مقالات پذیرفته شده در نهمین کنگره بین المللی زیست پزشکی
Improved Metastatic Risk-Stratification in Cutaneous Squamous Cell Carcinoma with the 40-Gene Expression Profile Test
Improved Metastatic Risk-Stratification in Cutaneous Squamous Cell Carcinoma with the 40-Gene Expression Profile Test
Shirin Fakhar,1Asiyeh Jebelli,2,*
1. Department of Cell and Molecular Biology, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran 2. Department of Cell and Molecular Biology, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran
Introduction: The second most common form of skin cancer is Cutaneous squamous cell carcinoma (cSCC), whose incidence rate has increased worldwide in recent years. Most of the cSCC patients can be treated with surgical excision, while a subset of cSCCs carry an increased risk of metastasis and mortality. There is a correlation between early detection of metastasis and the use of effective treatment strategies for improved outcomes. Therefore, it is pivotal to predict metastasis risk with high precision. Staging systems, such as the BWH (Brigham and Women’s Hospital) and AJCC8 (American Joint Committee on Cancer Staging Manual, 8th Edition), indicate a direct relationship between T-stage and metastasis. However, these staging systems identify substantial numbers of SCC tumors as low-risk metastasis. Therefore, the tumor staging systems based on clinicopathologic factors are considered insufficient due to their low positive predictive value (PPV), which can lead to limitations in identifying high-risk patients. The gene expression pattern can be used as an invaluable method for stratifying tumors. The expression level of mRNAs in a cell population can be measured by gene expression profiling, known as transcriptomics. Three essential techniques used for gene expression profiling include: (1) quantitative Reverse Transcriptase PCR (RT-qPCR), (2) microarrays, and (3) NGS (RNA-Seq). RT-qPCR discovers putative biomarkers for disease diagnosis and prediction. DNA microarray methods use a reference genome/transcriptome for designing the microarray probes based on the hybridization of fluorescently labeled probes. RNA-seq data analysis encompasses several steps: trimming, alignment, counting and normalization of the sequenced reads, and, very often, differential expression (DE) analysis across conditions. The 40-gene expression profile (40-GEP) test enables the simultaneous measurement of the activity of 40 genes, including 34 discriminant genes, along with 6 additional genes chosen as controls, providing a comprehensive analysis of cellular function at the molecular level by utilizing RT-PCR. In fact, SCC tumors are stratified into three classes by 40-GEP assay: Low (Class 1), moderate (Class 2A), and high (Class 2B) risk groups for regional or distant metastasis. The majority of metastatic events take place in Class 2A and Class 2B categories. Thus, in contrast to staging systems, the validated 40-GEP molecular test is used as a valuable tool with high PPV for tumor stratification and metastatic risk prognosis based on the biology of the tumor.
Methods: To design the 40-GEP test, scientists collected formalin-fixed paraffin-embedded (FFPE) samples from primary cSCC lesions (either biopsy or definitive surgery) and relative clinicopathologic data from patients according to institutional review board approval. The 3-year metastasis-free survival (MFS) was the primary endpoint of the study. Then, all the samples were analyzed by 40-GEP implemented in a clinical laboratory with personnel blinded to the patient's result. Finally, the accuracy of metastasis-risk prediction for the 40-GEP in combination with staging systems was assessed by measuring PPV and negative predictive value (NPV) with the help of statistical analyses.
Results: The results demonstrate 40-GEP is more accurate in comparison to AJCC8 and BWH staging systems. Moreover, the sensitivity, specificity, and NPV of a 40-GEP Class 2 (Class 2A and Class 2B) are significantly higher relative to AJCC8 T3/T4 and BWH T2b/T3 stages. 40-GEP test used the expression of 6 “housekeeping” genes (BAG6, FXR1, KMT2C, KMT2D, MDM2, and MDM4) to normalize the expression of the 34 discriminant genes.
Conclusion: In conclusion, 40-GEP enhances metastatic risk stratification of patients with cutaneous squamous cell carcinoma both independently and integrated with other clinicopathologic risk factors as well as conventional staging systems. Therefore, the 40-GEP test in combination with current methods can lead to better-informed decision-making for treatment and follow-up health care, especially in the high-risk Class 2B group.