to suppress both antibody-mediated and cell-mediated immune responses. The pro-inflammatory M1 macrophages, a phenotype of tumor-associated macrophages (TAMs), correlated to a better prognosis in CRC patients for its tumor-suppressing function[27, 28]. By triggering an anti-immune or anti-inflammatory response, autophagy might promote polarization or re-polarization towards the M2 phenotype spontaneously and thus lead to the decrease of M1 infiltration observed. Previous studies have described the link between autophagy and macrophage polarization in tumor microenvironment. For example, mTOR which functions as a conserved kinase protein in the regulation of autophagy also participates in the polarization of monocytes into TAMs. More new strategies targeting TAM polarization as well as autophagy await exploration and further studies are needed to clarify the above speculations.
However, our prognostic ATG signature relies on the gene expression profiles from microarray platforms, which make it too expensive and time-consuming to popularize in clinical application. In addition to the dataset limitations from retrospective studies, further prospective clinical tests are recommended to validate our results. Despite the limitations, our research proposes a novel perspective to predicting the prognosis of early CRC patients and offers valuable insights into the relationship between autophagy, immune/ inflammatory response, and tumorigenesis.