Identification of Risk Loci for Radiotoxicity in Prostate Cancer by Comprehensive Genotyping of TGFB1 and TGFBR1

Abstract

Genetic variability in transforming growth factor beta pathway (TGFB) was suggested to affect adverse events of radiotherapy. We investigated comprehensive variability in TGFB1 (gene coding for TGFβ1 ligand) and TGFBR1 (TGFβ receptor-1) in relation to radiotoxicity. Prostate cancer patients treated with primary radiotherapy (n = 240) were surveyed for acute and late toxicity. Germline polymorphisms (n = 40) selected to cover the common genetic variability in TGFB1 and TGFBR1 were analyzed in peripheral blood cells. Human lymphoblastoid cell lines (LCLs) were used to evaluate a possible impact of TGFB1 and TGFBR1 genetic polymorphisms to DNA repair capacity following single irradiation with 3 Gy. Upon adjustment for multiplicity testing, rs10512263 in TGFBR1 showed a statistically significant association with acute radiation toxicity. Carriers of the Cytosine (C)-variant allele (n = 35) featured a risk ratio of 2.17 (95%-CI 1.41-3.31) for acute toxicity ≥ °2 compared to Thymine/Thymine (TT)-wild type individuals (n = 205). Reduced DNA repair capacity in the presence of the C-allele of rs10512263 might be a mechanistic explanation as demonstrated in LCLs following irradiation. The risk for late radiotoxicity was increased by carrying at least two risk genotypes at three polymorphic sites, including Leu10Pro in TGFB1. Via comprehensive genotyping of TGFB1 and TGFBR1, promising biomarkers for radiotoxicity in prostate cancer were identified.

Keywords: LCL; Leu10Pro; SNP; TGBF1; TGFB; biomarkers; irradiation; prostate cancer; radiotherapy; rs10512263; side effects; toxicity.

Figure 1

Flowchart of patient selection for study cohort. LDR = low dose rate radiotherapy (i.e., brachytherapy with permanent seeds).

Figure 2

Late radiation toxicity stratified by none or mild (<°2) versus medium or high sequelae independence on the numbers of three risk genotypes (Guanine/Guanine at TGFB1 rs10417924, Cytosine/Cytosine at TGFB1 rs1800470, and Thymine/Thymine at TGFBR1 rs78471739). (a) Numbers of patients, (b), relative distribution of risk genotypes.

Figure 3

TGFB1 (panel (a)) and TGFBR1 (panel (b)) genetic polymorphisms in relation to residual γH2AX foci. Upon single irradiation of 3 Gy (with 3 µM 5-fluorouracil used as radiosensitizer), with (filled triangles) or without (open squares) 5 ng/mL TGFβ1, LCLs were subsequently incubated for 6 h at 37 °C to allow for DNA repair prior to γH2AX staining. Data were each normalized to total DNA content (by co-staining with allophycocyanin) and to only medium-treated control cells. (c,d) Visual depiction of residual γH2AX foci rate with TGFβ1 stimulation dependent on genotypic configurations at TGFBR1 rs10512263 and rs34733091. The boxplots indicate the data distributions as follows: The rectangle represents 50% of the values of a given distribution with the lower horizontal line reflecting the 25%- (Q₂₅), and the upper the 75%-(Q₇₅) quartile. The difference of these two delimiters is called the interquartile distance (IQA). Values within 1.5-times of the IQA below Q₂₅ or 1.5-times above Q₇₅ are depicted by the whiskers of the blot (vertical line limited by short horizontal). Values out of this range are either marked by circles (>1.5-times, but ≤ 3-times of IQA referred to Q₂₅ or Q₇₅) or asterisks (beyond 3-times of IQA on either side). Abbreviations: ATG = initiation codon; bp = base pairs, w/o = without; TT = Thymine/Thymine; TC = Thymine/Cytosine; CC = Cytosine/Cytosine.