Document text
Principal Investigator: Karen S. Anderson
Organization: YALE UNIVERSITY
Fiscal Year: 2024
Award: $432,279
Funding agency: National Institute of Dental and Craniofacial Research
SUMMARY
Human papillomavirus (HPV)-associated neck squamous cell carcinoma (HNSCC) represents an increasing
proportion of HNSCC. The incidence of HPV+ HNSCC has dramatically increased over the last 2 decades and
in 2012 surpassed uterine cervical cancer as the most common HPV-related malignancy in the U.S. Despite
the HPV vaccine, it is estimated that the “epidemic” of HNSCC caused by HPV will not diminish until 2060.
HPV+ HNSCCs occur in younger individuals and prognosis for patients with these tumors is better compared
to patients with classical HNSCC; however, ~25% of patients recur with few effective therapeutic options.
Based on observed hypermethylation of HPV+ HNSCC from TCGA, and understanding that HPV uses
hypermethylation to impede the innate immune response, effects of the demethylating agent, 5-azacytidine (5-
azaC), were tested on HPV+ HNSCC. We found that HPV+ HNSCC cells in culture and xenografts are
sensitive to 5-azaC, and that 5-azaC caused double strand breaks (DSB) that were not observed after 5-azaC
therapy in HPV-negative HNSCC, even with much higher doses. We found that following 5-azaC therapy,
APOlipoprotein B mRNA-Editing enzyme Catalytic polypeptide 3B (APOBEC3B) was associated with
chromatin in HPV+ HNSCC, but not HPV-negative cells. CRISPR knockdown of A3B prevented DSB and
protected cells from 5-azaC-induced death. Despite being required for DSBs and cellular toxicity caused by 5-
azaC, A3B was also required for clonogenic survival of untreated HPV+ HNSCC. These data showing that A3B
is required for survival of HPV+ HNSCC cells, but that following demethylation A3B mediates toxicity and DSB.
In addition, 5-azaC therapy increased type I interferon signaling as measured by increased expression of
interferon-stimulated genes. These exciting pre-clinical data led to a window trial of 5days of 5-azaC. Analysis
of tumor specimens confirmed in vitro data showing that 5-azaC resulted in cellular toxicity. Immunofluorescent
staining of an HPV+ patient tumors pre- and post-5-azaC showed a marked increase in tumor-associated
lymphocytes, possibly driven through activation of type I interferon combined with increased expression of
neoantigens. In this YHN-SPORE project, we hypothesize 5-azaC therapy will enhance response to nivolumab
(Nivo) through its ability to cause cell death, increase neoantigen expression, increase A3B-driven mutational
load, and enhance T cell infiltration through increased type I interferon signaling. These hypotheses will be
tested using established and novel in vitro assays, as well as through examination of pre- and post-therapy
tumor specimens from a 3-armed clinical trial. In Aim 1, tumor specimens from the SPORE window trial will be
analyzed to determine effects of 5-azaC, Nivo, or the combination on cell death, cell proliferation, immune
infiltration and immune activation. Aim 2 will employ standard and novel assays to explore the role of A3B in
cellular toxicity exposed by 5-azaC therapy. In Aim 3, we will determine effects of 5-azaC on activators of
immune recognition and response in the presence or absence of Nivo.
Terms: <5 AZC><5-AC><5-Aza-cytidine><5-Azacytidine><AXIN1 protein><AZC><After Care><After-Treatment><Aftercare><Assay><Axin 1><Axis Inhibitor 1><Azacitidine><Azacytidine><Bio-Informatics><Bioassay><Bioinformatics><Biological Assay><Biometrics><Biometry><Biostatistics><CRISPR><CRISPR/Cas system><Cancer Cause><Cancer Etiology><Cancers><Cell Body><Cell Communication and Signaling><Cell Death><Cell Growth in Number><Cell Line><Cell Multiplication><Cell Proliferation><Cell Protection><Cell Signaling><Cell Survival><Cell Viability><CellLine><Cells><Cellular Proliferation><Cessation of life><Characteristics><Checkpoint inhibitor><Chromatin><Clinical Trials><Clonal Expansion><Clustered Regularly Interspaced Short Palindromic Repeats><Cytidine Aminohydrolase><Cytidine Deaminase><Cytoprotection><DNA><DNA Damage><DNA Injury><Data><Death><Deoxyribonucleic Acid><Dependence><Diagnosis><Disease><Disorder><Dose><Epidemic><Epidermoid Carcinoma><Flow Cytofluorometries><Flow Cytofluorometry><Flow Cytometry><Flow Microfluorimetry><Flow Microfluorometry><GAGE><Gene Transcription><Genes><Genetic Alteration><Genetic Change><Genetic Transcription><Genetic defect><Genetics-Mutagenesis><Genomics><HNSCC><HPV><HPV Vaccine><HPV analysis><HPV associated HNSCC><HPV driven HNSCC><HPV driven head and neck cancer><HPV induced cancer><HPV malignancy><HPV(+) HNSCC><HPV(+) head and neck squamous cell carcinoma><HPV(-) HNSCC><HPV(-) head and neck squamous cell carcinoma><HPV+ HNSCC><HPV+ cancer><HPV+ head and neck cancers><HPV- HNSCC><HPV-Related Malignancy><HPV-associated cancer><HPV-associated head and neck cancer><HPV-associated head and neck squamous cell carcinoma><HPV-associated malignancy><HPV-negative HNSCC><HPV-negative head and neck cancer><HPV-negative head and neck squamous cell carcinoma><HPV-positive HNSCC><HPV-positive head and neck cancers><HPV-related HNSCC><HPV-related cancer><HPV-related head and neck squamous cell carcinoma><Head and Neck Cancer><Head and Neck Carcinoma><Head and Neck Squamous Cell Carcinoma><Health><Heterograft><Heterologous Transplantation><Human Papilloma Virus><Human Papilloma Virus Vaccine><Human Papilloma Virus-Related Malignancy><Human Papilloma Virus-Related Malignant Neoplasm><Human Papilloma Virus-associated cancer><Human Papilloma Virus-associated malignancy><Human Papilloma Virus-related cancer><Human Papillomavirus><Human papillomavirus Vaccine><Human papillomavirus cancer><Human papillomavirus driven HNSCC><Human papillomavirus driven head and neck cancer><Human papillomavirus induced cancer><Human papillomavirus malignancy><Human papillomavirus-Related Malignancy><Human papillomavirus-Related Malignant Neoplasm><Hypermethylation><IFN><Immune><Immune Cell Activation><Immune checkpoint inhibitor><Immune infiltrates><Immune response><Immunes><Immunofluorescence><Immunofluorescence Immunologic><Immunological response><In Vitro><Incidence><Individual><Infectious Human Wart Virus><Infiltration><Innate Immune Response><Interferon Type I><Interferons><Intracellular Communication and Signaling><Lymphatic cell><Lymphocyte><Lymphocytic><Lytotoxicity><Malignant Cell><Malignant Head and Neck Neoplasm><Malignant Neoplasms><Malignant Tumor><Measures><Mediating><Methylation><Molecular><Morbidity><Morbidity - disease rate><Mutagenesis><Mutagenesis Molecular Biology><Mutate><Mutation><Neck><Nivolumab><Opdivo><Outcome><PD 1><PD-1><PD1><Patients><Planocellular Carcinoma><Prognosis><RNA Expression><RNA Seq><RNA sequencing><RNAseq><Recurrence><Recurrent><Recurrent disease><Relapsed Disease><Research Specimen><Role><SCCHN><Sampling><Signal Pathway><Signal Transduction><Signal Transduction Systems><Signaling><Specimen><Squamous Carcinoma><Squamous Cell Epithelioma><Squamous cell carcinoma><Staining method><Stains><Strains Cell Lines><Supporting Cell><T cell infiltration><T-Cells><T-Lymphocyte><TCGA><Testicles><Testing><Testis><The Cancer Genome Atlas><Toxic effect><Toxicities><Transcription><Xenograft><Xenograft procedure><Xenotransplantation><apo B mRNA editing protein><apolipoprotein B mRNA editing enzyme><biological signal transduction><cancer cell><cancer microenvironment><cancer/testis antigen><cell killing><cervical and uterine cancer><cultured cell line><cytoprotective><cytotoxic><cytotoxicity><demethylating epigenetic therapy><demethylating therapy><demethylation><effective therapy><effective treatment><efficacy testing><flow cytophotometry><genome mutation><head and neck squamous carcinoma><head and neck squamous cell cancer><head/neck cancer><host response><human papilloma virus+ head and neck squamous cell carcinoma><human papillomavirus (-) head and neck squamous cell carcinoma><human papillomavirus - head and neck squamous cell carcinoma><human papillomavirus analysis><human papillomavirus associated head and neck cancer><human papillomavirus associated head and neck squamous cell carcinoma><human papillomavirus associated malignancy><human papillomavirus driven head and neck squamous cell carcinoma><human papillomavirus induced head and neck squamous cell carcinoma><human papillomavirus negative head and neck cancer><human papillomavirus negative head and neck squamous cell carcinoma><human papillomavirus positive HNSCC><human papillomavirus positive head and neck cancers><human papillomavirus positive head and neck squamous cell carcinoma><human papillomavirus related head and neck squamous cell carcinoma><human papillomavirus- head and neck squamous cell carcinoma><human papillomavirus-associated cancer><human papillomavirus-related cancer><immune activation><immune activator><immune cell infiltrate><immune check point inhibitor><immune system response><immunoresponse><in vitro Assay><knock-down><knockdown><ladakamycin><lymph cell><malignancy><malignant head and neck tumor><necrocytosis><neo-antigen><neo-epitopes><neoantigens><neoepitopes><neoplasm/cancer><new drug treatments><new drugs><new pharmacological therapeutic><new therapeutics><new therapy><next generation therapeutics><non-HPV HNSCC><non-human papillomavirus head and neck squamous cell carcinoma><novel><novel drug treatments><novel drugs><novel pharmaco-therapeutic><novel pharmacological therapeutic><novel therapeutics><novel therapy><oral HPV-positive HNSCC><oral HPV-positive head and neck cancers><oral human papillomavirus positive head and neck cancers><oral human papillomavirus positive head and neck squamous cell carcinoma><overexpress><overexpression><participant enrollment><patient enrollment><patient population><patient prognosis><polypeptide><post treatment><pre-clinical><preclinical><prevent><preventing><programmed cell death 1><programmed cell death protein 1><programmed death 1><resistance to therapy><resistant to therapy><response><sensor><side effect><sle2><social role><systemic lupus erythematosus susceptibility 2><therapeutic resistance><therapeutically effective><therapy resistant><thymus derived lymphocyte><transcriptome sequencing><transcriptomic sequencing><treatment resistance><tumor><tumor microenvironment><tumor xenograft><uterine and cervical cancer><wart virus><xeno-transplant><xeno-transplantation>