B cell-intrinsic function of TAPP adaptors in controlling germinal center responses and autoantibody production in mice

Control of B-cell signal transduction is critical to prevent production of pathological autoantibodies. Tandem PH domain containing proteins (TAPPs) specifically bind PI(3,4)P2, a phosphoinositide product generated by PI 3-kinases and the phosphatase SHIP. TAPP KI mice bearing PH domain-inactivating mutations in both TAPP1 and TAPP2 genes, uncoupling them from PI(3,4)P2, exhibit increased BCR-induced activation of the kinase Akt and develop lupus-like characteristics including anti-DNA antibodies and deposition of immune complexes in kidneys. Here we find that TAPP KI mice develop chronic germinal centers (GCs) with age and show abnormal expression of B cell activation and memory markers. Upon immunization with T-dependent Ag, TAPP KI mice develop functional but abnormally large GCs, associated with increased GC B cell survival. Disruption of chronic GCs in TAPP KI mice by deletion of the costimulatory molecule ICOS abrogate anti-DNA and anti-nuclear antibody production in TAPP KI mice, indicating an essential role for GCs. Moreover, TAPP KI B cells are sufficient to drive chronic GC responses and recapitulate the autoimmune phenotype in bone marrow chimeric mice. Our findings demonstrate a B cell-intrinsic role of TAPP-PI(3,4)P2 interaction in regulating GC responses and autoantibody production and suggest that uncontrolled Akt activity in B cells can drive autoimmunity. This article is protected by copyright. All rights reserved.</p

Real-world safety and tolerability of rapid infusion obinutuzumab in chronic lymphocytic leukemia, small lymphocytic lymphoma and non-Hodgkin lymphoma: a provincial review

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Interaction of TAPP adapters with the phosphoinositide PI(3,4)P2 regulates B cell activation and differentiation

Phosphoinositide 3-kinase is a family of lipid kinases that function by phosphorylating the D3 position of phosphoinositide (PI) lipids generating PI(3)P, PI(3,4)P2 and PI(3,4,5)P3. These D3 phosphoinositides regulate various cellular processes through the recruitment of effector proteins containing lipid specific pleckstrin homology (PH) domains. PI phosphatases such as PTEN and SHIP function to restrain PI3K signaling by limiting the amount of D3 PI available for binding. Deletion of either PTEN or SHIP significantly alters B cell function and humoral immune responses. TAPP1 and TAPP2 are dual PH domain containing adaptors which selectively bind the phosphoinositide PI(3,4)P2 via their C-terminal PH domains. PI(3,4)P2 is a lipid messenger generated by PI3K and through the inositol phosphatase activity of SHIP. The function of PI(3,4)P2 remains incompletely understood. To identify the functional role of TAPP-PI(3,4)P2 interactions, we utilized a knock-in (KI) mouse bearing mutations within the PI-binding pocket of both TAPPs. Our study assessed the effect of PI3K dependent KI mutation on B lymphocyte development, activation and antibody production. Flow cytometry analyses of lymphoid tissues found that TAPP KI mice develop relatively normal frequencies of mature B cell populations with the exception of peritoneal B1 cells, which are increased by approximately 50%. Strikingly, TAPP KI mice developed substantially elevated serum antibody levels. TAPP KI mice were able to generate high affinity antigen-binding antibodies upon immunization with NP-OVA in alum adjuvant; however, total immunoglobulin production was markedly increased under this immunization condition. We further assessed the germinal centre (GC) response, which are known to require PI3K signaling and a hallmark of T cell dependent (TD) antibody responses. TAPP KI mice generated larger germinal centers (GC) upon immunization, which was associated with increased GC B cell survival. We further assessed whether uncoupling of TAPPs from PI(3,4)P2 alters B cell signaling and functional responses in vitro. B cells purified from TAPP KI mice were found to have altered functional responses in vitro, with significantly increased survival and cell division following antigen receptor cross-linking. Consistent with increased cell survival, TAPP KI B cells show increased Akt phosphorylation on Ser473 and Thr308 after antigen receptor cross-linking. However, reconstitution of B cell deficient mice with either WT or TAPP KI B cells was found to generate similar GC responses, suggesting that activation of other cells may contribute to the enhanced in vivo responses. Consistently, when we examined the CD4+ T follicular helper cells, a subset providing critical cues to GC responses, we found increased expression of ICOS activation marker. Our results indicate the interactions of TAPP adapters with PI(3,4)P2 serve to restrain lymphocyte activation and limit antibody production, providing the first in vivo evidence that this interaction is important for immune function.February 201

Interaction of TAPP adapter proteins with phosphatidylinositol (3,4)-bisphosphate regulates germinal centre response and development of autoimmunity (P1107)

Abstract Activation of PI3-kinase enzymes is essential for lymphocyte development and function. Active PI3Ks generate several types of D3 phosphoinositides, including PI(3,4,5)P3 and PI(3,4)P2. Tandem PH domain containing proteins (TAPPs) are adaptor molecules which bind specifically to PI(3,4)P2, but their functions in immune cells are largely unknown. We have performed studies on TAPP mutant mice, in which a mutation was introduced to the C terminal domain of both TAPP1 and TAPP2, to determine the effect of uncoupling TAPPs from PI(3,4)P2. We found that these TAPP KI mice exhibit elevated serum antibody levels and also develop autoantibodies and kidney pathology resembling that of lupus. TAPP KI mice generated elevated germinal center (GC) responses upon immunization with T-dependent antigen, suggesting an inhibitory role associated with TAPPs in lymphocytes. The GC B cell population was increased in frequency and showed reduced expression of apoptosis markers, and expansion of follicular helper T cell subset. We hypothesize that TAPP KI mice have disruption in a regulatory mechanism within the GC leading to sustained B cell activation and survival, and contributing to development of autoimmunity. TAPP KI B cells were found to be hyper-responsive to BCR cross-linking in vitro, and T cell responses are currently being assessed. Our goal is to define the B cell and T cell intrinsic signaling mechanisms enhanced as a result of disrupted inhibitory signaling mediated by TAPPs.</jats:p

The landscape of T-cell engagers for the treatment of follicular lymphoma

Follicular lymphoma (FL), the second most common subtype of non-Hodgkin lymphoma, relies on interactions with immune elements in the tumor microenvironment, including T-follicular helper cells and follicular dendritic cells, for its survival and progression. Despite its initial responsiveness to chemoimmunotherapy, FL is generally considered incurable. Strategies to improve immune-mediated control of FL could significantly benefit this population, particularly as it includes many elderly and comorbid patients. Immune cell engagers, especially bispecific antibodies (BsAbs), are crucial in targeting FL by bridging tumor and effector cells, thereby triggering T-cell activation and cytotoxic killing. CD3 × CD20 BsAbs have shown the most promise in clinical development for B-NHL patients, with structural variations affecting their target affinity and potency. This review summarizes the current clinical trials of BsAbs for relapsed/refractory FL, highlighting the approval of some agents, their role in first-line treatment or combination therapies, their toxicity profiles, and the future of this therapeutic approach compared to other immune cell therapies

Interaction of TAPP adapter proteins with phosphatidylinositol (3,4)-bisphosphate regulates B cell activation and autoantibody production (159.5)

Abstract Activation of PI3-kinase enzymes through the BCR is essential for B cell development and function. Active PI3Ks generate D3 phosphoinositide PI(3,4)P2 that bind to signaling molecules such as TAPP (Tandem PH domain containing protein) adaptor via their C terminal PH domain. This interaction leads to translocation of TAPPs to the plasma membrane. However, the role of TAPP adaptors in regulating PI3K signaling in B cell is not completely understood. We have performed studies on TAPP mutant mice, where a mutation was introduced to the C terminal domain of both TAPP1 and TAPP2 to determine the effect of uncoupling TAPPs from PI(3,4)P2. The studies showed an enhanced B cell activation and elevated serum antibody levels in these mice, suggesting an inhibitory role associated with TAPPs in B cells. To test whether the aberrant responses observed in B cells leads to autoimmunity in these animals, we analyzed their serum and observed a progressive increase in anti-dsDNA antibodies in the aging cohort. This along with other observed characteristics such as the presence of anti-nuclear antibodies within the serum, progressive development of lymphopenia and glomerulonephritis in aged mice is reminiscent of lupus. We are currently working to define the cellular interactions involved in this autoimmune phenotype and the signaling mechanism associated with the inhibitory role of TAPPs.</jats:p

PD-1 Inhibition in Malignant Melanoma and Lack of Clinical Response in Chronic Lymphocytic Leukemia in the Same Patients: A Case Series

Chronic lymphocytic leukemia (cll) is the most common adult leukemia in the Western world. Unfortunately, affected patients are often immunosuppressed and at increased risk of infection and secondary malignancy. Previous meta-analysis has found that patients with cll have a risk of melanoma that is increased by a factor of 4 compared with the general population. Recent advances in the understanding of the PD receptor pathway have led to immunotherapies that target cancer cells. The use of PD-1 inhibitors is now considered first-line treatment for BRAF wild-type metastatic melanoma. Interestingly, early preclinical data suggest that inhibition of that pathway could also be used in the treatment of cll; however, recent clinical data did not support the effectiveness of that approach. In this case series, we highlight 2 cases in which patients with cll and concurrent malignant melanoma underwent treatment with PD-1 inhibitors and were found to experience reductions in their white blood cell counts without improvement in their hemoglobin. Those cases further illustrate that treatment of cll with PD-1 inhibitors is ineffective