Survival, neurocognitive function, and health-related quality of life outcomes after rituximab-methotrexate, BCNU, teniposide, and prednisolone for primary CNS lymphoma: Final results of the HOVON 105/ALLG NHL 24 study.

BACKGROUND: Studies on the efficacy of rituximab in primary CNS lymphoma (PCNSL) reported conflicting results. Our international randomized phase 3 study showed that the addition of rituximab to high-dose methotrexate, BCNU, teniposide, and prednisolone (MBVP) in PCNSL was not efficacious in the short term. Here we present long-term results after a median follow-up of 82.3 months. METHODS: One hundred and ninety-nine eligible newly diagnosed, nonimmunocompromised patients with PCNSL aged 18-70 years with WHO performance status 0-3 was randomized between treatment with MBVP chemotherapy with or without rituximab, followed by high-dose cytarabine consolidation in responding patients, and reduced-dose WBRT in patients aged ≤ 60 years. Event-free survival was the primary endpoint. Overall survival rate, neurocognitive functioning (NCF), and health-related quality of life (HRQoL) were additionally assessed, with the IPCG test battery, EORTC QLQ-C30 and QLQ-BN20 questionnaires, respectively. RESULTS: For event-free survival, the hazard ratio was 0.85, 95% CI 0.61-1.18, P = .33. Overall survival rate at 5 years for MBVP and R-MBVP was 49% (39-59) and 53% (43-63) respectively. In total, 64 patients died in the MBVP arm and 55 in the R-MBVP arm, of which 69% were due to PCNSL. At the group level, all domains of NCF and HRQoL improved to a clinically relevant extent after treatment initiation, and remained stable thereafter up to 60 months of follow-up, except for motor speed which deteriorated between 24 and 60 months. Although fatigue improved initially, high levels persisted in the long term. CONCLUSIONS: Long-term follow-up confirms the lack of added value of rituximab in addition to MBVP and HD-cytarabine for PCNSL.

The clinical features, management and outcomes of lymphoma in pregnancy: A multicentre study by the Australasian Lymphoma Alliance.

Lymphoma in pregnancy (LIP) presents unique clinical, social and ethical challenges; however, the evidence regarding this clinical scenario is limited. We conducted a multicentre retrospective observational study reporting on the features, management, and outcomes of LIP in patients diagnosed between January 2009 and December 2020 at 16 sites in Australia and New Zealand for the first time. We included diagnoses occurring either during pregnancy or within the first 12 months following delivery. A total of 73 patients were included, 41 diagnosed antenatally (AN cohort) and 32 postnatally (PN cohort). The most common diagnoses were Hodgkin lymphoma (HL; 40 patients), diffuse large B-cell lymphoma (DLBCL; 11) and primary mediastinal B-cell lymphoma (PMBCL; six). At a median follow up of 2.37 years, the 2- and 5-year overall survival (OS) for patients with HL were 91% and 82%. For the combined DLBCL and PMBCL group, the 2-year OS was 92%. Standard curative chemotherapy regimens were successfully delivered to 64% of women in the AN cohort; however, counselling regarding future fertility and termination of pregnancy were suboptimal, and a standardised approach to staging lacking. Neonatal outcomes were generally favourable. We present a large multicentre cohort of LIP reflecting contemporary practice and identify areas in need of ongoing research.

Capturing the lived experiences of women with lymphoma in pregnancy: a qualitative study.

Lymphoma in pregnancy is a rare and challenging diagnosis that complicates ∼1:6000 pregnancies; posing a series of unique therapeutic, social, and ethical challenges to the patient, her family, and the medical professionals involved. These difficulties are compounded by the paucity of real-world data on the management of LIP, and a lack of relevant support systems for women in this setting. We conducted a retrospective multicenter qualitative study, interviewing women aged ≥18 years of age diagnosed with Hodgkin (HL) or non-Hodgkin lymphoma (NHL) during pregnancy or within 12 months postpartum, between 1 January 2009 and 31 December 2020 from 13 Australasian sites. Semi-structured telephone interviews were conducted, recorded, and analyzed using QSR Int NVivo 12 Pro (March 2020, USA) to quantify salient themes. Of the 32 women interviewed, 20 (63%) were diagnosed during pregnancy (16, 34, and 13% in the 1st, 2nd, and 3rd trimesters, respectively), while 12 (37%) were diagnosed post-partum. Women recalled that their chief concerns at diagnosis were the welfare of their child (n = 13, 41%) and a fear of dying (n = 9, 28%). Perceived diagnostic delay attributed to pregnancy was reported by 41% of participants. Other key themes were communication, educational materials, psychosocial supports, and oncofertility issues. To our knowledge this is the first report capturing the lived experiences of survivors of lymphoma during pregnancy, affording a unique opportunity to consider the management, psychosocial supports, and delivery of care to meet the needs of these women.What is the NEW aspect of your work? To our knowledge, this is the first report capturing and analyzing the healthcare experiences of survivors of Lymphoma in Pregnancy (LIP).What is the CENTRAL finding of your work? Women valued clear and empathic communication, provision of tailored educational materials, access to psychosocial supports (particularly childcare and financial supports), and timely oncofertility management in their healthcare journey.What is (or could be) the SPECIFIC clinical relevance of your work? Women's personal accounts of positive and negative experiences of LIP care provide insights into their specific concerns and needs which can shape healthcare policy and development of a specific framework for managing and supporting patients with LIP (and other cancers).

Proinflammatory microenvironment promotes lymphoma progression in mice with high megakaryocyte and TPO levels.

Platelets have been shown to enhance the survival of lymphoma cell lines. However, it remains unclear whether they play a role in lymphoma. Here, we investigated the potential role of platelets and/or megakaryocytes in the progression of Eµ-myc lymphoma. Eµ-myc tumor cells were transplanted into recipient wild-type (WT) control, Mpl-/-, or TpoTg mice, which exhibited normal, low, and high platelet and megakaryocyte counts, respectively. TpoTg mice that underwent transplantation exhibited enhanced lymphoma progression with increased white blood cell (WBC) counts, spleen and lymph node weights, and enhanced liver infiltration when compared with WT mice. Conversely, tumor-bearing Mpl-/- mice had reduced WBC counts, lymph node weights, and less liver infiltration than WT mice. Using an Mpl-deficient thrombocytopenic immunocompromised mouse model, our results were confirmed using the human non-Hodgkin lymphoma GRANTA cell line. Although we found that platelets and platelet-released molecules supported Eµ-myc tumor cell survival in vitro, pharmacological inhibition of platelet function or anticoagulation in WT mice transplanted with Eµ-myc did not improve disease outcome. Furthermore, transient platelet depletion or sustained Bcl-xL-dependent thrombocytopenia did not alter lymphoma progression. Cytokine analysis of the bone marrow fluid microenvironment revealed increased levels of the proinflammatory molecule interleukin 1 in TpoTg mice, whereas these levels were lower in Mpl-/- mice. Moreover, RNA sequencing of blood-resident Eµ-myc lymphoma cells from TpoTg and WT mice after tumor transplantation revealed the upregulation of hallmark gene sets associated with an inflammatory response in TpoTg mice. We propose that the proinflammatory microenvironment in TpoTg mice promotes lymphoma progression.

Cancer during pregnancy: A qualitative study of healthcare experiences of Australian women.

OBJECTIVES: To identify features enhancing the quality of healthcare experiences for women with gestational cancer, and explore the impact of the heterogeneous Australian healthcare system on those experiences. METHODS: Semi-structured, qualitative interviews were conducted with women diagnosed with any cancer during pregnancy in the last five years. Recruitment occurred during 2018-2019 via social media and professional, clinical and community networks. Questions related to women's experiences of their healthcare, wellbeing and psychosocial needs. Interviews were analysed thematically. RESULTS: Study participants (n = 23) received treatment in the private sector (n = 10), public sector (n = 8), or both (n = 5). Five interview themes were found: Control over healthcare; Trust in clinicians, hospitals and systems; Coordination of care; An uncommon diagnosis; Holistic, future-oriented care. Women were most likely to have had a positive healthcare experience when (a)care was well-coordinated and adjusted to meet their unique needs/challenges, and (b)women perceived their care went beyond their immediate medical needs and encompassed future psychosocial wellbeing, including preparation for postpartum challenges. CONCLUSION: Existing 'usual care' in the public and/or private sector for both the pregnancy and the cancer is insufficient to meet these women's needs. Prioritising psychological wellbeing including psychosocial needs, and communication and planning around fertility and postnatal challenges are essential for this population.

Open-Label, phase 2 study of blinatumomab as second salvage therapy in adults with relapsed/refractory aggressive B-cell non-Hodgkin lymphoma.

The phase 2 portion of this open-label phase 2/3 study assessed the efficacy and safety of blinatumomab as second salvage for aggressive relapsed or refractory (r/r) aggressive B-cell non-Hodgkin lymphoma (B-NHL) following platinum-based first salvage chemotherapy. Forty-one patients with aggressive disease (32% relapsed; 68% refractory) enrolled and received stepwise blinatumomab (9-28-112 µg/day) in a 70-day cycle 1 and an optional 28-day cycle 2; 19 (46%) completed cycle 1 and 3 (7%) completed cycle 2. The overall response rate after 12 weeks was 37%, including 9 (22%) complete metabolic responses. Eight (20%) patients (all responders) subsequently received stem cell transplants. Grade ≥3 adverse events were reported in 29 (71%) patients. Grade 3 cytokine release syndrome occurred in one patient. Grade 3 neurologic events occurred in 10 (24%) patients; all resolved. Blinatumomab monotherapy appears effective as second salvage therapy in patients with r/r aggressive B-NHL. Trial registration: NCT02910063.

Venetoclax plus R- or G-CHOP in non-Hodgkin lymphoma: results from the CAVALLI phase 1b trial.

Novel strategies, such as chemosensitization with targeted agents, that build on the success of standard immunochemotherapy show promise for the treatment of non-Hodgkin lymphoma (NHL). Here, we report a phase 1b study investigating dose escalation of the BCL2 inhibitor, venetoclax, in combination with rituximab or obinutuzumab and cyclophosphamide, doxorubicin, vincristine, and prednisone (R-/G-CHOP) chemotherapy in B-cell NHL. Objectives included safety assessment and determination of a recommended phase 2 dose (RP2D). Fifty-six patients were enrolled, most with follicular lymphoma (43%) or diffuse large B-cell lymphoma (DLBCL; 32%). Dose-limiting toxicities were reported in 3/14 patients at the first venetoclax dose (200 mg/d), after which dosing was changed from daily to 10 days per cycle and escalated to 800 mg. A further reduction to 5 days per cycle occurred at the 800-mg dose level in the G-CHOP arm. Cytopenias were predominant among grade 3/4 events and reported at a higher rate than expected, particularly in the G-CHOP arm; however, safety was manageable. Overall response rates were 87.5% (R-CHOP and G-CHOP combinations); complete response (CR) rates were 79.2% and 78.1%, respectively. Most double-expressor (BCL2+ and MYC+) DLBCL patients (87.5%; n = 7/8) achieved CR. Although the maximum tolerated dose was not reached, the RP2D for venetoclax with R-CHOP was established at 800 mg days 4 to 10 of cycle 1 and days 1 to 10 of cycles 2 to 8; higher doses were not explored, and this dosing schedule demonstrated an acceptable safety profile. This regimen is subsequently being evaluated in first-line DLBCL in the phase 2 portion of the study. This trial was registered at www.clinicaltrials.gov as #NCT02055820.

Rituximab in patients with primary CNS lymphoma (HOVON 105/ALLG NHL 24): a randomised, open-label, phase 3 intergroup study.

BACKGROUND: The prognosis for primary CNS lymphoma has improved with the use of high-dose methotrexate-based chemotherapy, but patient outcomes remain poor. Rituximab, a chimeric monoclonal antibody that targets the CD20 cell surface protein, has substantial activity in systemic CD20-positive diffuse large B-cell lymphoma, but its efficacy in primary CNS lymphoma is unknown and low penetration of the large rituximab molecule through the blood-brain barrier could limit its effect. We aimed to investigate the addition of rituximab to a high-dose methotrexate-based chemotherapy regimen in patients with newly diagnosed primary CNS lymphoma. METHODS: This intergroup, multicentre, open-label, randomised phase 3 study was done at 23 hospitals in the Netherlands, Australia, and New Zealand. Non-immunocompromised patients aged 18-70 years with newly diagnosed primary CNS lymphoma were randomly assigned (1:1) to receive methotrexate-based chemotherapy with or without intravenous rituximab. We used a web-based randomisation system with stratification by centre, age, and Eastern Cooperative Oncology Group-WHO performance status, and a minimisation procedure. All group assignment was open label and neither investigators nor patients were masked to allocation. All patients were treated with two 28-day cycles of induction chemotherapy, consisting of intravenous methotrexate 3 g per m2 on days 1 and 15, intravenous carmustine 100 mg per m2 on day 4, intravenous teniposide 100 mg per m2 on days 2 and 3, and oral prednisone 60 mg per m2 on days 1-5, with (R-MBVP) or without (MBVP) intravenous rituximab 375 mg per m2 on days 0, 7, 14, and 21 in cycle one and days 0 and 14 in cycle two. Patients with response at the end of induction subsequently received high-dose cytarabine and, in patients aged 60 years or younger, low-dose whole-brain radiotherapy. The primary endpoint was event-free survival, with events defined as not reaching complete response or complete response unconfirmed at the end of treatment, or progression or death after response; analysis was adjusted for age and performance score. Patients were analysed on a modified intention-to-treat basis. This trial is registered with the Nederlands Trial Register, number NTR2427, and the Australian New Zealand Clinical Trials Registry, number ACTRN12610000908033. The trial was closed on May 27, 2016, after achieving complete accrual, and follow-up is ongoing. FINDINGS: Between Aug 3, 2010, and May 27, 2016, we recruited 200 patients (109 men and 91 women; median age was 61 years [IQR 55-67]). We randomly assigned 100 patients to MBVP and 99 patients to R-MBVP. One patient was randomly assigned to the R-MBVP group but found to be ineligible because of an incorrect diagnosis and was excluded from all analyses. After a median follow-up of 32·9 months (IQR 23·9-51·5), 98 patients had had an event (51 in the MBVP group and 47 in the R-MBVP group), of whom 79 had died (41 in the MBVP group and 38 in the R-MBVP group). Event-free survival at 1 year was 49% (95% CI 39-58) in the MBVP group (no rituximab) and 52% (42-61) in the R-MBVP group (with rituximab; hazard ratio 1·00, 95% CI 0·70-1·43, p=0·99). Grade 3 or 4 adverse events occurred in 58 (58%) patients in the MBVP group and 63 (64%) patients in the R-MBVP group, with infections (24 [24%] patients receiving MBVP vs 21 [21%] patients receiving R-MBVP), haematological toxicity (15 [15%] vs 12 [12%]), and nervous system disorders (ten [10%] vs 15 [15%]) being the most common. Life-threatening or fatal serious adverse events occurred in 12 (12%) patients in the MBVP group and ten (10%) patients in the R-MBVP group, and five (5%) patients in the MBVP group and three (3%) in the R-MBVP group died from treatment-related causes. INTERPRETATION: We found no clear benefit of addition of rituximab to methotrexate, carmustine, teniposide, and prednisone chemotherapy in primary CNS lymphoma. Therefore, the results of this study do not support the use of rituximab as a component of standard treatment in primary CNS lymphoma. FUNDING: Roche, the Dutch Cancer Society, and Stichting STOPhersentumoren.

Altered B-lymphopoiesis in mice with deregulated thrombopoietin signaling.

Thrombopoietin (TPO) is the master cytokine regulator of megakaryopoiesis. In addition to regulation of megakaryocyte and platelet number, TPO is important for maintaining proper hematopoietic stem cell (HSC) function. It was previously shown that a number of lymphoid genes were upregulated in HSCs from Tpo -/- mice. We investigated if absent or enhanced TPO signaling would influence normal B-lymphopoiesis. Absent TPO signaling in Mpl -/- mice led to enrichment of a common lymphoid progenitor (CLP) signature in multipotential lineage-negative Sca-1+c-Kit+ (LSK) cells and an increase in CLP formation. Moreover, Mpl -/- mice exhibited increased numbers of PreB2 and immature B-cells in bone marrow and spleen, with an increased proportion of B-lymphoid cells in the G1 phase of the cell cycle. Conversely, elevated TPO signaling in Tpo Tg mice was associated with reduced B-lymphopoiesis. Although at steady state, peripheral blood lymphocyte counts were normal in both models, Mpl -/- Eµ-myc mice showed an enhanced preneoplastic phase with increased numbers of splenic PreB2 and immature B-cells, a reduced quiescent fraction, and augmented blood lymphocyte counts. Thus, although Mpl is not expressed on lymphoid cells, TPO signaling may indirectly influence B-lymphopoiesis and the preneoplastic state in Myc-driven B-cell lymphomagenesis by lineage priming in multipotential progenitor cells.

Autoreactive T cells induce necrosis and not BCL-2-regulated or death receptor-mediated apoptosis or RIPK3-dependent necroptosis of transplanted islets in a mouse model of type 1 diabetes.

AIMS/HYPOTHESIS: Type 1 diabetes results from T cell-mediated destruction of pancreatic beta cells. The mechanisms of beta cell destruction in vivo, however, remain unclear. We aimed to test the relative roles of the main cell death pathways: apoptosis, necrosis and necroptosis, in beta cell death in the development of CD4(+) T cell-mediated autoimmune diabetes. METHODS: We altered expression levels of critical cell death proteins in mouse islets and tested their ability to survive CD4(+) T cell-mediated attack using an in vivo graft model. RESULTS: Loss of the B cell leukaemia/lymphoma 2 (BCL-2) homology domain 3-only proteins BIM, PUMA or BID did not protect beta cells from this death. Overexpression of the anti-apoptotic protein BCL-2 or combined deficiency of the pro-apoptotic multi-BCL2 homology domain proteins BAX and BAK also failed to prevent beta cell destruction. Furthermore, loss of function of the death receptor Fas or its essential downstream signalling molecule Fas-associated death domain (FADD) in islets was also not protective. Using electron microscopy we observed that dying beta cells showed features of necrosis. However, islets deficient in receptor-interacting serine/threonine protein kinase 3 (RIPK3), a critical initiator of necroptosis, were still normally susceptible to CD4(+) T cell-mediated destruction. Remarkably, simultaneous inhibition of apoptosis and necroptosis by combining loss of RIPK3 and overexpression of BCL-2 in islets did not protect them against immune attack either. CONCLUSIONS/INTERPRETATION: Collectively, our data indicate that beta cells die by necrosis in autoimmune diabetes and that the programmed cell death pathways apoptosis and necroptosis are both dispensable for this process.

Mitochondrial apoptosis is dispensable for NLRP3 inflammasome activation but non-apoptotic caspase-8 is required for inflammasome priming.

A current paradigm proposes that mitochondrial damage is a critical determinant of NLRP3 inflammasome activation. Here, we genetically assess whether mitochondrial signalling represents a unified mechanism to explain how NLRP3 is activated by divergent stimuli. Neither co-deletion of the essential executioners of mitochondrial apoptosis BAK and BAX, nor removal of the mitochondrial permeability transition pore component cyclophilin D, nor loss of the mitophagy regulator Parkin, nor deficiency in MAVS affects NLRP3 inflammasome function. In contrast, caspase-8, a caspase essential for death-receptor-mediated apoptosis, is required for efficient Toll-like-receptor-induced inflammasome priming and cytokine production. Collectively, these results demonstrate that mitochondrial apoptosis is not required for NLRP3 activation, and highlight an important non-apoptotic role for caspase-8 in regulating inflammasome activation and pro-inflammatory cytokine levels.

Proapoptotic Bak and Bax guard against fatal systemic and organ-specific autoimmune disease.

Dysregulation of the "intrinsic" apoptotic pathway is associated with the development of cancer and autoimmune disease. Bak and Bax are two proapoptotic members of the Bcl-2 protein family with overlapping, essential roles in the intrinsic apoptotic pathway. Their activity is critical for the control of cell survival during lymphocyte development and homeostasis, best demonstrated by defects in thymic T-cell differentiation and peripheral lymphoid homeostasis caused by their combined loss. Because most bak(-/-)bax(-/-) mice die perinatally, the roles of Bax and Bak in immunological tolerance and prevention of autoimmune disease remain unclear. We show that mice reconstituted with a Bak/Bax doubly deficient hematopoietic compartment develop a fatal systemic lupus erythematosus-like autoimmune disease characterized by hypergammaglobulinemia, autoantibodies, lymphadenopathy, glomerulonephritis, and vasculitis. Importantly, these mice also develop a multiorgan autoimmune disease with autoantibodies against most solid glandular structures and evidence of glandular atrophy and necrotizing vasculitis. Interestingly, similar albeit less severe pathology was observed in mice containing a hematopoietic compartment deficient for only Bak, a phenotype reminiscent of the disease seen in patients with point mutations in BAK. These studies demonstrate a critical role for Bak and an ancillary role for Bax in safeguarding immunological tolerance and prevention of autoimmune disease. This suggests that direct activators of the intrinsic apoptotic pathway, such as BH3 mimetics, may be useful for treatment of diverse autoimmune diseases.

ABT-199, a potent and selective BCL-2 inhibitor, achieves antitumor activity while sparing platelets.

Proteins in the B cell CLL/lymphoma 2 (BCL-2) family are key regulators of the apoptotic process. This family comprises proapoptotic and prosurvival proteins, and shifting the balance toward the latter is an established mechanism whereby cancer cells evade apoptosis. The therapeutic potential of directly inhibiting prosurvival proteins was unveiled with the development of navitoclax, a selective inhibitor of both BCL-2 and BCL-2-like 1 (BCL-X(L)), which has shown clinical efficacy in some BCL-2-dependent hematological cancers. However, concomitant on-target thrombocytopenia caused by BCL-X(L) inhibition limits the efficacy achievable with this agent. Here we report the re-engineering of navitoclax to create a highly potent, orally bioavailable and BCL-2-selective inhibitor, ABT-199. This compound inhibits the growth of BCL-2-dependent tumors in vivo and spares human platelets. A single dose of ABT-199 in three patients with refractory chronic lymphocytic leukemia resulted in tumor lysis within 24 h. These data indicate that selective pharmacological inhibition of BCL-2 shows promise for the treatment of BCL-2-dependent hematological cancers.

Megakaryocytes possess a functional intrinsic apoptosis pathway that must be restrained to survive and produce platelets.

It is believed that megakaryocytes undergo a specialized form of apoptosis to shed platelets. Conversely, a range of pathophysiological insults, including chemotherapy, are thought to cause thrombocytopenia by inducing the apoptotic death of megakaryocytes and their progenitors. To resolve this paradox, we generated mice with hematopoietic- or megakaryocyte-specific deletions of the essential mediators of apoptosis, Bak and Bax. We found that platelet production was unperturbed. In stark contrast, deletion of the prosurvival protein Bcl-x(L) resulted in megakaryocyte apoptosis and a failure of platelet shedding. This could be rescued by deletion of Bak and Bax. We examined the effect on megakaryocytes of three agents that activate the intrinsic apoptosis pathway in other cell types: etoposide, staurosporine, and the BH3 mimetic ABT-737. All three triggered mitochondrial damage, caspase activation, and cell death. Deletion of Bak and Bax rendered megakaryocytes resistant to etoposide and ABT-737. In vivo, mice with a Bak(-/-) Bax(-/-) hematopoietic system were protected against thrombocytopenia induced by the chemotherapeutic agent carboplatin. Thus, megakaryocytes do not activate the intrinsic pathway to generate platelets; rather, the opposite is true: they must restrain it to survive and progress safely through proplatelet formation and platelet shedding.

Bcl-xL-inhibitory BH3 mimetics can induce a transient thrombocytopathy that undermines the hemostatic function of platelets.

BH3 mimetics are a new class of proapo-ptotic anticancer agents that have shown considerable promise in preclinical animal models and early-stage human trials. These agents act by inhibiting the pro-survival function of one or more Bcl-2-related proteins. Agents that inhibit Bcl-x(L) induce rapid platelet death that leads to thrombocytopenia; however, their impact on the function of residual circulating platelets remains unclear. In this study, we demonstrate that the BH3 mimetics, ABT-737 or ABT-263, induce a time- and dose-dependent decrease in platelet adhesive function that correlates with ectodomain shedding of the major platelet adhesion receptors, glycoprotein Ibα and glycoprotein VI, and functional down-regulation of integrin α(IIb)ß(3). Analysis of platelets from mice treated with higher doses of BH3 mimetics revealed the presence of a subpopulation of circulating platelets undergoing cell death that have impaired activation responses to soluble agonists. Functional analysis of platelets by intravital microscopy revealed a time-dependent defect in platelet aggregation at sites of vascular injury that correlated with an increase in tail bleeding time. Overall, these studies demonstrate that Bcl-x(L)-inhibitory BH3 mimetics not only induce thrombocytopenia but also a transient thrombocytopathy that can undermine the hemostatic function of platelets.

BH3 mimetics antagonizing restricted prosurvival Bcl-2 proteins represent another class of selective immune modulatory drugs.

Death by apoptosis shapes tissue homeostasis. Apoptotic mechanisms are so universal that harnessing them for tailored immune intervention would seem challenging; however, the range and different expression levels of pro- and anti-apoptotic molecules among tissues offer hope that targeting only a subset of such molecules may be therapeutically useful. We examined the effects of the drug ABT-737, a mimetic of the killer BH3 domain of the Bcl-2 family of proteins that induces apoptosis by antagonizing Bcl-2, Bcl-X(L), and Bcl-W (but not Mcl-1 and A1), on the mouse immune system. Treatment with ABT-737 reduced the numbers of selected lymphocyte and dendritic cell subpopulations, most markedly in lymph nodes. It inhibited the persistence of memory B cells, the establishment of newly arising bone marrow plasma cells, and the induction of a cytotoxic T cell response. Preexisting plasma cells and germinal centers were unaffected. Notably, ABT-737 was sufficiently immunomodulatory to allow long-term survival of pancreatic allografts, reversing established diabetes in this model. These results provide an insight into the selective mechanisms of immune cell survival and how this selectivity avails a different strategy for immune modulation.

Two distinct pathways regulate platelet phosphatidylserine exposure and procoagulant function.

Procoagulant platelets exhibit hallmark features of apoptotic cells, including membrane blebbing, microvesiculation, and phosphatidylserine (PS) exposure. Although platelets possess many well-known apoptotic regulators, their role in regulating the procoagulant function of platelets is unclear. To clarify this, we investigated the consequence of removing the essential mediators of apoptosis, Bak and Bax, or directly inducing apoptosis with the BH3 mimetic compound ABT-737. Treatment of platelets with ABT-737 triggered PS exposure and a marked increase in thrombin generation in vitro. This increase in procoagulant function was Bak/Bax- and caspase-dependent, but it was unaffected by inhibitors of platelet activation or by chelating extracellular calcium. In contrast, agonist-induced platelet procoagulant function was unchanged in Bak(-/-)Bax(-/-) or caspase inhibitor-treated platelets, but it was completely eliminated by extracellular calcium chelators or inhibitors of platelet activation. These studies show the existence of 2 distinct pathways regulating the procoagulant function of platelets.

In vivo efficacy of the Bcl-2 antagonist ABT-737 against aggressive Myc-driven lymphomas.

Deregulated Myc expression drives many human cancers, including Burkitt's lymphoma and a highly aggressive subset of diffuse large cell lymphomas. Myc-driven tumors often display resistance to chemotherapeutics because of acquisition of mutations that impair the apoptosis pathway regulated by the Bcl-2 protein family. Given the need to identify new therapies for such lymphomas, we have evaluated the efficacy of ABT-737, a small molecule that mimics the action of the BH3-only proteins, natural antagonists of the prosurvival Bcl-2 proteins. ABT-737 selectively targets certain prosurvival proteins (Bcl-2, Bcl-x(L), and Bcl-w) but not others (Mcl-1 and A1). We treated mice transplanted with lymphomas derived either from Emu-myc transgenic mice or Emu-myc mice that also expressed an Emu-bcl-2 transgene. As a single agent, ABT-737 significantly prolonged the survival of mice transplanted with the myc/bcl-2 lymphomas but was ineffective for the myc lymphomas, probably because of the relatively higher Mcl-1 levels found in the latter. Strikingly, when combined with low-dose cyclophosphamide, ABT-737 produced sustained disease-free survival of all animals transplanted with two of three myc/bcl-2 lymphomas tested. The combination therapy was also more effective against some myc lymphomas than treatment with either agent alone. Our data suggest that antagonism of Bcl-2 with small organic compounds is an attractive approach to enhance the efficacy of conventional therapy for the treatment of Myc-driven lymphomas that over-express this prosurvival molecule.