Small cell lung cancer (SCLC) subtypes, defined by the expression of lineage-specific transcription factors (TFs), are thought to be mutually exclusive, with intra-tumoral heterogeneities. This study investigated the mechanism underlying this phenomenon with the aim of identifying a novel vulnerability of SCLC. We profiled the expression status of ASCL1, NEUROD1, POU2F3, and YAP1 in 151 surgically obtained human SCLC samples. On subtyping, a high degree of mutual exclusivity was observed between ASCL1 and NEUROD1 expression at the cell, but not tissue, level. Inducible co-expression models of all combinations of ASCL1, NEUROD1, POU2F3, YAP1, and ATOH1 using SCLC cell lines showed that some expression combinations, such as ASCL1 and NEUROD1, exhibited mutual repression and caused growth inhibition and apoptosis. Gene expression and ATAC-seq analyses of the ASCL1 and NEUROD1 co-expression models revealed that co-expression of ASCL1 in NEUROD1-driven cells, and of NEUROD1 in ASCL1-driven cells, both (although more efficiently by the former) reprogrammed the cell lineage to favor the ectopically expressed factor, with rewiring of chromatin accessibility. Mechanistically, co-expressed NEUROD1 in ASCL1-driven SCLC cells caused apoptosis by downregulating BCL2, likely in a MYC-independent manner. In conclusion, lethal co-expression intolerance underlies the mutual exclusivity between these pioneer TFs, ASCL1 and NEUROD1, in an SCLC cell. Further investigation is warranted to enable therapeutic targeting of this vulnerability.