Journal des biomarqueurs moléculaires et du diagnostic

Background: The ROBUST trial (CC-5013-DLC-002), is a phase 3 randomized, double-blind, placebo controlled, multicenter study to compare the efficacy and safety of Lenalidomide (CC-5013) plus R-CHOP (rituximab plus cyclophosphamide, doxorubicin, vincristine and prednisone) chemotherapy (R²-CHOP) versus placebo plus R-CHOP chemotherapy in subjects with previously untreated activated B-cell (ABC) type Diffuse Large B-cell Lymphoma (DLBCL). The most commonly utilized immunohistochemistry (IHC) algorithm for DLBCL subtyping, developed by Hans, et. al has been shown to have approximately an overall percent agreement of 80% with the gene expression profiling (GEP) classification of DLBCL into the germinal center B-cell–like (GCB) and non-GCB subtypes. The non-GCB subtype includes both the ABC subtype as well as the unclassified or indeterminate subtype which is neither ABC or GCB as defined by gene expression. New antibodies and algorithms specific to ABC tumors have been proposed with an aim to improve the performance of the IHC algorithm for specifically detecting the ABC subtype. This article describes one such new IHC algorithm which could be deployed for ABC subtype determination in clinic.

Methods: We analyzed 100 cases of newly diagnosed DLBCL with CD20, CD10, Bcl-6, MUM1, FOXP1, Bcl-2, Ki-67 and CD5 IHC assays using laboratory developed tests (LDT) and compared different combinations of the IHC assay results to the GEP classification. Statistical analyses were applied to evaluate the possible effect of inter-laboratory and inter-observer variations for the IHC assays and instead of using a decision tree algorithm approach to determine GCB and ABC, a novel approach was taken by using a weighted composite score algorithm. A new IHC algorithm using CD10, Bcl-6, MUM1, and FOXP1 was derived to identify ABC versus non-ABC tumors that closely approximated the GEP classification in this training set. The algorithm was assessed independently and also in conjunction with the Hans IHC algorithm to enhance testing performance. A separate set of 100 independent newly diagnosed DLBCL cases were used to validate the algorithms using LDTs developed independently from two different laboratories using different antibodies, different instrument systems and a total of four pathologists. It should be noted, the LDTs selected for use in this study were not pre-evaluated for performance.

Results: Statistical analyses indicated that the IHC assays and the algorithms for subtyping of DLBCL were robust and reproducible within the range of inter-laboratory and inter-observer variations. For the validation data set, comparing the GEP classification results and the IHC results which was derived using two independently developed LDTs per IHC marker and four pathologists, the new IHC algorithm using CD10, Bcl-6, MUM1, and FOXP1 achieved 81-91% concordance in identifying ABC tumors and 79%-86% concordance in overall classification between the individual pathologists’ calls and the GEP classification. When used in conjunction with the Hans algorithm, the IHC results achieved 89-97% concordance in identifying ABC tumors and 84-89% concordance in overall classification to the GEP results in the validation data set, simulating the predictive power of the GEP classification.

Conclusion: The use of the new IHC algorithm alone and in combination with the Hans algorithm can accurately predict ABC tumors of DLBCL and facilitate subtyping of DLBCL using standard pathology materials and routinely validated IHC assays.

The expression of N-glycolyl GM3 ganglioside (NGcGM3) in breast cancer was previously reported. However, the role of this molecule in the biological behavior of breast cancer still remains unclear. The aim of the present work was to assess a possible relationship between the expression of the evolutionary-fixed tumor neoantigen NGcGM3 ganglioside with clinicopathological parameters and the aggressiveness of breast tumors. 126 stage II/III breast tumors, obtained from patients participating in a phase III clinical trial with the GlycoVaxGM3 cancer vaccine, were clinically, molecularly and pathologically classified. An immunohistochemical study was performed using the 14F7 antibody, specific for NGcGM3, in order to measure the staining intensity and percentage of positive cells. The NGcGM3 expression in estrogen and progesterone receptors positive tumors was not associated with age, race, menopausal status, tumor size, lymph node status and stage. Nevertheless a significant increase in the presence of this cancer neoantigen in tumor tissue samples was mostly found in tumors displaying higher nuclear grades II and III when compared with grade I ones. Overall these results indicate a relationship between NGcGM3 expression and a more aggressive biological behavior of estrogen and progesterone receptors positive breast tumors, additionally sustaining this molecule as an attractive target for breast cancer immunotherapy.

Background: The intracellular mitogen-activated protein kinase (MAPK) cascade regulates intracellular processes that modulate cardiovascular disease progression. We explored the time-course of serum biomarkers that stimulate the MAPK-cascade in post-acute coronary syndrome (ACS) patients, prior to a recurrent coronary event.

Methods: BIOMArCS is a high-frequent repeated blood sampling study in post-ACS patients. We performed a nested case-control study selecting the 45 patients who experienced a recurrent event (cases) and 2 matched event-free controls per case during 1-year follow-up. Olink Proteomics ’immunoassay was used to measure 25 serum biomarkers. Results are expressed in the arbitrary Normalized Protein eXpression (NPX) unit on the 2log-scale. Linear mixed-effects models were applied to examine time-courses and differences between cases and controls.

Results: Mean age was 66 ± 12 years and 80% were men, with no differences between cases and controls. Early cases had significantly higher levels of ANG-1 (difference 0.95 NPX (95%CI 0.36-1.54), PAR-1 (difference 0.50 NPX (95%CI 0.22-0.77) and BMP-6 (difference 0.55 NPX (95%CI 0.21-0.90) than controls. No differences in biomarker levels were observed between late cases and matching controls. In particular, in cases, no increase was observed prior to the moment of the recurrent event.

Conclusion: Patients with an early recurrent coronary event after an index-ACS had higher levels of ANG-1, PAR-1 and BMP-6 than patients who remained event-free.