John R. Saunders

Prognostic significance of human papillomavirus in oropharyngeal squamous cell carcinomas

The human papillomavirus (HPV) has been identified as a causative factor in 20% to 25% of all head and neck squamous cell carcinomas (HNSCC). Ongoing research suggests that the presence of HPV DNA in HNSCC predicts a positive prognosis with respect to disease‐free and overall survival. However, most studies have been limited by the heterogeneity in treatment regimens and/or anatomic subsites of tumor origin. In this study, we correlate clinical outcomes with HPV status for patients with oropharyngeal carcinomas who were uniformly treated with a concurrent chemoradiation treatment protocol.

Gene mutations in saliva as molecular markers for head and neck squamous cell carcinomas.

BACKGROUND Cancer is caused by the accumulation of mutations that activate proto-oncogenes and inactivate tumor suppressor genes. The result is a clonal expansion of genetically identical daughter cells that eventually become clinical malignancies. The specific mutations acquired by the progenitor cell are like a fingerprint carried by each cell of the tumor. These mutations can serve as very specific markers for the presence of tumor cells in a background of normal cells. METHODS Mutations in the p53 gene recovered from head and neck squamous cell carcinomas were sequenced, and these altered DNA sequences were used retrospectively as tumor-specific genetic markers for cancer cells in the patients saliva. Cloned p53 sequences amplified by the polymerase chain reaction from DNA extracted from banked preoperative saliva specimens were screened for the presence of tumor-specific mutations using radiolabeled oligonucleotide probes. RESULTS We identified tumor-specific mutations in preoperative saliva samples of 5 of the 7 patients evaluated (71%). CONCLUSIONS These results suggest a potential for clinical applications of this novel approach to cancer detection using gene mutations as molecular markers for carcinomas.

Administration of HPV DNA vaccine via electroporation elicits the strongest CD8+ T cell immune responses compared to intramuscular injection and intradermal gene gun delivery

DNA vaccines are an attractive approach to eliciting antigen-specific immunity. Intracellular targeting of tumor antigens through its linkage to immunostimulatory molecules such as calreticulin (CRT) can improve antigen processing and presentation through the MHC class I pathway and increase cytotoxic CD8+ T cell production. However, even with these enhancements, the efficacy of such immunotherapeutic strategies is dependent on the identification of an effective route and method of DNA administration. Electroporation and gene gun-mediated particle delivery are leading methods of DNA vaccine delivery that can generate protective and therapeutic levels of immune responses in experimental models. In this study, we perform a head-to-head comparison of three methods of vaccination--conventional intramuscular injection, electroporation-mediated intramuscular delivery, and epidermal gene gun-mediated particle delivery--in the ability to generate antigen-specific cytotoxic CD8+ T cell responses as well as anti-tumor immune responses against an HPV-16 E7 expressing tumor cell line using the pNGVL4a-CRT/E7(detox) DNA vaccine. Vaccination via electroporation generated the highest number of E7-specific cytotoxic CD8+ T cells, which correlated to improved outcomes in the treatment of growing tumors. In addition, we demonstrate that electroporation results in significantly higher levels of circulating protein compared to gene gun or intramuscular vaccination, which likely enhances calreticulins role as a local tumor anti-angiogenesis agent. We conclude that electroporation is a promising method for delivery of HPV DNA vaccines and should be considered for DNA vaccine delivery in human clinical trials.

Considering the spinal accessory nerve in head and neck surgery.

Loss of trapezius muscle function represents the single most important source of long-term morbidity from a radical neck dissection. Its preservation has been one of the central features of the conservative or modified neck dissection. We recently undertook an evaluation of 100 consecutive patients who had undergone composite resection for head and neck cancer and examined them with particular emphasis on the function of the trapezius muscle. The mean interval from the time of radical neck dissection to the time of this evaluation was 6.2 years. The operations included radical neck dissection with sacrifice of the spinal accessory nerve, radical neck dissection with preservation of the spinal accessory nerve, and radical neck dissection with interpositioned cable graft reconstruction. The survey showed that 67 percent of the patients who underwent radical neck dissection with sacrifice of the spinal accessory nerve, although they showed profound atrophy of the trapezius muscle, had few symptoms related to this deficit. Similarly, 47 percent of patients who underwent radical neck dissection with preservation of the spinal accessory nerve showed some signs of muscle atrophy, and 20 percent showed little or no function of the muscle. Interpositioned nerve grafts appeared to function well in 66 percent of the patients. The survey showed that a surprising number of patients treated with a standard radical neck dissection and sacrifice of the spinal accessory nerve had few postoperative symptoms related to the loss of trapezius muscle function. Also unexpected was the number of patients with signs of muscle dysfunction despite nerve preservation.

Tadalafil augments tumor specific immunity in patients with head and neck squamous cell carcinoma

Purpose: To determine if phosphodiesterase 5 (PDE5) inhibitors can augment immune function in patients with head and neck cancer through inhibition of myeloid-derived suppressor cells (MDSC). Experimental Design: We performed a randomized, prospective, double blinded, placebo controlled, phase II clinical trial to determine the in vivo effects of systemic PDE5 inhibition on immune function in patients with head and neck squamous cell carcinoma (HNSCC). Results: Tadalafil augmented immune response, increasing ex vivo T-cell expansion to a mean 2.4-fold increase compared with 1.1-fold in control patients (P = 0.01), reducing peripheral MDSC numbers to mean 0.81-fold change compared with a 1.26-fold change in control patients (P = 0.001), and increasing general immunity as measured by delayed type hypersensitivity response (P = 0.002). Tumor-specific immunity in response to HNSCC tumor lysate was augmented in tadalafil-treated patients (P = 0.04). Conclusions: These findings demonstrate that tadalafil augments general and tumor-specific immunity in patients with HNSCC and has therapeutic potential in HNSCC. Evasion of immune surveillance and suppression of systemic and tumor-specific immunity is a significant feature of head and neck cancer development. This study demonstrates that a PDE5 inhibitor, tadalafil, can reverse tumor-specific immune suppression in patients with head and neck cancer, with potential for therapeutic application. Clin Cancer Res; 21(1); 30–38. ©2015 AACR.

Low-dose cyclophosphamide administered as daily or single dose enhances the antitumor effects of a therapeutic HPV vaccine.

Although therapeutic HPV vaccines are able to elicit systemic HPV-specific immunity, clinical responses have not always correlated with levels of vaccine-induced CD8+ T cells in human clinical trials. This observed discrepancy may be attributable to an immunosuppressive tumor microenvironment in which the CD8+ T cells are recruited. Regulatory T cells (Tregs) are cells that can dampen cytotoxic CD8+ T-cell function. Cyclophosphamide (CTX) is a systemic chemotherapeutic agent, which can eradicate immune cells, including inhibitory Tregs. The optimal dose and schedule of CTX administration in combination with immunotherapy to eliminate the Treg population without adversely affecting vaccine-induced T-cell responses is unknown. Therefore, we investigated various dosing and administration schedules of CTX in combination with a therapeutic HPV vaccine in a preclinical tumor model. HPV tumor-bearing mice received either a single preconditioning dose or a daily dose of CTX in combination with the pNGVL4a-CRT/E7(detox) DNA vaccine. Both single and daily dosing of CTX in combination with vaccine had a synergistic antitumor effect as compared to monotherapy alone. The potent antitumor responses were attributed to the reduction in Treg frequency and increased infiltration of HPV16 E7-specific CD8+ T cells, which led to higher ratios of CD8+/Treg and CD8+/CD11b+Gr-1+ myeloid-derived suppressor cells (MDSCs). There was an observed trend toward decreased vaccine-induced CD8+ T-cell frequency with daily dosing of CTX. We recommend a single, preconditioning dose of CTX prior to vaccination due to its efficacy, ease of administration, and reduced cumulative adverse effect on vaccine-induced T cells.

Surgical salvage improves overall survival for patients with HPV‐positive and HPV‐negative recurrent locoregional and distant metastatic oropharyngeal cancer

Human papillomavirus (HPV) tumor status and surgical salvage are associated with improved prognosis for patients with recurrent oropharyngeal squamous cell carcinoma (OPSCC). Current data regarding types of surgery and the impact of surgery for patients with distant metastatic disease are limited.

Characterization of the methylation patterns in human papillomavirus type 16 viral DNA in head and neck cancers.

Human papillomavirus (HPV) type 16 can integrate into the host genome, thereby rendering the viral coding genes susceptible to epigenetic modification. Using bisulfite genomic sequencing, we determined the methylation status of all 110 CpG sites within the viral epigenome in advanced stage III/IV HPV-16–associated head and neck cancers. We found that the viral genome was hypomethylated in the majority of head and neck cancers, in particular within the viral regulatory region, long control region (LCR), which controls transcription of the E6 and E7 oncogenes. The hypomethylation status of LCR correlated with detectable levels of E6 and E7 expression, which suggests that the tumors may still be dependent on these viral oncogenes to maintain the malignant phenotype. In addition to the methylation status of LCR, we report other potential factors which may influence intratumoral E6 and E7 expression including viral copy number and integration site. We were able to detect the viral epigenetic alterations in sampled body fluids, such as serum and saliva, which correlated with the changes observed in the primary tumors. Because viral epigenetic changes occur in the setting of viral integration into the human genome, the detection of methylated HPV genes in the serum and/or saliva may have diagnostic potential for early detection strategies of viral integration and assessment of risk for cancer development in high-risk individuals. Our findings also support continued targeting of the E6 and/or E7 antigens through various vaccine strategies against HPV-associated cancers. Cancer Prev Res; 4(2); 207–17. ©2011 AACR.

Definitive mandibular replacement using reconstruction plates.

Mandibular defects following radical cancer surgery continue to provide challenges to head and neck surgeons. Twenty-seven patients with advanced oral cancer underwent primary mandibular replacement with metal reconstruction plates without the use of bone. Twenty-one patients (78%) had successful reconstruction with primary soft tissue healing. Six patients required removal of the plate in the postoperative period. Two of these patients had their reconstruction plates replaced as a secondary procedure following soft tissue healing. Thus, 23 of 27 patients (85%) had final mandibular reconstruction and were followed for an average of 19 months. Functional and cosmetic results were satisfactory. For patients with advanced disease, this technique compares favorably with microvascular transfer in terms of operating time and donor defect. Despite problems with plate exposure, the initial and overall success rates of 78% and 85%, respectively, make the use of these plates a reasonable choice for immediate reconstructive needs in patients with difficult tumors.

Selective versus comprehensive neck dissection after chemoradiation for advanced oropharyngeal squamous cell carcinoma

OBJECTIVE: To determine whether a comprehensive neck dissection (CND) or a selective neck dissection (SND) is indicated as planned post–primary chemoradiation treatment (CRT) for patients with advanced oropharyngeal squamous cell carcinoma (OPSCC). STUDY DESIGN: Case series with chart review. SETTING: A community teaching hospital. SUBJECTS: Patients with advanced OPSCC who received a uniform CRT protocol at Greater Baltimore Medical Center (GBMC). METHODS: Medical records of patients treated with primary CRT for locoregionally advanced OPSCC at GBMC between 2001 and 2007 were reviewed. All patients received 7000 to 7500, 6000, and 5000 cGy to primary disease sites, involved cervical lymphatics, and uninvolved cervical and supraclavicular lymphatics, respectively, with concomitant cisplatin (12 mg/m2/1 h) and 5-fluorouracil (600 mg/m2/20 h) given on days one through five and 29 through 33. RESULTS: Seventy-six patients received CRT, and 41 met the criteria for neck dissection. Forty-eight neck dissections were performed (34 unilateral and 7 bilateral), of which 23 (48%) were CNDs and 25 (52%) were SNDs. Residual carcinoma was found in six (26%) of the CND and five (20%) of the SND heminecks. The CND group had six (26%) complications, whereas the SND group had two (8%). CONCLUSION: The high rate of residual disease demonstrated in this study supports the need for post-CRT neck dissection. Although complication rates were not significantly different between the two groups, the trend in this study indicates that SND results in less morbidity. The presumed reduced morbidity and equivalent regional control rate suggest that SND is an appropriate surgical option for OPSCC patients after primary CRT.