Somatic Mutation Profiling in Premalignant Lesions of Vulvar Squamous Cell Carcinoma

Abstract

Vulvar squamous cell carcinoma (VSCC) originates from the progression of either a high-grade squamous intraepithelial lesion (HSIL) or differentiated-type vulvar intraepithelial neoplasia (dVIN), often in a background of lichen sclerosus (LS). The mechanisms leading to the progression of these premalignant lesions to VSCC are elusive. This study aims to identify pathogenic mutations implicated in VSCC development. Using next-generation sequencing, 38 HSIL, 19 dVIN, 20 LS, of which 10 were solitary lesions and 10 with adjacent VSCC, and 10 VSCC adjacent to LS, were screened for hotspot mutations in 50 genes covered by the Ion AmpliSeq Cancer Hotspot Panel v2 Kit (Thermo Fisher Scientific). Pathogenic mutations of TP53 were the most common genetic alterations identified in 53% and 24% of dVIN and HSIL cases, respectively, followed by CDKN2A (p16) mutated in 42% and 0% of dVIN and HSIL, respectively. Seven (70%) and three (30%) of 10 cases of VSCC associated with LS carried TP53 and CDKN2A mutations, respectively, whereas neither solitary LS nor LS associated with VSCC cases harbored mutations in these genes. It appears that TP53 mutations are early events during VSCC carcinogenesis, being present in both HSIL and dVIN lesions. Our preliminary data do not support a genetic background for the notion of LS as the VSCC premalignant lesion.

Keywords: CDKN2A; HPV; HSIL; NGS; TP53; dVIN; lichen sclerosus; vulvar squamous cell carcinoma.

Figure 1

Examples of immunohistochemical CDKN2A (middle panel) and TP53 (lower panel) staining performed on tissue sections of high-grade squamous intraepithelial lesion (HSIL) and differentiated-type vulvar intraepithelial neoplasia (dVIN). H&E—hematoxylin and eosin staining (upper panel); 40x magnification—(A,B); 20x magnification—(C–F).

Figure 2

Distribution of pathogenic mutations and variants of uncertain significance in vulvar premalignant lesions. Samples were classified as HSIL (n = 38) and dVIN (n = 19) based on histopathological and IHC assessment. Each column corresponds to an individual tumor case, while each row corresponds to the mutated gene. Samples with no mutations detected are depicted in grey, missense mutations are highlighted in green, whereas red hits indicate nonsense mutations, indel mutation are violet and blue. hrHPV-positive and hrHPV-negative samples are marked in the bottom line with orange and yellow color, respectively.

Figure 3

Distribution of pathogenic mutations in solitary lichen sclerosus (LS) (n = 10) and LS (n=10) associated with VSCC (n = 10). Each column corresponds to an individual LS or VSCC case, while each row corresponds to the mutated gene. Samples with no mutations detected are depicted in grey, missense mutations are highlighted in green, whereas red hits indicate nonsense mutations. hrHPV-positive and hrHPV-negative samples are marked in the bottom line with orange and yellow color, respectively.

Figure 4

Mutation prevalence of the most often mutated genes in dVIN, HSIL, VSCC and LS samples.