The Role and Therapeutic Target of Base Excision Repair Genes in Multiple Myeloma (MM)

Background: Dysregulated DNA repair genes play a pathological role in diverse forms of malignancies, including multiple myeloma (MM). High expression of some DNA repair enzymes has been reported in various tumors and associated with their progression and drug resistance. Here, we investigated the association between MM and base excision repair (BER) system, which is known as a DNA repair pathway for the processing of small base lesions derived from oxidation and alkylation damages. To elucidate the roles of BER in the pathogenesis of MM, we investigated the distribution of functional polymorphisms of BER, the expression level of BER genes, and the effectiveness of BER inhibitors and knockdown BER genes for MM cell lines.

Material and methods: Genotyping of OGG1 Ser326Cys, MUTYH Gln324His, APE1 Asp148Glu, XRCC1 Arg399Gln, and PARP1 Val762Ala in 146 MM patients and 202 race-matched healthy controls was determined by using the PCR-restriction fragment length polymorphism (PCR-RFLP) method. Mononuclear cells were separated from bone marrow aspirates and CD138+ cells were collected using EasySep. The mRNA expression of BER genes in MM cell lines (KMM1, KMS12PE, KMSE12BM, KMS11, RPMI8226, and OPM2), primary MM cells, and normal plasma cells was assessed by real-time PCR method. The protein level of BER genes was assessed by western blot analysis. BER inhibitors, including APE1 inhibitor and PARP inhibitor were used. Inducible knockdown system was used for suppressed OGG1 expression. Genotype and allele frequencies were compared between the study groups using Χ2-test. Probability values <0.05 were considered statistically significant. This study was approved by the Institutional Review Board of Gunma University Hospital.

Results: The PARP1 Val762Ala single nucleotide polymorphism, which is reported to increase cancer risk, was associated with susceptibility to MM. In the patient group, the Val/Val genotype frequency was lower (32.9% compared with 43.6%, P = 0.044) and the Val/Ala genotype frequency was higher (56.8% compared with 43.1%, P = 0.011) than those in the control groups. In contrast to PARP1 polymorphism, the distributions of other BER polymorphisms, including OGG1 Ser326Cys, MUTYH Gln324His, APE1 Asp148Glu, and XRCC1 Arg399Gln, showed no significant difference between MM patients and control groups. Next, we examined the association between the polymorphisms and the clinical characteristics of MM patients. No significant difference was observed in the allele frequency as well as in sex, age, other clinical settings, and overall survival. The BER genes, such as OGG1, APE1, and PARP1, were detected in all MM cell lines. Protein expression of OGG1, APE1, and PARP1 correlated with mRNA expression in MM cell lines. The mRNA levels of OGG1, APE1, and PARP1 of malignant plasma cells of MM patients were significantly higher than those of normal plasma cells (p < 0.05). The mRNA levels of OGG1, APE1, and PARP1 genesof plasma cells of newly diagnosed MM patients were significantly higher than those of refractory or relapsed MM patients. We exposed MM cell lines to increasing concentrations of APE-1 inhibitor, methoxamine, and PARP1-selective inhibitor, BYK204165, which demonstrated potent and selective growth inhibition in vitro. To determine the role of OGG1 gene in MM growth and survival, we suppressed its expression using an inducible knockdown system and observed significant inhibition of MM cell lines (KMS11 and KMM1) proliferation.

Conclusion: The PARP1 Val762Ala polymorphism may be associated with the susceptibility to MM. High expression of BER genes such as OGG1, APE1, and PARP1 was detected in malignant plasma cells, and the inhibition of these BER genes reduced MM cell growth. According to these results, inhibition of BER genes may be a potential therapeutic target of MM.