Effect of the intratumoral microbiota on spatial and cellular heterogeneity in cancer.

The tumour-associated microbiota is an intrinsic component of the tumour microenvironment across human cancer types1,2. Intratumoral host-microbiota studies have so far largely relied on bulk tissue analysis1-3, which obscures the spatial distribution and localized effect of the microbiota within tumours. Here, by applying in situ spatial-profiling technologies4 and single-cell RNA sequencing5 to oral squamous cell carcinoma and colorectal cancer, we reveal spatial, cellular and molecular host-microbe interactions. We adapted 10x Visium spatial transcriptomics to determine the identity and in situ location of intratumoral microbial communities within patient tissues. Using GeoMx digital spatial profiling6, we show that bacterial communities populate microniches that are less vascularized, highly immuno­suppressive and associated with malignant cells with lower levels of Ki-67 as compared to bacteria-negative tumour regions. We developed a single-cell RNA-sequencing method that we name INVADEseq (invasion-adhesion-directed expression sequencing) and, by applying this to patient tumours, identify cell-associated bacteria and the host cells with which they interact, as well as uncovering alterations in transcriptional pathways that are involved in inflammation, metastasis, cell dormancy and DNA repair. Through functional studies, we show that cancer cells that are infected with bacteria invade their surrounding environment as single cells and recruit myeloid cells to bacterial regions. Collectively, our data reveal that the distribution of the microbiota within a tumour is not random; instead, it is highly organized in microniches with immune and epithelial cell functions that promote cancer progression.

Oncologic outcomes of human papillomavirus-associated oropharynx carcinoma treated with surgery alone: A 12-institution study of 344 patients.

BACKGROUND: The oncologic outcomes of surgery alone for patients with American Joint Committee on Cancer 7th edition (AJCC 7th) pN2a and pN2b human papillomavirus-associated oropharynx squamous cell carcinoma (HPV+OPSCC) are not clear. METHODS: The authors performed a 12-institution retrospective study of 344 consecutive patients with HPV+OPSCC (AJCC 7th pT0-3 N3 M0) treated with surgery alone with 6 months or more of follow-up using univariate and multivariate analyses. RESULTS: The 2-year outcomes for the entire cohort were 91% (182 of 200) disease-free survival (DFS), 100% (200 of 200) disease-specific survival (DSS), and 98% (200 of 204) overall survival (OS). The 18 recurrences within 2 years were 88.9% (16 of 18) local and/or regional recurrences and 11.1% (2 of 18) distant metastases. Recurrences were not significantly associated with smoking, pT stage, or pN stage. The 16 patients with locoregional recurrences within 2 years all underwent successful salvage treatments (median follow-up after salvage: 13.1 months), 43.8% (7 of 16) of whom underwent salvage surgery alone for a 2-year overall salvage radiation need of 4.5% (9 of 200). The 2-year outcomes for the 59 evaluable patients among the 109 AJCC 7th pT0-2 N2a-N2b patients with 1 to 3 pathologic lymph nodes (LNs) were as follows: local recurrence, 3.4% (2 of 59); regional recurrence, 8.4% (5 of 59); distant metastases, 0%; DFS, 88.1% (52 of 59); DSS, 100% (59 of 59); OS, 96.7% (59 of 61); and salvage radiation, 5.1% (3 of 59). CONCLUSIONS: With careful selection, surgery alone for AJCC 7th pT0-T2N0-N2b HPV+OPSCC with zero to 3 pathologic LNs without perineural invasion, extranodal extension, or positive margins results in high DFS, DSS, OS, and salvage treatment success. Because of the short-term follow-up, these data support further investigation of treatment de-escalation in this population.

Surgical Options for Locally Advanced Oropharyngeal Cancer.

OPINION STATEMENT: Oropharyngeal squamous cell carcinoma (OPSCC) incidence rates have been steadily increasing over the past several decades, and this has been largely attributed to human papillomavirus (HPV)-related OPSCC. The rise of HPV-related OPSCC and the observed distinct survival advantage it offers compared to HPV-unrelated OPSCC have resulted in the development of a new staging system specifically for OPSCC in the eighth edition of the AJCC Staging Manual for head and neck cancer. The observations on HPV-related OPSCC and its prognostic implications have coincided with increasing utilization of transoral surgical approaches to oropharyngeal tumors, such as transoral laser microsurgery (TLM) and transoral robotic surgery (TORS). These approaches were once thought to only be applicable to patients with low T-stage OPSCC tumors; however, they are being increasingly utilized in locally advanced OPSCC cases as several studies have shown that both of these transoral approaches are oncologically sound alternatives to concurrent chemoradiation therapy (CCRT), which was previously the standard-of-choice treatment in patients with locally advanced disease. Moreover, these transoral approaches have displayed better long-term swallowing outcomes compared to CCRT, as severe dysphagia is often the most bothersome functional impairment to OPSCC survivors who have undergone CCRT. While open surgical approaches were previously not utilized in the locally advanced OPSCC setting due to the risk of severe surgical complications compared to the potential benefits of organ preservation with CCRT and comparable survival rates after either treatment regimen, these approaches are still reasonable options for select patients in the salvage surgery setting, as they allow for maximum exposure to the deep oropharyngeal anatomy. Data from multiple clinical trials evaluating the potential for TORS to de-escalate radiation dose or CCRT regimen in certain settings will inform clinical decision-making for OPSCC patients for the next decade and allow for more personalized treatments tailored to an individual patient's disease burden.

Methods to reduce postoperative surgical site infections after head and neck oncology surgery.

Head and neck cancer surgery is often a complex multi-step procedure that includes major resections, vascularised tissue reconstruction, and extensive neck dissection. The upper aerodigestive tract mucosal lining is often disrupted during surgery, which requires the management of a clean-contaminated field and the need to reconstruct the mucosal lining. With bacterial contamination, surgical site infections (SSI) are a serious complication that can result in delayed wound healing, wound breakdown, fistula formation, and compromised tissue reconstruction. Methods to reduce SSI in patients with head and neck cancer have been intensely researched, yielding evolving and varied practice patterns. In this Review, we outline the data supporting perioperative antibiotic prophylaxis for clean-contaminated surgeries, which suggest that clindamycin is an inadequate prophylactic antibiotic therapy in the reduction of SSI, and that prolonged antibiotic courses have no established benefit. For salvage laryngectomy after radiotherapy with or without chemotherapy, reconstruction with vascularised tissue reduces the frequency and severity of pharyngocutaneous fistula formation. These evidence-based recommendations have been shown to reduce the chance of SSI after head and neck surgery.

The association between T-stage and clinical nodal metastasis In HPV-positive oropharyngeal cancer.

PURPOSE: To evaluate the association between T-stage and primary tumor subsite with clinical nodal metastasis in HPV-positive oropharyngeal squamous cell carcinomas. STUDY DESIGN: Retrospective medical record review. MATERIALS AND METHODS: A retrospective analysis of all previously untreated patients with HPV-positive oropharyngeal squamous cell carcinoma evaluated by the senior author in a tertiary-care center over a 5-year period was performed. Medical records were evaluated for HPV status, clinical T-stage, clinical nodal stage, and anatomic location of primary oropharyngeal subsite. RESULTS: 83% (59/71) of patients presented with clinical nodal disease. T-stage was not associated with the absence of regional disease (cN0) in tonsillar complex or base of tongue cancers. However, early T1/T2 tonsillar complex cancers were less likely to present with cN2c disease compared to T3/T4 lesions (0% vs. 18-33%, p=0.03). Further, base of tongue cancers were more likely to present with cN2c disease compared to tonsillar complex tumors (35% vs. 7%, p=0.01). CONCLUSION: In HPV-positive tonsillar complex cancers, T-stage does not appear to predict the presence of clinical nodal metastasis (N0 vs. N1-N3), but may predict the extent of metastasis when present. This association does not appear to affect base of tongue cancers. Further, base of tongue cancers have a greater incidence of cN2c disease compared to tonsillar complex cancers.

Early, precise, and safe clinical evaluation of the pharmacodynamic effects of novel agents in the intact human tumor microenvironment.

Introduction: Drug development is systemically inefficient. Research and development costs for novel therapeutics average hundreds of millions to billions of dollars, with the overall likelihood of approval estimated to be as low as 6.7% for oncology drugs. Over half of these failures are due to a lack of drug efficacy. This pervasive and repeated low rate of success exemplifies how preclinical models fail to adequately replicate the complexity and heterogeneity of human cancer. Therefore, new methods of evaluation, early in the development trajectory, are essential both to rule-in and rule-out novel agents with more rigor and speed, but also to spare clinical trial patients from the potentially toxic sequelae (high risk) of testing investigational agents that have a low likelihood of producing a response (low benefit). Methods: The clinical in vivo oncology (CIVO®) platform was designed to change this drug development paradigm. CIVO precisely delivers microdose quantities of up to 8 drugs or combinations directly into patient tumors 4-96 h prior to planned surgical resection. Resected tissue is then analyzed for responses at each site of intratumoral drug exposure. Results: To date, CIVO has been used safely in 6 clinical trials, including 68 subjects, with 5 investigational and 17 approved agents. Resected tissues were analyzed initially using immunohistochemistry and in situ hybridization assays (115 biomarkers). As technology advanced, the platform was paired with spatial biology analysis platforms, to successfully track anti-neoplastic and immune-modulating activity of the injected agents in the intact tumor microenvironment. Discussion: Herein we provide a report of the use of CIVO technology in patients, a depiction of the robust analysis methods enabled by this platform, and a description of the operational and regulatory mechanisms used to deploy this approach in synergistic partnership with pharmaceutical partners. We further detail how use of the CIVO platform is a clinically safe and scientifically precise alternative or complement to preclinical efficacy modeling, with outputs that inform, streamline, and de-risk drug development.

Prospective head and neck cancer research: a four-decade bibliometric perspective.

BACKGROUND: It is unknown whether changes in study sponsorship have affected the proportion of prospective research on surgery, radiotherapy, and pharmacotherapy for head and neck squamous cell carcinoma (HNSCC) being published over time. PATIENTS AND METHODS: We examined prospective studies from PubMed, Ovid MEDLINE, EMBASE, and the Cochrane Central Register of Controlled Trials from 1980, 1985, 1990, 1995, 2000, 2005, and 2010. Chi-squared tests were used to identify significant associations between sponsorship and authorship, treatments within study protocols, and presentation of results, whereas time-based trends were analyzed using the Cochran-Armitage test. RESULTS: Among 309 articles, industry (70, 22.7%) and the U.S. government (65, 21%) were the most common sponsors. There was a significant increase in the proportion of industry-sponsored research (p for trend = .013) and a decline in U.S. government-sponsored research (p for trend = .001) over time. The inclusion of surgery in treatment protocols declined over the past four decades (p for trend = .003). Protocols incorporating pharmacotherapy were more likely to have industry support than those without pharmacotherapy (p = .001), whereas protocols with radiotherapy (p = .003) or surgery (p = .002) were less likely to have industry support. CONCLUSION: Industry is the predominant sponsor of prospective HNSCC research, with an emphasis on pharmacotherapy.

Clarification of the margin status by the multidisciplinary tumor board following transoral robotic surgery for p16 positive oropharyngeal squamous cell carcinoma.

Objectives: Margin status interpretation following transoral robotic surgery (TORS) for oropharyngeal squamous cell carcinoma (OPSCC) is challenging. This study aims to assess the discrepancy between status of margins as reported by the pathologist versus as determined by multi-disciplinary team review (MDTB). Methods: A retrospective study of 57 patients with OPSCC who underwent TORS from January 2010 to December 2016 was conducted. Our primary outcome measure was the discrepancy between the surgical specimen margins as described in the pathology report versus final margin status that was determined after the multi-disciplinary team discussion. Fisher's exact test was used. Results: Based on the pathologist-report, 29 subjects (51%) had positive margins, compared to 2 (4%) after multi-disciplinary team discussion. Receipt of chemotherapy correlated with final margin status as determined by MDTB, not with initial main specimen margins (p = .02 and p = .08, respectively). With a median follow up of 28.4 months, two subjects (4%) had loco-regional recurrence. Conclusion: Following TORS, there was a significant discrepancy between status of margins as reported by the pathologist versus as determined by MDTB review. Chemotherapy was avoided in 93.1% of cases that were originally reported as positive margins by the pathologist with an acceptably low recurrence rate. Level of evidence: 4.

Radiation therapy for low- and high-risk perineural invasion in head and neck cutaneous squamous cell carcinoma: Clinical outcomes and patterns of failure.

BACKGROUND: Perineural invasion (PNI) in head and neck squamous cell carcinoma (HNSCC) portends poor prognosis. Extent of treatment of nerve pathways with varying degrees of PNI and patterns of failure following elective neural radiotherapy (RT) remain unclear. METHODS: Retrospective review of HNSCC patients with high-risk (clinical/gross, large-nerve, extensive) or low-risk (microscopic/focal) PNI who underwent curative-intent treatment from 2010 to 2021. RESULTS: Forty-four patients (mean follow-up 22 months; 59% high-risk, 41% low-risk PNI) were included. Recurrence following definitive treatment occurred in 31% high-risk and 17% low-risk PNI patients. Among high-risk patients, 69% underwent surgery with post-operative RT and 46% underwent elective neural RT. Local control (83% low-risk vs. 75% high-risk), disease-free, and overall survival did not differ between groups. CONCLUSIONS: High local control rates were achieved in high-risk PNI patients treated with adjuvant or primary RT, including treatment of both involved and uninvolved, communicating cranial nerves, with few failures in electively treated regions.

Neutrophil to lymphocyte ratio and peripheral blood biomarkers correlate with survival outcomes but not response among head and neck and salivary cancer treated with pembrolizumab and vorinostat.

BACKGROUND: Associations between peripheral blood biomarkers and oncologic outcomes were explored in recurrent/metastatic (R/M) head and neck squamous cell carcinoma (HN) and salivary gland cancer (SGC) treated with pembrolizumab and vorinostat on a phase II trial (NCT02538510). EXPERIMENTAL DESIGN: Twenty-five HN and 25 SGCs were treated with pembrolizumab and vorinostat. Baseline peripheral blood was available in 21 HN and 20 SGCs and evaluated for associations with grade ≥3 adverse events (G ≥ 3AE) by CTCAEv4, objective response rate (ORR), overall survival (OS), and progression-free survival (PFS). RESULTS: Higher pretreatment neutrophil-to-lymphocyte ratio (NLR) and neutrophils, as well as lower pretreatment lymphocytes and T helper cells correlated with worse OS and PFS. Higher NLR further predicted increased rates of G ≥ 3AEs. No correlations with ORR were observed. CONCLUSIONS: In a prospectively evaluated cohort of HN and SGCs treated with pembrolizumab and vorinostat, we observed novel associations between peripheral blood biomarkers and oncologic outcomes and toxicities.