pyrimidinones and Squamous-Cell-Carcinoma-of-Head-and-Neck

In a head and neck squamous cell carcinoma (HNSCC) "window of opportunity" clinical trial, we reported that trametinib reduced MEK-Erk1/2 activation and resulted in tumor responses in a subset of patients. Here, we investigated resistance to trametinib and molecular correlates in HNSCC cell lines and patient samples.. HNSCC cell lines were treated with trametinib to generate resistant lines. Candidate bypass pathways were assessed using immunoblotting, CRISPR knockout, and survival assays. Effectiveness of combined trametinib and verteporfin targeting was evaluated. Patient-derived xenografts (PDXs) from responder patients were treated with trametinib and resistant tumors were analyzed. Window trial clinical samples were subjected to whole-exome and RNA sequencing.. HNSCC cell lines developed resistance (CAL27-TR and HSC3-TR) after prolonged trametinib exposure. Downstream effectors of the Hippo pathway were activated in CAL27-TR and HSC3-TR, and combined trametinib and verteporfin treatment resulted in synergistic treatment response. We defined the Hippo pathway effector Yap1 as an induced survival pathway promoting resistance to trametinib in HSC3-TR. Yap1 was necessary for HSC3-TR trametinib resistance, and constitutively active Yap1 was sufficient to confer resistance in parental HSC3. Analysis of trametinib neoadjuvant trial patient tumors indicated canonical MEK-Erk1/2 pathway activating mutations were infrequent, and Yap1 activity increased following trametinib treatment. Trametinib treatment of a PDX from a responder patient resulted in evolution of resistance with increased Yap1 expression and activity.. These studies identify a Yap1-dependent resistance to trametinib therapy in HNSCCs. Combined Yap1 and MEK targeting may represent a strategy to enhance HNSCC response.

Topics: Animals; Biopsy; Cell Line, Tumor; Drug Resistance, Neoplasm; Exome Sequencing; Female; Gene Expression Regulation, Neoplastic; Gene Knockout Techniques; Head and Neck Neoplasms; Hippo Signaling Pathway; Humans; MAP Kinase Signaling System; Mice; Pyridones; Pyrimidinones; RNA-Seq; Squamous Cell Carcinoma of Head and Neck; Xenograft Model Antitumor Assays; YAP-Signaling Proteins

Adavosertib (AZD1775) is an inhibitor of the Wee1 kinase. The authors conducted a phase 1b trial to evaluate the safety of adavosertib in combination with definitive chemoradiotherapy for patients with newly diagnosed, intermediate-risk/high-risk, locally advanced head and neck squamous cell carcinoma (HNSCC).. Twelve patients with intermediate-risk/high-risk HNSCC were enrolled, including those with p16-negative tumors of the oropharynx, p16-positive tumors of the oropharynx with ≥10 tobacco pack-years, and tumors of the larynx/hypopharynx regardless of p16 status. All patients were treated with an 8-week course of concurrent intensity-modulated radiotherapy at 70 grays (Gy) (2 Gy daily in weeks 1-7), cisplatin 30 mg/m. Three patients (25%) experienced a dose-limiting toxicity, including febrile neutropenia (n = 2) and grade 4 thromboembolism (n = 1). Two dose-limiting toxicities occurred with adavosertib at 150 mg. The median follow-up was 14.7 months. The 12-week posttreatment objective response rate determined by positron emission tomography/computed tomography was 100%. The 1-year progression-free and overall survival rates were both 90%. The maximum tolerated dose of adavosertib was 100 mg.. Adavosertib 100 mg (twice daily on Monday, Tuesday, and Wednesday of weeks 1, 2, 4, 5, 7, and 8), in combination with 70 Gy of intensity-modulated radiotherapy and cisplatin 30 mg/m

Topics: Antineoplastic Combined Chemotherapy Protocols; Chemoradiotherapy; Cisplatin; Head and Neck Neoplasms; Humans; Pyrazoles; Pyrimidinones; Squamous Cell Carcinoma of Head and Neck

Patients with head and neck squamous cell carcinoma with locally advanced disease often require multimodality treatment with surgery, radiotherapy and/or chemotherapy. Adjuvant radiotherapy with concurrent chemotherapy is offered to patients with high-risk pathological features postsurgery. While cure rates are improved, overall survival remains suboptimal and treatment has a significant negative impact on quality of life.Cell cycle checkpoint kinase inhibition is a promising method to selectively potentiate the therapeutic effects of chemoradiation. Our hypothesis is that combining chemoradiation with a WEE1 inhibitor will affect the biological response to DNA damage caused by cisplatin and radiation, thereby enhancing clinical outcomes, without increased toxicity. This trial explores the associated effect of WEE1 kinase inhibitor adavosertib (AZD1775).. This phase I dose-finding, open-label, multicentre trial aims to determine the highest safe dose of AZD1775 in combination with cisplatin chemotherapy preoperatively (group A) as a window of opportunity trial, and in combination with postoperative cisplatin-based chemoradiation (group B).Modified time-to-event continual reassessment method will determine the recommended dose, recruiting up to 21 patients per group. Primary outcomes are recommended doses with predefined target dose-limiting toxicity probabilities of 25% monitored up to 42 days (group A), and 30% monitored up to 12 weeks (group B). Secondary outcomes are disease-free survival times (groups A and B). Exploratory objectives are evaluation of pharmacodynamic (PD) effects, identification and correlation of potential biomarkers with PD markers of DNA damage, determine rate of resection status and surgical complications for group A; and quality of life in group B.. Research Ethics Committee, Edgbaston, West Midlands (REC reference 16/WM/0501) initial approval received on 18/01/2017. Results will be disseminated via peer-reviewed publication and presentation at international conferences.. ISRCTN76291951 and NCT03028766.

Topics: Adolescent; Adult; Aged; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Cell Cycle Proteins; Chemoradiotherapy, Adjuvant; Cisplatin; Clinical Protocols; Disease-Free Survival; Dose-Response Relationship, Drug; Enzyme Inhibitors; Female; Follow-Up Studies; Head and Neck Neoplasms; Humans; Male; Middle Aged; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidinones; Squamous Cell Carcinoma of Head and Neck; Treatment Outcome; Young Adult

In this study, we report a differential response of mitogen-activated protein kinase-kinase (MEK) inhibitor trametinib in 20 head and neck squamous cell carcinoma (HNSCC) patients' tumor-derived cell cultures. Relatively sensitive and resistant cases to trametinib were identified using high throughput metabolic assays and validated in extended dose response studies in vitro. High throughput metabolic assays exploring combination therapies with trametinib were subjected to synergy models and maximal synergistic dose analyses. These yielded several candidates, including axtinib, GDC-0032, GSK-690693, and SGX-523. The combination regimen of trametinib and AXL/MET/VEGFR inhibitor glesatinib showed initial efficacy both in vitro and in vivo (92% reduction in tumor volume). Sensitivity was validated in vivo in a patient-derived xenograft (PDX) model in which trametinib as a single agent effected reduction in tumor volume up to 72%. Reverse Phase Protein Arrays (RPPA) demonstrated differentially expressed proteins and phosphoproteins upon trametinib treatment. Furthermore, resistant cell lines showed a compensatory mechanism via increases in MAPK and non-MAPK pathway proteins that may represent targets for future combination regimens. Intrinsic-targeted options have potential to address paucity of medical treatment options for HNSCC cancer patients, enhance response to extrinsic targeted agents, and/or reduce morbidity as neoadjuvant to surgical treatments.

Topics: Head and Neck Neoplasms; Humans; Proteomics; Pyridones; Pyrimidinones; Squamous Cell Carcinoma of Head and Neck

Topics: Apoptosis; Cell Line, Tumor; Head and Neck Neoplasms; Humans; Hydroxamic Acids; Pyrazoles; Pyrimidines; Pyrimidinones; Squamous Cell Carcinoma of Head and Neck; Tumor Suppressor Protein p53

Despite advances in surgery, chemotherapy, and radiation, there are limited treatment options for advanced head and neck squamous cell carcinoma (HNSCC) and survival remains very poor. Therefore, effective therapies are desperately needed. Recently, selective exploitation of DNA damage and replication stress responses has become a novel approach for cancer treatment. Wee1 kinase and Rad51 recombinase are two proteins involved in regulating replication stress and homologous recombination repair in cancer cells. In this study, we investigated the combined effect of Rad51 inhibitor (B02) and Wee1 inhibitor (AZD1775)

Topics: Animals; Apoptosis; Cell Cycle; Cell Cycle Proteins; Cell Line, Tumor; Cell Survival; Cells, Cultured; Computational Biology; Disease Models, Animal; DNA Damage; DNA Repair; Dose-Response Relationship, Drug; Drug Synergism; Gene Expression Profiling; Homologous Recombination; Humans; Mice; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidinones; Rad51 Recombinase; Radiation-Sensitizing Agents; Squamous Cell Carcinoma of Head and Neck; Xenograft Model Antitumor Assays

Head and neck squamous cell carcinoma (HNSCC) associated with high-risk human papilloma virus (HPV) infection is a growing clinical problem. The WEE1 kinase inhibitor AZD1775 (WEE1i) overrides cell cycle checkpoints and is being studied in HNSCC regimens. We show that the HPV16 E6/E7 oncoproteins sensitize HNSCC cells to single-agent WEE1i treatment through activation of a FOXM1-CDK1 circuit that drives mitotic gene expression and DNA damage. An isogenic cell system indicated that E6 largely accounts for these phenotypes in ways that extend beyond p53 inactivation. A targeted genomic analysis implicated FOXM1 signaling downstream of E6/E7 expression and analyses of primary tumors and The Cancer Genome Atlas (TCGA) data revealed an activated FOXM1-directed promitotic transcriptional signature in HPV+ versus HPV- HNSCCs. Finally, we demonstrate the causality of FOXM1 in driving WEE1i sensitivity. These data suggest that elevated basal FOXM1 activity predisposes HPV+ HNSCC to WEE1i-induced toxicity and provide mechanistic insights into WEE1i and HPV+ HNSCC therapies.

Topics: CDC2 Protein Kinase; Cell Cycle Checkpoints; Cell Cycle Proteins; Cell Line, Tumor; DNA Damage; Forkhead Box Protein M1; Head and Neck Neoplasms; Humans; Oncogene Proteins, Viral; Papillomavirus E7 Proteins; Papillomavirus Infections; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidinones; Repressor Proteins; Squamous Cell Carcinoma of Head and Neck; Up-Regulation

Hyperactivation of phosphatidylinositol 3-kinase (PI3K) pathway occurs frequently in head and neck squamous cell carcinoma (HNSCC). However, clinical outcomes of targeting the PI3K pathway have been underwhelming. In present study, we investigated the resistant mechanisms and potential combination therapeutic strategy to overcome adaptive resistance to PI3K inhibitor in HNSCC. Treatment of NVP-BKM120, a pan-PI3K inhibitor, led to upregulation of interleukin-6 (IL-6) and subsequent activation of either extracellular signal-regulated kinase (ERK) or signal transducers and activators of transcription 3 (STAT3), causing modest antitumor effects on the growth of HNSCC cells. Blockade of autocrine IL-6 signaling with siRNA or neutralizing antibody for IL-6 receptor (IL-6R) completely abolished NVP-BKM120-induced activation of ERK and STAT3 as well as expression of c-Myc oncogene, which resulted in enhanced sensitivity to NVP-BKM120. Moreover, when compared with a pharmacologic inhibitor or silencing of STAT3, trametinib, a MEK inhibitor, in combination with NVP-BKM120 yielded more potent anti-proliferative effects by inhibiting S phase transition, arresting cells at G0/G1 phase, and downregulating IL-6 and c-Myc expression. Furthermore, as compared with either agent alone, combination of NVP-BKM120 with trametinib or tocilizumab, a humanized anti-IL-6R antibody, significantly suppressed tumor growth in NVP-BKM120-resistant patient-derived tumor xenograft (PDTX) models, which was also confirmed in PDTX-derived cell lines. Collectively, these results suggested that IL-6/ERK signaling is closely involved in adaptive resistance of NVP-BKM120 in HNSCC cells, providing a rationale for a novel combination therapy to overcome resistance to PI3K inhibitors.

Topics: Aminopyridines; Animals; Antibodies, Monoclonal, Humanized; Antineoplastic Combined Chemotherapy Protocols; Autocrine Communication; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Drug Resistance, Neoplasm; Extracellular Signal-Regulated MAP Kinases; Female; Head and Neck Neoplasms; Humans; Interleukin-6; MAP Kinase Signaling System; Mice; Mice, Inbred NOD; Morpholines; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Protein Kinase Inhibitors; Pyridones; Pyrimidinones; Receptors, Interleukin-6; RNA, Small Interfering; Squamous Cell Carcinoma of Head and Neck; STAT3 Transcription Factor; Xenograft Model Antitumor Assays

Topics: Animals; Apoptosis; Carcinoma, Squamous Cell; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; DNA Damage; DNA Replication; Drug Synergism; Female; Head and Neck Neoplasms; Histone Deacetylase Inhibitors; Humans; Hydroxamic Acids; Mice; Mutation; Nuclear Proteins; Phosphorylation; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones; Risk Factors; S Phase; Squamous Cell Carcinoma of Head and Neck; Tumor Suppressor Protein p53; Vorinostat

Despite therapeutic advancements, there has been little change in the survival of patients with head and neck squamous cell carcinoma (HNSCC). Recent results suggest that cancer-associated fibroblasts (CAF) drive progression of this disease. Here, we report that autophagy is upregulated in HNSCC-associated CAFs, where it is responsible for key pathogenic contributions in this disease. Autophagy is fundamentally involved in cell degradation, but there is emerging evidence that suggests it is also important for cellular secretion. Thus, we hypothesized that autophagy-dependent secretion of tumor-promoting factors by HNSCC-associated CAFs may explain their role in malignant development. In support of this hypothesis, we observed a reduction in CAF-facilitated HNSCC progression after blocking CAF autophagy. Studies of cell growth media conditioned after autophagy blockade revealed levels of secreted IL6, IL8, and other cytokines were modulated by autophagy. Notably, when HNSCC cells were cocultured with normal fibroblasts, they upregulated autophagy through IL6, IL8, and basic fibroblast growth factor. In a mouse xenograft model of HNSCC, pharmacologic inhibition of Vps34, a key mediator of autophagy, enhanced the antitumor efficacy of cisplatin. Our results establish an oncogenic function for secretory autophagy in HNSCC stromal cells that promotes malignant progression.

Topics: Animals; Autophagy; Cancer-Associated Fibroblasts; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Chloroquine; Culture Media, Conditioned; Cytokines; Drug Resistance, Neoplasm; Female; Fibroblast Growth Factor 2; Head and Neck Neoplasms; Humans; Interleukin-6; Interleukin-8; Male; Mice; Mice, SCID; Neoplasm Invasiveness; Pyridines; Pyrimidinones; Squamous Cell Carcinoma of Head and Neck; Xenograft Model Antitumor Assays

Topics: Animals; Antineoplastic Agents; Apoptosis; Carcinoma, Squamous Cell; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Checkpoint Kinase 1; Checkpoint Kinase 2; Dose-Response Relationship, Drug; G2 Phase Cell Cycle Checkpoints; Genotype; Head and Neck Neoplasms; Humans; LIM Domain Proteins; Mice, Nude; Molecular Targeted Therapy; Mutation; Nuclear Proteins; Phenotype; Polo-Like Kinase 1; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Protein-Tyrosine Kinases; Proto-Oncogene Proteins; Pteridines; Pyrazoles; Pyrimidines; Pyrimidinones; ras Proteins; RNA Interference; Signal Transduction; Smad4 Protein; Squamous Cell Carcinoma of Head and Neck; Thiophenes; Time Factors; Transfection; Tumor Burden; Urea; Xenograft Model Antitumor Assays

Topics: Cell Cycle; Cell Cycle Proteins; Cell Line, Tumor; Enzyme Inhibitors; Humans; Nuclear Proteins; Practice Guidelines as Topic; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones; Squamous Cell Carcinoma of Head and Neck

The PIK3CA mutation is one of the most common mutations in head and neck squamous cell carcinoma (HNSCC). Through this research we attempt to elicit the role of oncogene dependence and effects of targeted therapy on this PIK3CA mutation.. (1) To determine the role of oncogene dependence on PIK3CA-one of the more common and targetable oncogenes in HNSCC, and (2) to evaluate the consequence of this oncogene on the effectiveness of newly developed targeted therapies.. This was a cell culture-based, in vitro study performed at an academic research laboratory assessing the viability of PIK3CA-mutated head and neck cell lines when treated with targeted therapy.. PIK3CA-mutated head and neck cell lines were treated with 17-AAG, GDC-0941, trametinib, and BEZ-235.. Assessment of cell viability of HNSCC cell lines characterized for PIK3CA mutations or SCC25 cells engineered to express the PIK3CA hotspot mutations E545K or H1047R.. Surprisingly, in engineered cell lines, the hotspot E545K and H1047R mutations conferred increased, rather than reduced, IC50 assay measurements when treated with the respective HSP90, PI3K, and MEK inhibitors, 17-AAG, GDC-0941, and trametinib, compared with the SCC25 control cell lines. When treated with BEZ-235, H1047R-expressing cell lines showed increased sensitivity to inhibition compared with control, whereas those expressing E545K showed slightly increased sensitivity of unclear significance.. (1) The PIK3CA mutations within our engineered cell model did not lead to enhanced oncogene-dependent cell death when treated with direct inhibition of the PI3K enzyme yet did show increased sensitivity compared with control with dual PI3K/mTOR inhibition. (2) Oncogene addiction to PIK3CA hotspot mutations, if it occurs, is likely to evolve in vivo in the context of additional molecular changes that remain to be identified. Additional study is required to develop new model systems and approaches to determine the role of targeted therapy in the treatment of PI3K-overactive HNSCC tumors.

Topics: Benzoquinones; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Survival; Class I Phosphatidylinositol 3-Kinases; Head and Neck Neoplasms; Humans; Imidazoles; Indazoles; Inhibitory Concentration 50; Lactams, Macrocyclic; Molecular Targeted Therapy; Mutation; Phosphatidylinositol 3-Kinases; Protein Serine-Threonine Kinases; Pyridones; Pyrimidinones; Quinolines; Squamous Cell Carcinoma of Head and Neck; Sulfonamides; TOR Serine-Threonine Kinases; Tumor Cells, Cultured

Although the majority of patients with HPV(+) oropharyngeal cancers have a favorable prognosis, there are some patients with tumors that are resistant to aggressive chemoradiotherapy with unusual patterns of locoregional and systemic recurrences. Therefore, more effective therapies are needed. In this study, we investigated the chemosensitizing efficacy of the selective Wee-1 kinase inhibitor, AZD-1775, in HPV(+) head and neck squamous cell carcinoma (HNSCC).. Clonogenic survival assays and an orthotopic mouse model of HPV(+) oral cancer were used to examine the in vitro and in vivo sensitivity of HPV(+) HNSCC cell lines to AZD-1775 in combination with cisplatin, respectively. Cell-cycle analysis, DNA damage (γH2AX), homologous recombination (HR), and apoptosis were examined to dissect molecular mechanisms.. We found that AZD-1775 displays single-agent activity and enhances the response of HPV(+) HNSCC cells to cisplatin both in vitro and in vivo. The sensitivity of the HPV(+) HNSCC cells to AZD-1775 alone or in combination with cisplatin was associated with G2 checkpoint abrogation, persistent DNA damage, and apoptosis induction. This finding of AZD-1775 increasing the sensitivity of HPV(+) HNSCC cells to cisplatin through apoptosis was not seen previously in the HPV(-) HNSCC cancer cells and is accompanied by a decreased expression of the antiapoptotic proteins, MCl-1and XIAP, which appear to be cleaved following AZD-1775 treatment.. AZD-1775 selectively sensitizes HPV(+) HNSCC cells and orthotopic oral xenografts to cisplatin through apoptosis and support the clinical investigation of AZD-1775 in combination with cisplatin particularly in patients with advanced and recurrent metastatic HPV(+) HNSCC tumors.

Topics: Animals; Antineoplastic Agents; Apoptosis; Carcinoma, Squamous Cell; Caspases; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Cisplatin; Disease Models, Animal; Drug Resistance, Neoplasm; Drug Synergism; G2 Phase Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Genes, p53; Head and Neck Neoplasms; Humans; Inhibitory Concentration 50; Male; Mice; Myeloid Cell Leukemia Sequence 1 Protein; Nuclear Proteins; Papillomavirus Infections; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Pyrazoles; Pyrimidines; Pyrimidinones; Squamous Cell Carcinoma of Head and Neck; Tumor Burden; X-Linked Inhibitor of Apoptosis Protein; Xenograft Model Antitumor Assays