No abstract

Our data show that an increase in FADD expression is associated with a higher incidence of lymph node metastasis at presentation, and with shorter DMFI when lymph node metastases are present. High FADD expression in the primary tumour could be a useful marker to select patients for systemic treatment strategies that reduce the risk of distant metastases.

This study showed that expression of phosphorylated FADD is a new prognostic biomarker for better local control after radiotherapy in patients with early-stage glottic carcinomas.

BACKGROUND: For locally advanced squamous cell carcinoma of the head and neck (HNSCC), the recurrence rate after surgery and postoperative radiotherapy is between 20 and 40%, and the 5-year overall survival rate is B50%. Presently, no markers exist to accurately predict treatment outcome. Expression of proteins in the human epidermal growth factor receptor (EGFR) pathway has been reported as a prognostic marker in several types of cancer. METHODS: The aim of this study was to investigate the prognostic value of proteins in the EGFR pathway in HNSCC. For this purpose, we collected surgically resected tissue of 140 locally advanced head and neck cancer patients, all treated with surgery and postoperative radiotherapy. RESULTS: In a multivariate analysis, expression of the phosphatase and tensin homologue deleted on chromosome 10 (PTEN) was significantly related to worse locoregional control (LRC; HR: 2.2, 95% CI: 1.1 -4.6; P ¼ 0.03), independent of lymph node metastases (HR: 5.6, 95% CI: 1.2 -27.4; P ¼ 0.03) and extranodal spread (HR: 2.7; 95% CI: 1.2 -6.5; P ¼ 0.02). In vitro clonogenic radiosensitivity assays confirmed that overexpression of PTEN resulted in increased radioresistance. CONCLUSION: Our study is the first report showing that expression of PTEN mediates radiosensitivity in vitro and that increased expression in advanced HNSCC predicts worse LRC.

The measurement of HER2 (ERBB2) gene amplification guides both the prognosis and treatment of breast cancer, as amplification correlates with worse outcomes and the recommendation for anti-HER2 treatment. Digital PCR (dPCR) can serve as a useful tool for the direct quantification of copy number variants (CNV) in this gene, without the need for calibration standards as required for qPCR, or labor-intensive histological techniques such as FISH. Furthermore, increasing the number of available partitions in a digital PCR reaction can yield substantial gains in precision, thus offering the potential for ultra-high resolution discrimination of copy number changes. In this study, we evaluated the utility of a multiplex dPCR assay for the high-resolution detection of HER2 copy number variation in clinically relevant samples. The precision achievable by digital PCR correlates with the number of partitions, and the resolution of small CNV differences greatly relies on high precision. We utilized a dPCR setup with ~100,000 physical partitions per reaction, which allowed for a theoretical precision (relative confidence) of 1%, assuming optimal input and template. Five single-plex primer and probe sets, published by the National Institute of Standards and Technology, were assembled and optimized into a multiplex configuration. The final 5-plex consisted of four reference genes and HER2 as the target of interest. The accuracy and precision of the optimized five-plex assay was tested in nine breast cancer cell lines with established HER2 amplification ratios, ranging from 1.3 to 70 fold. The assay demonstrated the precise and accurate measurement of the HER2 amplification ratio across all nine cell lines over a wide dynamic range of sample input. Along with increasing the reaction partition number, the use of multiple reference genes also improved the precision of the HER2 amplification ratio measurement and avoided inaccurate copy number calls due to outlier references. The resolution of the assay was evaluated using admixed samples simulating low tumor fraction. A minimum of a 1.03 fold difference (3% difference from wild type) was detected with statistical significance. Assay concordance was evaluated against IHC-characterized HER2+ FFPE donor tissue with concordant FISH/CISH results. The assay demonstrated robust performance in clinically relevant sample types, supporting further investigation of dPCR as a complementary or even alternative diagnostic approach to current methods. Citation Format: Tyler Landrith, Samantha Smith, Jennifer Chan, Mari Christensen, Yu Chuan Tai, Megan Gonzales, William Yee, Nicolas Newton, Wouter Pattje, Patrick Bogard, Wei Yang. High resolution detection of HER2 copy number variation in admixed breast cancer cell lines and HER2+ FFPE donor tissue using a digital PCR multigene reference panel [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6162.

Non-small cell lung cancer (NSCLC) patients harboring activating mutations in epidermal growth factor receptor (EGFR) are benefited from Tyrosine Kinase Inhibitor (TKI) targeted therapy. However, more than 50% of patients receiving the first or second generations of TKI develop a resistance mutation, EGFR-T790M, and they will need to switch to the third-generation TKI (Osimertinib). Upon treatment, a second acquired mutation, EGFR-C797S, leads to the resistance to Osimertinib. If the C797S and T790M mutations are located in trans, a combination of the first and third-generation of TKI(s) may offer some benefit to the patient. The impact of the allelic context to the subsequent treatment is yet to be established and thus an assay identifying C797S and T790M allelic context may serve as a useful research tool. In recent years, digital PCR has emerged to be an ultra-sensitive method in non-invasive identification and monitoring of cancer mutations. However, compared to qPCR and NGS methods, current digital PCR assays are limited by the number of biomarkers that can be incorporated in a single assay and the maximum sample volume that can be screened in one test. In this study, we describe a multiplex digital PCR assay for quantitative detection of EGFR sensitizing and resistance mutations, and identification of C797S and T790M allelic context, using a 6-color digital PCR system that allows high sample volume utilization. The sensitivity and specificity of this multiplex dPCR assay were tested using a panel of contrived cell-free DNA samples. Individual mutations can be detected at as low as 0.2% fractional abundance level. Furthermore, we compared the performance of the multiplex with the 2-color droplet digital PCR Expert Design Assays on the Bio-Rad QX200 ddPCR system, using a commercial multiplex cfDNA reference standard set. The multiplexed EGFR mutation assay on the 6-color Digital LightCycler® System is a promising method for rapid and sensitive monitoring of EGFR mutations in cell-free DNA. Citation Format: Wei Yang, Mari Christensen, Jennifer K. Chan, Julie Tsai, Nancy Patten, Ha B. Tran, Grant R. Hillman, Yu Chuan Tai, Patrick Bogard, Claudia M. Litterst, Victoria H. Brophy, Nick Newton, Christopher D. Nelson, Wouter J. Pattje. Multiplexed EGFR mutation detection with C797S and T790M allelic context using six-color digital PCR [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5156.