Clinical responses to anticancer therapies in advanced soft tissue sarcoma (STS) are unluckily restricted to a small subgroup of patients. Much of the inter-individual variability in treatment efficacy is as result of polymorphisms in genes encoding proteins involved in drug pharmacokinetics and pharmacodynamics. The nucleotide excision repair (NER) system is the main defense mechanism for repairing DNA damage caused by carcinogens and chemotherapy drugs. Single nucleotide polymorphisms (SNPs) of NER pathway key genes, altering mRNA expression or protein activity, can be significantly associated with response to chemotherapy, toxicities, tumor relapse or risk of developing cancer. In the present study, in a cohort of STS patients, we performed DNA extraction and genotyping by SNP assay, RNA extraction and quantitative real-time reverse transcription PCR (qPCR), a molecular dynamics simulation in order to characterize the NER pathway in STS. We observed a severe deregulation of the NER pathway and we describe for the first time the effect of SNP rs1047768 in the ERCC5 structure, suggesting a role in modulating single-stranded DNA (ssDNA) binding. Our results evidenced, for the first time, the correlation between a specific genotype profile of ERCC genes and proficiency of the NER pathway in STS.