Resistance to preoperative chemoradiotherapy (CRT) is a major obstacle to cancer treatment in patients with locally advanced rectal cancer. This study was to explore genome alterations in rectal cancer under CRT stress. Methods: Whole-exome sequencing (WES) was performed on 28 paired tumors collected before and after CRT from the same patients who did not respond to CRT treatment. Somatic point mutations and copy number variations were detected by VarScan2 and Exome CNVs respectively using paired tumor and blood samples. Somatic alterations associated with CRT resistance were inferred considering differences in significantly mutated genes, mutation counts and cancer cell fraction between matched pre- and post-CRT tumors. We employed SignatureAnalyzer to infer mutation signatures and PyClone to decipher clonal evolution and examine intratumoral heterogeneity in tumors before and after CRT. The associations between intratumoral heterogeneity and patients' survival were analyzed using the log-rank test and the Cox regression model. Results: (i) Recurrent mutations in CTDSP2, APC, KRAS, TP53 and NFKBIZ confer selective advantages on cancer cells and made them resistant to CRT treatment. (ii) CRT alters the genomic characteristics of tumors at both the somatic mutation and the copy number variation levels. (iii) CRT-resistant tumors exhibit either a branched or a linear evolution pattern. (iv) Different recurrent mutation signatures in pre-CRT and post-CRT patients implicate mutational processes underlying the evolution of CRT-resistant tumors. (v) High intratumoral heterogeneity in pre- or post-CRT is associated with poor patients' survival. Conclusion: Our study reveals genome landscapes in rectal cancer before and after CRT and tumors evolution under CRT stress. The treatment-associated characteristics are useful for further investigations of CRT resistance in rectal cancer.

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