Methods for Sample Acquisition and Processing of Serial Blood and Tumor Biopsies for Multicenter Diffuse Large B-cell Lymphoma Clinical Trials

Nielsen, Torsten Holm; Diaz, Zuanel; Christodoulopoulos, Rosa; Charbonneau, Fredrick; Qureshi, Samia; Rousseau, Caroline; Benlimame, Naciba; Camlioglu, Errol; Constantin, André; Oros, Kathleen Klein; Krumsiek, Jan; Crump, Michael; Morin, Ryan D.; Cerchietti, Leandro; Johnson, Nathalie A.; Petrogiannis-Haliotis, Tina; Miller, Wilson H.; Assouline, Sarit; Mann, Koren K.

doi:10.1158/1055-9965.epi-14-0549

Cited by 4 publications

(2 citation statements)

References 10 publications

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“…One core was fixed in formalin and embedded in paraffin (FFPET) for protein detection by immunohistochemistry (IHC). The remaining three cores were pooled in RPMI medium and DLBCL cells were enriched using a magnetic bead-negative selection technique (human B-cell enrichment cocktail without CD43; Stem Cell technologies) as previously described (25). DNA and RNA from purified B cells and peripheral blood mononuclear cells were isolated using the AllPrep kit (Qiagen).…”

Section: Sample Preparation For the Qcroc-2 Trialmentioning

confidence: 99%

Genetic Landscapes of Relapsed and Refractory Diffuse Large B-Cell Lymphomas

Morin

Assouline

Alcaide

et al. 2016

Clinical Cancer Research

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Purpose: Relapsed or refractory diffuse large B-cell lymphoma (rrDLBCL) is fatal in 90% of patients, and yet little is known about its biology.Experimental Design: Using exome sequencing, we characterized the mutation profiles of 38 rrDLBCL biopsies obtained at the time of progression after immunochemotherapy. To identify genes that may be associated with relapse, we compared the mutation frequency in samples obtained at relapse to an unrelated cohort of 138 diagnostic DLBCLs and separately amplified specific mutations in their matched diagnostic samples to identify clonal expansions.Results: On the basis of a higher frequency at relapse and evidence for clonal selection, TP53, FOXO1, MLL3 (KMT2C), CCND3, NFKBIZ, and STAT6 emerged as top candidate genes implicated in therapeutic resistance. We observed individual examples of clonal expansions affecting genes whose mutations had not been previously associated with DLBCL including two regulators of NF-kB: NFKBIE and NFKBIZ. We detected mutations that may be affect sensitivity to novel therapeutics, such as MYD88 and CD79B mutations, in 31% and 23% of patients with activated B-cell-type of rrDLBCL, respectively. We also identified recurrent STAT6 mutations affecting D419 in 36% of patients with the germinal center B (GCB) cell rrDLBCL. These were associated with activated JAK/STAT signaling, increased phospho-STAT6 protein expression and increased expression of STAT6 target genes.Conclusions: This work improves our understanding of therapeutic resistance in rrDLBCL and has identified novel therapeutic opportunities especially for the high-risk patients with GCB-type rrDLBCL.

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Section: Sample Preparation For the Qcroc-2 Trialmentioning

confidence: 99%

Genetic Landscapes of Relapsed and Refractory Diffuse Large B-Cell Lymphomas

Morin

Assouline

Alcaide

et al. 2016

Clinical Cancer Research

Self Cite

196

199

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“…Details on collection and preparation of these are reported elsewhere. 22 For every biopsy, a section was stained with hematoxylin and eosin and tumor content/viability assessed. Immunostaining for BCL2, BCL6, MYC, CD20, and acetylated H3 was performed (see supplemental Methods, available on the Blood Web site, for details).…”

Section: Correlative Tissue Biopsies and Analysesmentioning

confidence: 99%

Phase 2 study of panobinostat with or without rituximab in relapsed diffuse large B-cell lymphoma

Assouline

Nielsen

et al. 2016

Blood

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124

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Key Points• Panobinostat induces responses in 28% of patients with relapsed and refractory DLBCL that are typically durable off therapy.• MEF2B mutations predicted for response whereas early increase in ctDNA abundance was a strong predictor of subsequent treatment failure.The majority of diffuse large B-cell lymphoma (DLBCL) tumors contain mutations in histone-modifying enzymes (HMEs), indicating a potential therapeutic benefit of histone deacetylase inhibitors (HDIs), and preclinical data suggest that HDIs augment the effect of rituximab. In this randomized phase 2 study, we evaluated the response rate and toxicity of panobinostat, a pan-HDI administered 30 mg orally 3 times weekly, with or without rituximab, in 40 patients with relapsed or refractory de novo (n 5 27) or transformed (n 5 13) DLBCL. Candidate genes and whole exomes were sequenced in relapse tumor biopsies to search for molecular correlates, and these data were used to quantify circulating tumor DNA (ctDNA) in serial plasma samples. Eleven of 40 patients (28%) responded to panobinostat (95% confidence interval [CI] 14.6-43.9) and rituximab did not increase responses. The median duration of response was 14.5 months (95% CI 9.4 to "not reached"). At time of data censoring, 6 of 11 patients had not progressed. Of the genes tested for mutations, only those in MEF2B were significantly associated with response. We detected ctDNA in at least 1 plasma sample from 96% of tested patients. A significant increase in ctDNA at day 15 relative to baseline was strongly associated with lack of response (sensitivity 71.4%, specificity 100%). We conclude that panobinostat induces very durable responses in some patients with relapsed DLBCL, and early responses can be predicted by mutations in MEF2B or a significant change in ctDNA level at 15 days after treatment initiation. This clinical trial was registered at www.ClinicalTrials.gov (#NCT01238692). (Blood. 2016;128(2):185-194)

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PIK3CD promoted proliferation in diffuse large B cell lymphoma through upregulation of c-myc

Cui

et al. 2016

Tumor Biol.

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Despite PIK3CD has been extensively reported in cancers, however, little evidence has been available regarding its role in the setting of diffuse large B cell lymphoma (DLBCL). In the present study, to investigate the role of PIK3CD in DLBCL, relevant experiments were carried out on both in vivo clinical tissue level and in vitro cell line level. Prognostic and clinicopathological significance were analyzed after immunohistochemical assay of PIK3CD expression on DLBCL tissue microarray. MTT assay and flow cytometry were employed to evaluate the proliferative variation, cell cycle, and apoptosis. Athymic nude mice xenografted with DLBCL cell line were employed to confirm the role of PIK3CD. It was found that there was a significant difference between expression of PIK3CD and international prognosis index (IPI), performance state (PS), and inferior overall prognosis. Furthermore, PIK3CD can promote proliferation and prevent apoptosis in DLBCL cells in vitro through upregulation of c-myc and p-AKT and in contrast downregulation of p21 and p27. In nude mice model, knock-down of PIK3CD was shown to be able to suppress the proliferation of DLBCL but not significantly compared with control group. Taken together, our study showed that PIK3CD can promote proliferation of DLBCL cells both in vitro and in vivo, suggesting that PIK3CD could be druggable in the therapy of DLBCL.

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Methods for Sample Acquisition and Processing of Serial Blood and Tumor Biopsies for Multicenter Diffuse Large B-cell Lymphoma Clinical Trials

Cited by 4 publications

References 10 publications

Genetic Landscapes of Relapsed and Refractory Diffuse Large B-Cell Lymphomas

Genetic Landscapes of Relapsed and Refractory Diffuse Large B-Cell Lymphomas

Phase 2 study of panobinostat with or without rituximab in relapsed diffuse large B-cell lymphoma

PIK3CD promoted proliferation in diffuse large B cell lymphoma through upregulation of c-myc

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