Immunophenotypic studies of monoclonal gammopathy of undetermined significance

<p>Abstract</p> <p>Background</p> <p>Monoclonal gammopathy of undetermined significance (MGUS) is a common plasma cell dyscrasia, comprising the most indolent form of monoclonal gammopathy. However, approximately 25% of MGUS cases ultimately progress to plasma cell myeloma (PCM) or related diseases. It is difficult to predict which subset of patients will transform. In this study, we examined the immunophenotypic differences of plasma cells in MGUS and PCM.</p> <p>Methods</p> <p>Bone marrow specimens from 32 MGUS patients and 32 PCM patients were analyzed by 4-color flow cytometry, using cluster analysis of ungated data, for the expression of several markers, including CD10, CD19, CD20, CD38, CD45, CD56 and surface and intracellular immunoglobulin light chains.</p> <p>Results</p> <p>All MGUS patients had two subpopulations of plasma cells, one with a "normal" phenotype [CD19(+), CD56(-), CD38(bright +)] and one with an aberrant phenotype [either CD19(-)/CD56(+) or CD19(-)/CD56(-)]. The normal subpopulation ranged from 4.4 to 86% (mean 27%) of total plasma cells. Only 20 of 32 PCM cases showed an identifiable normal subpopulation at significantly lower frequency [range 0–32%, mean 3.3%, p << 0.001]. The plasma cells in PCM were significantly less likely to express CD19 [1/32 (3.1%) vs. 13/29 (45%), p << 0.001] and more likely to express surface immunoglobulin [21/32 (66%) vs. 3/28 (11%), p << 0.001], compared to MGUS. Those expressing CD19 did so at a significantly lower level than in MGUS, with no overlap in mean fluorescence intensities [174 ± 25 vs. 430 ± 34, p << 0.001]. There were no significant differences in CD56 expression [23/32 (72%) vs. 18/29 (62%), p = 0.29], CD45 expression [15/32 (47%) vs. 20/30 (67%), p = 0.10] or CD38 mean fluorescence intensities [6552 ± 451 vs. 6365 ± 420, p = 0.38]. Two of the six MGUS cases (33%) with >90% CD19(-) plasma cells showed progression of disease, whereas none of the cases with >10% CD19(+) plasma cells evolved to PCM.</p> <p>Conclusion</p> <p>MGUS cases with potential for disease progression appeared to lack CD19 expression on >90% of their plasma cells, displaying an immunophenotypic profile similar to PCM plasma cells. A higher relative proportion of CD19(+) plasma cells in MGUS may be associated with a lower potential for disease progression.</p

Results of the randomized phase IIB ADMIRE trial of FCR with or without mitoxantrone in previously untreated CLL

ADMIRE was a multi-center, randomized-controlled, open, phase IIB superiority trial in previously untreated Chronic Lymphocytic Leukemia (CLL). Conventional frontline therapy in fit patients is fludarabine, cyclophosphamide and rituximab (FCR). Initial evidence from non-randomized Phase II trials suggested that the addition of mitoxantrone to FCR (FCM-R) improved remission rates. 215 patients were recruited to assess the primary endpoint of complete remission (CR) rates according to IWCLL criteria. Secondary endpoints were progression-free survival (PFS), overall survival (OS), overall response rate, minimal residual disease (MRD) negativity and safety. At final analysis, CR rates were 69.8% FCR vs 69.3% FCM-R [adjusted odds ratio (OR): 0.97; 95%CI: (0.53-1.79), P=0.932]. MRD-negativity rates were 59.3% FCR vs 50.5% FCM-R [adjusted OR: 0.70; 95% CI: (0.39-1.26), P=0.231]. During treatment, 60.0% (n=129) of participants received G-CSF as secondary prophylaxis for neutropenia, a lower proportion on FCR compared with FCM-R (56.1 vs 63.9%). The toxicity of both regimens was acceptable. There are no significant differences between the treatment groups for PFS and OS. The trial demonstrated that the addition of mitoxantrone to FCR did not increase the depth of response. Oral FCR was well tolerated and resulted in impressive responses in terms of CR rates and MRD negativity compared to historical series with intravenous chemotherapy

Results of the randomized phase IIB ARCTIC trial of low dose Rituximab in previously untreated CLL

ARCTIC was a multi-center, randomized-controlled, open, phase IIB non-inferiority trial in previously untreated Chronic Lymphocytic Leukemia (CLL). Conventional frontline therapy in fit patients is fludarabine, cyclophosphamide and rituximab (FCR). The trial hypothesized that including mitoxantrone with low-dose rituximab (FCM-miniR) would be non-inferior to FCR. 200 patients were recruited to assess the primary endpoint of complete remission (CR) rates according to IWCLL criteria. Secondary endpoints were progression-free survival (PFS), overall survival (OS), overall response rate, minimal residual disease (MRD) negativity, safety and cost-effectiveness. The trial closed following the pre-planned interim analysis. At final analysis, CR rates were 76% FCR vs 55% FCM-miniR [adjusted odds-ratio: 0.37; 95% CI: 0.19–0.73]. MRD-negativity rates were 54% FCR vs 44% FCM-miniR. More participants experienced Serious Adverse Reactions with FCM-miniR (49%) compared to FCR (41%). There are no significant differences between the treatment groups for PFS and OS. FCM-miniR is not expected to be cost-effective over a lifetime horizon. In summary, FCM-miniR is less well tolerated than FCR with an inferior response and MRD-negativity rate and increased toxicity, and will not be taken forward into a confirmatory trial. The trial demonstrated that oral FCR yields high response rates compared to historical series with intravenous chemotherapy

Association of Minimal Residual Disease With Superior Survival Outcomes in Patients With Multiple Myeloma: A Meta-analysis

Importance: Numerous studies have evaluated the prognostic value of minimal residual disease (MRD) in patients with multiple myeloma (MM). Most studies were small and varied in terms of patient population, treatment, and MRD assessment methods. Objective: To evaluate the utility of MRD detection in patients with newly diagnosed MM. Data Sources: A Medline search was conducted for articles published in English between January 1990 and January 2016. Study Selection: Eligible studies reported MRD status and progression-free survival (PFS) or overall survival (OS) in 20 or more patients following treatment. Among 405 articles identified, 21 met the initial eligibility criteria and were included in the analysis. Data Extraction and Synthesis: Information on patient characteristics, treatment, MRD assessment, and outcomes were extracted using a standard form. Main Outcomes and Measures: The impact of MRD status on PFS and OS was assessed by pooling data from relevant trials. Data were adjusted to allow for different proportions of patients with MRD in different studies, and analyzed using the Peto method. Forest plots were created based on Cox model analysis. Other prespecified research questions were addressed qualitatively. Results: Fourteen studies (n = 1273) provided data on the impact of MRD on PFS, and 12 studies (n = 1100) on OS. Results were reported specifically in patients who had achieved conventional complete response (CR) in 5 studies for PFS (n = 574) and 6 studies for OS (n = 616). An MRD-negative status was associated with significantly better PFS overall (hazard ratio [HR], 0.41; 95% CI, 0.36-0.48; P < .001) and in studies specifically looking at CR patients (HR, 0.44; 95% CI, 0.34-0.56; P < .001). Overall survival was also favorable in MRD-negative patients overall (HR, 0.57; 95% CI, 0.46-0.71; P < .001) and in CR patients (HR, 0.47; 95% CI, 0.33-0.67; P < .001). Tests of heterogeneity found no significant differences among the studies for PFS and OS. Conclusions and Relevance: Minimal residual disease-negative status after treatment for newly diagnosed MM is associated with long-term survival. These findings provide quantitative evidence to support the integration of MRD assessment as an end point in clinical trials of MM

Ibrutinib induces chromatin reorganisation of chronic lymphocytic leukaemia cells

Chronic lymphocytic leukaemia (CLL) is the most common leukaemia in Western countries. It has recently been shown that the homogeneity of the chromatin landscape between CLL cells contrasts with the important observed genetic heterogeneity of the disease. To gain further insight into the consequences of disease evolution on the epigenome’s plasticity, we monitored changes in chromatin structure occurring in vivo in CLL cells from patients receiving continuous Ibrutinib treatment. Ibrutinib, an oral inhibitor of the Bruton’s tyrosine kinase (BTK) has proved to be remarkably efficient against treatment naïve (TN), heavily pre-treated and high-risk chronic lymphocytic leukaemia (CLL), with limited adverse events. We established that the chromatin landscape is significantly and globally affected in response to Ibrutinib. However, we observed that prior to treatment, CLL cells show qualitative and quantitative variations in chromatin structure correlated with both EZH2 protein level and cellular response to external stimuli. Then, under prolonged exposure to Ibrutinib, a loss of the two marks associated with lysine 27 (acetylation and trimethylation) was observed. Altogether, these data indicate that the epigenome of CLL cells from the peripheral blood change dynamically in response to stimuli and suggest that these cells might adapt to the Ibrutinib “hit” in a process leading toward a possible reduced sensitivity to treatment

Predicting and managing primary and secondary non-response to rituximab using B-cell biomarkers in systemic lupus erythematosus

Objective: To assess factors associated with primary and secondary non-response to rituximab in systemic lupus erythematosus (SLE) and evaluate management of secondary non-depletion non-response (2NDNR). Methods: 125 patients with SLE treated with rituximab over 12 years were studied prospectively. A major clinical response was defined as improvement of all active British Isles Lupus Assessment Group (BILAG)-2004 domains to grade C/better and no A/B flare. Partial responders were defined by one persistent BILAG B. B-cell subsets were measured using highly sensitive flow cytometry. Patients with 2NDNR, defined by infusion reaction and defective depletion, were treated with ocrelizumab or ofatumumab. Results: 117 patients had evaluable data. In cycle 1 (C1), 96/117 (82%) achieved BILAG response (major=50%, partial=32%). In multivariable analysis, younger age (OR 0.97, 95% CI 0.94 to 1.00) and B-cell depletion at 6 weeks (OR 3.22, 95% CI 1.24 to 8.33) increased the odds of major response. Complete depletion was predicted by normal complement and lower pre-rituximab plasmablasts and was not associated with increased serious infection post-rituximab. Seventy-seven (with data on 72) C1 responders were retreated on clinical relapse. Of these, 61/72 (85%) responded in cycle 2 (C2). Of the 11 C2 non-responders, nine met 2NDNR criteria (incidence=12%) and tested positive for anti-rituximab antibodies. Lack of concomitant immunosuppressant and higher pre-rituximab plasmablasts predicted 2NDNR. Five were switched to ocrelizumab/ofatumumab, and all depleted and responded. Conclusion: Treatment with anti-CD20 agents can be guided by B-cell monitoring and should aim to achieve complete depletion. 2NDNR is associated with anti-rituximab antibodies, and switching to humanised agents restores depletion and response. In SLE, alternative anti-CD20 antibodies may be more consistently effective

Results of the randomized phase IIB ADMIRE trial of FCR with or without mitoxantrone in previously untreated CLL

ADMIRE was a multi-center, randomized-controlled, open, phase IIB superiority trial in previously untreated Chronic Lymphocytic Leukemia (CLL). Conventional frontline therapy in fit patients is fludarabine, cyclophosphamide and rituximab (FCR). Initial evidence from non-randomized Phase II trials suggested that the addition of mitoxantrone to FCR (FCM-R) improved remission rates. 215 patients were recruited to assess the primary endpoint of complete remission (CR) rates according to IWCLL criteria. Secondary endpoints were progression-free survival (PFS), overall survival (OS), overall response rate, minimal residual disease (MRD) negativity and safety. At final analysis, CR rates were 69.8% FCR vs 69.3% FCM-R [adjusted odds ratio (OR): 0.97; 95%CI: (0.53-1.79), P=0.932]. MRD-negativity rates were 59.3% FCR vs 50.5% FCM-R [adjusted OR: 0.70; 95% CI: (0.39-1.26), P=0.231]. During treatment, 60.0% (n=129) of participants received G-CSF as secondary prophylaxis for neutropenia, a lower proportion on FCR compared with FCM-R (56.1 vs 63.9%). The toxicity of both regimens was acceptable. There are no significant differences between the treatment groups for PFS and OS. The trial demonstrated that the addition of mitoxantrone to FCR did not increase the depth of response. Oral FCR was well tolerated and resulted in impressive responses in terms of CR rates and MRD negativity compared to historical series with intravenous chemotherapy

GA101 (obinutuzumab) monocLonal Antibody as Consolidation Therapy In CLL (GALACTIC) trial: study protocol for a phase II/III randomised controlled trial

Background: Chronic lymphocytic leukaemia (CLL) is the most common adult leukaemia. Achieving minimal residual disease (MRD) negativity in CLL is an independent predictor of survival even with a variety of different treatment approaches and regardless of the line of therapy. Methods/design: GA101 (obinutuzumab) monocLonal Antibody as Consolidation Therapy In CLL (GALACTIC) is a seamless phase II/III, multi-centre, randomised, controlled, open, parallel-group trial for patients with CLL who have recently responded to chemotherapy. Participants will be randomised to receive either obinutuzumab (GA-101) consolidation or no treatment (as is standard). The phase II trial will assess safety and short-term efficacy in order to advise on continuation to a phase III trial. The primary objective for phase III is to assess the effect of consolidation therapy on progression-free survival (PFS). One hundred eighty-eight participants are planned to be recruited from forty research centres in the United Kingdom. Discussion: There is evidence that achieving MRD eradication with alemtuzumab consolidation is associated with improvements in survival and time to progression. This trial will assess whether obinutuzumab is safe in a consolidation setting and effective at eradicating MRD and improving PFS. Trial registration: ISRCTN, 64035629. Registered on 12 January 2015. EudraCT, 2014-000880-42. Registered on 12 November 2014

Predicting Sustained Clinical Response to Rituximab in Moderate to Severe Systemic Manifestations of Primary Sjögren Syndrome

Objective To assess outcomes of repeat rituximab cycles and identify predictors of sustained clinical response in systemic manifestations of primary Sjögren syndrome (pSS). Methods An observational study was conducted in 40 rituximab-treated patients with pSS. Clinical response was defined as a 3-point or more reduction in the European League Against Rheumatism (EULAR) Sjögren Disease Activity Index (ESSDAI) at 6 months from baseline. Peripheral blood B cells were measured using highly sensitive flow cytometry. Predictors of sustained response (within two rituximab cycles) were analyzed using penalized logistic regression. Results Thirty-eight out of 40 patients had moderate to severe systemic disease (ESSDAI >5). Main domains were articular (73%), mucocutaneous (23%), hematological (20%), and nervous system (18%). Twenty-eight out of 40 (70%) patients were on concomitant immunosuppressants. One hundred sixty-nine rituximab cycles were administered with a total follow-up of 165 patient-years. In cycle 1 (C1), 29/40 (73%) achieved ESSDAI response. Of C1 responders, 23/29 received retreatment on clinical relapse, and 15/23 (65%) responded. Of the 8/23 patients who lost response, these were due to secondary non-depletion and non-response (2NDNR; 4/23 [17%] as we previously observed in systemic lupus erythematosus with antirituximab antibodies, inefficacy = 2/23, and other side effects = 2/23). Within two cycles, 13/40 (33%) discontinued therapy. In multivariable analysis, concomitant immunosuppressant (odds ratio 7.16 [95% confidence interval: 1.37–37.35]) and achieving complete B-cell depletion (9.78 [1.32–72.25]) in C1 increased odds of response to rituximab. At 5 years, 57% of patients continued on rituximab. Conclusion Our data suggest that patients with pSS should be co-prescribed immunosuppressant with rituximab, and treatment should aim to achieve complete depletion. About one in six patients develop 2NDNR in repeat cycles. Humanized or type 2 anti-CD20 antibodies may improve clinical response in extra-glandular pSS