Genetic alterations in metastatic cutaneous squamous cell carcinoma (CSCC) which might serve as prognostic biomarkers are not well investigated. We investigated the mutation status and protein expression of the CDKN2A (INK4a-ARF) and TP53 genes in metastatic CSCCs and correlated this with clinicopathological variables, HPV presence, and survival data. Sequence analysis was performed on formalin-fixed and paraffin-embedded tissue of 35 metastases and their primary tumors, and was correlated with immunohistochemical stainings for p53, p16 and p14. Beta-PV and alpha-PV DNA was detected using PCRbased assays. Kaplan-Meier and Cox regression methods were used for survival assessment. CDKN2A was mutated in 31% of the metastases and their primary tumors, while the TP53 gene was mutated in 51% of the metastases. P53 protein expression was significantly associated with missense type of mutations (p 5 0.002). No persistent HPV types were detected. CDKN2A mutations were significantly associated with disease-specific death (p 5 0.001). A significant difference was observed in disease-specific survival between patients with or without a CDKN2A mutation (p 5 0.010), while this was not the case for TP53. At univariate Cox's regression analysis tumor size (p 5 0.010), invasion depth (p 5 0.030) and CDKN2A mutations (p 5 0.040) were significantly related to shorter disease-specific survival. At multivariate Cox's regression only tumor size had an adverse effect on survival (p 5 0.002). In conclusion, our study indicates that the CDKN2A mutation status might be of prognostic value in metastatic CSCCs. In most cases, CDKN2A and TP53 mutations are early genetic events in CSCC tumorigenesis. The possible role of HPV in metastatic CSCC needs further exploration.Cutaneous squamous cell carcinoma (CSCC) is the second most common malignancy in Caucasians, after basal cell carcinoma, and its incidence is still rising. 1 Generally the risk of metastasis is reported up to 5%, while immunocompromised patients have a larger tendency to metastasize (5-10%). 1,2 The outcome of these patients once metastasized is poor, with a 5-year survival rate between 25 and 50%. 3,4 Genetic alterations in CSCCs which might serve as prognostic biomarkers are not well investigated.TP53 mutations are the most frequent genetic alterations in CSCC with incidences of up to 50%. 5,6 TP53 mutations are early events in skin carcinogenesis, with a similar mutation frequency of 50% in actinic keratosis, a precancerous skin lesion that potentially can progress to CSCC. 7 Mutations in the CDKN2A (INK4a-ARF) locus are reported in CSCC with frequencies ranging up to 24% in sporadic CSCC. [8][9][10] This gene maps to chromosome 9p21, which by alternative transcripts encodes for 2 cell cycle regulatory proteins, p16 INK4a (Exons 1a, 2 and 3) and p14 ARF (Exons 1b, 2 and 3). 10,11 The exact role of the CDKN2A gene in skin carcinogenesis is not well understood. Especially in metastatic CSCCs alterations in the CDKN2A gene have not been investigated. In a small previous stud...