chromosome 6q23-q24 to profile DNA methylation was used to test the hypothesis that abnormal promoter methylation could help pinpoint the location of a candidate tumor suppressor in regions of LOH. 6q23-q24 was the chosen chromosomal region due to frequently described LOH in human head and neck squamous cell carcinomas (HNSCC) and non-small-cell lung cancers (NSCLC) as well as in other tumor types, but with no identified tumor suppressor. Hypermethylation was found to occur frequently in the same RLGS loci in HNSCC and NSCLC. Sodium bisulfite sequencing further identified tumor-specific methylation of TCF21 when compared to normal controls. RNA samples were isolated from tumor tissues and analyzed to correlate the amount of TCF21 mRNA and DNA methylation in the samples. Overall, tumor samples with higher levels of CpG island hypermethylation had decreased TCF21 expression than normal controls. Exogenous expression of TCF21 in cells with silenced endogenous TCF21 loci resulted in a reductions in tumor properties both in vitro and in vivo. Based on these results, it was concluded that TCF21 is a tumor suppressor gene, often silenced by hypermethylation in cancer. TCF21 has also been linked to metastatic melanoma progression through the inhibition of the KISS1 metastasis-suppressor gene. DNA methylation analysis in melanoma patient biopsies has demonstrated downregulation of TCF21 due to promoter hypermethylation, which also correlates with decreased survival in patients suffering from metastatic skin melanoma. TCF21, together with E12 and TCF12, bind the KISS1 promoter, sustaining its activity. Without TCF21 to interact with the KISS1 promoter, KISS1 expression is lost. Melanoma cells overexpressing TCF21 have also been found to display reduced motility compared with vector-only control cells. Tobacco-induced lung cancer research has found TCF21 to be among the genes identified as highly methylated at both high and low concentrations of cigarette smoke condensate (CSC). In the presence genistein, one of the soy-derived bioactive isoflavones, methylation of TCF21 is significantly reduced. Genistein has been known to affect tumorigenesis through epigenetic regulation, such as chromatin configuration and DNA methylation, activating other tumor suppressor genes that affect cancer cell survival. These findings support the hypothesis that increasing hypermethylated tumor suppressor genes such as TCF21 is a potential chemopreventative pathway in tobacco-induced lung cancer. TCF21 may have also therapeutic potential for breast cancer treatment, as downregulation of TCF21 has been implicated in breast cancer tumorigenesis and proliferation. TCF21 mRNA expression is very low in breast cancer cells compared with normal breast epithelial cells.