RESUMEN

BACKGROUND: Human papillomavirus (HPV) infection has become an important etiological driver of oropharyngeal squamous cell carcinoma (OPSCC), leading to unique tumor characteristics. However, the interplay between HPV-associated tumor cells and tumor microenvironment (TME) remains an enigma. METHODS: We performed a single-cell RNA-sequencing (scRNA-seq) on HPV-positive (HPV+) and HPV-negative (HPV‒) OPSCC tumors, each for three samples, and one normal tonsil tissue. Ex vivo validation assays including immunofluorescence staining, cell line co-culture, and flow cytometry analysis were used to test specific subtypes of HPV+ tumor cells and their communications with T cells. RESULTS: Through a comprehensive single-cell transcriptome analysis, we uncover the distinct transcriptional signatures between HPV+ and HPV‒ OPSCC. Specifically, HPV+ OPSCC tumor cells manifest an enhanced interferon response and elevated expression of the major histocompatibility complex II (MHC-II), potentially bolstering tumor recognition and immune response. Furthermore, we identify a CXCL13+CD4+ T cell subset that exhibits dual features of both follicular and pro-inflammatory helper T cells. Noteworthily, HPV+ OPSCC tumor cells embrace extensive intercellular communications with CXCL13+CD4+ T cells. Interaction with HPV+ OPSCC tumor cells amplifies CXCL13 and IFNγ release in CD4+T cells, fostering a pro-inflammatory TME. Additionally, HPV+ tumor cells expressing high MHC-II and CXCL13+CD4+ T cell prevalence are indicative of favorable overall survival rates in OPSCC patients. CONCLUSIONS: Together, our study underscores a synergistic inflammatory immune response orchestrated by highly immunogenic tumor cells and CXCL13+CD4+ T cells in HPV+ OPSCC, offering useful insights into strategy development for patient stratification and effective immunotherapy in OPSCC.

Asunto(s)

Linfocitos T CD4-Positivos , Quimiocina CXCL13 , Inmunoterapia , Neoplasias Orofaríngeas , Infecciones por Papillomavirus , Microambiente Tumoral , Humanos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Quimiocina CXCL13/metabolismo , Quimiocina CXCL13/genética , Antígenos de Histocompatibilidad Clase II/inmunología , Antígenos de Histocompatibilidad Clase II/metabolismo , Inmunoterapia/métodos , Activación de Linfocitos , Neoplasias Orofaríngeas/inmunología , Neoplasias Orofaríngeas/virología , Neoplasias Orofaríngeas/terapia , Papillomaviridae , Infecciones por Papillomavirus/inmunología , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/complicaciones

RESUMEN

Introduction: The human leukocyte antigen complex (HLA) is essential for inducing specific immune responses to cancer by presenting tumor-associated peptides (TAP) to T cells. Overexpressed tumor associated antigens, mainly cancer-testis antigens (CTA), are outlined as essential targets for immunotherapy in oropharyngeal squamous cell carcinoma (OPSCC). This study assessed the degree to which presentation, gene expression, and antibody response (AR) of TAP, mainly CTA, are correlated in OPSCC patients to evaluate their potential as immunotherapy targets. Materials and methods: Snap-frozen tumor (NLigand/RNA=40), healthy mucosa (NRNA=6), and healthy tonsils (NLigand=5) samples were obtained. RNA-Seq was performed using Illumina HiSeq 2500/NovaSeq 6000 and whole exome sequencing (WES) utilizing NextSeq500. HLA ligands were isolated from tumor tissue using immunoaffinity purification, UHPLC, and analyzed by tandem MS. Antibodies were measured in serum (NAb=27) utilizing the KREX™ CT262 protein array. Data analysis focused on 312 proteins (KREX™ CT262 panel + overexpressed self-proteins). Results: 183 and 94 of HLA class I and II TAP were identified by comparative profiling with healthy tonsils. Genes from 26 TAP were overexpressed in tumors compared to healthy mucosa (LFC>1; FDR<0.05). Low concordance (r=0.25; p<0.0001) was found between upregulated mRNA and class I TAP. The specific mode of correlation of TAP was found to be dependent on clinical parameters. A lack of correlation was observed both between mRNA and class II TAP, as well as between class II tumor-unique TAP (TAP-U) presentation and antibody response (AR) levels. Discussion: This study demonstrates that focusing exclusively on gene transcript levels fails to capture the full extent of TAP presentation in OPSCC. Furthermore, our findings reveal that although CTA are presented at relatively low levels, a few CTA TAP-U show potential as targets for immunotherapy.

Asunto(s)

Antígenos de Neoplasias , Neoplasias Orofaríngeas , Humanos , Neoplasias Orofaríngeas/inmunología , Neoplasias Orofaríngeas/genética , Antígenos de Neoplasias/inmunología , Antígenos de Neoplasias/genética , Masculino , Femenino , Persona de Mediana Edad , Presentación de Antígeno/inmunología , Anciano , Regulación Neoplásica de la Expresión Génica , Formación de Anticuerpos/genética , Formación de Anticuerpos/inmunología , Adulto , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Secuenciación del Exoma , Multiómica

RESUMEN

BACKGROUND: Oropharyngeal squamous cell carcinoma (OPSCC) induced by human papillomavirus (HPV-positive) is associated with better clinical outcomes than HPV-negative OPSCC. However, the clinical benefits of immunotherapy in patients with HPV-positive OPSCC remain unclear. METHODS: To identify the cellular and molecular factors that limited the benefits associated with HPV in OPSCC immunotherapy, we performed single-cell RNA (n=20) and T-cell receptor sequencing (n=10) analyses of tonsil or base of tongue tumor biopsies prior to immunotherapy. Primary findings from our single-cell analysis were confirmed through immunofluorescence experiments, and secondary validation analysis were performed via publicly available transcriptomics data sets. RESULTS: We found significantly higher transcriptional diversity of malignant cells among non-responders to immunotherapy, regardless of HPV infection status. We also observed a significantly larger proportion of CD4+ follicular helper T cells (Tfh) in HPV-positive tumors, potentially due to enhanced Tfh differentiation. Most importantly, CD8+ resident memory T cells (Trm) with elevated KLRB1 (encoding CD161) expression showed an association with dampened antitumor activity in patients with HPV-positive OPSCC, which may explain their heterogeneous clinical outcomes. Notably, all HPV-positive patients, whose Trm presented elevated KLRB1 levels, showed low expression of CLEC2D (encoding the CD161 ligand) in B cells, which may reduce tertiary lymphoid structure activity. Immunofluorescence of HPV-positive tumors treated with immune checkpoint blockade showed an inverse correlation between the density of CD161+ Trm and changes in tumor size. CONCLUSIONS: We found that CD161+ Trm counteracts clinical benefits associated with HPV in OPSCC immunotherapy. This suggests that targeted inhibition of CD161 in Trm could enhance the efficacy of immunotherapy in HPV-positive oropharyngeal cancers. TRIAL REGISTRATION NUMBER: NCT03737968.

Asunto(s)

Inmunoterapia , Neoplasias Orofaríngeas , Infecciones por Papillomavirus , Análisis de la Célula Individual , Humanos , Neoplasias Orofaríngeas/inmunología , Neoplasias Orofaríngeas/virología , Neoplasias Orofaríngeas/terapia , Inmunoterapia/métodos , Infecciones por Papillomavirus/inmunología , Infecciones por Papillomavirus/virología , Masculino , Femenino , Persona de Mediana Edad , Anciano , Subfamilia B de Receptores Similares a Lectina de Células NK

RESUMEN

Although several oropharyngeal cancer (OPC) susceptibility loci have been identified, most previous studies lacked detailed information on human papillomavirus (HPV) status. We conduct a genome-wide analysis by HPV16 serology status in 4,002 oral cancer cases (OPC and oral cavity cancer (OCC)) and 5,256 controls. We detect four susceptibility loci pointing to a distinct genetic predisposition by HPV status. Our most notable finding in the HLA region, that is now confirmed to be specific of HPV(+)OPC risk, reveal two independent loci with strong protective effects, one refining the previously reported HLA class II haplotype association. Antibody levels against HPV16 viral proteins strongly implicate the protective HLA variants as major determinants of humoral response against L1 capsid protein or E6 oncoprotein suggesting a natural immune response against HPV(+)OPC promoted by HLA variants. This indicates that therapeutic vaccines that target E6 and attenuate viral response after established HPV infections might protect against HPV(+)OPC.

Asunto(s)

Antígenos HLA/inmunología , Papillomavirus Humano 16/inmunología , Inmunidad Humoral , Neoplasias de la Boca/inmunología , Neoplasias Orofaríngeas/inmunología , Infecciones por Papillomavirus/inmunología , Anciano , Anticuerpos Antivirales/biosíntesis , Proteínas de la Cápside/genética , Proteínas de la Cápside/inmunología , Estudios de Casos y Controles , Femenino , Expresión Génica , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Antígenos HLA/clasificación , Antígenos HLA/genética , Haplotipos , Papillomavirus Humano 16/patogenicidad , Humanos , Masculino , Metaanálisis como Asunto , Persona de Mediana Edad , Neoplasias de la Boca/genética , Neoplasias de la Boca/patología , Neoplasias de la Boca/virología , Proteínas Oncogénicas Virales/genética , Proteínas Oncogénicas Virales/inmunología , Neoplasias Orofaríngeas/genética , Neoplasias Orofaríngeas/patología , Neoplasias Orofaríngeas/virología , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/patología , Infecciones por Papillomavirus/virología , Sitios de Carácter Cuantitativo , Proteínas Represoras/genética , Proteínas Represoras/inmunología , Factores de Riesgo , Fumar/fisiopatología

RESUMEN

Peptide vaccines exhibit great potential in cancer therapy via eliciting antigen-specific host immune response and long-term immune memory to defend cancer cells. However, the low induced immune response of many developing vaccines implies the imperatives for understanding the favorable structural features of efficient cancer vaccines. Herein, we report on the two groups of self-adjuvanting peptide vaccines with distinct morphology and investigate the relationship between the morphology of peptide vaccines and the induced immune response. Two nanofibril peptide vaccines were created via co-assembly of a pentapeptide with a central 4-aminoproline residue, with its derivative functionalized with antigen epitopes derived from human papillomavirus E7 proteins, whereas utilization of a pentapeptide with a natural proline residue led to the formation of two nanoparticle peptide vaccines. The immunological results of dendritic cell (DCs) maturation and antigen presentation induced by the peptide assemblies implied the self-adjuvanting property of the resulting peptide vaccines. In particular, cellular uptake studies revealed the enhanced internalization and elongated retention of the nanofibril peptide vaccines in DCs, leading to their advanced performance in DC maturation, accumulation at lymph nodes, infiltration of cytotoxic T lymphocytes into tumor tissues, and eventually lysis of in vivo tumor cells, compared to the nanoparticle counterparts. The antitumor immune response caused by the nanofibril peptide vaccines was further augmented when simultaneously administrated with anti-PD-1 checkpoint blockades, suggesting the opportunity of the combinatorial immunotherapy by utilizing the nanofibril peptide vaccines. Our findings strongly demonstrate a robust relationship between the immune response of peptide vaccines and their morphology, thereby elucidating the critical role of morphological control in the design of efficient peptide vaccines and providing the guidance for the design of efficient peptide vaccines in the future.

Asunto(s)

Adyuvantes Inmunológicos/farmacología , Vacunas contra el Cáncer/farmacología , Neoplasias Orofaríngeas/terapia , Papillomaviridae/efectos de los fármacos , Infecciones por Papillomavirus/terapia , Vacunas de Subunidad/farmacología , Adyuvantes Inmunológicos/síntesis química , Adyuvantes Inmunológicos/química , Animales , Presentación de Antígeno/efectos de los fármacos , Presentación de Antígeno/inmunología , Vacunas contra el Cáncer/síntesis química , Vacunas contra el Cáncer/química , Línea Celular , Humanos , Inmunoterapia , Ensayo de Materiales , Ratones , Estructura Molecular , Neoplasias Experimentales/inmunología , Neoplasias Experimentales/terapia , Neoplasias Orofaríngeas/inmunología , Papillomaviridae/inmunología , Infecciones por Papillomavirus/inmunología , Vacunas de Subunidad/síntesis química , Vacunas de Subunidad/química

RESUMEN

Human papillomavirus (HPV) is recognized as a major cause of oropharyngeal cancer (OPC) in Western countries. Less is known regarding its contribution to the OPC occurring in Asia. The current study aimed to investigate the association between antibody responses to HPV16 E7 and head and neck cancer (HNC) risk in a hospital-based case-control study conducted in Taiwan with 693 HNC cases and 1,035 controls. A positive association was observed between seropositivity to HPV16 E7 and OPC risk, whereas no significant association was found in the non-OPC cases. The increased OPC risk associated with seropositivity to HPV16 E7 was more significant among nonbetel quid or noncigarette users. Seropositivity to HPV16 E7 showed moderate agreement with P16 expression in OPC. OPC patients that were seropositive to HPV16 E7 or p16 positive were more highly educated and less likely to use alcohol, betel quids, and cigarettes compared to HPV16 E7 seronegative or p16 negative OPC patients. Furthermore, patients with p16 positive OPC were more likely to be women compared to patients with p16 negative OPC, likely owing to the low prevalence of alcohol, betel quid, and cigarette users among women. Overall, this study suggested that similar to Western countries, HPV may also be an important risk factor of OPC in Taiwan. With the declining consumption of betel quids and cigarettes in Taiwan, a higher percentage of OPC cases in Taiwan will be attributed to HPV in the future. Public health measures, including HPV vaccination, need to be implemented to prevent the occurrence of HPV-positive OPC.

Asunto(s)

Anticuerpos Antivirales/sangre , Papillomavirus Humano 16/inmunología , Neoplasias Orofaríngeas/virología , Proteínas E7 de Papillomavirus/inmunología , Consumo de Bebidas Alcohólicas/efectos adversos , Consumo de Bebidas Alcohólicas/epidemiología , Areca/efectos adversos , Estudios de Casos y Controles , Femenino , Neoplasias de Cabeza y Cuello/inmunología , Neoplasias de Cabeza y Cuello/virología , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Orofaríngeas/inmunología , Factores de Riesgo , Factores Sexuales , Fumar/efectos adversos , Fumar/epidemiología , Taiwán

RESUMEN

Human papillomavirus (HPV) infection drives tumorigenesis in the majority of cervical, oropharyngeal, anal, and vulvar cancers. Genetic and epidemiologic evidence has highlighted the role of immunosuppression in the oncogenesis of HPV-related malignancies. Here we review how HPV modulates the immune microenvironment and subsequent therapeutic implications. We describe the landscape of immunotherapies for these cancers with a focus on findings from early-phase studies exploring antigen-specific treatments, and discuss future directions. Although responses across these studies have been modest to date, a deeper understanding of HPV-related tumor biology and immunology may prove instrumental for the development of more efficacious immunotherapeutic approaches. SIGNIFICANCE: HPV modulates the microenvironment to create a protumorigenic state of immune suppression and evasion. Our understanding of these mechanisms has led to the development of immunomodulatory treatments that have shown early clinical promise in patients with HPV-related malignancies. This review summarizes our current understanding of the interactions of HPV and its microenvironment and provides insight into the progress and challenges of developing immunotherapies for HPV-related malignancies.

Asunto(s)

Neoplasias Orofaríngeas/inmunología , Papillomaviridae/inmunología , Infecciones por Papillomavirus/inmunología , Neoplasias del Cuello Uterino/inmunología , Femenino , Humanos , Inmunoterapia , Vacunas contra Papillomavirus

RESUMEN

OBJECTIVES: Limited information exists regarding the associations between pre-existing immune parameters in the tumor immune microenvironment (TIM) and either chemoradiosensitivity or prognosis for patients with squamous cell carcinoma of the nasopharynx or oropharynx involving virus-related or nonvirus-related tumors. STUDY DESIGN: Retrospective cohort study. METHODS: We retrospectively reviewed 141 patients with EBV+, p16+, or EBV- and p16- statuses who are receiving chemoradiotherapy. We performed immunohistochemistry using pretreatment biopsy specimens to analyze the programed death ligand 1 (PD-L1) levels in tumor and immune cells and CD8+ tumor-infiltrating lymphocyte (TIL) density. We evaluated chemoradiosensitivity and prognosis with respect to these immune-related parameters. RESULTS: Virus-related tumors showed associations with both PD-L1 expression and high CD8+ TIL density. Patients with higher CD8+ TIL density or greater numbers of PD-L1+ tumor and immune cells showed significant rates of favorable local recurrence-free survival (LRFS), progression-free survival (PFS), and overall survival (OS). Multivariate analyses demonstrated that higher CD8+ TIL density is an independent, significant, and favorable predictive factor for LRFS (P = .005) and OS (P = .003), although it is not a significant predictor of PFS (P = .077). CONCLUSIONS: Higher CD8+ TIL density represents a useful and favorable biomarker of chemoradiosensitivity in patients receiving chemoradiotherapy for nasopharyngeal or oropharyngeal cancer. LEVEL OF EVIDENCE: 3 Laryngoscope, 131:E1179-E1189, 2021.

Asunto(s)

Linfocitos T CD8-positivos/inmunología , Quimioradioterapia , Linfocitos Infiltrantes de Tumor/inmunología , Neoplasias Nasofaríngeas/inmunología , Neoplasias Nasofaríngeas/terapia , Neoplasias Orofaríngeas/inmunología , Neoplasias Orofaríngeas/terapia , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Microambiente Tumoral/inmunología

RESUMEN

Programmed death-ligand 1 (PD-L1) expression has been approved as an immune checkpoint inhibitor (ICI) response predictive biomarker; however, the clinicopathological and molecular features of HPV-positive oropharyngeal squamous cell carcinoma [HPV(+)OPSCC] based on PD-L1 expression are not well studied. We aimed to characterize clinicopathological, tumor immune microenvironmental, and molecular features of HPV(+)OPSCC with different PD-L1 expression scored by combined positive score (CPS). A total of 112 cases were collected from 2008-2021 and received PD-L1 and CD8 immunohistochemistry (IHC) staining. 71 samples received DNA sequencing out of which 32 samples received RNA sequencing for immune-related gene alterations or expression analysis. The 32 samples were also subjected to analysis of CD20, CD4, CD8, CD68, Foxp3 and P16 by multiplex immunofluorescence (mIF) staining, and the immune markers were evaluated in the tumor body (TB), tumor margin (TM) and normal stroma (NS) regions separately. Our results showed that of 112 HPV(+)OPSCC tumors, high(CPS≥20), intermediate(1≤CPS<20), and low(CPS<1) PD-L1 expression was seen in 29.5%, 43.8% and 26.8% cases respectively. Non-smoking patients and patients with tumors occurring at the tonsils or having rich lymphocytes infiltration had significantly higher PD-L1 expression. Patients with CPS≥20 had significantly higher tumor mutation burden (TMB, p=0.0058), and PD-L1 expression correlated significantly with CD8+ T cells infiltration, which were ample in tumor regions than in NS in mIF. CD20+, CD4+, CD68+, Foxp3+CD4+ cells were demonstrated to infiltrate higher in TM while CD20+ and CD68+ cells were also enriched in NS and TB regions respectively. However, none of them showed correlations with PD-L1 expression. ARID1A, STK11 alterations were enriched in the low PD-L1 group significantly, while anti-viral immune associated APOBEC mutation signature and immune-related genes expression such as XCL1 and IL11 were positively associated with PD-L1 expression (p<0.05). This is a comprehensive investigation revealing immune and molecular features of HPV(+)OPSCC based on PD-L1 expression. Our study suggested that 73.2% of HPV(+)OPSCC patients may benefit from immunotherapy, and high PD-L1 expression reflects immune-active status of HPV(+)OPSCC accompanied by higher immune effect factors such as TMB, CD8+ cytotoxic T cells and immune-related genomic alterations. Our study offers valuable information for understanding the immune features of HPV(+)OPSCC.

Asunto(s)

Antígeno B7-H1/inmunología , Carcinoma de Células Escamosas/inmunología , Reparación de la Incompatibilidad de ADN/inmunología , Inestabilidad de Microsatélites , Neoplasias Orofaríngeas/inmunología , Infecciones por Papillomavirus/inmunología , Antígeno B7-H1/biosíntesis , Antígeno B7-H1/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/inmunología , Biomarcadores de Tumor/metabolismo , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Reparación de la Incompatibilidad de ADN/genética , Femenino , Regulación Neoplásica de la Expresión Génica/inmunología , Humanos , Inmunohistoquímica/métodos , Linfocitos Infiltrantes de Tumor/inmunología , Linfocitos Infiltrantes de Tumor/metabolismo , Masculino , Persona de Mediana Edad , Mutación/inmunología , Neoplasias Orofaríngeas/genética , Neoplasias Orofaríngeas/metabolismo , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/metabolismo , RNA-Seq/métodos

RESUMEN

Epidermal growth factor (EGFR)-inhibitors have emerged as the primary therapy in advanced solid tumor malignancies because of improvement in survival with overall favorable side effect profile. However, 50–90% of patients treated with EGFR-inhibitors develop a follicular or acneiform rash, which can be symptomatic and source of psychosocial distress, negatively impacting quality of life. As this acneiform rash is a well-recognized cutaneous toxicity of EGFR-inhibitors, a treatment algorithm has been proposed for management based on severity. However, treatment options for EGFR-inhibitor induced rash may not be generalizable to African Americans whose differences in skin biology and sensitivity present pathophysiologic challenges. Herein, we present a case of an African American patient who developed this acneiform rash while on cetuximab. We also review the few cases that have been reported in the literature of EGFR-inhibitor rash in African Americans, highlighting important management considerations in this patient population. J Drugs Dermatol. 2020;19(9):894-896. doi:10.36849/JDD.2020.5275.

Asunto(s)

Antineoplásicos Inmunológicos/efectos adversos , Cetuximab/efectos adversos , Erupciones por Medicamentos/inmunología , Neoplasias Orofaríngeas/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello/tratamiento farmacológico , Negro o Afroamericano , Antibacterianos/uso terapéutico , Erupciones por Medicamentos/diagnóstico , Erupciones por Medicamentos/tratamiento farmacológico , Erupciones por Medicamentos/etnología , Quimioterapia Combinada , Receptores ErbB/antagonistas & inhibidores , Humanos , Masculino , Metilprednisolona/análogos & derivados , Metilprednisolona/uso terapéutico , Persona de Mediana Edad , Neoplasias Orofaríngeas/inmunología , Neoplasias Orofaríngeas/patología , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/secundario , Resultado del Tratamiento

RESUMEN

BACKGROUND: CMP-001 is a novel Toll-like receptor-9 agonist that consists of an unmethylated CpG-A motif-rich G10 oligodeoxynucleotide (ODN) encapsulated in virus-like particles. In situ vaccination of CMP-001 is believed to activate local tumor-associated plasmacytoid dendritic cells (pDCs) leading to type I interferon secretion and tumor antigen presentation to T cells and systemic antitumor T cell responses. This study is designed to investigate if CMP-001 would enhance head and neck squamous cell carcinoma (HNSCC) tumor response to anti-programmed cell death protein-1 (anti-PD-1) therapy in a human papilloma virus-positive (HPV+) tumor mouse model. METHODS: Immune cell activation in response to CMP-001±anti-Qß was performed using co-cultures of peripheral blood mononuclear cells and HPV+/HPV- HNSCC cells and then analyzed by flow cytometry. In situ vaccination with CMP-001 alone and in combination with anti-PD-1 was investigated in C57BL/6 mice-bearing mEERL HNSCC tumors and analyzed for anti-Qß development, antitumor response, survival and immune cell recruitment. The role of antitumor immune response due to CMP-001+anti-PD-1 treatment was investigated by the depletion of natural killer (NK), CD4+ T, and CD8+ T cells. RESULTS: Results showed that the activity of CMP-001 on immune cell (pDCs, monocytes, CD4+/CD8+ T cells and NK cells) activation depends on the presence of anti-Qß. A 2-week 'priming' period after subcutaneous administration of CMP-001 was required for robust anti-Qß development in mice. In situ vaccination of CMP-001 was superior to unencapsulated G10 CpG-A ODN at suppressing both injected and uninjected (distant) tumors. In situ vaccination of CMP-001 in combination with anti-PD-1 therapy induced durable tumor regression at injected and distant tumors and significantly prolonged mouse survival compared with anti-PD-1 therapy alone. The antitumor effect of CMP-001+anti-PD-1 was accompanied by increased interferon gamma (IFNγ)+ CD4+/CD8+ T cells compared with control-treated mice. The therapeutic and abscopal effect of CMP-001+ anti-PD-1 therapy was completely abrogated by CD8+ T cell depletion. CONCLUSIONS: These results demonstrate that in situ vaccination with CMP-001 can induce both local and abscopal antitumor immune responses. Additionally, the antitumor efficacy of CMP-001 combined with α-PD-1 therapy warrants further study as a novel immunotherapeutic strategy for the treatment of HNSCC.

Asunto(s)

Vacunas contra el Cáncer/farmacología , Inhibidores de Puntos de Control Inmunológico/farmacología , Neoplasias Orofaríngeas/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello/tratamiento farmacológico , Receptor Toll-Like 9/agonistas , Vacunas de Partículas Similares a Virus/farmacología , Animales , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Sinergismo Farmacológico , Femenino , Humanos , Ratones , Ratones Endogámicos C57BL , Neoplasias Orofaríngeas/inmunología , Receptor de Muerte Celular Programada 1 , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Receptor Toll-Like 9/inmunología

RESUMEN

BACKGROUND: We investigated T cell clonality (TCC) and T cell fraction (TCF) in human papilloma virus associated oropharyngeal squamous cell carcinoma (HPV(+)OPSCC) progressors [cases] vs. non-progressors [controls]. METHODS: This nested case-control study included patients undergoing intent-to-cure surgery ± adjuvant therapy from 6/1/2007-10/3/2016. Patients experiencing local/regional/distant disease (progressors), and a consecutive sample of non-progressors were matched (2 controls: 1 case) on tumor subsite, T-stage and number of metastatic lymph nodes. We performed imunosequencing of the CDR3 regions of human TCRß chains. RESULTS: 34 progressors and 65 non-progressors were included. There was no statistically significant difference in baseline TCF (range: 0.039-1.084) and TCC (range: 0.007-0.240) (p > 0.05). Female sex was associated with higher TCF (p = 0.03), while extranodal extension (ENE) was associated with lower TCF (p = 0.01). There was a positive correlation between tumor size and clonality (R = 0.34, p < 0.01). The strongest predictor of progression-free survival (PFS) was TCF (HR 0.80, 95%CI 0.66-0.96, p = 0.02). The strongest predictors of cancer specific survival (CSS) were TCF (HR0.69, 95%CI 0.47-1.00, p < 0.05) and Adult Comorbidity Evaluation-27 (ACE-27) score (p < 0.05). Similarly, the strongest predictors of overall survival (OS) were TCF (HR 0.62, 95%CI 0.43-0.91, p = 0.01) and ACE-27 score (p = 0.03). On multivariable modeling, TCF ≥ 0.4 was independently associated with PFS (HR 0.34, 95%CI 0.14-0.85, p = 0.02) while an ACE-27 score of ≥ 2 independently predicted CSS (HR 3.85, 95%CI 1.07-13.85, p = 0.04) and OS (HR 3.51, 95%CI 1.10-11.20, p = 0.03). CONCLUSIONS: In patients with HPV(+)OPSCC, TCF was higher in female patients and those without ENE, suggesting differential immune responses. Lower TCF was significantly and independently associated with disease progression. Better ACE-27 scores appear to predict improved oncologic control.

Asunto(s)

Alphapapillomavirus/inmunología , Neoplasias Orofaríngeas/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Linfocitos T/citología , Microambiente Tumoral/inmunología , Adulto , Factores de Edad , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Progresión de la Enfermedad , Femenino , Reordenamiento Génico de la Cadena beta de los Receptores de Antígenos de los Linfocitos T/inmunología , Papillomavirus Humano 16/inmunología , Humanos , Estimación de Kaplan-Meier , Ganglios Linfáticos/patología , Metástasis Linfática , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Neoplasias Orofaríngeas/mortalidad , Neoplasias Orofaríngeas/patología , Neoplasias Orofaríngeas/virología , Supervivencia sin Progresión , Factores Sexuales , Carcinoma de Células Escamosas de Cabeza y Cuello/mortalidad , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Carcinoma de Células Escamosas de Cabeza y Cuello/virología , Carga Tumoral

RESUMEN

Cellular immunotherapeutics targeting the human papillomavirus (HPV)-16 E6 and E7 proteins have achieved limited success in HPV-positive oropharyngeal cancer (OPC). Here we have conducted proteome-wide profiling of HPV-16-specific T cell responses in a cohort of 66 patients with HPV-associated OPC and 22 healthy individuals. Unexpectedly, HPV-specific T cell responses from OPC patients were not constrained to the E6 and E7 antigens; they also recognized E1, E2, E4, E5, and L1 proteins as dominant targets for virus-specific CD8+ and CD4+ T cells. Multivariate analysis incorporating tumor staging, treatment status, and smoking history revealed that treatment status had the most significant impact on HPV-specific CD8+ and CD4+ T cell immunity. Specifically, the breadth and overall strength of HPV-specific T cell responses were significantly higher before the commencement of curative therapy than after therapy. These data provide the first glimpse of the overall human T cell response to HPV in a clinical setting and offer groundbreaking insight into future development of cellular immunotherapies for HPV-associated OPC patients.

Asunto(s)

Antígenos de Neoplasias/inmunología , Papillomavirus Humano 16/inmunología , Neoplasias Orofaríngeas/inmunología , Infecciones por Papillomavirus/inmunología , Linfocitos T/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Estudios de Casos y Controles , Femenino , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Orofaríngeas/virología , Infecciones por Papillomavirus/virología

RESUMEN

OBJECTIVES: In oropharyngeal squamous cell carcinoma (OPSCC), toll-like receptors (TLR) 5 and 7 associate with the tumor's human papilloma virus (HPV) status (Jouhi et al., 2017). TLR 2, on the other hand, has been linked to head and neck squamous cell carcinoma (HNSCC), and to oral carcinogenesis (Farnebo et al., 2015; Binder Gallimidi et al., 2015). Here we investigated the presence of TLR 2 and 4 in HPV-positive and HPV-negative OPSCC, and their relationship to opportunistic oral pathogen Treponema denticola chymotrypsin-like protease (Td-CTLP) immunoexpression, clinical parameters, and patient outcome. MATERIALS AND METHODS: Clinicopathological data of 198 unselected consecutive OPSCC patients came from hospital registries. Immunoexpression of TLRs 2 and 4 we evaluated by immunohistochemistry, and earlier in this patient series we studied immunoexpression of Td-CTLP and HPV DNA, HPV mRNA, and p16 status. RESULTS: Immunoexpression of both TLRs 2 and 4 showed a significant association with HPV-status. Strong expression was associated with HPV-positivity and mild expression with HPV-negativity. Patients with strong TLR 2 immunoexpression in the HPV negative subgroup had significantly poorer 5-year DSS (58%) than did patients with mild TLR 2 expression (77%), and strong TLR 2 immunoexpression remained as an independent factor linked to increased disease mortality in the multivariable setting (P = 0.019). No association existed between TLR 2 or 4 and Td-CTLP expression. CONCLUSION: Our results support the role of TLR 2 receptor as a possible target for development of therapeutics as earlier proposed (Farnebo et al., 2015). The involvement of Td and other oral pathogens in carcinogenesis of OPSCC, remains open and calls for further study.

Asunto(s)

Carcinoma de Células Escamosas/mortalidad , Neoplasias Orofaríngeas/mortalidad , Receptor Toll-Like 2/inmunología , Carcinoma de Células Escamosas/inmunología , Femenino , Humanos , Masculino , Neoplasias Orofaríngeas/inmunología

RESUMEN

BACKGROUND: Oropharyngeal cancer (OPC) exhibits diverse immunological properties; however, their implications for immunotherapy are unknown. METHODS: We analysed 37 surgically resected and nine recurrent or metastatic anti-programmed death-1 (PD-1)/programmed death-ligand 1 (PD-L1)-treated OPC tumours. OPCs were classified into immune-rich (IR), mesenchymal (MS) and xenobiotic (XB) subtypes based on RNA-sequencing data. RESULTS: All IR type tumours were human papillomavirus (HPV) positive, most XB types were HPV negative, and MS types showed mixed HPV status. The IR type showed an enriched T cell exhaustion signature with PD-1+ CD8+ T cells and type I macrophages infiltrating the tumour nest on multiplex immunohistochemistry. The MS type showed an exclusion of CD8+ T cells from the tumour nest and high MS and tumour growth factor-ß signatures. The XB type showed scant CD8+ T cell infiltration and focal CD73 expression. The IR type was associated with a favourable response signature during anti-PD-1/PD-L1 therapy and showed a high APOBEC mutation signature, whereas the MS and XB types showed resistance signature upregulation. Among anti-PD-1/PD-L1-treated OPC patients, the IR type showed a favourable clinical response (3/4 patients), whereas the XB type showed early progression (3/3 patients). CONCLUSION: Our analysis classified OPCs into three subtypes with distinct immune microenvironments that are potentially related to the response to anti-PD-1/PD-L1 therapy.

Asunto(s)

Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Neoplasias Orofaríngeas/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Microambiente Tumoral/inmunología , Humanos , Linfocitos Infiltrantes de Tumor/inmunología , Neoplasias Orofaríngeas/tratamiento farmacológico , Neoplasias Orofaríngeas/genética , Carcinoma de Células Escamosas de Cabeza y Cuello/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Transcriptoma

RESUMEN

PURPOSE: In oropharyngeal squamous cell carcinoma (OPC), high CD8+ tumor-infiltrating lymphocyte (CD8+TIL) density confers improved prognosis. We compared neoadjuvant durvalumab (PD-L1 inhibitor) with durvalumab + tremelimumab (CTLA-4 inhibitor) in terms of impact on CD8+TIL density, safety, and efficacy in patients with OPC. PATIENTS AND METHODS: Patients with newly diagnosed stage II-IVA OPC or locoregionally recurrent OPC amenable to resection were included. Patients were randomized to two cycles of durvalumab or durvalumab + tremelimumab before surgery. The primary endpoint was change between baseline and resection specimen in CD8+TIL density between arms. Secondary endpoints included safety, response rate per RECIST, major pathologic response (MPR; ≤10% viable tumor cells) rate, and patient-reported outcomes. RESULTS: Of 28 eligible patients (14/arm), 20 (71%) had newly diagnosed OPC, and 24 (86%) were p16-positive. The posttreatment to pretreatment median CD8+TIL density ratio was 1.31 for durvalumab and 1.15 for combination treatment (P = 0.97; 95% CI: -1.07-2.28). In each group, 6 patients (43%, 95% CI: 17.66-71.14) had a response. Eight patients (29%) had a MPR at the primary tumor and/or nodal metastases. Neither baseline CD8+TIL density nor PD-L1 expression level correlated with overall response, but a trend toward greater CD8+TIL change in patients with a MPR was seen (P = 0.059; 95% CI: -0.33-3.46). Four patients (14%) had grade ≥3 adverse events. At median follow-up time of 15.79 months, all patients were alive, and one had an additional recurrence. CONCLUSIONS: Durvalumab + tremelimumab did not increase CD8+TIL density more than durvalumab alone did. The observed safety and activity support further investigation of neoadjuvant checkpoint inhibitor for OPC.

Asunto(s)

Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Recuento de Linfocitos , Neoplasias Orofaríngeas/tratamiento farmacológico , Neoplasias Orofaríngeas/inmunología , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales Humanizados/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Biomarcadores de Tumor , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Progresión de la Enfermedad , Femenino , Humanos , Inmunohistoquímica , Masculino , Terapia Neoadyuvante , Neoplasias Orofaríngeas/mortalidad , Neoplasias Orofaríngeas/patología , Calidad de Vida , Factores de Riesgo , Resultado del Tratamiento

RESUMEN

OBJECTIVE: The incidence of oropharyngeal squamous cell carcinoma (OPSCC), especially human papillomavirus (HPV)-associated, is increasing worldwide. Immunotherapy become available for patients with carcinomas in the head and neck region, however without ideal biomarker. Markers like PD-L1 vary in the clone of the antibody used, and the method of evaluation. Adequate and reliable immune cells characterization and evaluation is still not found. Furthermore, studies analyzing representativeness of different tissue samples are scarce. We analyzed small biopsy, lymph node (LN) metastasis and resected OPSCC, in regards of tumor infiltrating lymphocyte (TIL) density, PD-L1 and p16 expression. MATERIAL AND METHODS: Patients with OPSCC diagnosed from 2000 to 2016, with small biopsy, resection specimen and LN metastasis samples were selected. We analyzed TILs on hematoxylin-eosin stain, and PD-L1 and p16 expression in tumor cells. Concordance between different tumor locations was evaluated. RESULTS: 93 patients, with 65 small biopsies, 72 resection specimens, and 70 LN metastases were included. TILs, p16 and PD-L1 demonstrated very high concordance. Additionally, PD-L1 expression in the small biopsies was more representative of the PD-L1 expression in the resection specimens, than the LN samples. CONCLUSION: TILs density can be reliably assessed using hematoxylin-eosin stain with high concordance between the small biopsy, resection specimen and LN metastasis. Evaluation of concordance of p16 expression is very high, nevertheless some cases might be misdiagnosed on a small biopsy or lymph node metastasis. Evaluation of PD-L1 expression is very reliable on the biopsy specimen. Different PD-L1 clones and methods of evaluation still remain to be addressed.

Asunto(s)

Antígeno B7-H1/biosíntesis , Inhibidor p16 de la Quinasa Dependiente de Ciclina/biosíntesis , Linfocitos Infiltrantes de Tumor/inmunología , Neoplasias Orofaríngeas/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Antígeno B7-H1/inmunología , Biopsia , Estudios de Cohortes , Inhibidor p16 de la Quinasa Dependiente de Ciclina/inmunología , Femenino , Humanos , Inmunohistoquímica , Metástasis Linfática , Masculino , Persona de Mediana Edad , Neoplasias Orofaríngeas/patología , Neoplasias Orofaríngeas/cirugía , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Carcinoma de Células Escamosas de Cabeza y Cuello/cirugía , Adulto Joven

RESUMEN

INTRODUCTION: Human papilloma virus (HPV)-associated oropharyngeal squamous cell carcinoma (HPV-OPSCC) has been recognized to have an excellent response to treatment and has a distinct pathologic staging. For this reason, HPV testing is recommended in cytology specimens of metastatic OPSCC, although the guidelines for testing are not clearly defined. The aim of the current study was to establish a threshold for interpretation of p16 in aspirates from metastatic OPSCC. MATERIALS AND METHODS: Cases diagnosed as metastatic SCC by cytology in neck lymph nodes with concurrent p16 on cytology and on paired surgical specimen or an in situ hybridization (ISH) for HPV were included in the study. Stain intensity and percentage positive cells for p16 was compared with p16 on paired surgical pathology specimens and/or ISH RNA for HPV on cytology specimens. RESULTS: Of the 52 cases diagnosed as metastatic SCC on neck aspirates, paired surgical pathology specimens and/or ISH HPV was available in 17 cases. A p16 expression in ≥10%-15% cells resulted in a sensitivity and negative predictive value of 66% and 37%, respectively. However, when even minimal expression in tumor fragments is considered positive, the negative predictive value increases to 100%. CONCLUSIONS: We recommend that even minimal nuclear expression for p16 in viable tumor fragments must be considered as positive in cytology specimens. Expression limited to only background single tumor cells or in a necrotic specimen must be interpreted with caution.

Asunto(s)

Alphapapillomavirus/aislamiento & purificación , Inhibidor p16 de la Quinasa Dependiente de Ciclina , Metástasis Linfática , Carcinoma de Células Escamosas de Cabeza y Cuello , Biopsia con Aguja Fina , Inhibidor p16 de la Quinasa Dependiente de Ciclina/inmunología , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Humanos , Inmunohistoquímica , Hibridación in Situ , Metástasis Linfática/inmunología , Metástasis Linfática/patología , Neoplasias Orofaríngeas/inmunología , Neoplasias Orofaríngeas/patología , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/patología

RESUMEN

INTRODUCTION: Human papillomavirus with a high oncogenic potential (HR-HPV) is responsible for more than a half of squamous cell carcinomas of the oropharynx. The HR-HPV-dependent cases of this tumour have a better prognosis compared to the HR-HPV-negative cases, despite the usually more advanced disease at the time of diagnosis. In addition to genetic and epigenetic factors, the causes of this more favourable course of the disease are also seen in the participation of the tumour microenvironment, including the patient's immune system. Macrophages are one of the most important elements of the immunocompetent cells landscape that make up the tumour microenvironment. Traditionally, they are divided into 2 groups: inflammatory macrophages with the M1 phenotype and tumour-associated macrophages known as M2 phenotype macrophages. OBJECTIVE: The aim of this study was to investigate the impact of the macrophage infiltrates intensity of the M1/M2 and M2 phenotype separately on the clinical outcome of patients with squamous cell carcinoma of the oropharynx (OPSCC), taking into account the HR-HPV status of tumours. METHODS: The study involved 85 patients with OPSCC in which HR-HPV status in tumour tissue was determined using a double-check algorithm including the detection of viral DNA by RT-PCR method with subsequent confirmation of its biological activity by immunohistochemical demonstrating the P16INK4A protein overexpression. In each of the groups formed on the basis of HR-HPV status, macrophages were discriminated using CD68 and CD163 proteins as markers of pan-macrophage and M2 phenotype. The intensity of infiltrates was quantified by means of computer-assisted analysis in digital images of whole slides (virtual slides) separately in tumour tissue and stroma. RESULTS: In HPV-positive patients, significantly more intense infiltration of both M1/M2 and M2 macrophages was found in the tumour stroma compared to HPV-negative patients. The infiltrates from both types of macrophages in the tumour tissue were less intense and did not differ between these groups. Intensive infiltration of CD68+ macrophages in the tumour front was associated with higher rate of nodal failures and a shorter nodal control in both HR-HPV groups. In the group of HR-HPV-negative patients, heavy infiltration of CD163+ macrophages was associated with significantly shorter: loco-regional control (LRC), metastasis-free survival and overall survival (OS). These parameters and prognosis in patients with scanty CD163+ infiltration were similar to favourable outcomes in HR-HPV-positive patients. The relative risk of local-regional recurrence, distant metastases and disease-related death in HR-HPV-negative patients with intense CD163+ infiltrates was, respectively, 4.7, 5.4 and 5.7 compared to patients with scanty infiltrates. CONCLUSIONS: Tumours with a positive HR-HPV status demonstrate intense infiltrations of total pool M1/M2 and M2 macrophages. In the group of HPV-negative patients, intensive M1/M2 macrophage infiltrates correlate with higher risk of nodal failures, and intensive M2 infiltrates are an adverse prognostic factor for LRC, metastasis-free survival and OS.

Asunto(s)

Movimiento Celular/inmunología , Infecciones por Papillomavirus/complicaciones , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/virología , Macrófagos Asociados a Tumores/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Alphapapillomavirus , Antígenos CD/inmunología , Antígenos de Diferenciación Mielomonocítica/inmunología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Orofaríngeas/inmunología , Neoplasias Orofaríngeas/virología , Infecciones por Papillomavirus/inmunología , Adhesión en Parafina , Pronóstico , Receptores de Superficie Celular/inmunología , Microambiente Tumoral/inmunología , Macrófagos Asociados a Tumores/clasificación

RESUMEN

Oropharyngeal squamous cell carcinoma (OPSCC) incidence is increasing at a nearly epidemic rate, largely driven by the human papillomavirus (HPV). Despite the generally favorable clinical outcomes of patients with HPV driven (HPV+) OPSCC, a significant subset of HPV tumors associated with tobacco exposure have diminished treatment response and worse survival. The tumor immune microenvironment (TIME) has been shown to be a critical driver of treatment response and oncologic outcomes in OPSCC generally and HPV+ OPSCC more specifically. However, the impact of tobacco exposure on the TIME in OPSCC patients remains unclear. We analyzed the relationship between TIME, tobacco exposure and clinical outcomes in OPSCC patients (n = 143) with extensive tobacco exposure (median pack-years = 40). P16 overexpression, a surrogate marker of HPV association, was a strong predictor of relapse-free (RFS) and overall survival (OS) (p < 0.001, p < 0.001 respectively) regardless of tobacco exposure and associated strongly with differential infiltration of the tumor by both CD3 and CD8 lymphocytes measured via immunohistochemistry (p < 001, p < 0.001 respectively). CD3 and CD8 infiltration was a strong predictor of RFS and OS and associated strongly with disease stage (AJCC 8th Edition Staging Manual). Tobacco exposure correlated significantly (p < 0.001) with decreased CD8 infiltration in p16+ OPSCC tumors. Our findings demonstrate that the HPV+ OPSCC clinical outcomes are strongly correlated with the TIME, which is potentially modulated by tobacco exposure. Immunomodulatory strategies targeting this disease in smokers must take into consideration the potential modifying effects of tobacco exposure on treatment effectiveness and clinical outcomes.

Asunto(s)

Antígenos CD8/metabolismo , Nicotiana/efectos adversos , Neoplasias Orofaríngeas/inducido químicamente , Neoplasias Orofaríngeas/metabolismo , Carcinoma de Células Escamosas/inducido químicamente , Carcinoma de Células Escamosas/inmunología , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/virología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Orofaríngeas/inmunología , Neoplasias Orofaríngeas/virología , Papillomaviridae/fisiología , Estudios Retrospectivos , Riesgo , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/inmunología