Computerized tumor multinucleation index (MuNI) is prognostic in p16+ oropharyngeal carcinoma

BACKGROUNDPatients with p16+ oropharyngeal squamous cell carcinoma (OPSCC) are potentially cured with definitive treatment. However, there are currently no reliable biomarkers of treatment failure for p16+ OPSCC. Pathologist-based visual assessment of tumor cell multinucleation (MN) has been shown to be independently prognostic of disease-free survival (DFS) in p16+ OPSCC. However, its quantification is time intensive, subjective, and at risk of interobserver variability.METHODSWe present a deep-learning-based metric, the multinucleation index (MuNI), for prognostication in p16+ OPSCC. This approach quantifies tumor MN from digitally scanned H&E-stained slides. Representative H&E-stained whole-slide images from 1094 patients with previously untreated p16+ OPSCC were acquired from 6 institutions for optimization and validation of the MuNI.RESULTSThe MuNI was prognostic for DFS, overall survival (OS), or distant metastasis-free survival (DMFS) in p16+ OPSCC, with HRs of 1.78 (95% CI: 1.37-2.30), 1.94 (1.44-2.60), and 1.88 (1.43-2.47), respectively, independent of age, smoking status, treatment type, or tumor and lymph node (T/N) categories in multivariable analyses. The MuNI was also prognostic for DFS, OS, and DMFS in patients with stage I and stage III OPSCC, separately.CONCLUSIONMuNI holds promise as a low-cost, tissue-nondestructive, H&E stain-based digital biomarker test for counseling, treatment, and surveillance of patients with p16+ OPSCC. These data support further confirmation of the MuNI in prospective trials.FUNDINGNational Cancer Institute (NCI), NIH; National Institute for Biomedical Imaging and Bioengineering, NIH; National Center for Research Resources, NIH; VA Merit Review Award from the US Department of VA Biomedical Laboratory Research and Development Service; US Department of Defense (DOD) Breast Cancer Research Program Breakthrough Level 1 Award; DOD Prostate Cancer Idea Development Award; DOD Lung Cancer Investigator-Initiated Translational Research Award; DOD Peer-Reviewed Cancer Research Program; Ohio Third Frontier Technology Validation Fund; Wallace H. Coulter Foundation Program in the Department of Biomedical Engineering; Clinical and Translational Science Award (CTSA) program, Case Western Reserve University; NCI Cancer Center Support Grant, NIH; Career Development Award from the US Department of VA Clinical Sciences Research and Development Program; Dan L. Duncan Comprehensive Cancer Center Support Grant, NIH; and Computational Genomic Epidemiology of Cancer Program, Case Comprehensive Cancer Center. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH, the US Department of VA, the DOD, or the US Government

HPV-Positive EBV-Negative Nasopharyngeal Cancer: Prevalence and Impact on Outcomes in a Non-Endemic Population

Purpose/Objective(s): To determine the prevalence of high-risk human papillomavirus (HPV) in non-endemic nasopharyngeal cancer (NPC), its association with p16 status, and potential influence on clinical outcomes in a cohort treated with definitive chemoradiotherapy (CRT). Materials/Methods: We identified 24 patients from a prospectively-maintained database treated with CRT for NPC from 1997 to 2014. All patients had paraffin-embedded tumor specimens on which Epstein-Barr virus-encoded small RNAs (EBER) in-situ hybridization and p16 immunohistochemistry (IHC) were performed. All specimens were then reviewed by an experienced head and neck pathologist who isolated and reverse transcribed total RNA from tumor regions, then performed quantitative PCR for E6 and E7 of 13 different high-risk HPV types. Log-rank tests and Cox proportional hazard models were performed to evaluate the impact of clinical factors on patient outcomes. Survival estimates were derived via the Kaplan Meier method. Results: Of the 24 tumors, 7 were HPV-positive/EBV-negative (29%), 15 were HPV-negative/EBV-positive (63%), and 2 were negative for both HPV and EBV (8%). All tumors positive for HPV mRNA expression were also positive for p16 IHC, and all tumors negative for HPV were also negative for p16, resulting in a 100% sensitivity and 100% specificity of p16 as a surrogate for high-risk HPV expression. Median age of diagnosis was 48 (19 – 68). All but 1 HPV-positive tumor was WHO II and no patients with HPV-positive tumors were WHO III. All patients received concurrent chemotherapy, with 3 patients also receiving neoadjuvant and 16 receiving adjuvant chemotherapy. Median doses to the primary and neck were 70 Gy (69.96 – 72) and 56 Gy (50.4 – 64.6), respectively. Median follow-up was 5.9 years (0.9 – 18.0) and was not different when stratified by HPV status. Local-regional control at 5 years was 100% for HPV-positive versus 81.9% for HPV-negative patients (p=0.171). Distant control at 5 years was 83.3% for HPV-positive versus 70.1% for HPV-negative patients (p=0.414). Overall survival at 5 years was 100% for HPV-positive versus 74.5% for HPV-negative patients (p=0.044). Multivariable analysis revealed that older age (HR 1.15, 95% CI 1.01-1.28) and advanced nodal stage (HR 33, 95% CI 1.19-91.44) remained as independent predictors of OS. Conclusion: We revealed that in a group of patients diagnosed with NPC in the midwest United States, HPV-driven NPC comprised a significant proportion of NPC cases, and was mutually exclusive from EBV positivity. Importantly, we discovered that p16 IHC is a strong surrogate marker for HPV-positivity in NPC. Patients with HPV-positive NPC had significantly improved overall survival in our cohort

Activation of the mTOR Pathway in Primary Medullary Thyroid Carcinoma and Lymph Node Metastases.

Understanding the molecular pathogenesis of medullary thyroid carcinoma (MTC) is prerequisite to the design of targeted therapies for patients with advanced disease.We studied by immunohistochemistry the phosphorylation status of proteins of the RAS/MEK/ERK and PI3K/AKT/mTOR pathways in 53 MTC tissues (18 hereditary, 35 sporadic), including 51 primary MTCs and 2 cases with only lymph node metastases (LNM). We also studied 21 autologous LNMs, matched to 21 primary MTCs. Staining was graded on a 0 to 4 scale (S score) based on the percentage of positive cells. We also studied the functional relevance of the mTOR pathway by measuring cell viability, motility, and tumorigenicity upon mTOR chemical blockade.Phosphorylation of ribosomal protein S6 (pS6), a downstream target of mTOR, was evident (S ≥ 1) in 49 (96\%) of 51 primary MTC samples. This was associated with activation of AKT (phospho-Ser473, S > 1) in 79\% of cases studied. Activation of pS6 was also observed (S ≥ 1) in 7 (70\%) of 10 hereditary C-cell hyperplasia specimens, possibly representing an early stage of C-cell transformation. It is noteworthy that 22 (96\%) of 23 LNMs had a high pS6 positivity (S ≥ 3), which was increased compared with autologous matched primary MTCs (P = 0.024). Chemical mTOR blockade blunted viability (P < 0.01), motility (P < 0.01), and tumorigenicity (P < 0.01) of human MTC cells.The AKT/mTOR pathway is activated in MTC, particularly, in LNMs. This pathway sustains malignant features of MTC cell models. These findings suggest that targeting mTOR might be efficacious in patients with advanced MTC. Clin Cancer Res; 18(13); 3532-40. ©2012 AACR