PURPOSE:STAT3 is important in promoting a pro-oncogenic tumor microenvironment. This first randomized trial of a STAT3-targeting ASO with pembrolizumab assessed whether this first-line combination would improve outcomes compared with pembrolizumab alone in recurrent/metastatic (R/M) head and neck squamous cell carcinoma (HNSCC). PATIENTS AND METHODS:This multicenter, randomized, open-label, controlled study (NCT05814666) enrolled patients with previously untreated R/M-HNSCC with positive PD-L1 expression (combined positive score [CPS] ≥1). Patients were stratified by CPS (1-19; ≥20) and randomized 2:1 to danvatirsen 3mg/kg IV weekly (with 2 loading doses in week 1) plus pembrolizumab 200mg IV every 3 weeks or pembrolizumab alone. Patients received 21-day treatment cycles until progressive disease, unacceptable toxicity, or voluntary withdrawal. Primary endpoint was overall response rate (ORR: partial + complete response per RECIST-v1.1). RESULTS:69 patients were randomized and 66 treated (51 [77.3%] male), 45 with danvatirsen + pembrolizumab, 21 with pembrolizumab alone. Enrolment was stopped due to lack of response in the CPS 1-19 cohort after ≥1 scan (~6 weeks of treatment) and in the CPS ≥20 cohort after ≥2 scans (~12 weeks). In the final analysis, ORRs were 15.6% and 14.3%, respectively (risk difference 1.3, P=1.0). The treatment-emergent adverse-event rate was increased in the combination arm (97.8% vs 85.7%), and the combination side effect profile was similar to previous danvatirsen studies. CONCLUSIONS:Combination danvatirsen + pembrolizumab did not improve ORR compared to pembrolizumab alone in first-line treatment of R/M-HNSCC. Further exploration of STAT3-targeted approaches is needed within the context of immunotherapy.

BACKGROUND:Human papillomavirus-positive oropharyngeal squamous cell carcinoma (HPV+ OPC) patients typically exhibit reduced rates of second primary malignancies (SPMs) compared to HPV-negative head and neck cancer patients. While HPV+ OPC patients may be predisposed to SPMs in HPV-associated anatomical sites, the metachronous presentation of an HPV+ OPC and subsequent glioblastoma multiforme (GBM) has not been previously documented. CASES:We present two cases of HPV+ OPC patients who developed GBMs within a short period after definitive therapy. METHODS:Comprehensive whole exome sequencing (WES) was performed on paired GBM, oropharyngeal, and matched normal tissues from two HPV+ OPC patients who developed GBMs within a short period after definitive therapy, with the goal of identifying shared somatic and germline variants. Bioinformatic analyses included variant calling, annotation, and pathway enrichment. RESULTS:WES revealed a shared CEP104 missense mutation among both oropharyngeal tumors as well as a potential KIR2DL4 germline variant in the second patient, suggesting a possible role for disrupted NK cell immunity in driving these cancers. CONCLUSION:Although these cases were likely random events, they illustrate the diagnostic and therapeutic challenges of GBM following HPV+ OPC and underscore the importance of personalized genomic assessment in HPV+ OPC survivors, who may develop non-head and neck SPMs unrelated to field cancerization.

Supplementary Figure S1: Case examples of false negative TTMV-HPV DNA results identified during surveillance

6089 Background: De-escalation therapy for HPVOPC remains undefined and controversial. Definitive and postoperative chemoradiation are associated with significant toxicity. Efforts to de-intensify treatment with CRT only, such as HN002 and HN005, were unsuccessful. A previous investigation by this group (SIRS 1.0) and E3311 have highlighted the potential value of TORS with pathological risk stratification of patients for de-escalated adjuvant therapy. Our early work demonstrated that TORS, in combination with reduced-dosed adjuvant therapy for early and intermediate HPVOPC, resulted in equivalent progression-free survival (PFS) and overall survival (OS) while reducing toxicity in the first two years of treatment. Here we report the 5-year results for SIRS. Methods: This is a nonrandomized phase II trial for early-stage molecularly confirmed HPVOPC patients (n=63) treated with TORS followed by reduced-dose adjuvant therapy based on pathological risk-stratification into one of three groups: Group 1 (n=31) with no adjuvant therapy, Group 2 (n=15) with 50-Gy radiotherapy, and Group 3 (n=17) with 56-Gy chemoradiotherapy concurrent with weekly cisplatin. Patient demographics, baseline tumor characteristics, clinical outcomes, and adverse events during treatment and surveillance were recorded across the full 5-year study period. Results: Among the 63 total patients, median follow-up is 58 months (IQR 43—76 months). In the first two years following TORS, 5 patients experienced locoregional recurrence (7.9%); of these, one later had distant metastasis and one had a second HPV+ recurrence in the ipsilateral neck. All were salvaged by TORS, neck dissection, and/or chemoradiation. No patients had an HPVOPC recurrence between years 2 to 5 of follow-up. Two patients, one each from Group 1 and Group 2, developed a molecularly proven HPV+ contralateral second primary tumor at the tonsils approximately 5 years following TORS. Both patients were successfully salvaged and remain disease-free. Two patients (one each in Group 2 and 3) died from causes unrelated to cancer. The five-year OS is 96.8% (61/63) and the disease-specific survival (DSS) is 100% (63/63). Five-year PFS is 87.1% (27/31) for Group 1, 86.7% (13/15) for Group 2, 94.1% (16/17) for Group 3, and 88.9% (56/63) for the full cohort. Conclusions: Long-term follow-up of SIRS demonstrates de-escalation TORS and pathologic risk stratification is safe and effective in molecularly proven HPVOPC and reduces the lethal and morbid long-term side effects of full dose radiation and CRT. All recurrences occurred in the first two years postoperatively and were salvaged. Multi-disciplinary decision-making utilizes the benefits of each specialty to optimize outcomes in de-escalation. Clinical trial information: NCT02072148 .

Importance:Despite the favorable prognosis for HPV-positive oropharyngeal squamous cell carcinoma (HPV+ OPSCC), efforts to de-escalate treatment intensity, while maintaining low recurrence and mortality rates, have proven challenging. Identifying appropriate prognostic factors remains elusive; however, the association of pretreatment circulating tumor tissue viral-modified HPV (TTMV-HPV) DNA level with known characteristics of disease burden-clinical staging, characteristics of pretreatment imaging, and aggressive histopathologic features of surgical specimen-may offer insights that could shift treatment paradigms for HPV+ OPSCC. Objective:To investigate the association of pretreatment TTMV-HPV DNA levels with clinical, radiologic, histopathologic, and outcome metrics in patients with HPV+ OPSCC. Design, Setting, and Participants:This cohort study of patients with HPV+ OPSCC and positive test results for pretreatment TTMV-HPV DNA fragment levels used data from a single tertiary center from April 2020 to September 2023. TTMV-HPV DNA fragments levels were categorized into 3 cohorts: low (≤99 fragments/mL), moderate (100-999/mL), and high (≥1000/mL). Main Outcomes and Measures:Association of clinical tumor (cT) and nodal (cN) staging with TTMV-HPV DNA fragment level. Secondary outcomes included the association between TTMV-HPV DNA fragment level and positive emission tomography-computed tomography (PET-CT) characteristics as well as histopathologic features of surgical specimen. The association of pretreatment fragment level with receiving adjuvant therapy for surgical patients was also analyzed. Recurrence-free survival and disease-specific survival were also assessed. Results:The study population included 203 patients (mean [SD] age, 62 [10] years; 24 [12%] females and 179 males [88%]), 58 (29%) of whom were in the low, 73 (36%) in the moderate, and 72 (35%) in the high TTMV-HPV DNA fragment-level cohort. Compared to patients with cT0/1 stage, those with cT2 stage and cT3/4 stage had increased odds of higher TTMV-HPV DNA levels, with adjusted odds ratios (aORs) of 2.33 (95% CI, 1.24-4.46) and 2.51 (95% CI, 1.17-5.46), respectively. Compared to patients with cN0 stage, those with cN1 stage and cN2/3 stage also had increased odds of higher TTMV-HPV DNA levels, with aORs of 4.26 (95% CI, 1.82-10.34) and 3.64 (95% CI, 1.46-9.36), respectively. In adjusted analysis of pretreatment PET-CT characteristics, total primary tumor plus nodal volume was associated with higher TTMV-HPV DNA levels, with an aOR of 1.04 (95% CI, 1.02-1.07). Among 94 surgical patients, no significant association was found between pretreatment fragment level and lymphovascular invasion, perineural invasion, pathologic T stage, number of positive nodes, or extranodal extension on pathological analysis of surgical specimen. No significant differences in recurrence-free survival or disease-specific survival were found. Conclusion and Relevance:This cohort study found that higher pretreatment TTMV-HPV DNA fragment levels were associated with more advanced clinical staging and higher aggregate primary and cervical nodal volume on PET-CT results. Future studies are needed to explore how pretreatment fragment level may influence treatment decisions.

SIGNIFICANCE:The PGV001 platform is feasible, safe, and immunogenic. The OpenVax pipeline predicted immunogenic neoantigens in tumors with wide-ranging mutational burdens. Data from this study prompted three additional PGV001 trials, one in newly diagnosed glioblastoma, one in urothelial cancer in combination with an ICI, and another in prostate cancer.

e18116 Background: To enhance patients' quality of life and prolong their survival, exploring an effective, low-toxicity version of the induction regimen for LA SCCHN is crucial. The addition of immunotherapy to cisplatin and docetaxel (TP) is a less toxic and potentially more effective alternative to the historic chemotherapy induction regimens. Methods: Patients with LA SCCHN were enrolled in a phase 1, NCT05376553, non-randomized study with 2 cohorts. Cohort A received 100 mg/m² cisplatin and 75 mg/m² docetaxel IV on day 1 of each cycle, followed by 350 mg cemiplimab on day 14 for 3 cycles. Cohort B received one additional dose of cemiplimab on D-7. Both cohorts underwent concurrent chemoradiation (CRT) or surgical resection +/- CRT as per standard care. All patients were to receive adjuvant cemiplimab 350 mg every 3 weeks for 8 cycles. With no dose escalation, cemiplimab was given with TP using a standard 3+3 dosing scheme. Dose-limiting toxicity (DLT) was evaluated in the first cycle for the initial 6 patients in each scheduling cohort. Only DLTs related to cemiplimab were considered, excluding known chemotherapy-related adverse events. A total of 24 previously untreated stage IV LA SCCHN patients were enrolled, 21 men and 3 women. Tumors were in the oropharynx (15, HPV + 9/15), larynx (2), oral cavity (3), nasopharynx (1, HPV +), and hypopharynx (3). Results: All 24 eligible patients completed ICI and 17 of 24 patients have completed the study. 7 are still receiving treatment, and 1 death occurred before study drug was given and was not included in the assessment. 16/17 patients who completed the study remain in remission. One patient progressed systemically in the lungs 4 months after radiation, and is undergoing systemic therapy. No DTLs were observed during ICI in all 24 treated patients. Out of the 24 patients that have completed ICI the overall response rate (ORR) to ICI was 87.5% with 12 partial responses, 9 complete responses, 2 stable disease, and 1 progression of disease. The disease control rate is 95.8%, with a relapse rate of 4.2% (1/24). 2/3 oral cancer patients underwent resection, 1 had complete pathological response, 1 with 85% pathological tumor necrosis and 1 pending surgical resection. Median follow-up was 18.06 months (8-29). A full assessment of toxicity with CRT and adjuvant cemiplimab will take place when study treatments are complete. Conclusions: The combination of cemiplimab given with TP appears to be safe and well tolerated in both cohorts. These clinically meaningful findings with high ORR and only one systemic relapse supports the promising role of cemiplimab based ICI for LA SCCHN and may have the potential to be practice-changing if confirmed in larger studies. Clinical trial information: NCT05376553 .

e18057 Background: Tumor tissue-modified viral (TTMV) HPV DNA is a sensitive, highly specific biomarker for human papillomavirus-positive (HPV+) head and neck squamous cell carcinoma (HNSCC). However, TTMV clearance patterns during induction chemotherapy (IC) or chemoimmunotherapy (ICI) have not been well described.We monitored the kinetics of TTMV responses to IC(I) and analyzed the results for predictive implications for treatment outcomes. Methods: This is a retrospective cohort study of patients treated at a single institution between June 2020 and December 2024. Adult patients with previously untreated, locally advanced, unresectable molecularly identified HPV+ HNSCC with high-risk features (e.g. extracapsular extension, T4, or >N2c) treated with IC(I) were identified. Patientswere treated with 3 cycles of IC (docetaxel-cisplatin-fluorouracil) or anti-PD1 ICI (docetaxel-cisplatin-Cemiplimab or docetaxel-cisplatin-Pembrolizumab). Depending on clinical trial enrollment, patients were followed by standard- or reduced-dose CRT alone or with adjuvant therapy. TTMV testing was done pre-treatment, after each cycle and CRT, and throughout surveillance and analyzed retrospectively. Patients were required to have a positive pretreatment TTMV test. TTMV clearance kinetics throughout IC and CRT with post cycle TTMV testing were correlated with clinical outcomes during post-treatment surveillance and TTMV testing during surveillance was monitored and is reported. Results: Twenty-eight subjects (26 male, 13 smokers) had a median pre-treatment TTMV of 1321 (range 27-90770, IQR 200-3217). Patients had a mean 33.7 months of follow-up and a mean 10.1 TTMV tests over the study period. 10/28 patients reached full TTMV clearance after one IC cycle (IC1), 8/28 after two cycles (IC2), 3/28 after three cycles (IC3), and 1/28 remained TTMV+ throughout IC and cleared after CRT. 6/28 had unknown kinetics due to missing data. All patients were TTMV-negative following completion of CRT. During surveillance, 6/28 patients developed recurrences and all were both TTMV+ and eventually PET+ (1 locoregional, 5 metastatic); of these, 2/6 had reached TTMV clearance after IC2, 2/6 after IC3, 1/6 after CRT, and 1/6 unknown. Rapid clearance after IC1 was associated with having no evidence of disease (p=0.046) and lower baseline TTMV (p=0.049). TTMV detection of recurrence had a mean lead-time of 115 days (range 0-340) over PET detection. Conclusions: Recurrences occurred in patients who failed to clear plasma TTMV after one IC(I) cycle. Delayed TTMV clearance strongly predicted systemic failure and may help identify patients requiring more systemic therapy. Future studies should validate these findings in larger cohorts.

e18056 Background: This is a retrospective study of tumor-modified HPV DNA (TTMV) as a tool for post-treatment surveillance in patients with HPV+ head and neck squamous cancer (HPV+HNC). HPV+HNC, has a better prognosis and lower recurrence rates than smoking and alcohol-related cancers. Approximately 80% of recurrences occur within the first 2 years post-treatment. Early identification of recurrence is likely to improve the outcomes of salvage therapies in this disease. TTMV is a sensitive and specific biomarker for HPV+HNC. This study aims to inform best practices for TTMV use in surveillance to enhance early recurrence detection and timely intervention. Methods: The study analyzed 54 HPV+HNC patients treated for cure with induction chemotherapy or chemoimmunotherapy (IC) followed by chemoradiotherapy (CRT) (30 patients) or CRT alone (24 patients). All patients were confirmed HPV+ by molecular pathology and had a +TTMV prior to treatment. TTMV was performed during and after treatment; routine follow-up assessments were done using physical exam (PE), imaging (PET), and biopsy (bx). TTMV results were compared to traditional surveillance methods for prediction of recurrence. Results: Post-treatment TTMV status: 53 out of 54 patients had an initial -TTMV post-treatment. Recurrence detection: With a mean follow-up of 25 months (3.5–51 mo). 12 out of 54 patients showed signs of recurrence or persistent disease, identified by +TTMV, imaging, symptoms, and/or PE. Specifically: Group 1: 4/12 patients had positive imaging, symptoms and/or PE and -TTMV. All 4 patients remain disease-free (NED) after further investigation, a median of 23.5 months (4-34 mo) post findings. Group 2: 7/12 patients had a + post treatment TTMV with a median of 3 months (1-9 mo) from treatment end. 6 /7 (87.5%) had confirmed HPV+ disease via biopsy . 3/7 (43%) patients with + post-treatment TTMV had initial positive imaging. 4/7 (57%) with +TTMV and negative initial imaging developed positive imaging with amedian time of 3 months (2-11 mo) after +TTMV. One patient with a positive PET scan 11 months after the first +TTMV has a biopsy pending. Group 3: 1/12 (8.33%) patient with a -TTMV developed HPV+ metastases by + PET and + bx 9 months post therapy. Conclusions: A positive TTMV post-treatment was 100% predictive of recurrence, demonstrating its strong utility in early detection. TTMV can guide the timing of imaging as TTMV + recurrence may not be immediately detectable on scans. TTMV kinetics can be used alongside PE and imaging to improve surveillance accuracy. While TTMV cannot replace imaging, due to its inability to confirm disease location or burden, it can help guide decision-making which is especially useful when interpreting complex post-treatment changes in the head and neck region.TTMV could be efficiently incorporated into surveillance guidelines for HPV+ HNC, as a complementary tool for detecting recurrence and justifying imaging in resource-limited settings.

Patients with head and neck cancer often present with complex challenges due to a substantial comorbidity burden, including substance use disorders, and the tumor's location in regions that are both cosmetically and anatomically sensitive. These challenges can be categorized into 6 areas, that is, overall health (eg, performance status, biological age), physiological life stages (eg, aging), organ dysfunctions (including autoimmune comorbidities, organ transplants, and psychiatric disorders), previous and concurrent malignancies, previous and current therapies, and adherence to therapy. We provide a practical guide to help physicians understand and address the phenotypic multitude of potential complications in the management of these patients. The process has 4 main phases involving identification of the clinical challenge, understanding the reasons for ineligibility (contraindications), assessment of the risk to benefit ratio, and finally making informed decisions about systemic treatment. Proactive interventions, including prehabilitation, are crucial for optimizing patient outcomes and reversing some ineligibility issues. The evidence supporting contraindications is drawn from both clinical trials and real-world data, each with its strengths and limitations. These contraindications are applied as absolute or relative and further refined by expert opinions and consensus statements. There are 2 main reasons for ineligibility for a given treatment, absence of supporting data or negative outcome data. In these cases, careful interpretation using all levels of clinical evidence, including extrapolation and preclinical rationale, is essential. By mastering these skills, that may in the future be enhanced by artificial intelligence methods, significant advancements in patient care can be achieved.

OBJECTIVE:Circulating tumor DNA (ctDNA) has emerged as a promising tool in the treatment of HPV-associated oropharyngeal squamous cell cancer (OPSCC). This systematic review sought to answer the question: what is the current role of ctDNA in the diagnosis, treatment, and surveillance of HPV-associated OPSCC? DATA SOURCES:Medline (Ovid), Embase (Ovid), Scopus. REVIEW METHODS:Original articles studying the role of ctDNA in the diagnosis or surveillance of HPV-associated OPSCC were eligible for inclusion. Two authors independently reviewed studies for inclusion and abstracted data, including study design, characterization of liquid biopsy technology, and diagnostic outcomes. RESULTS:After a preliminary screening of 441 studies, 23 were selected for inclusion. Ten studies were conducted retrospectively, and 13 were conducted prospectively. In these studies, diagnostic testing included plasma-based droplet digital polymerase chain reaction (ddPCR, n = 13), quantitative PCR (qPCR, n = 4), digital PCR (dPCR, n = 3), next-generation sequencing (NGS) (n = 3), or a ctDNA detection kit (n = 1). Diagnostic outcomes were reported for pre-diagnosis (n = 1), pre-treatment (n = 17), during treatment (n = 6), and surveillance/recurrence (n = 11) timepoints. Test sensitivities ranged from 20.6 %-100 % pre-treatment and 72 %-100 % during surveillance, while test specificities ranged from 95 %-100 % pre-treatment and 87.2 %-100 % during surveillance. CONCLUSION:The majority of studied ctDNA technologies allow for detection of HPV-associated OPSCC with high diagnostic accuracy. However, heterogeneity is introduced by test type and assay used. These findings highlight the utility, as well as limitations, of ctDNA in the diagnosis, treatment monitoring, and surveillance of HPV-associated OPSCC. Future studies and clinical consensus will need to address acceptable diagnostic accuracy thresholds for clinical use.

INSM1 has rapidly emerged as a robust marker for neuroendocrine (NE) differentiation, prompting the use of INSM1 as a standalone marker for NE differentiation in various sites. INSM1 staining could be an especially practical tool for evaluating HPV-associated oropharyngeal squamous cell carcinoma (HPV-OPSCCs) because high-grade NE transformation may be exceedingly difficult to recognize on morphologic grounds alone, even though it portends highly aggressive clinical behavior. The purpose of this study was to determine the incidence of INSM1 expression in HPV-OPSCC and study its role in determining NE transformation and clinical aggressiveness. Forty-six consecutive nonselected HPV-OPSCCs were stained with INSM1 and synaptophysin. In addition, 2 clinically matched cohorts of aggressive (n=28) versus nonaggressive (n=26) HPV-OPSCCs were retrospectively stained with the INSM1 and synaptophysin to determine the prognostic significance of INSM1 staining. In all, 42% of HPV-OPSCC showed positive INSM1 staining, but synaptophysin was not co-expressed in even a single case. In selected cases based on clinical behavior, INSM1 staining was observed more frequently in nonaggressive than aggressive tumors (50.0% vs. 21.4%, P=0.03). INSM1 expression is a common finding in HPV-OPSCC and is usually not linked with true NE transformation or aggressive behavior. Indeed, INSM1 expression is more commonly observed in nonaggressive cancers. Reliance on INSM1 staining alone in recognizing NE differentiation in HPV-OPSCC could be calamitous.

BACKGROUND:Peltopepimut-S is a therapeutic vaccine, which induces specific expansion of both CD4+helper and CD8+cytotoxic T-cells against human papillomavirus type 16 (HPV16) E6/E7 oncoproteins. PATIENTS AND METHODS:In a randomized phase 2 trial, we evaluated the efficacy and safety of peltopepimut-S plus cemiplimab compared with cemiplimab alone as first-line or second-line therapy in recurrent/metastatic HPV16-positive head and neck cancer. The primary efficacy endpoint was the objective response rate (ORR) by an independent review (Response Evaluation Criteria in Solid Tumors version 1.1, RECIST v1.1), while the primary safety endpoint was frequency and severity of adverse events. Secondary endpoints included progression-free survival (PFS) and overall survival (OS). RESULTS:Overall, 198 anti-programmed cell death protein-1 therapy-naïve patients with confirmed HPV16-positive recurrent/metastatic oropharyngeal cancer were randomized to receive cemiplimab plus peltopepimut-S (n=100) or placebo (n=99). The trial did not meet its primary objective (ORR, 25.3% in peltopepimut-S arm vs 22.9% in placebo arm; p=0.735). The median OS (mOS) and PFS in the placebo arm were unexpectedly longer than in the peltopepimut-S arm (26.9 vs 15.8 months) and (20.3 vs 5.5 months), respectively. In predefined exploratory analyses this was associated with an excess death rate from progressive disease in patients with pre-treatment programmed death-ligand 1 (PD-L1) combined positive score (CPS) <20. In contrast, patients with CPS ≥20 had a higher ORR of 51.7% (95% CI, 32.5% to 70.6%) vs 25.8% (95% CI, 11.9% to 44.6%) and a longer mOS in the peltopepimut-S arm (34.8 vs 28.8 months) compared with the placebo arm, respectively. If patients had received all three vaccine or placebo doses, the ORR in patients with CPS ≥20 was 70.0% (95% CI, 45.7% to 88.1%) vs 29.2% (95% CI, 12.6% to 51.1%) in the peltopepimut-S arm and placebo arms, respectively, associated with mOS not reached (after 42 months) versus 23.3 months. The addition of peltopepimut-S to cemiplimab did not increase cemiplimab's toxicity. CONCLUSION:Adding peltopepimut-S to cemiplimab did not improve ORR and worsened mOS in the primary analysis. Divergent outcomes were seen in patients with pretreatment PD-L1 CPS <20 (worse ORR and mOS compared with placebo) and CPS ≥20 (higher ORR and longer mOS compared with placebo) values. Future drug development is justifiable in the CPS ≥20 patient population. TRIAL REGISTRATION NUMBER:NCT03669718.

Circulating tumor human papillomavirus DNA (ctHPVDNA) testing using digital-droplet polymerase chain reaction (PCR) detects fragments of tumor-modified human papillomavirus (HPV) in the plasma of patients with HPV-associated head and neck squamous cell carcinomas (HNSCCs). Its impact on tumor surveillance and primary diagnosis is limited by unresolved issues relating to sensitivity and specificity. The study population consisted of patients with HNSCC who had undergone ctHPVDNA testing. HPV status was determined by p16 immunohistochemistry and PCR-HPV genotyping on the tumor samples. For discrepant cases (HPV-positive/ctHPVDNA-negative), HPV status was confirmed by RNA in situ hybridization and, when possible, targeted single-nucleotide polymorphisms genotyping. A total of 167 patients had ctHPVDNA testing, and 141 tumors were HPV positive by p16 immunohistochemistry and PCR genotyping. Genotypes included types 16 (91.5%), 33 (4.3%), 35 (2.1%), and 18 (2.1%). ctHPVDNA was detected in 133 (94.3%) of HPV-positive HNSCCs but in none of the HPV-negative HNSCCs. Four of the 5 p16-positive cases that were negative by PCR and ctHPVDNA were positive by RNA in situ hybridization, and in 2 of these cases, rare high-risk genotypes were identified. ctHPVDNA had a sensitivity of 91.7%, specificity of 100%, positive predictive value of 100%, and negative predictive value of 63.6%. The likelihood that patients with HPV-positive HNSCC have detectable ctHPVDNA is high. Non-HPV16 genotypes contribute to discrepancies but only in a small subset of cases. This finding validates ongoing efforts to use ctHPVDNA as a surveillance tool, and even as a primary diagnostic assay in patients presenting with masses in the neck and/or oropharynx.

6003 Background: ISA101b (peltopepimut-S) is a therapeutic vaccine targeting the HPV16 E6/E7 oncoproteins. The synthetic long peptides of ISA101b induce specific expansion of both CD4+ T-helper cells and CD8+ cytotoxic T-cells against E6/7 oncogenes1. Combination of cemiplimab, an anti-PD-1 antibody with ISA101b elicits a synergistic anti-tumor effect2. Methods: First and second line anti-PD-1 naïve patients with confirmed HPV16+ R/M OPC were randomized to treatment with either ISA101b (subcutaneously 100µg/peptide on days 1, 29, and 50) or placebo, with cemiplimab (intravenously 350mg q/21 days) for up to 24 months or until disease progression or treatment withdrawal. The primary efficacy endpoint was ORR after ≥6 months of follow-up by independent review as per RECIST1.1. Data cut-off for this analysis was 5 July 2023. The primary safety endpoint was frequency and severity of AEs. Secondary endpoints included PFS and OS. For the latter 12-month survival data are shown. Combined Positive Score (CPS) analyses were planned subgroup analyses. A p-value <0.1 was defined as statistically significant. Predefined analysis sets are the full analysis set (FAS), i.e. all patients who received ≥1 dose of study drug, and the per protocol set (PPS) which includes patients with centrally confirmed HPV16-positivity who received all 3 doses of ISA101b/placebo, had at least 1 post-baseline tumor assessment and no major protocol deviations. Results: A total of 198 patients (mean age 62.8 ±9.6 years) received ≥1 dose of study drug: 173 (87.4%) male, and 25 (12.6%) female; 110 (55.6%) were treated in first, and 74 (37.4%) in second line. Baseline characteristics were well balanced. In the ISA101b arm, ORR was 25.3% compared to 22.9% in the control arm (NS, Table). SAEs occurred in 33.0% of patients in the ISA101b arm vs 31.6% in the control arm. Patients with a CPS ≥20 treated with cemiplimab and 3 doses of ISA101b had a significantly better ORR and OS compared to patients in the control arm (Table; mOS (95% CI) not reached (28.1, -) vs 23.3 (11.9, 30.1) months, P = 0.0232 (PPS)). Patients with CPS <20 had on average a shorter OS in the ISA101b arm. Conclusions: Whereas there was no advantage of the addition of ISA101b to cemiplimab regarding ORR on the overall population, in patients with CPS ≥20 ISA101b significantly improved the ORR. Median OS was better in patients with CPS ≥20 who completed a full course of ISA101b. In contrast, patients with lower CPS did not benefit. Toxicity was comparable between the 2 arms. Clinical trial information: NCT03669718 . [Table: see text]

Brain metastasis (BM) is a rare but severe complication of head and neck squamous cell carcinoma (HNSCC), with limited knowledge of molecular characteristics and immunogenicity. We analyzed 61 cases of HNSCC-BM from three academic institutions (n = 24) and Foundation Medicine Inc (FMI, n = 37). A subset of cases underwent next-generation sequencing, multiple immunofluorescence, and proximity ligation sequencing. Gene enrichment analysis compared alterations in FMI BM samples (n = 37) with local samples (n = 4082). Demographics included: median age of 59 years, 75

6036 Background: BM is a rare complication of HNSCC that carries a high rate of morbidity and poor prognosis. Clinical risk factors, molecular characteristics, and the immunogenicity of HNSCC BM are not well defined, leaving a critical knowledge gap in this field. We performed one of the largest multi-institutional analyses summarizing the clinical, molecular, and immunologic profile of 61 cases of BM-HNSCC. Methods: We conducted a pooled analysis of the clinical characteristics pertaining to BM-HNSCC from 3 academic institutions (n=24). Next-generation sequencing (NGS) and immune profiling (IP) of primary and BM specimens was conducted on a subset of cases (n=19 and n=16, respectively); there were 3 paired samples for NGS and 0 for IP. Four samples (2 BM and 2 non-BM) were submitted to Phase Genomics, Inc for evaluation of structural variants in BM genomes by proximity ligation sequencing (PLS). These results were complimented by a comparative analysis of genomic alterations in an additional cohort of BM (n=37) and local samples (n=4082) submitted for NGS at Foundation Medicine, Inc (FMI). Statistical comparisons were done using Fisher’s exact testing of 2x2 contingency tables with p-values controlled for FDR by the Benjamini-Hochberg procedure. Results: Clinical features were as follows: median age at diagnosis 59 years, 75% male, 55% current/former smokers, 75% oropharyngeal primary, and 84% HPV+ or p16+. The most frequently altered genes in BM specimens (62% HPV/p16+) were ATM (54%), KMT2A (54%), PTEN (46%), RB1 (46%), and TP53 (46%). BM and non-BM samples demonstrated significant levels of structural rearrangement ranging from 9 to 90 variants by PLS. IP identified lower densities of CD8+, PD1+, PDL1+, and FOXP3+ cells in BMs compared to primary tumors. PDL1 combined positive scores were <1% in 12/13 unpaired samples (92%; 10 BM and 2 primary). The FMI BM-HNSCC cohort (51% HPV+) identified CDKN2A (40.5%), TP53 (37.8%), and PIK3CA (27.0%) as the most frequently altered genes. Enrichment analysis of the FMI cohort showed MAP2K2 alterations significantly enriched in BM (11.8% vs 6.4%, P=0.005) and TSC1 alterations significantly enriched in the local site (67.3% vs 37.8%, P=0.008). HPV+ was also significantly enriched in the BM cohort (51.25% vs 26.11%, P=0.001). Overall survival from BM diagnosis was 6m (range 0-27m). Conclusions: HNSCC patients with BM have higher-than-expected proportions of oropharyngeal primary site and HPV/p16-positivity. The most frequent molecular alterations in BM samples are also commonly found in non-BM HNSCC, including targetable PIK3CA alterations. MAP2K2 alterations were significantly enriched in BM compared to non-BM samples, which warrants further investigation. BM samples also tended to have lower markers of immunogenicity. This latter finding could have important clinical implications when considering immunotherapy or immune-modulating drugs.

Purpose/Objective(s)As the population continues to age, there will be an increase in human papilloma virus (HPV) associated oropharyngeal carcinomas (OPSCC) in older adults. Frailty is a risk factor for worse disease and survival outcomes in the geriatric population. Biomarkers and frailty indices have been used to quantify and predict survival outcomes and complications. We seek to develop a frailty index using lab abnormalities and comorbidities to predict of mortality and morbidity in older adults with OPSCC.Materials/MethodsStudy participants were identified from a retrospective database of head and neck squamous cell carcinoma patients consecutively treated from 2007 to 2020 at our institution's cancer center. Patients aged ≥65 with newly diagnosed, nonrecurrent, nonmetastatic, curatively treated OPSCC, with a minimum follow-up of 1 month, were included in this study. Approval for this study was obtained from the IRB at our institution and informed consent was waived given the retrospective nature of the study. Pre-treatment lab values (basic metabolic panel) were used to extract data on albumin levels. The modified frailty index (mFi-5) was also calculated – it is a 5-item score validated from the NSQIP dataset that calculates frailty based on comorbidities including hypertension (on medications), diabetes mellitus, CHF, COPD/pneumonia, and functional status. Outcome measures included disease specific survival (DSS), treatment interruptions, prophylactic PEG tube placement, and overall survival. Subsequently, univariate analyses were adjusted for stage and Charlson Comorbidity Index. DSS, treatment interruptions, and prophylactic PEG tube placement were compared across frailty markers using the χ2 test and Fisher's exact test. Overall survival was analyzed using ANOVA.Results146 patients with OPSCC were identified with a median age of 71.5 (IQR: 67.9-76.1). The majority of patients were male (83%), white (63%), former/active smokers (90.4%), and HPV+ (68.6%). There was no association between mFI-5 score and any of the outcome measures. Low albumin levels at diagnosis (<3.5 g/dL) were associated with lower DSS (p=0.023) as with higher rates of prophylactic PEG tube placement (p=0.028). There was an association between albumin levels and acute mucositis (3 months after RT) (p=0.008). The relationship of albumin and prophylactic PEG tube placement remained significant after adjusting for comorbidities and stage.ConclusionThis is one of the first studies to investigate the indexes of frailty (albumin and mFI-5) on DSS, PEG-tube placement, and treatment complications in older adults with OPSCC. Preliminary analysis shows that low albumin levels are associated with worse disease specific survival, higher rates of prophylactic PEG tube placement, and acute mucositis.

Abstract Purpose: Detecting minimal residual disease (MRD) has emerged as a key strategy in oncology for identifying those at high risk of relapse. Circulating tumor tissue modified viral (TTMV)-HPV DNA has emerged as a promising biomarker in HPV-driven oropharyngeal cancer. Prior studies have largely focused on detection beyond three months post-treatment. This study evaluated the ability of TTMV-HPV DNA to detect MRD immediately after initial or salvage treatment. Patients and Methods: This IRB-approved, retrospective cohort study included 171 patients across eight U.S. centers who had pathologically confirmed HPV-driven oropharyngeal cancer. Patients received at least one TTMV-HPV DNA test (NavDx®, Naveris, Inc.) done between zero and three months of treatment between February 2020 and April 2022. Minimum four months of follow-up was required for patient inclusion. Test results were correlated with physician-reported exams, imaging studies, and biopsies. Results: Patients were primarily White (94%) and male (86%) and were a median age at diagnosis of 60 years (range: 31 - 81). Primary tumors were located in the tonsil or base of tongue (94%) and were mostly T1-T2 (77%), N1 (66%), and M0 (98%). Patients had a median follow-up of 17 months after the completion of initial definitive treatment (range: 4-96 months). In this cohort, 198 tests conducted in 171 patients were evaluable. A total of 14 patients had both biopsy-confirmed pathologic recurrences and relevant post-treatment testing data in relation to the recurrence, with a median disease-free interval of three months (range: 1-16). Within this cohort, TTMV-HPV DNA testing demonstrated a per-patient sensitivity of 85.7% (12/14, 95% CI: 67-100). The positive predictive value (PPV) of a single TTMV-HPV DNA test within the first three months following treatment was 86.7% (13/15, 95% CI: 69.5-100). Six of the positive tests (46.2%) preceded clinical recurrence detection by a median lead-time of 36 days (range: 21-97). Test positivity following initial definitive treatment was significantly associated with worse recurrence-free survival (log-rank test, p<0.0001; hazard ratio: 21). The two patients with false positive test results had their TTMV-HPV DNA testing performed just four days after completing CRT. Their test scores returned to negative on the subsequent test, and both patients have remained disease-free. Of the 157 patients who remained TTMV-HPV DNA negative throughout the duration of follow-up, only two (1.3%) had biopsy-confirmed, HPV-positive recurrence before or within three months of a negative test. The specificity and negative predictive value (NPV) per-test were each 98.9% (181/183, 95% CI: 97.4-100). Conclusion: TTMV-HPV DNA demonstrates strong potential as a prognostic biomarker for detecting MRD in the immediate post-treatment setting, with a high PPV and NPV for recurrence. These findings support the integration of TTMV-HPV DNA into routine surveillance protocols to enhance early detection of recurrence and guide timely interventions. Citation Format: Scott A Roof, James Jabalee, Eleni M Rettig, Rocco Ferrandino, Sida Chen, Marshall R Posner, Krzysztof J Misiukiewicz, Eric M Gender, Raymond L Chai, John Sims, Elaine Thrash, Scott J Stern, Noah S Kalman, Sreenija Yarlagadda, Adam Raben, Lydia Clements, Abie Mendelsohn, John M Kaczmar, Yadav Pandey, Mihir Bhayani, Catherine DV Fitz, Glenn J Hanna. Immediate post-treatment detection of minimal residual disease in HPV-Driven oropharyngeal cancer is associated with high risk of recurrence [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr B022.