Refractory and Relapse Predictive Biomarkers in Hodgkin Lymphoma and Potential Directed-Therapy Targets
Authors: Gamboa-Cedeño Angélica María , Castillo Mariángeles , Otero Victoria , Schutz Natalia Paola , Fantl Dorotea Beatriz , Jauk Federico , García Rivello Hernán , Zerga Marta , Cugliari Silvana , Rojas-Bilbao Erica , Nuñez Myriam , Ranuncolo Stella 
Published: October 2019

Abstract
Despite 90% stage I Hodgkin Lymphoma (HL) patients can respond to current systemic therapy, this drops to 60%, when diagnosed in advanced stages. Nevertheless and independently of the lymphoma stage, the real challenge when treating these patients, is the refractory and relapsed disease. There is no molecular biomarker to identify patients that would be non-responsive to conventional treatment or that would relapse. Furthermore, rescue chemotherapy schemes for refractory and relapsed patients, associate with acute and late toxicity high risk. This highlights the need to deeper understand the HL molecular biology and the screening for predictive biomarkers as well as potential therapeutic directed-targets.
We have previously reported that HL relies on the alternative NFkB pathway, mediated by RelB and NIK, to survive. Depletion of either RelB or NIK by shRNAs or pharmacological NIK inhibitors induce HL cell death. ChIP-Seq analysis uncovered RelB target genes showing RelB bound to BCL2 promoter. A significant downregulation of BCL2 mRNA and protein levels, following RelB or NIK knockdown was observed, indicating that RelB regulates BCL2 expression in human HL cell lines.
Our molecular studies suggested that NFkB alternative pathway constitutive signaling could at least partially explain the non-responding HL cases. We aimed to analyze whether mediators of this pathway could be useful as predictive biomarkers and would represent potential targetable factors in both refractory and relapsed patients.
We analyzed NIK and BCL2 citoplasm expression in Hodgkin Reed-Sternberg cells (HRS) in lymphatic node biopsies of 96 patients by inmunohistochemistry [50 female Md age and (range) 59 (6-82), 46 male 42 (9-78)]. The univariate analysis showed no correlation between NIK or BCL2 expression and the prognosis clinical and pathological parameters, neither the molecular markers routinely assayed. A positive correlation was found between NIK and BCL2 expression (p=0.01). NIK and BCL2 correlated with lack of response to conventional therapy and both early and late disease progression.
The analysis of survival, applying the Kaplan-Meier Curves, showed > 60% NIK positive HRS cells associated with shorter Disease Free Survival (DFS) [Log Rank Test (p=0.000)] and predicted overall survival (OS) as well [Log Rank Test (p=0.01)]. Furthermore, > 60% BCL2 positive HRS cells correlated with poor prognosis in terms of OS [Log Rank Test (p=0.002)]. The statistical significance was maintained in the multivariate analysis [Cox Regression and Logistic Regression (p=0.001)].
NIK and BCL2 performed successfully as useful predictive markers to identify refractory or risk of relapse HL patients at diagnosis. They represent attractive molecules to further analyse their potential as directed-therapy targets, since we have already reported that HL is sensitive to NIK inhibitors and BCL2 blockers have already been approved for clinical use in other hematological pathologies.

Disclosures
Zerga: Bristol Myers Squibb: Other: Conference fees; Janssen: Other: Conference fees; Roche: Other: Conference fees; Takeda: Other: Conference fees.