Identification and Validation of a Multigene Predictor of Recurrence in Primary Laryngeal Cancer

Abstract

PurposeLocal recurrence is the major manifestation of treatment failure in patients with operable laryngeal carcinoma. Established clinicopathological factors cannot sufficiently predict patients that are likely to recur after treatment. Additional tools are therefore required to accurately identify patients at high risk for recurrence. This study attempts to identify and independently validate gene expression models, prognostic of disease-free survival (DFS) in operable laryngeal cancer.Materials and MethodsUsing Affymetrix U133A Genechips, we profiled fresh-frozen tumor tissues from 66 patients with laryngeal cancer treated locally with surgery. We applied Cox regression proportional hazards modeling to identify multigene predictors of recurrence. Gene models were then validated in two independent cohorts of 54 and 187 patients (fresh-frozen and formalin-fixed tissue validation sets, respectively).ResultsWe focused on genes univariately associated with DFS (p<0.01) in the training set. Among several models comprising different numbers of genes, a 30-probe set model demonstrated optimal performance in both the training (log-rank, p<0.001) and 1st validation (p = 0.010) sets. Specifically, in the 1st validation set, median DFS as predicted by the 30-probe set model, was 34 and 80 months for high- and low-risk patients, respectively. Hazard ratio (HR) for recurrence in the high-risk group was 3.87 (95% CI 1.28–11.73, Wald's p = 0.017). Testing the expression of selected genes from the above model in the 2nd validation set, with qPCR, revealed significant associations of single markers, such as ACE2, FLOT1 and PRKD1, with patient DFS. High PRKD1 remained an unfavorable prognostic marker upon multivariate analysis (HR = 2.00, 95% CI 1.28–3.14, p = 0.002) along with positive nodal status.ConclusionsWe have established and validated gene models that can successfully stratify patients with laryngeal cancer, based on their risk for recurrence. It seems worthy to prospectively validate PRKD1 expression as a laryngeal cancer prognostic marker, for routine clinical applications.

Positive cross talk between protein kinase D and β-catenin in intestinal epithelial cells: impact on β-catenin nuclear localization and phosphorylation at Ser552

“…For example, PRKD1 gene expression is increased in subpopulations of cancers of the colon, liver, and upper aero-digestive system, as revealed by searching COSMIC (catalog of somatic mutations in cancer). Recently, high PKD1 mRNA levels have been linked to lower cancer-specific survival in heavy smokers with esophageal squamous cell cancers (74) and predicts recurrence in laryngeal cancer (12). Indeed, the level of PRKD1 expression has been proposed as a laryngeal cancer prognostic marker for routine clinical applications.…”

Evaluation of PD-L1 Expression and Associated Tumor-Infiltrating Lymphocytes in Laryngeal Squamous Cell Carcinoma

“…H&E-stained slides from tissue blocks had been reviewed for adequacy of tumor tissue and estimation of the percentage of tumor cells in each case (9). Two hundred and sixty FFPE tissue blocks (cohort I) were histologically evaluated for tumor type, marked for tissue microarray (TMA) construction and, where necessary, for manual macrodissection.…”

“…RNA extraction from FFPE specimens was performed as previously described (9). Upon deparaffinization and overnight tissue fragment lysis with proteinase K at 56 C, RNA was extracted with TRIzol-LS Reagent (Life Technologies) and reverse transcribed with random primers and the Superscript III system (Life Technologies), according to the instructions of the manufacturer.…”

“…Given the promising results of PD-1/PD-L1 inhibitors in clinical trials for R/M HNSCC, we used a well-annotated cohort of 260 laryngeal cancers (9) to characterize PD-L1 protein expression and TIL level in association with clinical outcome. We also evaluated PD-L1 mRNA levels in the abovementioned formalin-fixed paraffin-embedded (FFPE) cohort as well as in a second independent well-annotated cohort of 89 fresh-frozen specimens of paired tumors and normal adjacent tissues.…”

Tumor cells PD-L1 expression as a favorable prognosis factor in nasopharyngeal carcinoma patients with pre-existing intratumor-infiltrating lymphocytes

“…15 All patients in this study have provided written informed consent before being subjected to tumor tissue samples. TMA sections were deparaffinized with xylene, rehydrated, microwaved in EDTA (1 mmol/L, pH D 9.0) antigen retrieval buffer and then boiled for 15 min in a microwave oven.…”