Toxicogenomic assessments in vitro might serve to generate novel toxicity biomarkers and support the replacement of animals in toxicity testing. Toxicity of formaldehyde (FA), a classified human carcinogen, was studied in a cell culture model for cancer development that included normal (NOK), immortalized (SVpgC2a) and malignant (SqCC/Y1) cells. Repeated 1h FA exposures of SVpgC2a induced transformation as indicated from cell crisis and the generation of a novel line (SVpgC3a) with altered morphology (e.g., multifocal, soft agar growth, but non-tumorigenic in athymic nude mice). Assessment of cytotoxicity (MMT method) and genotoxicity (DNA protein crosslinks) indicated similar initial damage levels in all lines whereas the longer-term consequences differed (cell number and cloning assessments indicated a sensitivity order of NOK>SVpgC2a>SVpgC3a≈SqCC/Y1). FA induced cell death primarily by terminal differentiation in NOK whereas differently SVpgC2a, SVpgC3a and SqCC/Y1 died by other means e.g., apoptosis. Interestingly, over a dose range, the highest levels of micronuclei in SVpgC2a were observed at the FA concentration that induced cell transformation. Proteomics and transcriptomics profiling of SVpgC3a versus SVpgC2a showed that cell transformation coupled with multiple changes in single genes, molecular networks and gene ontologies. Likewise, transcript profiling of the respective cell lines for up to 48h following 1h FA exposure indicated multiple genomic changes, some of which overlapped to results from cancer-inducing protocols in vivo. In conclusion, the sensitivity to formaldehyde toxicity might differ between stages in cancer development, and specific gene expression changes couple to the respective phenotypes. Generation of the SVpgC3a line extended the current cancer model of normal, immortal and malignant cells.