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PURPOSE Small-cell lung cancer (SCLC) is an aggressive malignancy with limited treatments. Delta-like ligand 3 (DLL3) is aberrantly expressed in most SCLC. Tarlatamab (AMG 757), a bispecific T-cell engager molecule, binds both DLL3 and CD3 leading to T-cellb–mediated tumor lysis. Herein, we report phase I results of tarlatamab in patients with SCLC. PATIENTS AND METHODS This study evaluated tarlatamab in patients with relapsed/refractory SCLC. The primary end point was safety. Secondary end points included antitumor activity by modified RECIST 1.1, overall survival, and pharmacokinetics. RESULTS By July 19, 2022, 107 patients received tarlatamab in dose exploration (0.003 to 100 mg; n = 73) and expansion (100 mg; n = 34) cohorts. Median prior lines of anticancer therapy were 2 (range, 1-6); 49.5% received antiprogrammed death-1/programmed death ligand-1 therapy. Any-grade treatment-related adverse events occurred in 97 patients (90.7%) and grade b % 3 in 33 patients (30.8%). One patient (1%) had grade 5 pneumonitis. Cytokine release syndrome was the most common treatment-related adverse event, occurring in 56 patients (52%) including grade 3 in one patient (1%). Maximum tolerated dose was not reached. Objective response rate was 23.4% (95% CI, 15.7 to 32.5) including two complete and 23 partial responses. The median duration of response was 12.3 months (95% CI, 6.6 to 14.9). The disease control rate was 51.4% (95% CI, 41.5 to 61.2). The median progression-free survival and overall survival were 3.7 months (95% CI, 2.1 to 5.4) and 13.2 months (95% CI, 10.5 to not reached), respectively. Exploratory analysis suggests that selecting for increased DLL3 expression can result in increased clinical benefit. CONCLUSION In patients with heavily pretreated SCLC, tarlatamab demonstrated manageable safety with encouraging response durability. Further evaluation of this promising molecule is ongoing.

Effective treatments for de novo and treatment-emergent small-cell/neuroendocrine (t-SCNC) prostate cancer represent an unmet need for this disease. Using metastatic biopsies from patients with advanced cancer, we demonstrate that delta-like ligand 3 (DLL3) is expressed in de novo and t-SCNC and is associated with reduced survival. We develop a PET agent, [89Zr]-DFO-DLL3-scFv, that detects DLL3 levels in mouse SCNC models. In multiple patient-derived xenograft models, AMG 757 (tarlatamab), a half-life–extended bispecific T-cell engager (BiTE) immunotherapy that redirects CD3-positive T cells to kill DLL3-expressing cells, exhibited potent and durable antitumor activity. Late relapsing tumors after AMG 757 treatment exhibited lower DLL3 levels, suggesting antigen loss as a resistance mechanism, particularly in tumors with heterogeneous DLL3 expression. These findings have been translated into an ongoing clinical trial of AMG 757 in de novo and t-SCNC, with a confirmed objective partial response in a patient with histologically confirmed SCNC. Overall, these results identify DLL3 as a therapeutic target in SCNC and demonstrate that DLL3-targeted BiTE immunotherapy has significant antitumor activity in this aggressive prostate cancer subtype. Significance: The preclinical and clinical evaluation of DLL3-directed immunotherapy, AMG 757, and development of a PET radiotracer for noninvasive DLL3 detection demonstrate the potential of targeting DLL3 in SCNC prostate cancer.

Effective treatments for de novo and treatment-emergent small-cell/neuroendocrine (t-SCNC) prostate cancer represent an unmet need for this disease. Using metastatic biopsies from patients with advanced cancer, we demonstrate that delta-like ligand 3 (DLL3) is expressed in de novo and t-SCNC and is associated with reduced survival. We develop a PET agent, [89Zr]-DFO-DLL3-scFv, that detects DLL3 levels in mouse SCNC models. In multiple patient-derived xenograft models, AMG 757 (tarlatamab), a half-life–extended bispecific T-cell engager (BiTE) immunotherapy that redirects CD3-positive T cells to kill DLL3-expressing cells, exhibited potent and durable antitumor activity. Late relapsing tumors after AMG 757 treatment exhibited lower DLL3 levels, suggesting antigen loss as a resistance mechanism, particularly in tumors with heterogeneous DLL3 expression. These findings have been translated into an ongoing clinical trial of AMG 757 in de novo and t-SCNC, with a confirmed objective partial response in a patient with histologically confirmed SCNC. Overall, these results identify DLL3 as a therapeutic target in SCNC and demonstrate that DLL3-targeted BiTE immunotherapy has significant antitumor activity in this aggressive prostate cancer subtype.Significance:The preclinical and clinical evaluation of DLL3-directed immunotherapy, AMG 757, and development of a PET radiotracer for noninvasive DLL3 detection demonstrate the potential of targeting DLL3 in SCNC prostate cancer.

FigureS7

Supplemental Movies 1a-1d: 22Rv1-LacZ and 22Rv1-DLL3 cells co-cultures. Time-lapse movies of 22Rv1-LacZ and 22Rv1-DLL3 cells (labeled with NuclightRed) co-cultured with T cells and either NT control BiTE® or AMG 757. T cells were plated at an E:T ratio of 5:1. Images were captured every 6 hours at 10x magnification. a. 22Rv1-LacZ cells and T cells treated with NT control BiTE®. b. 22Rv1-DLL3 cells and T cells treated with NT control BiTE®. c. 22Rv1-LacZ cells and T cells treated with AMG 757. d. 22Rv1-DLL3 cells and T cells treated with AMG 757.

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Background CD40, a TNF receptor family member, is expressed by a variety of immune cells and is involved in the activation of both adaptive and innate immune responses. Here, we used quantitative immunofluorescence (QIF) to evaluate CD40 expression on the tumor epithelium of solid tumors in large patient cohorts of lung, ovarian, and pancreatic cancers. Methods Tissue samples from nine different solid tumors (bladder, breast, colon, gastric, head and neck, non-small cell lung cancer (NSCLC), ovarian, pancreatic and renal cell carcinoma), constructed in tissue microarray format, were initially assessed for CD40 expression by QIF. CD40 expression was then evaluated on the large available patient cohorts for three of the tumor types demonstrating high CD40 positivity rate; NSCLC, ovarian and pancreatic cancer. The prognostic impact of CD40 expression on tumor cells was also investigated. Results CD40 expression on tumor cells was found to be common, with 80% of the NSCLC population, 40% of the ovarian cancer population, and 68% of the pancreatic adenocarcinoma population displaying some degree of CD40 expression on cancer cells. All of three of these cancer types displayed considerable intra-tumoral heterogeneity of CD40 expression, as well as partial correlation between expression of CD40 on tumor cells and on surrounding stromal cells. CD40 was not found to be prognostic for overall survival in NSCLC, ovarian cancer, or pancreatic adenocarcinoma. Conclusions The high percentage of tumor cells expressing CD40 in each of these solid tumors should be considered in the development of therapeutic agents designed to target CD40.

Cisplatin-based chemoradiation is an established organ-preserving strategy for locally advanced laryngeal cancer, but long-term survival remains suboptimal. Immunotherapy has been studied in the metastatic and unresectable recurrent settings. However, additional data are needed to assess its role in organ preservation for locally advanced laryngeal cancer.This trial was an open-label, single-arm, multi-institutional study with a Phase I run-in portion followed by a planned Phase II component, which closed early due to low accrual. Study patients had Stage III or IV (T2-3; N0-3; M0) laryngeal squamous cell carcinoma and were candidates for larynx preservation. Pembrolizumab was given 2-3 weeks prior to chemoradiation and then, q21 days concurrently with high-dose cisplatin and radiation prescribed to a total dose of 70 Gy. The primary endpoint of the trial was organ-preservation rate (OPR) at 18 months.A total of nine patients were enrolled with a median follow-up of 30.1 months. No patient required laryngectomy, resulting in 100% OPR at 18 months. The 12-month overall survival (OS) rate was 77.8% and the median duration of OS was not reached. All acute Grade 4 (UCCI-HN-15-02 demonstrated the safety of the addition of immunotherapy to definitive chemoradiation and the patient outcomes suggest the potential for improving long-term survival while minimizing negative impact from treatment. While results from this trial were promising, a randomized study with a larger number of patients and longer follow-up is warranted to verify this treatment approach prior to wider adoption. NCT #: NCT02759575.Level of evidence: 2b.

Background The 5-year overall survival (OS) rate remains at 50% for patients with locally advanced head and neck squamous cell carcinoma (LAHNSCC), thereby underscoring the need for improved treatments. An antidiabetic agent, metformin, was found in retrospective studies to improve survival in patients with HNSCC. Therefore, the authors conducted a phase 1 dose escalation study combining metformin with chemoradiotherapy in patients with LAHNSCC. Methods Nondiabetic patients with LAHNSCC were enrolled in the current study to receive escalating doses of metformin and CRT based on the modified toxicity probability interval design. Metformin cohort doses included 2000 mg, 2550 mg, and 3000 mg daily in divided doses in addition to cisplatin (at a dose of 100 mg/m2 on days 1, 22, and 43) and standard radiotherapy (70 grays). Adverse events were categorized as per the National Cancer Institute Common Terminology Criteria for Adverse Events (version 4.03). Results Twenty patients were enrolled, 2 of whom withdrew consent. The median age of the patients was 56 years and the majority were male (83%), were white (88%), had p16-positive disease (72%), and were tobacco users (61%). The median length of metformin exposure was 28.5 days. The most common grade ≥3 toxicities were nausea (11%), vomiting (11%), mucositis (6%), acute kidney injury (17%), anemia (6%), and leukopenia (11%). Dose-limiting toxicities included diarrhea and acute kidney injury. After a median follow-up of 19 months, the 2-year overall survival and progression-free survival rates were 90% and 84%, respectively. No hypoglycemia events or lactic acidosis were observed. Cisplatin administration did not appear to affect metformin pharmacokinetics. The maximum tolerated dose for metformin could not be determined given the limited number of patients who tolerated metformin during chemoradiotherapy. Conclusions To the authors' knowledge, the current study is the first phase 1 trial combining metformin with chemoradiotherapy. Rates of overall survival and progression-free survival were encouraging in this limited patient population, and warrant further investigation in a phase 2 trial.

Concurrent chemoradiation (CRT) with a platinum-containing regimen is considered standard of care for eligible unresectable stage III non-small-cell lung cancer (NSCLC) patients. Attempts to improve locoregional control, and thereby enhance overall survival (OS), by increasing the dose of radiation failed to do so in a contemporary phase III trial. Better understanding of the effects of radiation on the immune system, particularly absolute lymphocyte counts (ALC), may explain the poorer survival seen with high-dose (HD) radiation compared to low dose (LD). We hypothesized that treatment-induced lymphopenia affects outcomes in stage III NSCLC patients treated with concurrent CRT. The study is a retrospective multi-institution study. Stage III NSCLC patients who received CRT between 1994 and 2014 were categorized into LD RT (≤ 66 Gy) and HD RT (> 66 Gy) groups. Overall survival was assessed using Kaplan-Meier method with log-rank test and Cox proportional hazard regression model to estimate the effect of radiation dose (LD vs. HD groups) on OS. Hematologic values including ALC were evaluated at diagnosis and at intervals during treatment. Patients with pre-existing immunodeficiency states and autoimmune disease were excluded from the analysis. Of the patients, 182 were included in the analysis. Median age was 62 years (range 37–82). Seventy-seven percent were males, 67% Caucasians. Most common types of histology were adenocarcinoma (52%) and squamous cell carcinoma (41%). One-hundred-fifty-four patients received LD RT and 28 received HD RT. Medians (ranges) of pre-treatment ALC were 1730/mm3 (384, 4300) vs. 2065/mm3 (600, 4580) (p = 0.58) between LD and HD groups. Medians (ranges) of nadir ALC were 324/mm3 (20, 2410) vs. 279.5/mm3 (20–930) (p = 0.11) between LD and HD groups. Kaplan-Meier survival curves showed that LD has better OS than the HD group (median survival 31.2 months vs. 13.9 months, p

Objective The antifolate chemotherapy agent pemetrexed has been widely used to treat non-small-cell-lung-cancer (NSCLC), but there is no clinically validated biomarker to select patients likely to respond. The aim of this study was to assess two proteins involved in DNA repair mechanisms, uracil DNA glycosylase (UDG) and BRCA1 as potential prognostic biomarkers in NSCLC patients treated with pemetrexed-based chemotherapy. Material and methods Formalin-fixed-paraffin-embedded tumor specimens from 119 patients with advanced NSCLC treated with pemetrexed between 2004 and 2011 were retrospectively analyzed. Expression of UDG, BRCA1, and known prognostic factors ALK, TTF-1, thymidylate synthase and folylpolyglutamate synthase was assessed by immunohistochemistry using H-SCORE (product of percent stained cells and intensity of expression). Progression-free (PFS) and overall survival (OS) served as reference endpoint. Results Most NSCLC tumor samples had UDG positivity in at least 5% of tumor cells and 34% samples had more than 50% positive tumor cells. Using the median expression value as threshold, high UDG expression (H-SCORE≥75) was significantly associated with shorter median PFS (3-year PFS 7% vs. 37%, p = 0.045) and a trend for shorter OS (3-year OS 15% vs 42%, p = 0.066) compared to patients with low UDG. In multivariable Cox analysis, the association between high UDG and shorter PFS was close to statistically significant (p = 0.08) at a significance level of 0.05 after controlling for age, gender, ALK- and TTF1-status with hazard ratio of 2.1. Grouping patients according to combined UDG and BRCA1 expression, patients with a profile of UDGhigh/BRCA1high had the shortest PFS and OS compared to all other patient groups (p = 0.007 and 0.02, respectively). Conclusion Our results demonstrate an important prognostic role for high UDG expression in pemetrexed-treated NSCLC patients, in addition to its previously reported role in pemetrexed cytotoxicity. High UDG expression was predictive of shorter PFS and OS, and patients with a combined profile of UDGhigh/BRCA1high had the poorest outcome following pemetrexed treatment.

Purpose/Objective(s) Cisplatin-based chemoradiation is an established organ-preserving strategy for locally advanced laryngeal squamous cell carcinoma, but long-term survival remains suboptimal, warranting an improved approach. Immunotherapy has been studied in the metastatic and unresectable recurrent setting, but additional data are needed to assess its role in definitive therapy. The study hypothesis was that the use of immunotherapy in conjunction with chemoradiation would be safe and result in improved laryngectomy-free survival (LFS) compared to standard of care chemoradiation. Materials/Methods This trial was an open-label, single-arm, prospective, multi-institutional study. The study included a Phase I run-in portion to assess safety in the first 6 enrolled patients and a planned subsequent Phase II component. Due to slow accrual, the study was closed after enrollment of 9 patients and the data were allowed to mature. The primary endpoint of the Phase II portion was LFS at 18 months. Study patients had Stage III or IV (T1-3; N0-3; M0) laryngeal squamous cell carcinoma and were candidates for larynx preservation. Pembrolizumab was given as a 200 mg flat dose 3 weeks prior to the start of chemoradiation and was then given q21 days until the completion of chemoradiation. Cisplatin was given at a dose of 100 mg/m2 q21 days during radiation with allowed dose modifications for toxicities. Radiation was prescribed to a total dose of 70 Gy in 35 daily fractions using IMRT with an elective nodal dose of 56-63 Gy. Results A total of 9 patients with a median age of 54 were enrolled from 2017 to 2019. The median follow-up time was 30.1 months. None of the enrolled patients required laryngectomy, resulting in 100% LFS at 18 months for evaluable patients. The 18-month overall survival (OS) rate was 66.7%; of the 3 patient deaths, 2 were due to co-morbid conditions (diabetes, peripheral arterial disease) rather than malignancy. The remaining 6 patients were alive at the time of last follow-up and the median duration of OS was not reached. There were 27 Grade 3 toxicities, with 2 attributable to pembrolizumab. Nearly all Grade 4 (n = 4) and Grade 5 (n = 1) toxicities occurred in a single patient with poorly-controlled diabetes; the Grade 5 toxicity was due to diabetic ketoacidosis. One patient had late Grade 4 laryngeal edema requiring tracheostomy 8 months after chemoradiation; the edema has since resolved and the tracheostomy was reversed. Conclusion The use of concurrent pembrolizumab with chemoradiation for locally advanced larynx cancer resulted in high LFS. Most toxicities were attributable to chemoradiation and the majority of Grade 4 and 5 toxicities occurred in a patient with poorly-controlled diabetes. A case of late Grade 4 laryngeal edema could have been related to immunotherapy. Future studies with concurrent immunotherapy with chemoradiation should account for this possibility.

Background Delta-like ligand 3 (DLL3) is an inhibitory Notch ligand that is highly expressed in small cell lung cancer (SCLC) and minimally expressed in normal tissues.1 AMG 757, a half-life extended BiTE® immune therapy, binds to DLL3 on tumor cells and CD3 on T cells, resulting in T cell-dependent killing of tumor cells. We report initial safety and efficacy from the ongoing phase 1 study of AMG 757 in patients with SCLC. Methods AMG 757 was administered intravenously every two weeks (with/without step dose) at doses of 0.003–3.0 mg. Eligible patients had SCLC that progressed or recurred following ≥1 platinum-based regimen. Antitumor activity was assessed using modified RECIST 1.1. The study was approved by the Ethics Board at participating institutions. Results As of 1 June 2020, safety and efficacy data are available for 31 patients enrolled at the first seven dose levels (DL) with median age, 63 (44–74) years; ECOG PS: 0–1, n=30 (96.8%); median prior lines, 2.0 (1–6); and previous PD-1/PD-L1 treatment: n=12 (38.7%). Median treatment duration was 6.1 (0.1–59.4) weeks. Treatment-emergent adverse events (AEs) were reported for 30 (96.8%) patients. AMG 757-related AEs occurred in 25 (80.6%) patients, including 5 (16.1%) that were grade ≥3 and one (3.2%) grade 5 (pneumonitis in DL5 [0.3 mg]). Three AEs (dyspnea, pneumonitis, fatigue) led to treatment discontinuation. The most common AE was cytokine release syndrome (CRS), which was reported in 11 (35.5%) patients. CRS AEs were grade 1–2, consisted mainly of fever with/without hypotension, and occurred mostly within 24 hours of the first or second dose of AMG 757. CRS events were reversible, did not lead to treatment interruption or discontinuation, and were managed with supportive care, corticosteroids, and/or anti-IL 6 therapy. The MTD for AMG 757 has not yet been reached. AMG 757 exhibited dose proportional increase in exposures. Response to AMG 757 is shown (figure 1). Confirmed partial response was reported in 5 (16.1%) patients (1/12 [8.3%] in DL5, 1/8 [12.5%] in DL6, 3/7 [42.9%] in DL7), and stable disease in 8 (25.8%) of all treated patients. Most responses occurred after 8 weeks on treatment. All responders remain on treatment with duration of response ranging from 2.0+ to 7.4 months+. Conclusions AMG 757 administered at a dose of up to 3 mg every two weeks has an acceptable safety profile and shows anti-tumor activity in patients with relapsed/refractory SCLC. Further dose escalation is ongoing. Trial Registration NCT03319940 Ethics Approval The study was approved by the Ethics Board at participating institutions. Consent N/A Reference Leonetti A, Facchinetti F, Minari R, Cortellini A, Rolfo CD, Giovannetti E, Tiseo M. Cell Oncol (Dordr). 2019;42:261–273.

Background Enhancement of antitumor immunity through inhibition of the checkpoint PD-1 receptor has been effective in the treatment of many malignancies. AMG 404 is a monoclonal antibody (mAb) targeting PD-1. This phase 1, open-label, multicenter first-in-human study (NCT03853109) will evaluate the safety, tolerability, pharmacokinetics, and efficacy of AMG 404 monotherapy in adult patients with advanced solid tumors. Methods The primary study endpoint is dose-limiting toxicity (DLT) and safety; key secondary endpoints include pharmacokinetic parameters, objective response rate (assessed Q8W), duration of response, and progression-free survival. Key inclusion criteria include histologically or cytologically proven metastatic or locally advanced solid tumors not amenable to curative treatment with surgery or radiation for which standard therapies have been exhausted or not available. Prior anti-PD-(L)1 or other checkpoint inhibitors were not allowed. Five dose-finding cohorts, including 2 expansion cohorts, ranged from 3–20 patients each. AMG 404 was given until disease progression, intolerance, or consent withdrawal. Results As of the data cutoff date of May 4, 2020, 62 patients received at least 1 dose of AMG 404 and were included in the safety and efficacy analysis sets. Fifty percent were men, 72% had ECOG 1 performance status, median age was 62 years (range: 28–83), and 42% had ≥3 lines of prior anticancer therapy. Median AMG 404 exposure was ~3 months (maximum: ~12 months). No DLTs were observed. Treatment-related adverse events (TRAEs) were reported for 29 patients (47%): those reported for ≥2 patients were fatigue (n=7); hypothyroidism (n=6); increased blood thyroid stimulating hormone and nausea (n=4 each); increased aspartate aminotransferase, decreased appetite, and pyrexia (n=3 each); and increased alanine aminotransferase, arthralgia, diarrhea, and increased weight (n=2 each). AEs leading to withdrawal of AMG 404 were reported for 3 patients (5%); all were serious and considered to be not related to AMG 404. Sixteen (26%) patients died on study; no deaths were considered related to AMG 404. Preliminary pharmacokinetic results were consistent with those of other therapeutic anti-PD-1 mAbs. Three patients had a confirmed partial response (pancreatic cancer, clear cell cancer, and pleomorphic sarcoma); an additional 4 patients had one scan with a partial response and are pending a confirmatory scan (clear cell renal carcinoma, undifferentiated nasopharyngeal carcinoma, sarcomatoid carcinoma of unknown primary, and colon cancer). Conclusions AMG 404 is tolerable at the tested doses with no DLTs reported. All observed TRAEs are consistent with other anti-PD-1 therapies. Encouraging anti-tumor activity has been observed in heavily pretreated patients. The study is continuing enrollment into additional cohorts. Trial Registration NCT03853109 Ethics Approval The study was approved by the Ethics Board of each institution involved in this study and can be produced upon request.

Background Checkpoint inhibitors are a promising therapy for patients with solid tumors; however, many patients require additional therapies to maximize clinical benefit or overcome resistance.1 The type-1 cytokine interleukin-21 (IL-21) is a promising candidate for combination and has shown clinical activity in melanoma and renal cell cancer.2 IL-21 has also shown improved efficacy when combined with anti-programmed death (PD)-1 antibodies in preclinical models.3 4 AMG 256 is a mutated IL-21 cytokine fused to an anti-PD-1 antibody to combine IL-21 pathway stimulation with checkpoint inhibition—a strategy that is designed to prime and extend the activity of cytotoxic and memory T cells and induce anti-tumor immunity. This first-in-human (FIH) study will assess safety, tolerability, and estimated dosing of AMG 256 monotherapy in patients with advanced solid tumors. Methods This is a FIH, multicenter, non-randomized, open-label, phase 1 study (NCT04362748) of AMG 256 in patients with advanced solid tumors. The planned sample size is approximately 100 patients in two parts: part 1 will evaluate safety, tolerability, pharmacokinetics (PK), pharmacodynamics, and determine the maximum tolerated dose (MTD), part 2 will evaluate the MTD determined in part 1 to further characterize the safety profile and preliminary tumor response. AMG 256 will be delivered by intravenous (IV) infusion. Enrollment criteria include adults with life expectancy of > 3 months, ECOG performance status ≤ 2, histologically or cytologically confirmed metastatic or locally advanced solid tumors not amenable to curative treatment with surgery or radiation, and at least one measurable lesion ≥ 10 mm that has not undergone biopsy within 3 months of screening scan. Exclusion criteria include primary brain tumor, untreated or symptomatic brain metastases, currently receiving treatment in another investigational device or drug study, or less than 28 days since ending treatment on another investigational device or drug study, history of solid organ transplantation or major surgery within 28 days of study day 1, live vaccine therapy within 4 weeks prior to study day 1, and active infection requiring oral or IV therapy. The primary endpoints are incidence of dose-limiting toxicities and adverse events, MTD, and recommended phase 2 dose. Secondary objectives will evaluate PK parameters, preliminary antitumor activity (objective response, duration of response, progression-free survival, disease control rate, duration of stable disease, overall survival), and immunogenicity of AMG 256 via incidence of anti-AMG 256 antibodies. Results N/A Conclusions N/A Acknowledgements • The authors thank the investigators, patients, and study staff who are contributing to this study.• The study was sponsored and funded by Amgen Inc. • Medical writing support was provided by Christopher Nosala (Amgen Inc.). Trial Registration NCT04362748 Ethics Approval The study was approved by all institutional ethics boards. References Kluger HM, Tawbi HA, Ascierto ML, et al. Defining tumor resistance to PD-1 pathway blockade: recommendations from the first meeting of the SITC Immunotherapy Resistance Taskforce. J Immunother Cancer 2020;8:e000398. Thompson JA, Curti BD, Redman BG, et al. Phase I study of recombinant interleukin-21 in patients with metastatic melanoma and renal cell carcinoma. J Clin Oncol 2008;26:2034–2039. Lewis KE, Selby MJ, Masters G, et al. Interleukin-21 combined with PD-1 or CTLA-4 blockade enhances antitumor immunity in mouse tumor models. Oncoimmunology. 2017;7:e1377873. Shen S, Sckisel G, Sahoo A, et al. Engineered IL-21 cytokine muteins fused to anti-PD-1 antibodies can improve CD8+ T cell function and anti-tumor immunity. Front Immunol 2020;11:832.

Macroautophagy/autophagy is considered to play key roles in tumor cell evasion of therapy and establishment of metastases in breast cancer. High expression of LC3, a residual autophagy marker, in primary breast tumors has been associated with metastatic disease and poor outcome. FIP200/Atg17, a multi-functional pro-survival molecule required for autophagy, has been implicated in brain metastases in experimental models. However, expression of these proteins has not been examined in brain metastases from patients with breast cancer. In this retrospective study, specimens from 44 patients with brain metastases of infiltrating ductal carcinoma of the breast (IDC), unpaired samples from 52 patients with primary IDC (primary-BC) and 16 matched-paired samples were analyzed for LC3 puncta, expression of FIP200/Atg17, and p62 staining. LC3-puncta+ tumor cells and FIP200/Atg17 expression were detected in greater than 90% of brain metastases but there were considerable intra- and inter-tumor differences in expression levels. High numbers of LC3-puncta+ tumor cells in brain metastases correlated with a significantly shorter survival time in triple-negative breast cancer. FIP200/Atg17 protein levels were significantly higher in metastases that subsequently recurred following therapy. The percentages of LC3 puncta+ tumor cells and FIP200/Atg17 protein expression levels, but not mRNA levels, were significantly higher in metastases than primary-BC. Meta-analysis of gene expression datasets revealed a significant correlation between higher FIP200(RB1CC1)/Atg17 mRNA levels in primary-BC tumors and shorter disease-free survival. These results support assessments of precision medicine-guided targeting of autophagy in treatment of brain metastases in breast cancer patients.

Head and neck cancer (HNC) remains the sixth most common malignancy worldwide and survival upon recurrence and/or metastasis remains poor. HNSCC has traditionally been associated with alcohol and nicotine use, but more recently the Human Papilloma Virus (HPV) has emerged as a favorable prognostic risk factor for oropharyngeal HNSCC. However, further stratification with additional biomarkers to predict patient outcome continues to be essential. One candidate biomarker is the DEK oncogenic protein, which was previously detected in the urine of patients with bladder cancer and is known to be secreted by immune cells such as macrophages. Here, we investigated if DEK could be detected in human plasma and if DEK levels correlated with clinical and pathological variables of HNSCC. Plasma was separated from the peripheral blood of newly diagnosed, untreated HNSCC patients or age-matched normal healthy controls and analyzed for DEK protein using ELISA. Plasma concentrations of DEK protein were lower in p16-negative tumors compared to both normal controls and patients with p16-positive tumors. Patients with lower plasma concentrations of DEK were also more likely to have late stage tumors and a lower white blood cell count. Contrary to previously published work demonstrating a poor prognosis with high intratumoral DEK levels, we show for the first time that decreased concentrations of DEK in patient plasma correlates with poor prognostic factors, including HPV-negative status as determined by negative p16 expression and advanced tumor stage.

Introduction Checkpoint inhibitors are a promising therapy for patients with solid tumors; however, many patients require additional therapies to maximize clinical benefit or overcome resistance. The type-1 cytokine interleukin-21 (IL-21) is a promising candidate for combination and has shown clinical activity in melanoma and renal cell cancer. IL-21 has also shown improved efficacy when combined with anti-programmed death (PD)-1 antibodies in preclinical models. AMG 256 is a bifunctional fusion protein comprising a PD-1-targeting antibody and IL-21 mutein designed to deliver IL-21 pathway stimulation to PD-1+ cells—a strategy that is designed to prime and extend the activity of cytotoxic and memory T cells and induce anti-tumor immunity. This first-in-human (FIH) study will assess safety, tolerability, and estimated dosing of AMG 256 monotherapy in patients with advanced solid tumors. Methods This is a FIH, multicenter, non-randomized, open-label, phase 1 study (NCT04362748) of AMG 256 in patients with advanced solid tumors. The planned sample size is approximately 100 patients in two parts: part 1 will evaluate safety, tolerability, pharmacokinetics (PK), pharmacodynamics, and determine the maximum tolerated dose (MTD), part 2 will evaluate the MTD determined in part 1 to further characterize the safety profile and preliminary tumor response. AMG 256 will be delivered by intravenous (IV) infusion. Enrollment criteria include adults with life expectancy of > 3 months, ECOG performance status ≤ 2, histologically or cytologically confirmed metastatic or locally advanced solid tumors not amenable to curative treatment with surgery or radiation, and at least one measurable lesion ≥ 10 mm that has not undergone biopsy within 3 months of screening scan. Exclusion criteria include primary brain tumor, untreated or symptomatic brain metastases, currently receiving treatment in another investigational device or drug study, or less than 28 days since ending treatment on another investigational device or drug study, history of solid organ transplantation or major surgery within 28 days of study day 1, live vaccine therapy within 4 weeks prior to study day 1, and active infection requiring oral or IV therapy. The primary endpoints are incidence of dose-limiting toxicities and adverse events, MTD, and recommended phase 2 dose. Secondary objectives will evaluate PK parameters, preliminary antitumor activity (objective response, duration of response, progression-free survival, disease control rate, duration of stable disease, overall survival), and immunogenicity of AMG 256 via incidence of anti-AMG 256 antibodies. Citation Format: Greg Durm, Sophia Frentzas, Erik Rasmussen, Saltanat Najmi, Nooshin Hashemi Sadraei. Design and rationale of a phase 1 study evaluating AMG 256, a novel, targeted, PD-1 antibody x IL-21 mutein bifunctional fusion protein, in patients with advanced solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT205.

Head and neck squamous cell cancer (HNSCC) is the sixth most common malignancy worldwide, and despite advances in cytotoxic, surgical and radiation techniques, outcomes are still poor in those with both locally advanced and metastatic diseases. The need for development of better therapeutics along with a greater understanding of the relationship between the immune system and malignancies has led to a new therapeutic modality, immune modulators, particularly checkpoint inhibitors in HNSCC. It is now well recognized that HNSCC circumvents crucial pathways utilized by the immune system to escape surveillance. These hijacked pathways include impairing tumor antigen presentation machinery and co-opting checkpoint receptors. This understanding has led to the development of monoclonal antibodies targeting checkpoint receptors and has resulted in promising outcomes in HNSCC. This article describes the mechanisms that HNSCC utilizes to escape immune surveillance, clinical impact of checkpoint inhibitors (with a focus on pembrolizumab), ongoing studies, and future directions.

Approximately 5% of all cancer incidences result from human papillomavirus (HPV) infection. HPV infection most commonly leads to cancers of the anogenital region or oropharynx. It is unknown whether different HPV‐mediated cancers collectively share a molecular signature and it is important to determine if there are targetable alterations common to different types of HPV‐positive tumors. We analyzed 743 p53 wild‐type samples of anal, cervical, oropharyngeal, and vulvar squamous cell carcinomas which underwent multiplatform testing at a commercial molecular profiling service. Expression of 24 proteins was measured by immunohistochemistry (IHC), mutation of 48 genes was determined by next‐generation and Sanger sequencing, and copy number alteration for six genes was determined by in situ hybridization. The four cohorts had remarkably similar molecular profiles. No gene had a statistically significant difference in mutation frequency or copy number change between the four different types of squamous cell carcinomas. The only significant differences between cohorts were frequency of ERCC1 and SPARC loss as determined by IHC. In all four cancer types, oncogene mutation and PD‐L1 expression was relatively infrequent. The most commonly mutated gene was PIK3CA, with mutations most often affecting the helical domain of the protein and accompanied by concurrent lack of PTEN expression. Loss of MGMT and RRM1 was common among the four cohorts and may be predictive of response to cytotoxic therapies not currently being used to treat these cancer types. The similar molecular profiles of the four cohorts indicate that treatment strategies may be similarly efficacious across HPV‐positive cancers.

TPS5100 Background: NEPC is an aggressive cancer with poor prognosis. No standard treatment approach for NEPC exists and it remains an unmet need. NEPC is usually treatment-emergent, characterized by histological transformation from adenocarcinoma to a high-grade neuroendocrine tumor (NET), and may develop in 15%–20% of patients (pts) treated with standard prostate adenocarcinoma therapies, including novel hormonal therapies. The inhibitory Notch ligand, Delta-like ligand 3 (DLL3), is highly expressed on the surface of cancer cells, including NEPC cells, making it an attractive and a promising therapeutic target. AMG 757 is an HLE BiTE immuno-oncology therapy designed to redirect cytotoxic T cells to tumor cells by binding DLL3 on cancer cells and CD3 on T cells, resulting in T cell activation and expansion and T cell-dependent killing of tumor cells. AMG 757 showed in vitro activity in DLL3-expressing NETs, including NEPC. Preliminary results of an on-going first-in-human study suggest AMG 757 is safe and effective in pts with small cell lung cancer (NCT03319940), which prompted its study in NEPC. Methods: NCT04702737 is an open-label, phase 1b study evaluating AMG 757 infusion in pts with metastatic de novo or treatment-emergent NEPC, consisting of dose exploration and then dose expansion. Key eligibility criteria include adults (≥18 y) with NEPC whose disease progressed/recurred after ≥1 treatment course including a platinum-based regimen for de novo NEPC or an androgen signaling inhibitor, measurable disease per modified RECIST 1.1 per Prostate Cancer Working Group 3 modifications, ECOG performance status ≤2, life expectancy > 3 mo, adequate organ function, and no untreated/symptomatic brain metastases. Primary objectives are to evaluate safety and tolerability and determine the maximum tolerated dose or recommended phase 2 dose of AMG 757. Secondary objectives are to evaluate antitumor activity (ie, objective response, duration of response, progression-free survival, overall response) and characterize pharmacokinetics. The starting dose for dose exploration will be based on the dose deemed safe and tolerable in the ongoing trial of AMG 757 in SCLC. The study is open to enrollment. Clinical trial information: NCT04702737.

8510 Background: DLL3, an inhibitory Notch ligand, is a promising target as it is highly expressed in SCLC compared to normal tissue. AMG 757, a half-life extended BiTE immuno-oncology therapy, binds DLL3 on tumor cells and CD3 on T cells, leading to T cell-dependent killing of tumors. Results from the first nine dosing cohorts showing preliminary efficacy of AMG 757 (confirmed partial response [PR], 14% of pts) were previously presented. Here, updated safety, efficacy, and pharmacokinetic data from 10 cohorts from the ongoing phase 1 study of AMG 757 in SCLC are reported (NCT03319940). Methods: AMG 757 (0.003–100 mg) was administered IV every 2 weeks ± step dosing. Eligible patients (pts) had SCLC that progressed after ≥1 platinum-based regimen. Antitumor activity was assessed by modified RECIST 1.1. Results: As of 11 Jan 2021, 64 pts enrolled at 10 dose levels (DLs; 0.003–100 mg) had received ≥1 AMG 757 dose and were available for analyses. Median age was 64 (range, 32–80) y; 63 pts (98%) had ECOG PS 0–1 and median number of prior lines of therapy was 2 (range, 1–6), with 28 pts (44%) receiving prior PD-1/PD-L1 therapy. Median treatment duration was 6 (range, 0.1–71) wk. Treatment-related AEs occurred in 53 pts (83%): 16 (25%) ≥ grade (G) 3, 4 (6%) ≥G4, 1 (2%) G5 (pneumonitis; DL5 [0.3 mg]). AEs led to discontinuation in 1 pt (G3 encephalopathy, DL10 [100 mg]). Cytokine release syndrome (CRS; graded per Lee 2014 criteria) was reported in 27 pts (42%): G2 in 7 (11%), ≥G3 in 1 (2%). CRS presented mainly as fever (31%), tachycardia (17%), nausea (13%), fatigue (9%), and hypotension (9%). CRS was usually reversible and was managed with supportive care, corticosteroids, and/or anti-IL-6R. CRS did not lead to any treatment discontinuations. Sixty pts treated across 10 DLs, with a median follow-up of 4.2 (range, 0.2–18.6) mo, were evaluated for efficacy. Confirmed PR across all DLs was reported in 8/60 pts (13%), with 5/8 pts (63%) pts achieving unconfirmed PR at 100 mg (DL10). The median time to response was 1.7 (range, 1.2–3.7) mo. The estimated duration of response was >6 months in 71% pts (95% CI: 26, 92) with any PR. Disease control rate was 43%, with any tumor shrinkage in 23/60 pts (38%). AMG 757 serum exposures increased approximately dose proportionally within the evaluated dose range. Conclusions: AMG 757 has an acceptable safety profile at doses up to 100 mg. Responses were rapid and durable. Encouraging anti-tumor activity was seen across dose ranges, with ongoing unconfirmed PR in 5/8 pts (63%) at the highest DL. The study is ongoing; updated data, including response rates and duration of response, will be presented. Clinical trial information: NCT03319940.

Immune surveillance is well recognized as an important mechanism to prevent development or progression of head and neck cancers. Head and neck cancer cells can escape the immune system through multiple mechanisms including development of tolerance in T cells and inhibition of T-cell–related pathways, generally referred to as checkpoint inhibitors. This article highlights advances in immuno-oncology treatment approaches in recurrent and metastatic head and neck squamous cell carcinoma. Clinical trials are discussed in detail, with an emphasis on response dynamics, oncologic efficacy, safety, and tolerability of checkpoint inhibitors. In addition, developing concepts and ongoing studies in this setting are also reviewed.

Standard initial therapy for patients with pure and mixed anaplastic oligodendrogliomas (AO/MAO) includes chemotherapy and radiation therapy. Anaplastic oligodendrogliomas with 1p/19q co-deletion are more responsive to chemotherapy. There is concern for potential long-term CNS toxicity of radiation. Hence an approach using chemotherapy initially and reserving radiation for progressive disease is attractive. This multicenter phase II trial included patients with newly diagnosed AO/MAO with central pathology review and 1p/19q assay. Temozolomide was given 150 mg/m2 days 1–7 and 15–21, every 28 days for 8 cycles. The primary endpoint was progression free survival (PFS). Secondary endpoints included response rate, overall survival (OS), treatment toxicity and health-related quality of life (HRQL). Data from 62 patients enrolled between December 2001 and April 2007 at seven centers were analyzed. Among patients with measurable disease, 8 % achieved complete remission, 56 % had stable disease and 36 % had progression. The median PFS and OS were 27.2 months (95 % CI 11.9–36.3) and 105.8 months (95 % CI 51.5–N/A), respectively. Both 1p loss and 1p/19q co-deletion were positive prognostic factors for PFS (p

6037 Background: Metformin is a biguanide, widely used oral hypoglycemic agent. Metformin has also shown to inhibit tumor growth and progression in a wide variety of cancers including Head and Neck Squamous Cell Carcinoma (HNSCC). Metformin activates AMP protein kinase (AMPK) related pathways leading to inactivation of mammalian target of rapamycin (mTOR) and suppression of its downstream effectors. In addition, metformin is postulated to alter immune regulation in the tumor microenvironment leading to increased tumor cell killing. Here, we report our findings on the impact of metformin on T cells, NK cells and cytokines from patient peripheral blood mononuclear cells (PBMCs) from a phase I open-label single site dose escalation study combining metformin and chemoradiation (CRT) in HNSCC (NCT02325401). Methods: In this study, we evaluated the immune cell phenotypes and cytokine profiles of peripheral blood in patients before and after metformin treatment on trial by using flow cytometry and cytokine magnetic bead assays (Luminex). Cytokine profiles were further studied in co-culture experiments combining PBMCs, HNSCC cell lines, and metformin. Results: Patients who received metformin developed expanded NK cell populations, increased NKG2D expression, and a shift in their CD8+ T-cell memory phenotypes. Patient serum ELISA examination revealed increased anti-tumorigenic cytokine profiles. Metformin treatment of HNSCC cell lines in vitro as well as HNSCC PBMCs ex vivo resulted in downregulation of STAT3 compared to healthy controls. Downregulation of STAT3 may be a potential mechanism in which metformin stimulates NK cells. Conclusions: Here we show evidence that metformin treatment has a direct effect on the innate immune system in patients with HNSCC, inducing an anti-tumorigenic immune response suggesting that metformin continues to be a good candidate to yield improved clinical outcomes in patients with advanced stage HNSCC. Clinical trial information: NCT02325401.

Lessons Learned The combination of bevacizumab with docetaxel-gemcitabine resulted in unacceptable toxicity, particularly a high rate of pulmonary toxicity (30%). Despite promising efficacy, excessive toxicity of this regimen does not support its use in patients with advanced nonsquamous non-small cell lung cancer. Background Prior to immunotherapy, standard treatment for advanced non-small cell lung cancer (NSCLC) was platinum doublet chemotherapy. In a previous phase II study, docetaxel-gemcitabine demonstrated comparable efficacy and tolerability to platinum doublets. In this phase II trial, we evaluated the efficacy and tolerability of adding bevacizumab to docetaxel- gemcitabine in patients with advanced nonsquamous NSCLC. Methods Patients with untreated advanced nonsquamous NSCLC were treated with up to six cycles of docetaxel-gemcitabine-bevacizumab, followed by bevacizumab until progression. The primary endpoint for this study was 1-year progression-free survival (PFS); secondary endpoints were safety, overall response rate (ORR) and overall survival (OS). The planned sample size was 46 patients. Results A total of 13 patients were enrolled and received a median of six cycles of chemotherapy and four cycles of bevacizumab. The treatment was poorly tolerated, with five patients requiring dose reduction and four discontinuing treatment for toxicity. Grade 3–5 nonhematologic toxicity was seen in 10 patients, and 4 (30%) were hospitalized with pulmonary toxicity possibly related to study drugs. At this point, enrollment was halted for safety concerns. The 12-month PFS was 8%. In 11 evaluable patients, ORR was 72%, median PFS 6 months, and median OS was 11 months. Conclusion Docetaxel, gemcitabine, and bevacizumab at this dose and schedule resulted in excessive toxicity. Despite promising efficacy, in light of efficacious and safe alternative therapies, this regimen should not be used to treat advanced NSCLC.

Concurrent chemotherapy and radiation therapy remains the standard-of-care treatment in patients with unresectable stage III non-small-cell lung cancer. Most regimens include low doses of radiosensitizing agents. Because of concern for the presence of micrometastatic disease and the high rate of systemic failure, many trials have addressed the role of additional consolidation chemotherapy. Only a few of these studies have been performed in a randomized setting on a large number of patients, and the rest are smaller phase I and phase II trials that explore the safety and efficacy of different chemotherapy regimens. More recently, targeted agents have also been evaluated in such regimens, although molecular and histologic markers have not been fully incorporated in these studies. In this review, we discuss these trials and compare the different sequences and regimens of systemic doses of chemotherapy when delivered in addition to concurrent chemotherapy and radiation therapy.

The standard of care for the treatment of patients with advanced NSCLC includes 4-6 cycles of platinum-doublet chemotherapy with or without bevacizumab, with modest improvements in survival. To improve upon outcomes, recent studies have investigated the role of maintenance therapy after first-line chemotherapy. This concept can be divided into continuation and switch maintenance. The majority of studies have shown significant improvements in progression-free survival (PFS) with the addition of maintenance, but the improved PFS has not always resulted in an improvement of overall survival (OS). Two notable exceptions are erlotinib and, for non-squamous NSCLC, pemetrexed. For patients with non-squamous NSCLC who respond or remain stable after four cycles of platinum-doublet chemotherapy, either continuation of pemetrexed (if included in the induction regimen) or switch to pemetrexed as maintenance has been shown to improve OS compared with observation. Whether maintenance pemetrexed improves OS compared with treatment with pemetrexed at progression is unknown. Recent trials suggest that maintenance therapy benefits both patients with initial response and stable disease after chemotherapy. There is insufficient evidence to support recommending the combination of pemetrexed and bevacizumab over maintenance pemetrexed alone as a switch maintenance approach, although the combination seems to be more effective than bevacizumab alone. The ongoing ECOG 5508 trial is examining this question. For both squamous and non-squamous NSCLC, switch maintenance with erlotinib has been shown to improve both PFS and OS, although the improvement is modest. Switch maintenance with docetaxel or continuation maintenance with gemcitabine confers improvements in PFS regardless of histology but has failed to show improvements in OS. For this reason, switch maintenance with erlotinib can be considered in patients with squamous NSCLC. Overall, maintenance therapy may benefit patients with good performance status who complete four cycles of induction chemotherapy with manageable toxicity, but there is insufficient evidence to make this a blanket recommendation for everyone. Maintenance should remain an individual decision between patients and the treating oncologist.

6017Background: 2Despite aggressive adjuvant treatment, locally advanced HNSCC patients undergoing primary surgical resection with high risk [HR] features (extracapsular extension of lymph nodes (L...

6074Background: Up to 60% of HNSCC present as locally advanced disease (LAHNSCC). Although prognosis has improved significantly, 3 year PFS and OS remain at 62%, and 73% respectively (RTOG 0522) de...

Background: Head and neck squamous cell carcinoma (HNSCC) has traditionally been associated with alcohol and nicotine use, but more recently the Human Papilloma Virus (HPV) has emerged as a favorable prognostic risk factor for oropharyngeal HNSCC. However, further stratification with additional biomarkers to predict patient outcome continues to be essential. One candidate biomarker is the chromatin remodeling DEK protein, which is both an auto-antigen in autoimmune diseases and an oncogene in epithelial tissues. DEK is secreted by stimulated macrophages and neutrophils and was previously detected in the urine of patients with bladder cancer. Previously, we have reported that DEK mRNA and protein is upregulated in HNC tumor tissue and higher DEK levels are associated with poor prognoses in many types of solid tumors. We hypothesized that DEK could be detected in the plasma of HNC patients, either due to secretion from the tumor or as part of the antitumor immune response, and therefore may be a biomarker for disease status. Methods: We recruited 38 newly diagnosed HNSCC patients and 37 age-matched normal healthy controls into the study. Plasma isolated from peripheral blood was subjected to DEK specific ELISA and DEK concentration levels were compared to levels found in normal controls, and to clinical and pathological variables. Results: We show for the first time that DEK can be detected in human plasma. We did not find an association between DEK plasma concentrations and variables including sex, age, race, or drinking/smoking status. However, we detected decreased concentrations of DEK in HNC patients with p16-negative disease and in patients with larger tumor sizes, indicating an association between DEK levels and known prognostic markers. In addition, HNC patients with lower DEK concentrations had a decreased white blood cell count, largely due to differences in lymphocyte and eosinophil counts. This direct association between plasma DEK levels and white blood cell count was independent of p16 status. Conclusions: Together, the data suggest that lower levels of DEK in HNC patient plasma may be predictive of poor outcome. This is in direct contrast to what is observed with intratumoral levels of DEK protein, in which higher levels of DEK expression are an independent factor predicting poor prognosis. Future studies will investigate the role that secreted DEK, such as that found in the plasma, may have in the antitumor immune response. Citation Format: Trisha Wise-Draper, Arun Sendilnathan, Sarah Palackdharry, Nicholas Pease, Julianne Qualtieri, Randy Butler, Nooshin Hashemi Sadraei, John C. Morris, Yash Patil, Keith Wilson, Jonathan Mark, Keith Casper, Vinita Takiar, Adam Lane, Lisa M. Privette Vinnedge. Plasma concentrations of the DEK oncogene correlate with pathological variables in a case-control study of patients with HNSCC [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; April 23-25, 2017; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(23_Suppl):Abstract nr 50.

e23083 Background: Approximately 5% of all cancers result from HPV, commonly leading to head and neck (HN) and anogenital (AG) cancers. Immune therapy is used in many cancers including HN. Positive PDL1 and high tumor mutation load (TML) have been proposed as potential markers of immunogenicity among many tumors, but little is known specifically among HPV driven cancers. Methods: A retrospective analysis of patients with HPV related cancers was performed based on tissue availability. TP53 wild type status was used as a surrogate for HPV positivity. Among oropharynx cases with TP53 wt, p16 testing was performed for confirmation. Next generation Sequencing (NGS Sanger) was used for tumor mutation load (TML) calculation. High TML cut off was defined as ≥17 mutations/megabase based on prior publications. PDL1 status was reported based on IHC ≥5%. A 2-tail Fisher’s exact test was utilized for statistical analysis. Results: A total of 737 patients with HN and AG (cervical, anal, vulvar, penile) squamous cell carcinoma were identified. Of these, 251 had complete 592 gene panel NGS data. HPV + TP53 Wt disease was detected in oropharynx (70%), cervical (88%), and anal (82%) cancer. HPV+ HNSCC had a trend to a higher PDL1 compared to HPV neg (50% vs 31%, p = 0.16) but there were no significant differences among PDL1 between HPV+ and neg patients across the entire combined disease cohorts (41% vs 36%, p = 0.56). TML-high patients showed a trend to increased PDL1 positivity (60% vs 38%, p = 0.09). However, high TML was less common in HPV+ HNSCC vs HPV neg (2.2 vs. % 9.7% p = 0.19). Unlike some other malignancies, high TML was uncommon across HPV+ tumors in general (8%). Conclusions: Despite fewer TML high cases among HPV + patients, there was no difference among PDL1 expression. It is possible that a lower threshold may need to be used when determining TML high vs. low in viral-driven cancers. It is also possible that in view of viral immunogenicity, certain genomic alterations contribute more to higher PDL1 expression rather than the sum of total genomic alterations. We are currently pursuing both lines of investigation. These findings are hypothesis generating for future trial design rather than treatment decision-making, and need to be confirmed in large prospective cohorts.

e18139Background: Despite improved survival outcomes, HNC pts suffer significant impairment in QOL as a result of disease and treatment toxicity. We evaluated the association between QOL and oncolo...

TPS6109Background: Diabetic patients with head and neck squamous cell carcinoma (HNSCC) receiving the oral hypoglycemic, metformin, demonstrated improved survival compared to those not on metformin...

Lung cancer is the second most commonly occurring non-cutaneous cancer in the United States with the highest mortality rate among both men and women. In this study, we utilized three lung cancer microarray datasets generated by previous researchers to identify differentially expressed genes, altered signaling pathways, and assess the involvement of Hedgehog (Hh) pathway. The three datasets contain the expression levels of tens of thousands genes in normal lung tissues and squamous cell lung carcinoma. The datasets were combined and analyzed. The dysregulated genes and altered signaling pathways were identified using statistical methods. We then performed Fisher's exact test on the significance of the association of Hh pathway downstream genes and squamous cell lung carcinoma. 395 genes were found commonly differentially expressed in squamous cell lung carcinoma. The genes encoding fibrous structural protein keratins and cell cycle dependent genes encoding cyclin-dependent kinases were significantly up-regulated while the ones encoding LIM domains were down. Over 100 signaling pathways were implicated in squamous cell lung carcinoma, including cell cycle regulation pathway, p53 tumor-suppressor pathway, IL-8 signaling, Wnt-β-catenin pathway, mTOR signaling and EGF signaling. In addition, 37 out of 223 downstream molecules of Hh pathway were altered. The P-value from the Fisher's exact test indicates that Hh signaling is implicated in squamous cell lung carcinoma. Numerous genes were altered and multiple pathways were dysfunctional in squamous cell lung carcinoma. Many of the altered genes have been implicated in different types of carcinoma while some are organ-specific. Hh signaling is implicated in squamous cell lung cancer, opening the door for exploring new cancer therapeutic treatment using GLI antagonist GANT 61.

11107 Background: Several distinct cancers are caused by human papillomavirus (HPV)- including squamous cell carcinomas of the cervix (CSCC), anal canal (ASCC), and oropharynx (OSCC). Importantly, ...

e19043 Background: Pemetrexed (Pem) has been widely used in non-small cell lung cancer (NSCLC), but data of biomarkers predicting its clinical response and outcome is very limited. Thymidylate synt...

e18513 Lymphopenia as the predictor of survival in chemo radiation (CRT) treated stage III non-small cell lung cancer (NSCLC) Background: Despite potential curative CRT treatment for locally advanc...

e17088 Background: HNC pts suffer from symptoms of their disease and side effects of standard treatments that can significantly impact their QOL. There is paucity of data regarding impact of sympto...

6036 Background: Almost all survivors of FA, an inherited DNA repair deficiency syndrome, will develop H&N Ca. Preclinical data in H&N Ca cells harboring FA mut show increased sensitivity to chemo ...

e19025 Background: Pemetrexed (Pem) is the first agent showing different efficacy based on histology in the treatment (tx) of NSCLC. Our goal was to identify outcome predictors in patients (pts) with metastatic NSCLC (mNSCLC) treated with pem. Methods: Retrospective data of pts with mNSCLC who received pem were analyzed. Variables included demographics, ECOG performance status (PS), disease sites, pre/post-pem tx, and toxicities. Clinical benefit defined as complete response (CR), partial response (PR), and stable disease (SD) >6 months, progression-free survival (PFS), and overall survival (OS) were analyzed using logistic regression and Cox proportional hazards model. Results: 240 pts were included. 55% male, 84% smokers. 68% adenocarcinomas, 10% squamous/adenosquamous carcinomas. Median age 64 years (range 34-84). Pem was given for a median of 4 cycles (range 1-73), as 1st line in 20% and 2nd line in 50%, given as a single agent in 69%. The most common toxicities were constitutional (50%) and GI related (29%). 31% of pts achieved CR (n=4) or PR, 33% progressed, 36% had SD. Front-line pem use (p=.0003), adenocarcinoma histology (p=.05), and non-pulmonic metastatic sites ≤2 (p=.01) were independent predictors of clinical benefit, with response rate of 71% for pts with all three features, compared to only 7% for pts with none of these features. Multivariable analysis of survival data revealed ECOG PS >1, non-pulmonic metastatic sites>2, and squamous/poorly differentiated histology predicted poor PFS and OS. In addition, interval from diagnosis to start of pem 2 non-pulmonic metastatic sites, squamous/poorly differentiated histology predict poor PFS and OS. An analysis of biomarkers is ongoing.

e19012 Background: Bev in combination with chemotherapy (CT) is a standard treatment for pts with stage IV NSCLC. However, there are no established biomarkers to improve benefit-to-risk profile of therapy. CEC are a marker of vascular turnover. We hypothesized CEC may be associated with treatment outcome/toxicity. Methods: Stage IV NSCLC pts were treated with Bev along with CT of choice per clinician. CEC were measured at baseline and after 2 cycles of Bev-containing therapy. CellSearch (Veridex, New Jersey) was used to capture and quantify CEC via immunomagnetic and immunofluorescence techniques. CEC were enumerated as nucleated, CD146+/CD105+/CD45- cells in 4 mL of blood. For comparison, baseline CEC were also collected in 3 other groups; healthy subjects, stage I-III NSCLC, and advanced solid tumor pts. Chi-square tests and non-parametric methods were used to assess associations between CEC and pt/disease characteristics, toxicity, and proportional hazards models were used for comparisons of progression free and overall survival (PFS and OS). Results: Evaluated were 62 individuals: 29 stage IV NSCLC, 10 stage I-III NSCLC, 13 advanced solid tumors (melanoma, sarcoma, renal and adrenal cancers), and 10 healthy subjects. The median of CEC/ml blood in healthy subjects was 6. Based on its distribution, CEC >13/ml was deemed elevated. Elevated CEC was seen in 48% of stage IV NSCLC (median 12/ml), in comparison to 20% of stage I-III NSCLC (8/ml) and 23% of advanced solid tumors (6/ml) Baseline CEC in stage IV NSCLC was independent of patient/ disease characteristics including site of disease and tumor size. CEC increased during treatment in most patients (61%). CEC doubling between baseline and cycle2 was associated with more cytopenia and hemorrhage. (75% vs 20%, p=0.05 and p=0.03 for absolute and relative CEC changes). Baseline CEC and changes during treatment did not correlate with response (p≥ 0.32) and pts with elevated baseline CEC had a trend towards worse PFS (p=0.09) and OS (p=0.10). Conclusions: CEC are commonly elevated in advanced NSCLC. CEC increases may be predictive of adverse events from Bev-CT and when elevated at baseline may suggest worse PFS.