Augmentation of tumor expression of HLA-DR, CXCL9, and CXCL10 may improve olfactory neuroblastoma immunotherapeutic responses.

BACKGROUND: Olfactory neuroblastoma is a rare malignancy of the anterior skull base typically treated with surgery and adjuvant radiation. Although outcomes are fair for low-grade disease, patients with high-grade, recurrent, or metastatic disease oftentimes respond poorly to standard treatment methods. We hypothesized that an in-depth evaluation of the olfactory neuroblastoma tumor immune microenvironment would identify mechanisms of immune evasion in high-grade olfactory neuroblastoma as well as rational targetable mechanisms for future translational immunotherapeutic approaches. METHODS: Multispectral immunofluorescence and RNAScope evaluation of the tumor immune microenvironment was performed on forty-seven clinically annotated olfactory neuroblastoma samples. A retrospective chart review was performed and clinical correlations assessed. RESULTS: A significant T cell infiltration was noted in olfactory neuroblastoma samples with a stromal predilection, presence of myeloid-derived suppressor cells, and sparse natural killer cells. A striking decrease was observed in MHC-I expression in high-grade olfactory neuroblastoma compared to low-grade disease, representing a mechanism of immune evasion in high-grade disease. Mechanistically, the immune effector stromal predilection appears driven by low tumor cell MHC class II (HLA-DR), CXCL9, and CXCL10 expression as those tumors with increased tumor cell expression of each of these mediators correlated with significant increases in T cell infiltration. CONCLUSION: These data suggest that immunotherapeutic strategies that augment tumor cell expression of MHC class II, CXCL9, and CXCL10 may improve parenchymal trafficking of immune effector cells in olfactory neuroblastoma and augment immunotherapeutic responses.

Determining if T cell antigens are naturally processed and presented on HLA class I molecules.

BACKGROUND: Determining T cell responses to naturally processed and presented antigens is a critical immune correlate to determine efficacy of an investigational immunotherapeutic in clinical trials. In most cases, minimal epitopes and HLA restriction elements are unknown. RESULTS: Here, we detail the experimental use of ex vivo expanded autologous B cells as antigen presenting cells to overcome the limitation of unknown HLA restriction, and the use of electroporated full length mRNA encoding full length parental proteins to ensure that any observed T cell responses are specific for antigens that are naturally processed and presented. CONCLUSIONS: This technique can serve as useful experimental approach to determine the induction or enhancement of specific responses to naturally processed and presented antigens on HLA class I molecules in peripheral blood or tumor infiltrating T cells.

Discordance in patient and physician global assessment in relapsing polychondritis.

OBJECTIVES: Relapsing polychondritis (RP) is a rare, heterogeneous, systemic inflammatory disease that targets cartilage. Patient-reported outcome measures may differ from physician assessment. This study compared patient global assessment (PtGA) and physician global assessment (PhGA) scores in a prospective cohort of patients with RP. METHODS: Adult patients with RP underwent a standardized comprehensive evaluation at ∼6 month intervals. At each visit, three physicians scored PhGA by consensus. The patient independently completed four patient-reported outcomes: PtGA, 36-item Short Form Health Survey (SF-36), Brief Illness Perception Questionnaire (BIPQ) and Multidimensional Fatigue Inventory (MFI). Patient-physician discordance was defined as a difference between PtGA and PhGA of ≥3 on a 0-10 scale. RESULTS: A total of 76 patients were evaluated over 154 visits. The median PhGA was 3 [interquartile range (IQR) 2-3] and the median PtGA was 5 (IQR 4-7). PtGA and PhGA were concordant in 66 visits (42.9%) and patients scored disease severity ≥3 points higher than physicians scored disease activity (positive discordance) in 84 visits (54.5%). Compared with visits with concordance, visits with positive discordance were associated with significantly worse scores on the MFI, BIPQ, SF-36 physical component score and SF-36 mental component score. CONCLUSION: Patients with RP typically self-report high PtGA that does not align with PhGA. Discordance is likely driven by the high physical and psychological burden of illness experienced by patients. Multifaceted treatment approaches that address the burden of disease in RP from the patient perspective are needed.

Sicca Syndrome Associated with Immune Checkpoint Inhibitor Therapy.

BACKGROUND: The objective of this study was to characterize the clinicopathologic features of sicca syndrome associated with immune checkpoint inhibitor (ICI) therapy. SUBJECTS, MATERIALS, AND METHODS: Consecutive patients with new or worsening xerostomia in the setting of ICI treatment for benign or malignant neoplastic disease were evaluated, including labial salivary gland biopsy (LSGB). RESULTS: Twenty patients (14 male; median age 57 years) had metastatic melanoma (n = 10), metastatic carcinoma (n = 6), or recurrent respiratory papillomatosis (n = 4) and were being treated with avelumab (n = 8), nivolumab (n = 5), pembrolizumab (n = 4), nivolumab/ipilimumab (n = 2), and M7824, a biologic targeting programmed cell death ligand 1 (PD-L1) and transforming growth factor ß (n = 1). Four had pre-existing autoimmune disease. Nineteen had very low whole unstimulated saliva flow; six had new dry eye symptoms. The median interval between ICI initiation and dry mouth onset was 70 days. Rheumatoid factor and anti-Sjögren's Syndrome-related Antigen A (Anti-SSA) were both positive in two subjects. LSGB showed mild-to-severe sialadenitis with diffuse lymphocytic infiltration and architectural distortion. There were lymphocytic aggregates in eight patients, composed mainly of CD3+ T cells with a slight predominance of CD4+ over CD8+ T cells. ICI targets (e.g., programmed cell death 1 and PD-L1) were variably positive. In direct response to the advent of the sicca immune-related adverse event, the ICI was held in 12 patients and corticosteroids were initiated in 10. Subjective improvement in symptoms was achieved in the majority; however, salivary secretion remained very low. CONCLUSION: ICI therapy is associated with an autoimmune-induced sicca syndrome distinct from Sjögren's syndrome, often abrupt in onset, usually developing within the first 3 months of treatment, and associated with sialadenitis and glandular injury. Improvement can be achieved with a graded approach depending on severity, including withholding the ICI and initiating corticosteroids. However, profound salivary flow deficits may be long term. IMPLICATIONS FOR PRACTICE: Sicca syndrome has been reported as an immune-related adverse event (irAE) of immune checkpoint inhibitor therapy (ICI) for neoplastic diseases. Severe dry mouth (interfering with eating or sleeping) developed abruptly, typically within 90 days, after initiation of ICI therapy. Salivary gland biopsies demonstrated mild-to-severe sialadenitis distinct from Sjögren's syndrome, with diffuse T-cell lymphocytic infiltration and acinar injury. Recognition of the cardinal features of ICI-induced sicca will spur appropriate clinical evaluation and management, including withholding of the ICI and corticosteroid, initiation. This characterization should help oncologists, rheumatologists, and oral medicine specialists better identify patients that develop ICI-induced sicca to initiate appropriate clinical evaluation and therapy to reduce the likelihood of permanent salivary gland dysfunction.

Murray secretion scale and fiberoptic endoscopic evaluation of swallowing in predicting aspiration in dysphagic patients.

The objective of this retrospective review is to evaluate the ability of the Murray secretion scale to predict aspiration as determined by fiberoptic endoscopic evaluation of swallowing. Patients with dysphagia undergoing a fiberoptic endoscopic evaluation of swallowing study between January 2013 and November 2015 from a single, tertiary care institution were retrospectively reviewed. The Murray secretion scale and penetration aspiration scale on fiberoptic endoscopic evaluation of swallowing examination were determined. Spearman's correlation analysis, sensitivity, specificity, predictive values, and relative risk evaluating the relationship between the Murray secretion scale and aspiration on fiberoptic endoscopic evaluation of swallowing were calculated. Subgroups of head and neck cancer patients, penetration group, and aspiration group were also analyzed. The mean age of the cases (N = 212) was 62.4 years. Eighty percent were male. There was a strong correlation between Murray secretion scale grade and penetration aspiration scale score (r = 0.785, p < 0.001). The sensitivity and specificity of a Murray secretion scale grade 2 or higher in predicting aspiration were 74 and 90%, respectively. Individuals with a Murray secretion scale grade of 2 or higher were 13.6 times more likely to aspirate than patients with a lower Murray secretion scale grade. All subgroups showed similar trend. Determination of a Murray secretion scale grade, determined by flexible nasopharyngoscopy, may predict patients at high risk for aspiration. In clinical scenarios where more complete assessments of aspiration risk are immediately impossible or impractical, the Murray secretion scale grade may add valuable information to assist in clinical decision-making in patients with dysphagia.

Risk stratification in endoscopic airway surgery: is inpatient observation necessary?

PURPOSE: To compare post-operative complication rates between inpatient and outpatient endoscopic airway surgery in patients with laryngotracheal stenosis. Secondary objectives included characterization of a cohort of patients with this disease. METHODS: Retrospective review of patients with laryngotracheal stenosis in a tertiary care laryngology practice over a 5-year period. RESULTS: Ninety-one patients underwent 223 endoscopic airway surgeries. Of 114 outpatient interventions, 1 patient (0.8%) sought emergent medical care following discharge for respiratory distress. Of 109 procedures resulting in admission, no patients required transfer to a higher level of care, endotracheal intubation or placement of a surgical airway. There was no statistically significant difference in complication rates between patients treated as outpatients or inpatients (p=0.33, chi square). There were no cardiopulmonary events. There were no pneumothoraces despite frequent use of jet ventilation. The most common etiologic category was idiopathic (58%), followed by granulomatosis with polyangiitis (16%) and history of tracheotomy (12%). Most patients with idiopathic disease were female (p<0.001, Fisher's exact test). CONCLUSION: Patients undergoing endoscopic surgery for airway stenosis rarely have post-operative complications, and outpatient surgery appears to be a safe alternative to post-operative admission and observation.

Effectiveness and residual activity of four common insecticides used in the Mississippi Delta to control tarnished plant bugs in cotton.

The tarnished plant bug, (TPB) Lygus lineolaris Palisot de Beauvois (Hemiptera: Miridae) is a key pest of cotton in the midsouth region and some areas of the eastern United States. Its control methods have been solely based on chemical insecticides which has contributed to insecticidal resistance and shortened residual periods for control of this insect pest. This study was conducted over a two-year period and examined the efficacy and residual effect of four commercial insecticides including lambda-cyhalothrin (pyrethroid), acephate (organophosphate), imidacloprid (neonicotinoid), and sulfoxaflor (sulfoxamine). The effectiveness and residual effects of these insecticides were determined by application on cotton field plots on four different dates during each season using three different concentrations (high: highest labeled commercial dose (CD), medium: 1/10 of the CD, low: 1/100 of the CD) on field cotton plots. Four groups of cotton leaves were randomly pulled from each treated plot and control 0-, 2-, 4-, 7-, and 9-days post treatment (DPT) and exposed to a lab colony of TPB adults. One extra leaf sample/ plot/ spray /DPT interval (0-2-4-7-9-11) during 2016 was randomly collected from the high concentration plots and sent to Mississippi State Chemical Laboratory for residual analysis. Mortality of TPB adults was greatest for those placed on leaves sprayed with the organophosphate insecticide with mortalities (%) of 81.7±23.4 and 63.3±28.8 (SE) 1-day after exposure (DAE) on leaves 0-DPT with the high concentration for 2016 and 2017, respectively, reaching 94.5±9.5 and 95.4±7.6 6-DAE each year. Mortality to all insecticides continued until 9 and 4-DPT for high and medium concentrations, respectively. However, organophosphate (39.4±28.6) and pyrethroid (24.4±9.9) exhibited higher mortality than sulfoxamine (10.6±6.6) and the neonicotinoid (4.0±1.5) 7-DAE on 9-DPT leaves with the high concentration. Based on our results using the current assay procedure, TPB adults were significantly more susceptible to contact than systemic insecticides and due to its residual effect, organophosphate could kill over 80% of the TPB population 7-DPT.

Heterogeneous characterization of neutrophilic cells in head and neck cancers.

BACKGROUND: Neutrophilic cells are among the most abundant immune populations within the head and neck tumor microenvironment (TME) and harbor multiple mechanisms of immunosuppression. Despite these important features, neutrophilic cells may be underrepresented in contemporary studies that aim to comprehensively characterize the immune landscape of the TME due to discrepancies in tissue processing and analysis techniques. Here, we review the role of pathologically activated neutrophilic cells within the TME and pitfalls of various approaches used to study their frequency and function in clinical samples. METHODS: The literature was identified by searching PubMed for "immune landscape" and "tumor immune microenvironment" in combination with keywords describing solid tumor malignancies. Key publications that assessed the immune composition of solid tumors derived from human specimens were included. The tumor and blood processing methodologies in each study were reviewed in depth and correlated with the reported abundance of neutrophilic cells. RESULTS: Neutrophilic cells do not survive cryopreservation, and many studies fail to identify and study neutrophilic cell populations due to cryopreservation of clinical samples for practical reasons. Additional single-cell transcriptomic studies filter out neutrophilic cells due to low transcriptional counts. CONCLUSIONS: This report can help readers critically interpret studies aiming to comprehensively study the immune TME that fail to identify and characterize neutrophilic cells.

Complete tumor resection reverses neutrophilia-associated suppression of systemic anti-tumor immunity.

OBJECTIVES: Tumor infiltrating neutrophils suppress T cell function, but whether neutrophils in circulation contribute to systemic immunosuppression is unclear. We aimed to study whether peripheral neutrophils that accumulate with tumor progression contribute to systemic immunosuppression, and if observed suppression of systemic anti-tumor immunity could be reversed with complete surgical tumor removal. MATERIALS AND METHODS: Syngeneic murine oral cancers were established in immunocompetent mice. Proteomic and functional immune assays were used to study plasma cytokine concentration, peripheral immune frequencies, and systemic anti-tumor immunity with and without complete primary tumor resection. RESULTS: Ly6G+ neutrophilic cells, but not other myeloid cell types, accumulated in the periphery of mice with progressing tumors. This accumulation positively associated with plasma G-CSF concentration. Circulating neutrophils were functionally immunosuppressive. Complete surgical tumor removal reversed the observed neutrophilia, with neutrophil frequencies returning to baseline in 21 days. Multiple independent functional assays revealed enhanced systemic anti-tumor immunity in mice following tumor resection compared to tumor-bearing mice, and the observed enhanced systemic immunity could be reproduced with selective neutrophil depletion. CONCLUSIONS: Complete primary tumor resection can reverse neutrophilia that develops during tumor progression and result in enhanced systemic anti-tumor immunity. Primary tumor removal relieves neutrophil-driven systemic immunosuppression and may itself contribute to the clinical benefit observed with neoadjuvant immunotherapy.

Quantification and Functional Studies of Neutrophilic Cells Identifies Distinct Papilloma Phenotypes.

OBJECTIVES: To characterize the distribution of immune cell subsets within laryngeal papillomas and to study the function of potentially immunosuppressive neutrophilic and regulatory T cells (Tregs). METHODS: Fresh clinical papilloma specimens were collected at the time of surgery and studied with multiparameter flow cytometry. Papilloma infiltrating neutrophilic cells and Tregs were sorted and studied functionally with ex vivo T cell suppression assays. RESULTS: Flow cytometric analysis of fresh laryngeal papillomas samples from 18 adult patients with recurrent respiratory papillomatosis revealed patterns in immune constituency between patients. Clearly divergent phenotypes based primarily on the degree of neutrophilic and T cell infiltration were identified. Relative neutrophilic cell enrichment and T cell depletion were observed in 50% of samples and neutrophilic cell depletion and T cell enrichment were observed in the others. Greater papilloma neutrophilic cell enrichment was positively associated with the number of clinically indicated interventions required in the 12 months prior to sample collection, linking papilloma neutrophil inflammation to disease severity. Functional assays revealed the ability of both papilloma infiltrating neutrophilic and Tregs to suppress T cell function at roughly equal magnitudes, but substantially increased infiltration of neutrophilic cells compared to Tregs across samples. CONCLUSION: Neutrophilic cells are an important contributor to immunosuppression within the respiratory papilloma microenvironment. Given these data and the association between greater neutrophilic cell infiltration and lack of clinical response to therapeutic vaccination, additional study of strategies aimed at limiting neutrophilic cell infiltration or function within papillomas is warranted. LEVEL OF EVIDENCE: 4 Laryngoscope, 134:3238-3244, 2024.

Risk of recurrence after neoadjuvant chemotherapy and transoral robotic surgery in patients with oropharynx cancer that avoid adjuvant radiation.

BACKGROUND: De-escalation strategies for newly-diagnosed p16-positive oropharyngeal squamous cell carcinoma (p16+ OPSCC), aim to reduce treatment-related morbidity without compromising disease control. One strategy is neoadjuvant cisplatin and docetaxel chemotherapy (NAC + S) before transoral robotic surgery, with pathology-based risk-adapted adjuvant treatment. METHODS: We examined the recurrence-free survival (RFS) for patients who received NAC + S. RESULTS: Comparing outcomes in 103 patients between 2008 and 2023, 92% avoided adjuvant treatment and showed significantly higher 2-year recurrence-free survival (RFS) compared to those with adjuvant treatment (95.9% vs. 43.8%, p = 0.0049) CONCLUSION: Our findings suggest that pathology-based risk-adapted omission of adjuvant treatment following NAC + S does not appear to elevate recurrence risk and that NAC may identify patients with favorable tumor biology, yielding a 2-year RFS probability exceeding 95% without adjuvant treatment. Further, the study identifies a patient subset experiencing disease recurrence despite triple modality therapy. Despite limitations, including a retrospective design and modest sample size, the data advocate for controlled NAC + S studies.

Chordoma cancer stem cell subpopulation characterization may guide targeted immunotherapy approaches to reduce disease recurrence.

Introduction: Cancer stem cells (CSCs), a group of tumor-initiating and tumor-maintaining cells, may be major players in the treatment resistance and recurrence distinctive of chordoma. Characterizing CSCs is crucial to better targeting this subpopulation. Methods: Using flow cytometry, six chordoma cell lines were evaluated for CSC composition. In vitro, cell lines were stained for B7H6, HER2, MICA-B, ULBP1, EGFR, and PD-L1 surface markers. Eighteen resected chordomas were stained using a multispectral immunofluorescence (mIF) antibody panel to identify CSCs in vivo. HALO software was used for quantitative CSC density and spatial analysis. Results: In vitro, chordoma CSCs express more B7H6, MICA-B, and ULBP1, assessed by percent positivity and mean fluorescence intensity (MFI), as compared to non-CSCs in all cell lines. PD- L1 percent positivity is increased by >20% in CSCs compared to non-CSCs in all cell lines except CH22. In vivo, CSCs comprise 1.39% of chordoma cells and most are PD-L1+ (75.18%). A spatial analysis suggests that chordoma CSCs cluster at an average distance of 71.51 mm (SD 73.40 mm) from stroma. Discussion: To our knowledge, this study is the first to identify individual chordoma CSCs and describe their surface phenotypes using in vitro and in vivo methods. PD-L1 is overexpressed on CSCs in chordoma human cell lines and operative tumor samples. Similarly, potential immunotherapeutic targets on CSCs, including B7H6, MICA-B, ULBP1, EGFR, and HER2 are overexpressed across cell lines. Targeting these markers may have a preferential role in combating CSCs, an aggressive subpopulation likely consequential to chordoma's high recurrence rate.

A deep-learning framework to predict cancer treatment response from histopathology images through imputed transcriptomics.

Advances in artificial intelligence have paved the way for leveraging hematoxylin and eosin-stained tumor slides for precision oncology. We present ENLIGHT-DeepPT, an indirect two-step approach consisting of (1) DeepPT, a deep-learning framework that predicts genome-wide tumor mRNA expression from slides, and (2) ENLIGHT, which predicts response to targeted and immune therapies from the inferred expression values. We show that DeepPT successfully predicts transcriptomics in all 16 The Cancer Genome Atlas cohorts tested and generalizes well to two independent datasets. ENLIGHT-DeepPT successfully predicts true responders in five independent patient cohorts involving four different treatments spanning six cancer types, with an overall odds ratio of 2.28 and a 39.5% increased response rate among predicted responders versus the baseline rate. Notably, its prediction accuracy, obtained without any training on the treatment data, is comparable to that achieved by directly predicting the response from the images, which requires specific training on the treatment evaluation cohorts.

Endoscopic keel placement to treat and prevent anterior glottic webs.

We performed a retrospective chart review to examine and describe our clinical experience of use of the Lichtenberger technique to place silicone elastomer keels after lysis of existing webs or for prevention of webs following anterior commissure surgery in adults. Twenty-two patients were identified for inclusion, ranging in age from 24 to 80 years. For 18 patients with existing glottic webs,the surgical procedure involved laryngoscopy with complete lysis of the anterior glottic web by laser or sharp technique, followed by placement of a square of silicone elastomer that is sutured in place with the Lichtenberger needle holder and left in place for 3 to 5 weeks. The procedure was well tolerated, and successfully corrected the web in all but 2 cases. For 4 patients, the procedure was performed prophylactically at the time of anterior commissure surgery considered high-risk for web formation. The procedure does not require a tracheotomy, and patients can maintain a normal diet and have functional phonation while the keel is in place. This approach to treating anterior glottic webs offers several advantages over traditional open thyrotomy with keel placement and should be considered to treat or prevent anterior glottic webs.

Meet the authors: Dr. Clint Allen and Dr. Sandro Santagata.

In this Q&A, Cell Press Community Review Scientific Editor Leia Judge talks to Dr. Clint Allen and Dr. Sandro Santagata about their new papers "Phenotypic plasticity and reduced tissue retention of exhausted tumor-infiltrating T cells following neoadjuvant immunotherapy in head and neck cancer" and "Lymphocyte networks are dynamic cellular communities in the immunoregulatory landscape of lung adenocarcinoma" and their experiences with publishing through Cell Press Community Review.

Egress of resident memory T cells from tissue with neoadjuvant immunotherapy: Implications for systemic anti-tumor immunity.

INTRODUCTION: Resident memory T (TRM) cells are embedded in peripheral tissue and capable of acting as sentinels that can respond quickly to repeat pathogen exposure as part of an endogenous anti-microbial immune response. Recent evidence suggests that chronic antigen exposure and other microenvironment cues may promote the development of TRM cells within solid tumors as well, and that this TRM phenotype can sequester tumor-specific T cells into tumors and out of circulation resulting in limited systemic antitumor immunity. Here, we perform a review of the published English literature and describe tissue-specific mediators of TRM cell differentiation in states of infection and malignancy with special focus on the role of TGF-ß and how targeting TGF-ß signaling could be used as a therapeutical approach to promote tumor systemic immunity. DISCUSSION: The presence of TRM cells with antigen specificity to neoepitopes in tumors associates with positive clinical prognosis and greater responsiveness to immunotherapy. Recent evidence indicates that solid tumors may act as reservoirs for tumor specific TRM cells and limit their circulation - possibly resulting in impaired systemic antitumor immunity. TRM cells utilize specific mechanisms to egress from peripheral tissues into circulation and other peripheral sites, and emerging evidence indicates that immunotherapeutic approaches may initiate these processes and increase systemic antitumor immunity. CONCLUSIONS: Reversing tumor sequestration of tumor-specific T cells prior to surgical removal or radiation of tumor may increase systemic antitumor immunity. This finding may underlie the improved recurrence free survival observed with neoadjuvant immunotherapy in clinical trials.

Tumor cell HLA class I expression and pathologic response following neoadjuvant immunotherapy for newly diagnosed head and neck cancer.

OBJECTIVES: Biomarkers are needed to identify patients likely to respond to neoadjuvant immunotherapy (NIT) prior to receiving definitive treatment. MATERIALS AND METHODS: We hypothesized that expression of tumor cell HLA class I would correlate with pathologic response (PR) following NIT for primary untreated head and neck cancer. Multispectral immunofluorescence of pre- and post-treatment biopsy specimens from a neoadjuvant study of bintrafusp alfa, a dual TGF-ß and PD-L1 inhibitor, was performed. RESULTS: Discordant expression of tumor cell HLA class I and PD-L1 measured by multispectral immunofluorescence was observed with most positive tumor cells expressing HLA class I or PD-L1 but not both. Spatial analysis revealed colocalization between tumor parenchyma T cells and HLA class I positive tumors cells, but no clear colocalization between T cells and PD-L1 positive tumor cells. Greater pre-treatment tumor cell HLA class I expression, but not PD-L1 expression or tumor T cell infiltration, correlated with the development of a PR. Additionally, increased tumor cell HLA class I expression after NIT compared to before NIT correlated with development of a PR, whereas inconsistent changes in PD-L1 and T cell infiltration were observed after treatment in all patients. CONCLUSIONS: These data provide the rationale for the study of tumor cell HLA class I expression in larger prospective studies powered to determine the performance of biomarkers of PR in newly diagnosed HNSCC patients receiving NIT.

Neoadjuvant chemotherapy enhances tumor-specific T cell immunity in patients with HPV-associated oropharyngeal cancer.

BACKGROUND: Treatment of patients with newly diagnosed HPV-associated oropharyngeal squamous cell carcinoma (OPSCC) with neoadjuvant chemotherapy (NAC) results in a high rate of 5-year recurrence free survival with few patients requiring adjuvant treatment. We hypothesized that NAC enhances primary tumor HPV-specific T cell responses. METHODS: HPV-specific responses in tumor infiltrating lymphocytes (TILs) before and after NAC were determined using autologous co-culture assays. RESULTS: Greater HPV16-specific TIL responses, sometimes polyclonal, were observed after NAC compared to before in 8 of 10 patients (80%) with PCR-verified HPV16-positive tumors. A significant association was observed between net-negative change in HPV-specific TIL response and disease relapse (p = 0.04, Mann-Whitney test), whereas pathologic complete response at time of surgery did not correlate with recurrence. CONCLUSIONS: NAC induces HPV-specific tumor T cell responses in patients with newly diagnosed HPV-associated OPSCC; whereas lack of an increase following NAC may associate with risk of relapse.

Combined Inhibition of IAPs and WEE1 Enhances TNFα- and Radiation-Induced Cell Death in Head and Neck Squamous Carcinoma.

Head and neck squamous cell carcinoma (HNSCC) remains a prevalent diagnosis with current treatment options that include radiotherapy and immune-mediated therapies, in which tumor necrosis factor-α (TNFα) is a key mediator of cytotoxicity. However, HNSCC and other cancers often display TNFα resistance due to activation of the canonical IKK-NFκB/RELA pathway, which is activated by, and induces expression of, cellular inhibitors of apoptosis proteins (cIAPs). Our previous studies have demonstrated that the IAP inhibitor birinapant sensitized HNSCC to TNFα-dependent cell death in vitro and radiotherapy in vivo. Furthermore, we recently demonstrated that the inhibition of the G2/M checkpoint kinase WEE1 also sensitized HNSCC cells to TNFα-dependent cell death, due to the inhibition of the pro-survival IKK-NFκB/RELA complex. Given these observations, we hypothesized that dual-antagonist therapy targeting both IAP and WEE1 proteins may have the potential to synergistically sensitize HNSCC to TNFα-dependent cell death. Using the IAP inhibitor birinapant and the WEE1 inhibitor AZD1775, we show that combination treatment reduced cell viability, proliferation and survival when compared with individual treatment. Furthermore, combination treatment enhanced the sensitivity of HNSCC cells to TNFα-induced cytotoxicity via the induction of apoptosis and DNA damage. Additionally, birinapant and AZD1775 combination treatment decreased cell proliferation and survival in combination with radiotherapy, a critical source of TNFα. These results support further investigation of IAP and WEE1 inhibitor combinations in preclinical and clinical studies in HNSCC.

Oncogenic Ras and ΔNp63α cooperate to recruit immunosuppressive polymorphonuclear myeloid-derived suppressor cells in a mouse model of squamous cancer pathogenesis.

Introduction: Amplification of human chromosome 3q26-29, which encodes oncoprotein ΔNp63 among other isoforms of the p63 family, is a feature common to squamous cell carcinomas (SCCs) of multiple tissue origins. Along with overexpression of ΔNp63, activation of the protooncogene, RAS, whether by overexpression or oncogenic mutation, is frequently observed in many cancers. In this study, analysis of transcriptome data from The Cancer Genome Atlas (TCGA) demonstrated that expression of TP63 mRNA, particularly ΔNp63 isoforms, and HRAS are significantly elevated in advanced squamous cell carcinomas of the head and neck (HNSCCs), suggesting pathological significance. However, how co-overexpressed ΔNp63 and HRAS affect the immunosuppressive tumor microenvironment (TME) is incompletely understood. Methods: Here, we established and characterized an immune competent mouse model using primary keratinocytes with retroviral-mediated overexpression of ΔNp63α and constitutively activated HRAS (v-rasHa G12R) to evaluate the role of these oncogenes in the immune TME. Results: In this model, orthotopic grafting of wildtype syngeneic keratinocytes expressing both v-rasHa and elevated levels of ΔNp63α consistently yield carcinomas in syngeneic hosts, while cells expressing v-rasHa alone yield predominantly papillomas. We found that polymorphonuclear (PMN) myeloid cells, experimentally validated to be immunosuppressive and thus representing myeloid-derived suppressor cells (PMN-MDSCs), were significantly recruited into the TME of carcinomas arising early following orthotopic grafting of ΔNp63α/v-rasHa-expressing keratinocytes. ΔNp63α/v-rasHa-driven carcinomas expressed higher levels of chemokines implicated in recruitment of MDSCs compared to v-rasHa-initiated tumors, providing a heretofore undescribed link between ΔNp63α/HRAS-driven carcinomas and the development of an immunosuppressive TME. Conclusion: These results support the utilization of a genetic carcinogenesis model harboring specific genomic drivers of malignancy to study mechanisms underlying the development of local immunosuppression.