Fas (APO-1/CD95) is a 45-kDa membrane protein which regulates apoptosis in many lymphoid cell types. In the present study, FAS expression was examined in primary leukemic cells from infants with acute lymphoblastic leukemia (ALL). The cells were resistant to apoptosis induction by an anti-FAS antibody and expressed nearly undetectable amounts of FAS protein. Molecular analysis of FAS transcripts in these cells revealed no detectable expression of full-length Fas mRNA after a single round of reverse transcription and polymerase chain reaction (PCR) amplification (RT-PCR). However, a more sensitive nested RT-PCR analysis revealed alternatively spliced Fas transcripts in three of five infants (60%) with the remaining two infants showing no detectable Fas mRNA expression. The primary sequence variation of Fas mRNA seen in the samples was a previously described variant lacking exon 6 encoding soluble FAS. However, we also detected the presence of several novel alternatively spliced FAS transcripts in the ALL cells. In one patient, we observed a novel spliced form of soluble Fas, which not only lacked exon 6 but also contained an insertion of an alternative exon 7 (exon 7B). In another, a novel exon 4Del FAS mRNA variant was observed, which contained an additional 4-bp deletion at the exon 5/6-splice junction. These variants lack intact transmembrane domains and thus are predicted to encode soluble FAS variants. The low level of expression of functional full length FAS transcripts with corresponding low level of FAS protein expression in the ALL cells contribute to their resistance to CD95-mediated apoptosis.