Comparative molecular profiling of HPV-induced squamous cell carcinomas

Abstract

Approximately 5% of all cancer incidences result from human papillomavirus (HPV) infection. HPV infection most commonly leads to cancers of the anogenital region or oropharynx. It is unknown whether different HPV-mediated cancers collectively share a molecular signature and it is important to determine if there are targetable alterations common to different types of HPV-positive tumors. We analyzed 743 p53 wild-type samples of anal, cervical, oropharyngeal, and vulvar squamous cell carcinomas which underwent multiplatform testing at a commercial molecular profiling service. Expression of 24 proteins was measured by immunohistochemistry (IHC), mutation of 48 genes was determined by next-generation and Sanger sequencing, and copy number alteration for six genes was determined by in situ hybridization. The four cohorts had remarkably similar molecular profiles. No gene had a statistically significant difference in mutation frequency or copy number change between the four different types of squamous cell carcinomas. The only significant differences between cohorts were frequency of ERCC1 and SPARC loss as determined by IHC. In all four cancer types, oncogene mutation and PD-L1 expression was relatively infrequent. The most commonly mutated gene was PIK3CA, with mutations most often affecting the helical domain of the protein and accompanied by concurrent lack of PTEN expression. Loss of MGMT and RRM1 was common among the four cohorts and may be predictive of response to cytotoxic therapies not currently being used to treat these cancer types. The similar molecular profiles of the four cohorts indicate that treatment strategies may be similarly efficacious across HPV-positive cancers.

Keywords: HPV; Biomarkers; DNA sequencing; molecular profiling; protein expression; squamous cell carcinoma.

Figure 1

HPV‐positive squamous cell carcinomas have similar rates of expression profile alteration. (A) IHC revealed similar rates of biomarker detection in ASCC, CSCC, OSCC, and VSCC tumors. Positivity is indicative of high expression unless denoted. # indicates low or no expression as positive. (B) Frequency of gene amplification across SCCs as determined by ISH. P < 0.05 (*); P < 0.001 (**).

Figure 2

Mutation of biomarker genes occurs in similar proportions of different HPV+ squamous cell carcinomas. Mutations of clinical significance were identified through sequencing of 47 genes. Genes that were not mutated in any of the four SCC types were not included in this figure. No difference between cohorts was statistically significant.

Figure 3

Different HPV+ squamous cell carcinomas have similar PIK3CA mutational landscapes. (A) Frequency and distribution of mutations within the PIK3CA gene as determined by NGS and Sanger sequencing. Corresponding protein domains are shown below the graph. (B) Percent of PIK3CA‐mutant tumors with co‐occurring loss of PTEN as determined by IHC. (C) Percent of samples with PIK3CA mutation stratified by disease state. For primary tumors, n = 50 (ASCC), n = 113 (CSCC), n = 56 (OSCC), n = 29 (VSCC). For regional metastases, n = 35 (ASCC), n = 85 (CSCC), n = 12 (OSCC), n = 3 (VSCC). For samples from lymph nodes, n = 11 (ASCC), n = 25 (CSCC), n = 9 (OSCC), n = 14 (VSCC). For distant metastases, n = 27 (ASCC), n = 32 (CSCC), n = 13 (OSCC), n = 1 (VSCC). (D) Percent of PIK3CA‐mutant samples with co‐occurring loss of PTEN expression. For primary tumors, n = 10 (ASCC), n = 32 (CSCC), n = 8 (OSCC), n = 2 (VSCC). For regional metastases, n = 13 (ASCC), n = 28 (CSCC), n = 3 (OSCC), n = 0 (VSCC). For samples from lymph nodes, n = 2 (ASCC), n = 8 (CSCC), n = 4 (OSCC), n = 2 (VSCC). For distant metastases, n = 6 (ASCC), n = 7 (CSCC), n = 3 (OSCC), n = 1 (VSCC). No differences were statistically significant after correcting for multiple comparisons.

Figure 4

PD‐1 and PD‐L1 expression in different squamous cell carcinomas stratified by disease state. (A) Percent of samples positive for PD‐1 expression on tumor infiltrating lymphocytes as determined by IHC. For primary tumors, n = 25 (ASCC), n = 79 (CSCC), n = 31 (OSCC), n = 15 (VSCC). For regional metastases, n = 20 (ASCC), n = 54 (CSCC), n = 9 (OSCC), n = 2 (VSCC). For samples from lymph nodes, n = 5 (ASCC), n = 19 (CSCC), n = 11 (OSCC), n = 9 (VSCC). For distant metastases, n = 10 (ASCC), n = 22 (CSCC), n = 11 (OSCC), n = 2 (VSCC). (B) Percent of tumor samples positive for PD‐L1 expression by IHC. For primary tumors, n = 33 (ASCC), n = 90 (CSCC), n = 34 (OSCC), n = 16 (VSCC). For regional metastases, n = 23 (ASCC), n = 63 (CSCC), n = 10 (OSCC), n = 3 (VSCC). For samples from lymph nodes, n = 5 (ASCC), n = 24 (CSCC), n = 11 (OSCC), n = 9 (VSCC). For distant metastases, n = 10 (ASCC), n = 24 (CSCC), n = 11 (OSCC), n = 2 (VSCC).