Abstract
Effective treatments for recurrent/metastatic human papillomavirus-positive (HPV+) head and neck squamous cell cancer (HNSCC) are limited. To aid treatment development, we characterized a novel murine model of recurrent/metastatic HPV+ HNSCC. Further analysis of the parental tumor cell line and its four recurrent/metastatic derivatives led to preclinical testing of an effective treatment option for this otherwise fatal disease. Reverse phase protein arrays identified key signaling cascades in the parental and recurrent/metastatic cell lines. While protein expression profiles differed among the recurrent/metastatic cell lines, activated proteins associated with the mTOR signaling cascade were a commonality. Based on these data, mTOR inhibition was evaluated as an adjuvant treatment for recurrent/metastatic disease. mTOR activity and treatment response were assessed in vitro by western blot, Seahorse, proliferation, clonogenic, and migration assays. Standard-of-care cisplatin/radiation therapy (CRT) versus CRT/rapamycin were compared in vivo. Low-dose rapamycin inhibited mTOR signaling, decreasing proliferation (43%) and migration (62%) while it enhanced CRT-induced cytotoxicity (3.3 fold) in clonogenic assays. Furthermore, rapamycin re-sensitized CRT-resistant, metastatic tumors to treatment in vivo, improving long-term cures (0–30% improved to 78–100%, depending on the recurrent/metastatic cell line) and limiting lymph node metastasis (32%) and lung metastatic burden (30 fold). Studies using immune compromised mice suggested rapamycin's effect on metastasis is independent of the adaptive immune response. These data suggest a role of mTOR activation in HPV+ HNSCC recurrent/metastatic disease and that adjuvant mTOR inhibition may enhance treatment of resistant, metastatic cell populations at the primary site and limit distant metastasis.
Highlights
600,000 cases of head and neck squamous cell cancer (HNSCC) are diagnosed annually worldwide, accounting for approximately five percent of all cancers reported in the United States alone [1, 2]
Protein expression was significantly different across cell lines (Supplementary Figure 1); mTOR signaling was consistently activated in each (Figure 1). mTOR signaling proteins demonstrating activation included P-Rictor, P-Akt s473, P-S6K and P-S6, and P-NDRG1 (N-myc downstream regulated gene 1, a downstream target of mTOR induced by radiation and a key determinant in resistance to alkylating chemotherapeutic agents [15])
Full-length arrays are included in Supplementary Figure 1. These results suggest mTOR signaling may be implicated in this murine model of HPV+ oropharyngeal cancers specifically (OPSCC) and its recurrent/metastatic derivatives
Summary
600,000 cases of head and neck squamous cell cancer (HNSCC) are diagnosed annually worldwide, accounting for approximately five percent of all cancers reported in the United States alone [1, 2]. The natural disease course includes local recurrence and regional lymph node metastasis; spread to cervical lymph nodes is perhaps the most important indicator of poor prognosis, decreasing overall survival by nearly 50% [3]. Though the incidence of head and neck cancer has decreased with smoking prevalence [6], the distinct clinical subtype that is HPV-related is increasing rapidly [7, 8]. HPV+ cancers typically present at more advanced stages (III-IV), with nodal involvement, and in younger patients compared to HPV-negative (HPV- ) cancers, which are more often associated with smoking, drinking, and older patients [9]
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