A phase 2 study of venetoclax plus R-CHOP as first-line treatment for patients with diffuse large B-cell lymphoma.

The phase 2 CAVALLI (NCT02055820) study assessed efficacy and safety of venetoclax, a selective B-cell lymphoma-2 (Bcl-2) inhibitor, with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) in first-line (1L) diffuse large B-cell lymphoma (DLBCL), including patients demonstrating Bcl-2 protein overexpression by immunohistochemistry (Bcl-2 IHC+). Eligible patients were ≥18 years of age and had previously untreated DLBCL, Eastern Cooperative Oncology Group performance status ≤2, and International Prognostic Index 2 to 5. Venetoclax 800 mg (days 4-10, cycle 1; days 1-10, cycles 2-8) was administered with rituximab (8 cycles) and cyclophosphamide, doxorubicin, vincristine, and prednisone (6-8 cycles) in 21-day cycles. Primary end points were safety, tolerability, and research_plete response (CR) at end of treatment (EOT). Secondary end points were progression-free survival (PFS) and overall survival. Comparative analyses used covariate-adjusted R-CHOP controls from the GOYA/BO21005 study, an appropriate contemporary benchmark for safety and efficacy. Safety and efficacy analyses included 206 patients. CR rate at EOT was 69% in the overall population and was maintained across Bcl-2 IHC+ subgroups. With a median follow-up of 32.2 months, trends were observed for improved investigator-assessed PFS for venetoclax plus R-CHOP in the overall population (hazard ratio [HR], 0.61; 95% confidence interval [CI], 0.43-0.87) and Bcl-2 IHC+ subgroups (HR, 0.55; 95% CI, 0.34-0.89) vs R-CHOP. Despite a higher incidence of grade 3/4 hematologic adverse events (86%), related mortality was not increased (2%). Chemotherapy dose intensity was similar in CAVALLI vs GOYA. The addition of venetoclax to R-CHOP in 1L DLBCL demonstrates increased, but manageable, myelosuppression and the potential of improved efficacy, particularly in high-risk Bcl-2 IHC+ patient subgroups.

Venetoclax-rituximab with or without bendamustine vs bendamustine-rituximab in relapsed/refractory follicular lymphoma.

This open-label phase 2 study (CONTRALTO) assessed the safety and efficacy of BCL-2 inhibitor venetoclax (VEN) plus rituximab (R), and VEN plus bendamustine (B) and R, vs B + R (BR) alone in relapsed/refractory (R/R) follicular lymphoma. Patients in the chemotherapy-free arm (arm A: VEN + R) received VEN 800 mg/d plus R 375 mg/m2 on days 1, 8, 15, and 22 of cycle 1 and day 1 of cycles 4, 6, 8, 10, and 12. After a safety run-in with VEN 600 mg, patients in the chemotherapy-containing cohort were randomized to either VEN + BR (arm B; VEN 800 mg/d for 1 year + 6 cycles of BR [B 90 mg/m2 on days 1 and 2 and R 375 mg/m2 on day 1]) or 6 cycles of BR (arm C). Overall, 163 patients were analyzed (9 in the safety run-in and 52, 51, and 51 in arms A, B, and C, respectively). Complete metabolic/complete response rates were 17% (arm A), 75% (arm B), and 69% (arm C). Of patients in arm B, only 61% received ≥90% of the planned B dose vs 96% of patients in arm C. More frequent hematologic toxicity resulted in more reduced dosing/treatment discontinuation in arm B vs arm C. Rates of grade 3/4 adverse events were 51.9%, 93.9%, and 60.0% in arms A, B, and C, respectively. VEN + BR led to increased toxicity and lower dose intensity of BR than in arm C, but efficacy was similar. Optimizing dose and schedule to maintain BR dose intensity may improve efficacy and tolerability of VEN + BR, while VEN + R data warrant further study. This study was registered at www.clinicaltrials.gov as #NCT02187861.

Minimal Residual Disease Status Predicts Outcome in Patients With Previously Untreated Follicular Lymphoma: A Prospective Analysis of the Phase III GALLIUM Study.

PURPOSE: We report an analysis of minimal residual/detectable disease (MRD) as a predictor of outcome in previously untreated patients with follicular lymphoma (FL) from the randomized, multicenter GALLIUM (ClinicalTrials.gov identifier: NCT01332968) trial. PATIENTS AND METHODS: Patients received induction with obinutuzumab (G) or rituximab (R) plus bendamustine, or cyclophosphamide, doxorubicin, vincristine, prednisone (CHOP) or cyclophosphamide, vincristine, prednisone (CVP) chemotherapy, followed by maintenance with the same antibody in responders. MRD status was assessed at predefined time points (mid-induction [MI], end of induction [EOI], and at 4-6 monthly intervals during maintenance and follow-up). Patients with evaluable biomarker data at diagnosis were included in the survival analysis. RESULTS: MRD positivity was associated with inferior progression-free survival (PFS) at MI (hazard ratio [HR], 3.03 [95% CI, 2.07 to 4.45]; P < .0001) and EOI (HR, 2.25 [95% CI, 1.53 to 3.32]; P < .0001). MRD response was higher after G- versus R-chemotherapy at MI (94.2% v 88.9%; P = .013) and at EOI (93.1% v 86.7%; P = .0077). Late responders (MI-positive/EOI-negative) had a significantly poorer PFS than early responders (MI-negative/EOI-negative; HR, 3.11 [95% CI, 1.75 to 5.52]; P = .00011). The smallest proportion of MRD positivity was observed in patients receiving bendamustine at MI (4.8% v 16.0% in those receiving CHOP; P < .0001). G appeared to compensate for less effective chemotherapy regimens, with similar MRD response rates observed across the G-chemo groups. During the maintenance period, more patients treated with R than with G were MRD-positive (R-CHOP, 20.7% v G-CHOP, 7.0%; R-CVP, 21.7% v G-CVP, 9.4%). Throughout maintenance, MRD positivity was associated with clinical relapse. CONCLUSION: MRD status can determine outcome after induction and during maintenance, and MRD negativity is a prerequisite for long-term disease control in FL. The higher MRD responses after G- versus R-based treatment confirm more effective tumor cell clearance.

Npr2, yeast homolog of the human tumor suppressor NPRL2, is a target of Grr1 required for adaptation to growth on diverse nitrogen sources.

Npr2, a putative "nitrogen permease regulator" and homolog of the human tumor suppressor NPRL2, was found to interact with Grr1, the F-box component of the SCF(Grr1) (Skp1-cullin-F-box protein complex containing Grr1) E3 ubiquitin ligase, by mass spectrometry-based multidimensional protein identification technology. Npr2 has two PEST sequences and has been previously identified among ubiquitinated proteins. Like other Grr1 targets, Npr2 is a phosphoprotein. Phosphorylated Npr2 accumulates in grr1Delta mutants, and Npr2 is stabilized in cells with inactivated proteasomes. Phosphorylation and instability depend upon the type I casein kinases (CK1) Yck1 and Yck2. Overexpression of Npr2 is detrimental to cells and is lethal in grr1Delta mutants. Npr2 is required for robust growth in defined medium containing ammonium or urea as a nitrogen source but not for growth on rich medium. npr2Delta mutants also fail to efficiently complete meiosis. Together, these data indicate that Npr2 is a phosphorylation-dependent target of the SCF(Grr1) E3 ubiquitin ligase that plays a role in cell growth on some nitrogen sources.

Identification of a conserved F-box protein 6 interactor essential for endocytosis and cytokinesis in fission yeast.

The F-box domain is a degenerated motif consisting of approximately 40 amino acid residues that specifically bind Skp1, a core component of the SCF (Skp1-Cdc53/Cullin 1-F-box protein) ubiquitin ligase. Recent work, mainly performed in budding yeast, indicates that certain F-box proteins form non-SCF complexes together with Skp1 in the absence of cullins and play various roles in cell cycle and signalling pathways. However, it is not established whether these non-SCF complexes are unique to budding yeast or common in other eukaryotes. In the present paper, using TAP (tandem affinity purification) coupled to MudPIT (Multidimensional Protein Identification Technology) analysis, we have identified a novel conserved protein, Sip1, in fission yeast, as an interacting partner of an essential F-box protein Pof6. Sip1 is a large HEAT (huntingtin, elongation factor 3, the PR65/A subunit of protein phosphatase 2A and the lipid kinase Tor)-repeats containing protein (217 kDa) and forms a complex with Pof6 and Skp1. This complex does not contain cullins, indicating that it is a novel non-SCF complex. Like Pof6 and Skp1, Sip1 is essential for cell viability and temperature-sensitive sip1 mutants display cell division arrest as binucleate cells with septa. Sip1 localizes to the nucleus and dynamic cytoplasmic dots, which are shown in the present study to be endocytic vesicles. Consistent with this, sip1 mutants are defective in endocytosis. Furthermore, towards the end of cytokinesis, constriction of the actomyosin ring and dissociation of type II myosin and septum materials are substantially delayed in the absence of functional Sip1. These results indicate that the conserved Sip1 protein comprises a novel non-SCF F-box complex that plays an essential role in endocytosis, cytokinesis and cell division.

MRD response in relapsed/refractory FL after obinutuzumab plus bendamustine or bendamustine alone in the GADOLIN trial.

We report assessment of minimal residual disease (MRD) status and its association with outcome in rituximab-refractory follicular lymphoma (FL) in the randomized GADOLIN trial (NCT01059630). Patients received obinutuzumab (G) plus bendamustine (Benda) induction followed by G maintenance, or Benda induction alone. Patients with a clonal marker (t[14;18] translocation and/or immunoglobulin heavy or light chain rearrangement) detected at study screening were assessed for MRD at mid-induction (MI), end of induction (EOI), and every 6-24 months post-EOI/discontinuation by real-time quantitative PCR. At MI, 41/52 (79%) patients receiving G-Benda were MRD-negative vs. 17/36 (47%) patients receiving Benda alone (p = 0.0029). At EOI, 54/63 (86%) patients receiving G-Benda were MRD-negative vs. 30/55 (55%) receiving Benda alone (p = 0.0002). MRD-negative patients at EOI had improved progression-free survival (HR, 0.33, 95% CI, 0.19-0.56, p < 0.0001) and overall survival (HR, 0.39, 95% CI, 0.19-0.78, p = 0.008) vs. MRD-positive patients, and maintained their MRD-negative status for longer if they received G maintenance than if they did not. These results suggest that the addition of G to Benda-based treatment during induction can significantly contribute to the speed and depth of response, and G maintenance in MRD-negative patients potentially delays lymphoma regrowth.

Prognostic Impact of Natural Killer Cell Count in Follicular Lymphoma and Diffuse Large B-cell Lymphoma Patients Treated with Immunochemotherapy.

PURPOSE: Natural killer (NK) cells are key effector cells for anti-CD20 monoclonal antibodies (mAb), such as obinutuzumab and rituximab. We assessed whether low pretreatment NK-cell count (NKCC) in peripheral blood or tumor tissue was associated with worse outcome in patients receiving antibody-based therapy. PATIENTS AND METHODS: Baseline peripheral blood NKCC was assessed by flow cytometry (CD3-CD56+ and/or CD16+ cells) in 1,064 of 1,202 patients with follicular lymphoma treated with obinutuzumab or rituximab plus chemotherapy in the phase III GALLIUM trial (NCT01332968) and 1,287 of 1,418 patients with diffuse large B-cell lymphoma (DLBCL) treated with obinutuzumab or rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (G-CHOP or R-CHOP) in the phase III GOYA trial (NCT01287741). The prognostic value of tumor NK-cell gene expression, as assessed by whole-transcriptome gene expression using TruSeq RNA sequencing, was also analyzed. The association of baseline variables, such as treatment arm, was evaluated using multivariate Cox regression models using a stepwise approach. RESULTS: In this exploratory analysis, low baseline peripheral blood NKCC was associated with shorter progression-free survival (PFS) in both follicular lymphoma [hazard ratio (HR), 1.48; 95% confidence interval (CI), 1.02-2.14; P = 0.04] and DLBCL (HR, 1.36; 95% CI, 1.01-1.83; P = 0.04), and overall survival in follicular lymphoma (HR, 2.20; 95% CI, 1.26-3.86; P = 0.0058). Low tumor NK-cell gene expression was associated with shorter PFS in G-CHOP-treated patients with DLBCL (HR, 1.95; 95% CI, 1.22-3.15; P < 0.01). CONCLUSIONS: These findings indicate that the number of NK cells in peripheral blood may affect the outcome of patients with B-cell non-Hodgkin lymphoma receiving anti-CD20-based immunochemotherapy.

Regulation and recognition of SCFGrr1 targets in the glucose and amino acid signaling pathways.

SCFGrr1, one of several members of the SCF family of E3 ubiquitin ligases in budding Saccharomyces cerevisiae, is required for both regulation of the cell cycle and nutritionally controlled transcription. In addition to its role in degradation of Gic2 and the CDK targets Cln1 and Cln2, Grr1 is also required for induction of glucose- and amino acid-regulated genes. Induction of HXT genes by glucose requires the Grr1-dependent degradation of Mth1. We show that Mth1 is ubiquitinated in vivo and degraded via the proteasome. Furthermore, phosphorylated Mth1, targeted by the casein kinases Yck1/2, binds to Grr1. That binding depends upon the Grr1 leucine-rich repeat (LRR) domain but not upon the F-box or basic residues within the LRR that are required for recognition of Cln2 and Gic2. Those observations extend to a large number of Grr1-dependent genes, some targets of the amino acid-regulated SPS signaling system, which are properly regulated in the absence of those basic LRR residues. Finally, we show that regulation of the SPS targets requires the Yck1/2 casein kinases. We propose that casein kinase I plays a similar role in both nutritional signaling pathways by phosphorylating pathway components and targeting them for ubiquitination by SCFGrr1.

Grr1-dependent inactivation of Mth1 mediates glucose-induced dissociation of Rgt1 from HXT gene promoters.

In budding yeast, HXT genes encoding hexose permeases are induced by glucose via a mechanism in which the F box protein Grr1 antagonizes activity of the transcriptional repressor Rgt1. Neither the mechanism of Rgt1 inactivation nor the role of Grr1 in that process has been understood. We show that glucose promotes phosphorylation of Rgt1 and its dissociation from HXT gene promoters. This cascade of events is dependent upon the F-box protein Grr1. Inactivation of Rgt1 is sufficient to explain the requirement for Grr1 but does not involve Rgt1 proteolysis or ubiquitination. We show that inactivation of Mth1 and Std1, known negative regulators of HXT gene expression, leads to the hyperphosphorylation of Rgt1 and its dissociation from HXT promoters even in the absence of glucose. Furthermore, inactivation of Mth1 and Std1 bypasses the requirement for Grr1 for induction of these events, suggesting they are targets for inactivation by Grr1. Consistent with that proposal, Mth1 is rapidly eliminated in response to glucose via a mechanism that requires Grr1. Based upon these data, we propose that glucose acts via Grr1 to promote the degradation of Mth1. Degradation of Mth1 leads to phosphorylation and dissociation of Rgt1 from HXT promoters, thereby activating HXT gene expression.