EBV and HLA Associations In Classical Hodgkin Lymphoma Patients From Brazil
Authors: Kushekhar Kushi , Veenstra Rianne , Gualco Gabriela , Domeney Pollyanna , Hepkema Bouke , Visser Lydia , Bacchi Carlos E , Diepstra Arjan , Van Den Berg Anke 
Published: October 2019

Abstract
Abstract 4858

Background
The proportion of classical Hodgkin lymphoma that is associated with Epstein - Barr virus (EBV+ cHL) varies between geographic locations, ethnic groups and socio-economic status. In Western Europe, approximately 30% of cHL is EBV associated, while studies from Brazil have shown a 40–100% incidence of EBV+ cHL. We previously demonstrated that specific HLA-A alleles are associated with EBV+ cHL in the Western European population, with HLA-A1 being a risk allele and HLA-A2 being protective. We also demonstrated that in the northern Chinese population HLA-A*02:07 is a protective allele for EBV- and a risk allele for EBV+ cHL. In both China and Western Europe, EBV involvement is associated with a retained expression of HLA class I by the Hodgkin Reed-Sternberg (HRS) cells. The current study focuses on EBV, HLA class I expression and associations with HLA-A alleles in a group of Brazilian cHL patients.

Patients and Methods
A total of 158 cHL consultation cases (2005-2007), obtained throughout Brazil were available at the Consultoria em Patologia, Botucatu-São Paulo, Brazil. Data on sex and age were retrieved for all cases. Hematoxilin & Eosin stained tissue sections were used to reclassify the histological subtypes according to the WHO classification. EBV status was determined by EBER in-situ hybridization (ISH) using tissue micro arrays with clearly identifiable tumor cells. Membranous expression of HLA class I was determined by immunohistochemistry using HC10 (class-I heavy chain) and beta-2-microglobulin monoclonal antibodies. HLA-A1 and HLA-A2 qPCR was performed on DNA isolated from tissue sections. The qPCR was validated using known HLA-genotype samples to discriminate between non carriers and heterozygous or homozygous carriers for HLA-A1 and HLA-A2. Control data on HLA typing was available for 46,044 blood bank donors from Brazil.

Results
The median age of the patients was 25 years (3-83) with a male to female ratio of 1:2. The nodular Sclerosis (NS) histopathologic subtype was the most frequent (78%) and Mixed Cellularity (MC) was observed in 18% of the patients. Tumor cells in 34% of the patients were EBV positive (MC 79% and NS 25%). The age incidence was bimodal with a first prominent incidence peak between 11 and 30 years and a second peak in the 61–70 years age group. The first incidence peak was not associated with EBV (22% EBV+), while the second peak was (58% EBV+). In the pediatric age group (10 yr or less), 73% of the patients were EBV+.
Membranous expression of HLA class I was significantly more often retained in HRS cells of EBV+ cHL compared to EBV- cHL, 53% vs. 17% (p=0.0004). The HLA-A1 carrier frequency was significantly increased in EBV+ cHL (32%) compared to controls (18%, p=0.02). In contrast, the HLA-A2 carrier frequency in EBV+ cHL was significantly reduced compared to controls, 28% vs. 45% (p=0.02).

Conclusions
We observed a bimodal age incidence distribution with a prominent early peak in the second to third decade. EBV positivity was especially high in young children and MC subtype and was associated with HLA class I expression by the HRS cells. The genetic risk pattern was similar to the Western European population, with HLA-A1 as a risk and HLA-A2 as a protective allele for developing EBV+ cHL.

Disclosures:
No relevant conflicts of interest to declare.