Protocol for qRT-PCR analysis from formalin fixed paraffin embedded tissue sections from diffuse large b-cell lymphoma: Validation of the six-gene predictor score.

Abstract

As a part of an international study on the molecular analysis of Diffuse Large B-cell Lymphoma (DLBCL), a robust protocol for gene expression analysis from RNA extraction to qRT-PCR using Formalin Fixed Paraffin Embedded tissues was developed. Here a study was conducted to define a strategy to validate the previously reported 6-gene (LMO2, BCL6, FN1, CCND2, SCYA3 and BCL2) model as predictor of prognosis in DLBCL. To avoid variation, all samples were tested in a single centre and single platform. This study comprised 8 countries (Brazil, Chile, Hungary, India, Philippines, S. Korea, Thailand and Turkey). Using the Kaplan-Meier and log rank test on patients (n=162) and two mortality risk groups (with those above and below the mean representing high and low risk groups) confirmed that the 6-gene predictor score correlates significantly with overall survival (OS, p<0.01) but not with event free survival (EFS, p=0.18). Adding the International Prognostic Index (IPI) shows that the 6-gene predictor score correlates significantly with high IPI scores for OS (p<0.05), whereas those with low IPI scores show a trend not reaching significance (p=0.08). This study defined an effective and economical qRT-PCR strategy and validated the 6-gene score as a predictor of OS in an international setting.

• Authors: Nilgun Tekin, Nader Omidvar, Tim Peter Morris, Paulette Conget, Flavia Bruna, Botond Timar, Eva Gagyi, Ranjan Basak, Omkar Naik, Chirayu Auewarakul, Narongrit Sritana, Debora Levy, Juliano Julio Cerci, Sergio Paulo Bydlowski, Juliana Pereira, Mark Pierre Dimamay, Filipinas Natividad, June-Key Chung, Nevin Belder, Isinsu Kuzu, Diana Paez, Maurizio Dondi, Robert Carr, Hilal Ozdag, Rose Ann Padua
• Journal: Oncotarget (Oncotarget) Vol. 7 Issue 50 Pg. 83319-83329 (Dec 13 2016) ISSN: 1949-2553 [Electronic] United States
• PMID: 27825111 (Publication Type: Journal Article, Multicenter Study, Validation Study)

Chemical References

• Biomarkers, Tumor
• Fixatives
• Formaldehyde

Topics

• Aged
• Asia
• Biomarkers, Tumor (genetics)
• Biopsy
• Disease-Free Survival
• Europe
• Female
• Fixatives (chemistry)
• Formaldehyde (chemistry)
• Gene Expression Profiling (methods, standards)
• Genetic Predisposition to Disease
• Humans
• Kaplan-Meier Estimate
• Lymphoma, Large B-Cell, Diffuse (drug therapy, genetics, mortality, pathology)
• Male
• Middle Aged
• Paraffin Embedding
• Phenotype
• Predictive Value of Tests
• Proportional Hazards Models
• Real-Time Polymerase Chain Reaction (standards)
• Reproducibility of Results
• Reverse Transcriptase Polymerase Chain Reaction (standards)
• South America
• Time Factors
• Tissue Fixation (methods)
• Transcriptome