HPV Infection and Immunohistochemical Analysis of P16, P53, and PD-L1 Expression as Prognostic Biomarkers in Squamous Cell Anal Cancer Patients Receiving Definitive Radiotherapy/Chemoradiotherapy

Abstract

The treatment of anal squamous cell carcinoma (SCC) with definitive radiotherapy (RT)/chemoradiotherapy (CRT) has a high likelihood of success; nevertheless, treatment resistance and recurrence rates cannot be ignored. Therefore, we aimed to identify the relationship between immunohistochemical (IHC) evaluation, treatment response, and prognosis. This retrospective study included 42 patients with anal SCC treated with definitive RT/CRT at a single institution between 2006 and 2020. Detection of high-risk HPV-DNA and IHC analysis of p16, p53, and PD-L1 expression was performed from diagnostic formalin-fixed, paraffin-embedded (FFPE) biopsies. Positive staining was accepted as >5% in tumor cells for p16 and p53 expression in addition to ≥1% combined positive score (CPS) [(PD-L1 positive tumor, lymphocyte, macrophage count/total viable cell count) x100] for PD-L1 expression. Thirty patients (71.4%) had a complete response to definitive RT/CRT. Recurrence was observed in 16 (38.1%) patients, with 3 (7.1%) having locoregional recurrence (LRR), 6 (14.3%) having distant metastases (DM), and 7 (16.7%) having both LRR and DM. Twenty-four (57.1%) patients were alive. Thirty (71.4%) patients were HPV+, while 12 (28.6%) were HPV-. There was a significant correlation between HPV+ and p16+ status (p<0.001). HPV- status was associated with the male gender (p = 0.001). HPV- and p16- status were significantly associated with a lack of complete response to definitive RT/CRT (p<0.001, p<0.001, respectively). Furthermore, there was a significant relationship between lack of complete response and increased recurrence (p = 0.016) and distant metastases (p = 0.015). Ten of the 16 patients with recurrence were p53+; a significant correlation was found between recurrence and p53+ status (p = 0.006). Similarly, p53+ status was associated with increased LRR (p = 0.014). PD-L1 CPS ≥ 1% was found in 31 (73.8%) patients. PD-L1 positivity was significantly correlated with HPV+ (p = 0.026) and p16+ (p = 0.013) status. All 10 (23.8%) patients with LRR were PD-L1+; PD-L1 CPS ≥ 1% was associated with poor local control (p = 0.031). In univariate analysis, age [<65] (p = 0.049), complete response (p = 0.015), and HPV+ status (p = 0.010) were related to increased 5-year (y) overall survival (OS); complete response (p = 0.001), HPV+ status (p = 0.025) and p53- status (p = 0.010) were associated with increased 5-y disease-free survival (DFS). In multivariate analysis, age [<65] (p = 0.010) and HPV+ status (p = 0.002) were significant prognostic factors for 5-y OS, whereas complete response (p = 0.007) and p53- status (p = 0.038) were significant prognostic factors for 5-y DFS. Patients with HPV- status and/or poor prognostic biomarkers should be identified at diagnosis. Thus, better outcomes can be achieved with different treatment options, such as combining immunotherapy and standard CRT.