Background: Preoperative radiation therapy (RT) followed by surgery is the main effective treatment for colorectal cancer. However, some patients do not respond to preoperative RT due to the radioresistance of tumor cells, depending on their molecular characteristics, in particular on the expression of a certain group of genes. The aim of this work was to study the effect of expression of BRCA2, H2AX, RBBP8, CASP9 and BCL2 genes on the effectiveness of preoperative RT for rectal tumors. Methods: In the study, paired biopsy preparations (obtained by VCS before irradiation) of normal and tumor tissues of the rectum (adenocarcinoma G1-2) in 30 patients were used. RT was carried out on a Novalis TX linear accelerator (Varian, USA) (SFD = 2.4 Gy to TFD = 54 Gy). RNA isolation was performed by Chomczynski&Sacchi method. Reverta-L kits were used to synthesize cDNA libraries. Using the RT-qPCR method, the relative expression of 5 genes was determined: BRCA2, H2AX, CASP9, RBBP8 and BCL2 (reference GAPDH, ACTB and B2M). Statistical analysis was performed using the Shapiro–Wilk and Mann–Whitney criteria. Results: 2 clusters of patients were identified that differed in the expression of the BRCA2, H2AX, CASP9, RBBP8 and BCL2 genes. In cluster 1, 81% of patients had increased expression of the CASP9 gene and 100% decreased expression of the BRCA2, H2AX, RBBP8, and BCL2 genes. In cluster 2, the expression of the CASP9 gene was reduced in 100% of patients and the expression of the BRCA2, H2AX, RBBP8 and BCL2 genes was increased in 93%. A subsequent analysis of the results of RT showed that 16 patients had complete tumor regression after RT, while expression of the H2AX and RBBP8 genes was reduced by a factor of 2 (p < 0.05), and the expression of the CASP9 gene was increased by a factor of 4 (p < 0.005) relative to normal tissues. 8 patients showed slight tumor regression, and 6 patients had no regression at all, while their expression of the BRCA2, H2AX, RBBP8 and BCL2 genes was statistically significantly (p < 0.005) 2, 4, 6 and 3 times higher, respectively, and that of CASP9 gene was 4 times lower than in patients with complete tumor regression. Conclusions: The study found that the expression of BRCA2, H2AX, CASP9, RBBP8 and BCL2 genes affects the effectiveness of RT, enhancing it in patients with increased expression of CASP9 and reduced expression of H2AX and RBBP8, and vice versa, the effectiveness of therapy decreases with increased expression of H2AX, RBBP8, BRCA2 and BCL2.