Study Highlights Genetic Diversity and a Potential New Treatment Target in Diffuse Large B-Cell Lymphoma

In a new study classifying the genetic diversity of diffuse large B-cell lymphoma (DLBCL), researchers have identified a potentially important new treatment target for the most common form of lymphoma.

Sandeep Dave, MD, MS, of the Division of Hematologic Malignancies at Duke, presented findings from his gene profiling study at the American Society of Hematology's 59th Annual Meeting & Exposition, December 9 to 12, 2017, in Atlanta, GA.

Although DLBCL is characterized by striking clinical and molecular heterogeneity, treatment has followed a one-size-fits-all approach for many patients. The standard frontline treatment regimen of R-CHOP has not changed for the past 15 years. "A major reason for our slow progress in DLBCL has been our difficulty in recognizing the heterogeneity that underlies a single diagnosis," Dave says.

In the current study, Dave and his team sought to characterize the most common genetic mutations in DLBCL, with a focus on those that occur in at least 5% of patients. The group also worked to understand the connection between common genetic mutations and clinical outcomes in DLBCL.

DLBCL study population
The study population included 1,001 patients with DLBCL treated at multiple institutions around the world. All patients had been treated with standard R-CHOP therapy.

Historically, the vast majority of pathology specimens have been maintained using a standard formalin-fixed, paraffin-embedded (FFPE) method. Many genomic studies, however, have excluded FFPE samples in favor of frozen tissue. This approach excludes many centers from participating in genomic studies, thereby introducing a source of sample bias. Recent research supports the feasibility of next-generation sequencing techniques using FFPE samples.

In the lab directed by Dave, the team used whole-genome sequencing to analyze defined mutations, copy number alterations, and abnormalities in the FFPE specimens. RNA sequencing was used to evaluate gene expression profiles in DLBCL, including those in the activated B-cell (ABC) and germinal center B-cell (GCB) subgroups.

Genetic alterations in DLBCL
The analysis identified 150 genetic drivers present in at least 5% of patients, highlighting wide molecular diversity of DLBCL. Of these, many were novel DLBCL genes, such as SPEN, MGA, SETD1B, and KLHL14. Large networks of mutations were also identified, including groups of genes affecting apoptosis (n = 389), cell-cycle function (n = 208), the B-cell receptor (n = 166), and interferon signaling (n = 176).

Prognostic markers
In current clinical practice, few prognostic markers are available to stratify patients based on expected overall survival. Patients with poor prognosis include those who are classified as high risk by the International Prognostic Index (IPI), those with ABC DLBCL according to ABC/GCB molecular types, and those with high MYC and BCL2 expression ("double-expressor" DLBCL).

As part of the current study, the team examined the prognostic significance of individual gene alterations and clusters of genetic abnormalities. Results showed that mutations in the phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit delta (PIK3CD) gene were associated with improved overall survival (P = .03).

The team aims to develop a genomic model of survival in DLBCL based on the presence of high- and low-risk genetic alterations. In contrast with IPI risk categories, which lose their prognostic significance in patients surviving longer than 3 years, the proposed genomic risk model has the potential to differentiate long-term risk beyond 5 years of survival.

Role of RHOA in DLBCL
Research in tumor genomics often involves a search for drug-targetable mutations. In this study, the team identified multiple oncogenes that were potential candidates for drug therapy, including genes that occurred across DLBCL subtypes and genes that were specific to the ABC DLBCL and GCB DLBCL subtypes. Oncogenes were then validated as treatment targets with existing therapies, including everolimus and idelalisib.

During the process of target validation, the research team focused on mutations in RHOA, an oncogene in the B-cell lineage. Whereas deletions in RHOA were lethal to DLBCL cells, alterations that led to RHOA overexpression were associated with increased proliferation of DLBCL cells. Additional experiments in RHOA knockout mice confirmed the critical role of RHOA in B-cell development, germinal center maintenance, and DLBCL growth.

Overall, the lab findings highlight the diversity of DLBCL molecular subtypes and the therapeutic potential of characterizing drug-targetable mutations. "Our study provides a comprehensive starting point for exploring new risk-stratification models and new treatments in DLBCL," Dave concludes.

Source: Zhang J, Reddy A, Davis N, et al. Integrative analysis of 1001 diffuse large B cell lymphoma identifies novel oncogenic roles for RHOA. Presented at: American Society of Hematology 59th Annual Meeting; December 9-12, 2017; Atlanta, GA. Abstract 37.