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Letters to the Editor
Clinicopathological and genetic features of limited-stage diffuse large B-cell lymphoma with late relapse: targeted sequencing analysis of gene alterations in the initial and late relapsed tumors
According to the long-term analysis of several clinical studies, late relapse (LR) is more common in patients with limited-stage diffuse large B-cell lymphoma (DLBCL) than in those with advanced-stage DLBCL;1,2 therefore a unique but poorly understood tumor biology has been proposed for limited-stage DLBCL.3 Cell of ori- gin (COO), defined by gene expression profiling4 or immunohistology,5 and patterns of gene alterations, iden- tified by next-generation sequencing, have recently allowed the classification of DLBCL into biologically and clinically distinct subgroups.6,7 Several studies have evalu- ated the genetic alterations in DLBCL that occur between initial diagnosis and relapse;8,9 however, there is limited information concerning such alterations in LR. This study focused on patients with limited-stage DLBCL who developed LR and evaluated the clinicopathological and genetic features of paired specimens obtained at initial diagnosis and LR. We aimed to reveal the biological back- ground and mechanisms of LR.
The retrospective study protocol was approved by the Institutional Review Board of the National Cancer Center Hospital (NCCH). Detailed methods are available in the Online Supplementary Appendix. We defined LR as the first relapse of any B-cell lymphoma (including indolent B-cell lymphoma) that occurred at >5 years after the initial diag- nosis. Between 1997 and 2012, 334 consecutive patients with limited-stage de novo DLBCL were treated using CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisolone) with or without rituximab therapy at the NCCH. Regular recurrences were observed (Figure 1), and of the 334 patients, 16 developed LR during a median follow-up period of 8.9 years (interquartile range [IQR]: 6.2–12.3 years). Additionally, three patients developed
LR after initial treatment at other institutions and were subsequently referred to the NCCH. Then, 19 patients (patients #1-19) were subjected to the clinicopathological and genetic evaluations (Tables 1, Online Supplementary Table S1–2).
Immunohistochemical analysis of formalin-fixed, paraffin-embedded (FFPE) tissues was performed using antibodies against MYC, BCL2, programmed death-lig- and 1 (PD-L1), and COO markers (CD10, BCL6, MUM1) (Online Supplementary Table S3). Fluorescence in situ hybridization (FISH) was performed using probe map- ping 9p24.1 for specimens with PD-L1 tumor expression and commercial MYC break-apart probe.
Genomic DNA (gDNA) was extracted from (i) the pri- mary tumor, (ii) LR tumor, and (iii) control bone marrow without lymphoma infiltration in 12 patients (patient #1- 12) who were eligible for further genetic evaluations. Immunoglobulin H (IgH) clonality asssays and target sequencing of 64 lymphoma-related genes (Online Supplementary Table S4) were performed using the paired gDNA samples from the initial diagnosis and relapsed tumor. Mutations in the paired tumor specimens, which were defined as described in the Online Supplementary Appendix and Online Supplementary Table S5, were com- pared between samples, and shared/non-shared muta- tions among the samples in the pair were identified.
The median age at the initial diagnosis of the 19 patients was 60 years (range: 32–77 years), 13 (68%) patients had stage I disease, and all had low or low-inter- mediate International Prognostic Index (IPI) risk. The median duration from the initial diagnosis to LR was 8 years (range: 5–18 years). At LR, 13 (68%) patients had an advanced-stage disease, and 10 (53%) had high-inter- mediate or high IPI risk (Table 1).
At initial diagnosis, 14 of the 19 patients (78%) had a non-germinal center B-cell like (non-GCB) DLBCL, although 4 patients experienced relapse that involved a different COO subtype (from the initial diagnosis). At LR, three patients relapsed with indolent lymphoma as
Figure 1. Progression-free survival of the 334 patients with limited-stage diffuse large B-cell lymphoma who received CHOP with/without ritux- imab. No plateau was observed in the progression-free survival (PFS) curve. CHOP: cyclophosphamide, doxorubicin, vincristine, and prednisolone.
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