The pathogenesis of AIDS-related non-Hodgkin lymphomas (AIDS-NHLs) is associated with chromosomal translocations that deregulate the expression of various oncogenes. Recently, a novel mechanism of genetic lesion, termed aberrant hypermutation, has been identified in diffuse large B-cell lymphoma (DLBCL) of immunocompetent hosts. In these tumors, the somatic hypermutation (SHM) process that normally targets immunoglobulin V (IgV) genes in B cells appears to misfire and causes mutations in the 5' sequences of multiple proto-oncogenes, including PIM-1, PAX-5, RhoH/TTF, and c-MYC. To investigate whether aberrant hypermutation occurs also in AIDS-NHL, we studied the mutation profile of these 4 genes in various histologic subtypes. Mutations in 1 gene or more were detected in 19 of 39 (48.7%) AIDS-NHL cases (10 of 18 AIDS-diffuse large B-cell lymphoma; 4 of 11 AIDS-Burkitt lymphoma; 4 of 6 AIDS-primary effusion lymphoma; 1 of 4 AIDS-primary central nervous system lymphoma), with 9 of 39 (23.1%) cases carrying mutations in 2 or more genes. Overall, PIM-1 was mutated in 5 of 39 (12.8%), PAX-5 in 8 of 39 (20.5%), RhoH/TTF in 9 of 39 (23.1%), and c-MYC in 7 of 27 (25.9%) AIDS-NHL cases. Mutations were represented mainly by single base pair substitutions (n = 63) with rare deletions/insertions (n = 5) and displayed features typical of the IgV-associated SHM process. In addition, a number of mutations in PIM-1 and c-MYC were found to affect coding exons, leading to amino acid substitutions with likely functional consequences. Analysis of intraclonal heterogeneity documented that the aberrant hypermutation activity may be ongoing in at least some cases. These data indicate that aberrant hypermutation is associated with various subtypes of AIDS-NHL and may represent a major contributor to their pathogenesis.