Objective Given the recent characterization of ADAMTS‐5 as the main aggrecanase of cartilage destruction in mouse models, we explored whether genetic variation and, in particular, putative damaging polymorphisms in the ADAMTS‐5 gene modify susceptibility to osteoarthritis (OA). Methods Two likely deleterious nonsynonymous single‐nucleotide polymorphisms (SNPs) were identified in ADAMTS‐5 by bioinformatics analysis, rs2830585 in exon 5 affecting a thrombospondin 1 motif, and rs226794 in exon 7. Exploration of their role was carried out in 3 steps, discovery, extension, and replication, on samples obtained from 4 European Caucasian collections, comprising a total of 2,715 patients with knee, hip, or hand OA and 1,185 OA‐free controls. In addition, 6 tagSNPs were studied to fully evaluate genetic variation in the ADAMTS‐5 locus. Results Initial analyses of 2 sample collections (n = 277 and n = 159) showed a trend toward decreased frequency of the putative deleterious allele of rs226794 among patients with severe knee OA (P = 0.047 versus controls). However, results in patients with knee OA from 2 additional sample collections (n = 360 and n = 265) did not confirm this trend. No association was found with hip OA or hand OA. None of the other SNPs or haplotypes constructed with these SNPs showed a significant association with OA susceptibility. Conclusion Use of several collections of OA samples allowed us to obtain sound evidence against the participation of genetic variation in ADAMTS‐5 in OA susceptibility. These results indicate the need to further explore the function of this aggrecanase in human OA to determine whether it is as critical as has been observed in mouse models.