Treatment Of Early Stage Follicular Lymphoma With Involved Field Radiotherapy and Rituximab. Role Of Bcl-2 Molecular Monitoring
Authors: Pulsoni Alessandro , Della Starza Irene , D'Urso Pasqualina , D'Elia Gianna Maria , Annechini Giorgia , Stefanizzi Caterina , De Angelis Federico , Tricarico Sara , Grapulin Lavinia , Cavalli Marzia , FoÃ Robin 
Published: October 2019

Abstract

Background
Stage I or II follicular lymphoma (FL) is an uncommon disease, representing only 20% of FL. Conventional treatment is represented by local radiotherapy (RT), which allows eradication of the disease in about 50% of patients. Despite the negative bone marrow biopsy in all cases, most patients present Bcl-2 rearranged cells in the bone marrow (BM) and/or peripheral blood (PB). The aim of this study was to analyze the prognostic role of Bcl-2 molecular monitoring in a series of stage I-II FL cases followed at a single center.

Methods
Fifty-seven consecutive patients with a confirmed diagnosis of stage I/II FL were investigated at presentation by PCR in order to identify the presence of Bcl-2 rearranged cells in the BM and/or PB. All patients were treated with involved field RT (30-36 Gy). Subsequently, minimal residual disease (MRD) was evaluated every 6 months after RT in patients positive at baseline; patients negative at baseline were not retested. In part of the patients (after 2005) Rituximab was administered in case of persistently positive Bcl-2 cells in the BM or PB after radiotherapy. The PCR analysis of the Bcl-2/IgH rearrangement was performed according to published methods. It consists in a nested PCR that uses in the first round a couple of primers for the major breakpoint region (MBR) or for the minor cluster region (mcr). After this first step, the amplification products were re-amplified using oligonucleotide primers internal to the original ones. An aliquot of the PCR products was analysed on 2% agarose gel containing ethidium bromide in Tris-borate electrophoresis buffer and visualized under UV light. For MBR and mcr, a reproducible sensitivity level of 10-5 and 10-4 respectively, was obtained.

Results
1. Prognostic value of basal PCR in BM/PB: PCR analysis revealed Bcl-2 rearranged cells in the PB and/or BM in 38/57 patients (66.7%) at presentation. After a median follow-up of 55 months, 11 patients (19.3%) had a clinical relapse; of them, 10 belonged to the group with positive PCR at baseline, while only 1 patient with negative basal Bcl-2 (1.7%) experienced a clinical relapse (Pearson’s chi2= 0.058, Fisher exact test = 0.079). Among the 11 patients who showed a clinical relapse, 5 presented a positive Bcl-2 at relapse, 3 were negative (1 already at baseline),while in 3 this information is not evaluable. 2. Effect of local RT: After irradiation of the sole site of the disease, Bcl-2 rearranged cells disappeared in 19 of 38 patients positive at baseline (50%). In 17/38 (44.7%), MRD remained positive, while 2 patients refused to perform the analysis. A negative MRD after RT does not seem to correlate with a lower relapse probability. Only 1 patient died of breast cancer. 3. Effect of rituximab treatment in Bcl-2+ patients: Fourteen patients with persistently positive Bcl-2 after RT were treated with Rituximab 375 mg/m2 for 4 weekly administrations: 9 of them (64%) patients became negative. This result was only temporary in 4/9 cases (1 clinical relapse). Among persistently Bcl-2 positive patients after Rituximab, 1 clinical relapse was also observed.

Conclusions
In limited stage FL, despite a negative BM biopsy, Bcl-2/IgH rearranged cells can be found in the BM and/or PB, and they can disappear after local RT of the involved lymph node(s) in 50% of cases (19/38). The basal presence of Bcl-2+ cells in the BM/PB has a prognostic role: no clinical relapses were observed in Bcl-2 negative cases at baseline, except for 1 patient. Conversely, a negative MRD after radiotherapy does not seem to correlate with a better prognosis. -Rituximab therapy can induce a negativization of Bcl-2 in MRD-positive patients. Nevertheless, Rituximab treatment was only partially effective: negativization was observed in the majority of MRD-positive patients, but it was only temporary in a proportion of them. In Rituximab-treated patients, clinical relapses occurred only in the presence of MRD. -Not all clinical relapses were preceded by MRD positivity; further data are necessary to establish the usefullness of MRD monitoring over time. Prognosis of patients with early-stage FL treated with local RT + Rituximab in case of MRD persistence, is excellent: cause-specific survival=100%, EFS=70% projected at 10 years.

Disclosures:
No relevant conflicts of interest to declare.