“ABC, GCB and Double-Hit diffuse large B cell lymphoma: Does subtype make a

difference in Therapy Selection?”

Journal ClubModerator: Dr. Suresh Babu MC

Presenter: Dr. Gita R Bhat

Authors: Grzegorz S. Nowakowski, Myron S. Czuczman

ASCO 2015, Educational Book

Overview • Personalized therapy for treatment of patients with cancer is rapidly

approaching and is an achievable goal in the near future.• DLBCL is the most common NHL• 40% patients have refractory disease or disease that will relapse after

initial response• 2 major biologically distinct molecular subtypes of DLBCL: GCB and ABC• Double hit lymphomas (approx 5%-10%) of patients and double

expressor lymphomas are aggressive and associated with poor prognosis.

• Early clinical trials evaluating combination of novel targeted agents in combination with R-CHOP have shown encouraging results.

• Hence, molecular classification: Prognostication + personalization of therapy for DLBCL.

• Addition of rituximab (R) to CHOP in patients with DLBCL: dramatic improvements in PFS and OS.

• Inspite of this, 40% relapse or have refractory disease.

• Various strategies to improve outcomes: intensification of chemotherapy, use of maintenance therapy, novel agents.

• Alternate regimens for front-line R-CHOP :• Dose –dense R-CHOP 14: Phase III trial

showed no additional clinical benefit• Dose-adjusted R-EPOCH• R-CEOP 90• Phase III trial: 1080 patients – No additional

clinical benefit was observed in patients treated wih R-CHOP14 vs. R-CHOP21.

• Addition of novel agents (X) to R-CHOP:• XR-CHOP• “DLBCL has molecular heterogeneity”• “X – targets specific oncogenic pathways”

• Classification of DLBCL based on the cell of origin (COO):

• GCB (CD10, BCL6)• Non-GCB ABC (poor outcome) Primary mediastinal B-cell types• Associated with differences in clinical outcome

GCB DLBCL ABC DLBCL

Markers of germinal centre differentiation (CD10 and BCL6)

-

- NF-κB pathway is constitutively activeHigh expression of NF-κB genes

BCL2 + BCL2+ (> 4 fold higher than in GCB DLBCL)

Arises from germinal centre B cell Post-germinal centre B cell, blocked during plasmacytic differentiation

Outcome of GCB DLBCL and ABC DLBCL treated with RCHOP

Primary mediastinal B cell lymphoma

Arises from Thymic B cell

Predominantly in young women

Shares many features with CHL-NS

OCT-2 and BOB-1 (B cell transcription factors) are positive, Immunoglobulin production is defective

Agents predominantly active in Non-GCB (ABC) DLBCL

• Pathways that are constitutively activated in ABC DLBCL:

• B-cell receptor (BCR) pathway • Pathways downstream of BCR pathway• Constitutive activation of NF-kB genes

• Proteasome inhibitors• Immunomodulatory agents• B-cell receptor signaling pathway inhibitors

B-cell receptor pathways

• Plays an important role in proliferation and survival in B-NHL

• Targets: • Spleen tyrosine kinase (SyK): Survival• Bruton tyrosine kinase (BTK): BCR signaling

and maturation

Proteasome inhibitors

• Inhibit transcription factor NFkB• Downstream pathway of BCR pathway• Bortezomib combined with DA-EPOCH:• ORR: GCB DLBCL (13%) vs. ABC DLBCL (83%)• Median OS: GCB (10.8 months) vs. ABC ( 3.4

months)

IMiDs

• Structural and functional analogues of thalidomide• Regulate production of T-helper cells• Inhibit cytokine production• Inhibit production of TNFa• Induce G0/G1 cell cycle arrest• Inhibit angiogenesis through suppression of VEGF

and FGF• Decrease NFkB activity

• Single agent Lenalidomide in relapsed refractory/refractory NHL including DLBCL

• Phase II study, n= 217 patients• DLBCL subpopulation: median PFS (2.7

months) and Response duration (4.6 months)

B-cell receptor signaling pathway inhibitors - IBRUTINIB

• Bruton tyrosine kinase inhibitor

• Forms covalent bond with cysteine -481 in BTK

• High BTK specificity• Daily oral dosing produces 24-

hour BTK inhibition• Blocks NF-kB activation in

ABC DLBCL• Phase II study: ORR 22%, CR

5%, PFS: 1.6 months (relapsed/ refractory DLBCL)

• SYK inhibitor: FOSTAMATANIB

• Phase II study, 23 patients• Median PFS 2.7 months• ORR: 22%

Agents with potential activity in GCB DLBCL

• GCB DLBCL has better outcomes than ABC subtype

• 20% of patients with GCB DLBCL relapse after R-CHOP or R-CHOP like chemotherapy

• Associated with poor outcomes

• BCL6: highly expressed in GCB subtype• Key transcription factor• Translocations/ mutations enhance the

inhibitory effect of BCL6 on apoptotic stress response

• This leads to tumor proliferation and treatment failure

Therapeutic implications

A) Small molecule inhibitor of BCL-6:• 79-6 complex• Binds to the co-repressor binding groove of

the BCL6 domain and kills BCL-6 positive linesB) HDAC inhibitors to overcome the effects of

BCL6 repression on p53

• C) Etoposide: Topoisomerase II inhibition – ubiquitin mediated protein degradation and transcriptional inhibition --- downregulates BCL6

• DSHNHL study: better EFS in those who received CHOEP vs. CHOP alone

• GCB DLBCL has higher incidence in younger patients, hence they benefit more from the addition of Etoposide.

• D) DA-EPOCH-R• Inhibition of Topo II is optimised by continuous

delivery of drugs over 96 hours• This ensures steady state concentration• 5 years of follow-up: EFS (95% to 100%)• E) EZH2: EZH2 inhibitors• Gain of function mutations in EZH2 result in

increased H3K27 methylation

BCL2 inhibitors• Members of BCL-2 family (BCL-2, BCL-XL, BCL-w, MCL-1,

BFL1/A-1, and BCL-B):• Suppress apoptosis through interaction with, and

inactivation of, pro-apoptotic proteins such as BH3• BCL2 inhibitors are active in ABC and GCB DLBCL• In GCB: BCL2 is overexpressed as a result of translocation• In ABC: BCL2 is overexpressed at the protein level• ABT-737 and ABT-263: target BCL-2, BCL-Xl and BCL-w• ABT-199: potently and selectively inhibits BCL2

Front-line treatment: XR-CHOP

• Bor-RCHOP• R2-CHOP• IR-CHOP

Bor-RCHOP

• Untreated DLBCL or mantle cell lymphoma

• Ongoing Phase III RCT: CHOP vs. Bor-RCHOP in DLBCL

ORR – 100%

CR or Cru: 86%

2-year PFS: 64%

2-year OS: 70%

R2-CHOP

• R2-CHOP:• Lenalidomide-RCHOP• Improves the poor prognosis usually reported

in non-GCB DLBCL• Grade 3 and 4 AEs: Neutropenia (31%),

leucopenia (28%), thrombocytopenia (13%)

• Phase II trial: newly diagnosed DLBCL• Treated with R2-CHOP vs R-CHOP

• Addition of Lenalidomide can improve the poor prognosis in non-GCB population

2-year OS

GCB DLBCL Non-GCB DLBCL

R2-CHOP 75% 83%

R-CHOP 78% 46%

• IR-CHOP:• Phase I randomized trial, 33 patients• Newly diagnosed DLBCL (22 patients) , Mantle

cell lymphoma, Follicular lymphoma• ORR 100% (CR 64% and PR 36%)• Most common AEs: Neutropenia, nausea,

thrombocytopenia, vomiting, anemia

• IR-CHOP:• Bruton’s tyrosine kinase inhibitor• Ibrutinib• Phase Ib: Newly diagnosed DLBCL, mantle cell

lymphoma, follicular lymphoma

MYC-positive and Double-hit DLBCL

• MYC is a transcription factor• Potent proto-oncogene• Regulates 10%-15% of human genome• Member of the helix-loop-helix leucine zipper

family of nuclear transcription factors• Key to formation and maintenance of germinal

centres

• It can be activated via 3 modes: “Avalanche effect”

• Translocation (5% - 14%)• Copy gain (19% to 38%)• Amplification (2%)• Mutation (32%)

Target genes of MYCProcess involved Function Target genes induced Target genes repressed

Cell cycle Transit through cell cycle G0 to S transition

Cyclin D2, CDK4 P21, p15, GADD45

Differentiation Blocks many cellular systems

LDH, ribosomal proteins, EIF4E, EI2A

Growth and metabolism Increase in cell size and number

N-cadherin, integrin

Adhesion/migration Enables anchorage dependent growth

Thrombospondin

Angiogenesis Induces angiogenesis IL 16, mir 17-92

Chromosomal instability Telomere aggregation, ROS production

MAD2, TOP1, BUBR1, Cyclin B1

Stem cell self renewal Potentiates induced pluripotent stem cells

? ?

Transformation Drives tumorigenesis Several genes Several genes

MYC-associated pathways of regulation of proliferation and survival

MYC-driven lymphomagenesis

Neoplasms with MYC gene rearrangements

• Burkitt lymphoma• t(8;14)• Simple karyotype• Sole chromosomal

abnormality

• DLBCL (7% - 14%)• Unclassifiable B-cell

lymphoma (35%)• Plasmablastic lymphoma

(50%)• Plasma cell myeloma

(15% - 50%)• Mantle cell lymphoma• Complex karyotype• Secondary genetic events

Why is this important?

• MYC rearrangement predicts an inferior outcome in DLBCL

• These are seen in 58% - 83% of MYC-translocated DLBCL• OS when treated with RCHOP is ≤ 12 months

? Due to the MYC rearrangement itself

Double hit DLBCL• Concurrent BCL2 translocation• Less likely BCL6

Triple hit DLBCL• Concurrent translocations in MYC+ BCL2 +BCL6

Amazing survival advantage!

Concurrent MYC + BCL2

MYC1) Cell growth

2) Cell cycle transit3) Angiogenesis

BCL21) Increased anti-

apoptosis (drug resistance)

• Double-expressor lymphomas:• High percentage of MYC and BCL2 protein • By IHC staining• Tumor cells should express at least 40% MYC

and at least 50% to 70% BCL2 positivity• These are primarily ABC-like• R-CHOP or CHOP-like chemotherapy: inferior

OS and PFS

FISH vs IHC for detecting Double hit DLBCL

• ? Effect of MYC alone (FISH or IHC) without BCL2 on outcome

• BCL2+ MYC by IHC or FISH has worst outlook

“Assessment of MYC and BCL2 expression by IHC represents a robust, rapid, and

inexpensive approach to risk-stratify patients with DLBCL at diagnosis”

• DHL patients have several poor prognostic factors:

• Median age: 7th decade• Stage III/IV disease• IPI: High intermediate/high• Elevated LDH• High frequency of extra-nodal sites (excluding

CNS)

• Of the several intensive chemotherapy regimens used:

• R-EPOCH• (1) has curative potential in BL• (2) is better tolerated than most dose-

intensive regimen• (3) appears to have similar efficacy compared

to other dose-intensive therapies

Combination chemotherapy in Double-hit lymphoma

Initial therapy for Double hit lymphoma

• Petrich AM et al. Blood 2014; 124:2354-2361• Retrospective study of outcomes in 23 US centres

over 12 years:• 311 patients with newly diagnosed DLBCL (154),

BCLU (150), FL (7)• MYC-R and BCL2 (87%) or BCL6 (5%) by

FISH/cytogenetics• 76% raised lDH, 33% ≥3* ULN• 65% stage IV, 41% BM +ve, &% CNS +ve

Comparison of long-term, progression-free, and overall survival.

Adam M. Petrich et al. Blood 2014;124:2354-2361

©2014 by American Society of Hematology

Role of stem cell therapy

• Autologous SCT in those who achieve CR:• Does not significantly change clinical

outcomes• Inherent rapid tumor cell growth and inherent

drug resistant DHL cells (Minimal residual disease)

• Allogenic SCT is unlikely to have a major effect since:

• 1) limited data from a small number of selected patients

• 2) the risk of relapsed disease while awaiting graft-vs-lymphoma to occur

• 3) the need for a suitable HLA-compatible donor• 4) chronic GvHD

Overall survival by SCT versus observation in first complete remission.

Adam M. Petrich et al. Blood 2014;124:2354-2361

©2014 by American Society of Hematology

• Adverse factors for OS at diagnosis:• Leukocytosis• LDH >3*ULN• Advanced Ann Arbor stage• CNS involvement

• Hence,• DA-R-EPOCH induction + CNS prophylaxis is a

reasonable approach

• Further escalation of chemotherapy, especially in the salvage setting is unlikely to be of benefit…. Hence, novel agents…

Waiting in the wings….

ABT - 199 Platelet sparing BCL2 inhibitor (BH3 mimetic) restores apoptosis

Bromodomain inhibitors Down-regulation of MYC-associated transcription: Decreased cell proliferation and inhibition of MYC-driven neoplasms

CAR-T cells Autologous T-cell mediated killing of CD19-positive lymphoid neoplasm

Aurora kinase inhibitors (Alisertib)

Aurora kinase is required for tumor maintenance of MYC-driven lymphoma

mTor inhibition mTor plays an important role in tumor maintenance by MYC in B lymphocytes

Second generation Proteasome inhibitor (Ixazomib)

Degrade MYC and can induce lymphoma cell death

PI3K inhibition In GCB-DLBCL: Loss of PTEN leads to activation of PI3K/AKT pathway ___ MYC upregulation

Inhibition of mitochondrial peptide deformylase

Apoptosis in MYC-over expressing hematopoietic neoplasms

SIRT4 protein Suppresses tumor formation in MYC-induced B-cell lymphoma

Key points• ABC subset of DLBCL is biologically distinct• Associated with poor outcomes when treated with a standard therapy. • Activation of the clonic B-cell receptor pathway allows for therapeutic

targeting.• Targeted agents in relapsed DLBCL can be combined with R-CHOP in front-

line therapy of DLBCL. • The germinal center B-cell (GCB) subset of DLBCL is associated with better

outcomes and may require different therapeutic approaches.• Double-hit lymphoma (DHL) is responsible for a substantial number of

relapses in GCB DLBCL • All newly diagnosed DLBCL biopsy samples should be tested for DHL by

fluorescent in situ hybridization and by immunohistochemistry for double-expressor DLBCL

• Whenever possible, patients should be referred to participation in clinical trials

• DHL: DA-R-EPOCH plus central nervous system prophylaxis until more effective novel targeted agents for this lymphoma subtype are developed

Thank you!