Search

The reciprocal relationship between malignant T cells and lymphoma-associated macrophages (LAM) within the tumor microenvironment (TME) is unique, as LAMs are well poised to provide ligands for antigen, costimulatory, and cytokine receptors that promote T-cell lymphoma growth. Conversely, malignant T cells promote the functional polarization and homeostatic survival of LAM. Therefore, we sought to determine the extent to which LAMs are a therapeutic vulnerability in these lymphomas, and to identify effective therapeutic strategies for their depletion. We utilized complementary genetically engineered mouse models and primary peripheral T-cell lymphoma (PTCL) specimens to quantify LAM expansion and proliferation. A high-throughput screen was performed to identify targeted agents that effectively deplete LAM within the context of PTCL. We observed that LAMs are dominant constituents of the TME in PTCL. Furthermore, their dominance was explained, at least in part, by their proliferation and expansion in response to PTCL-derived cytokines. Importantly, LAMs are a true dependency in these lymphomas, as their depletion significantly impaired PTCL progression. These findings were extrapolated to a large cohort of human PTCL specimens where LAM proliferation was observed. A high-throughput screen demonstrated that PTCL-derived cytokines led to relative resistance to CSF1R selective inhibitors, and culminated in the identification of dual CSF1R/JAK inhibition as a novel therapeutic strategy to deplete LAM in these aggressive lymphomas. Malignant T cells promote the expansion and proliferation of LAM, which are a bone fide dependency in these lymphomas, and are effectively depleted with a dual CSF1R/JAK inhibitor. Significance: LAMs are a therapeutic vulnerability, as their depletion impairs T-cell lymphoma disease progression. Pacritinib, a dual CSF1R/JAK inhibitor, effectively impaired LAM viability and expansion, prolonged survival in preclinical T-cell lymphoma models, and is currently being investigated as a novel therapeutic approach in these lymphomas.

TME gene sets

Fig. S6: GSEA in LAM

Antibodies used

The reciprocal relationship between malignant T cells and lymphoma-associated macrophages (LAM) within the tumor microenvironment (TME) is unique, as LAMs are well poised to provide ligands for antigen, costimulatory, and cytokine receptors that promote T-cell lymphoma growth. Conversely, malignant T cells promote the functional polarization and homeostatic survival of LAM. Therefore, we sought to determine the extent to which LAMs are a therapeutic vulnerability in these lymphomas, and to identify effective therapeutic strategies for their depletion. We utilized complementary genetically engineered mouse models and primary peripheral T-cell lymphoma (PTCL) specimens to quantify LAM expansion and proliferation. A high-throughput screen was performed to identify targeted agents that effectively deplete LAM within the context of PTCL. We observed that LAMs are dominant constituents of the TME in PTCL. Furthermore, their dominance was explained, at least in part, by their proliferation and expansion in response to PTCL-derived cytokines. Importantly, LAMs are a true dependency in these lymphomas, as their depletion significantly impaired PTCL progression. These findings were extrapolated to a large cohort of human PTCL specimens where LAM proliferation was observed. A high-throughput screen demonstrated that PTCL-derived cytokines led to relative resistance to CSF1R selective inhibitors, and culminated in the identification of dual CSF1R/JAK inhibition as a novel therapeutic strategy to deplete LAM in these aggressive lymphomas. Malignant T cells promote the expansion and proliferation of LAM, which are a bone fide dependency in these lymphomas, and are effectively depleted with a dual CSF1R/JAK inhibitor.Significance:LAMs are a therapeutic vulnerability, as their depletion impairs T-cell lymphoma disease progression. Pacritinib, a dual CSF1R/JAK inhibitor, effectively impaired LAM viability and expansion, prolonged survival in preclinical T-cell lymphoma models, and is currently being investigated as a novel therapeutic approach in these lymphomas.

Peripheral T-cell lymphomas (PTCL) are agressive lymphomas that develop from mature T cells. The most common PTCLs are genetically, molecularly, and clinically diverse and are generally associated with dismal outcomes. While Notch signaling plays a critically important role in both the development of immature T cells and their malignant transformation, its role in PTCL is poorly understood, despite the increasingly appreciated function of Notch in regulating the proliferation and differentiation of mature T cells. Here, we demonstrate that Notch receptors and their Delta-like family ligands (DLL1/DLL4) play a pathogenic role in PTCL. Notch1 activation was observed in common PTCL subtypes, including PTCL-not otherwise specified (NOS). In a large cohort of PTCL-NOS biopsies, Notch1 activation was significantly associated with surrogate markers of proliferation. Complementary genetically engineered mouse models and spontaneous PTCL models were used to functionally examine the role of Notch signaling, and Notch1/Notch2 blockade and pan-Notch blockade using dominant-negative MAML significantly impaired the proliferation of malignant T cells and PTCL progression in these models. Treatment with DLL1/DLL4 blocking antibodies established that Notch signaling is ligand-dependent. Together, these findings reveal a role for ligand-dependent Notch signaling in driving peripheral T-cell lymphomagenesis.Significance:This work demonstrates that ligand-dependent Notch activation promotes the growth and proliferation of mature T-cell lymphomas, providing new therapeutic strategies for this group of aggressive lymphomas.

Supplementary Data from Notch Signaling Promotes Mature T-Cell Lymphomagenesis

Peripheral T-cell lymphomas (PTCL) are agressive lymphomas that develop from mature T cells. The most common PTCLs are genetically, molecularly, and clinically diverse and are generally associated with dismal outcomes. While Notch signaling plays a critically important role in both the development of immature T cells and their malignant transformation, its role in PTCL is poorly understood, despite the increasingly appreciated function of Notch in regulating the proliferation and differentiation of mature T cells. Here, we demonstrate that Notch receptors and their Delta-like family ligands (DLL1/DLL4) play a pathogenic role in PTCL. Notch1 activation was observed in common PTCL subtypes, including PTCL-not otherwise specified (NOS). In a large cohort of PTCL-NOS biopsies, Notch1 activation was significantly associated with surrogate markers of proliferation. Complementary genetically engineered mouse models and spontaneous PTCL models were used to functionally examine the role of Notch signaling, and Notch1/Notch2 blockade and pan-Notch blockade using dominant-negative MAML significantly impaired the proliferation of malignant T cells and PTCL progression in these models. Treatment with DLL1/DLL4 blocking antibodies established that Notch signaling is ligand-dependent. Together, these findings reveal a role for ligand-dependent Notch signaling in driving peripheral T-cell lymphomagenesis. Significance: This work demonstrates that ligand-dependent Notch activation promotes the growth and proliferation of mature T-cell lymphomas, providing new therapeutic strategies for this group of aggressive lymphomas.

Background Diabetic retinopathy is a microvascular disease that results from retinal vascular degeneration and defective repair due to diabetes-induced endothelial progenitor dysfunction. Objective Understanding key molecular factors involved in vascular degeneration and repair is paramount for developing effective diabetic retinopathy treatment strategies. We propose that diabetes-induced activation of acid sphingomyelinase (ASM) plays essential role in retinal endothelial and CD34+ circulating angiogenic cell (CAC) dysfunction in diabetes. Methods Human retinal endothelial cells (HRECs) isolated from control and diabetic donor tissue and human CD34+ CACs from control and diabetic patients were used in this study. ASM messenger RNA and protein expression were assessed by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. To evaluate the effect of diabetes-induced ASM on HRECs and CD34+ CACs function, tube formation, CAC incorporation into endothelial tubes, and diurnal release of CD34+ CACs in diabetic individuals were determined. Results ASM expression level was significantly increased in HRECs isolated from diabetic compared with control donor tissue, as well as CD34+ CACs and plasma of diabetic patients. A significant decrease in tube area was observed in HRECs from diabetic donors compared with control HRECs. The tube formation deficiency was associated with increased expression of ASM in diabetic HRECs. Moreover, diabetic CD34+ CACs with high ASM showed defective incorporation into endothelial tubes. Diurnal release of CD34+ CACs was disrupted with the rhythmicity lost in diabetic patients. Conclusion Collectively, these findings support that diabetes-induced ASM upregulation has a marked detrimental effect on both retinal endothelial cells and CACs.

Tight junctions (TJs) involve close apposition of transmembrane proteins between cells. Although TJ proteins have been studied in detail, the role of lipids is largely unknown. We addressed the role of very long-chain (VLC ≥26) ceramides in TJs using diabetes-induced loss of the blood-retinal barrier as a model. VLC fatty acids that incorporate into VLC ceramides are produced by elongase elongation of very long-chain fatty acids protein 4 (ELOVL4). ELOVL4 is significantly reduced in the diabetic retina. Overexpression of ELOVL4 significantly decreased basal permeability, inhibited vascular endothelial growth factor (VEGF)– and interleukin-1β–induced permeability, and prevented VEGF-induced decrease in occludin expression and border staining of TJ proteins ZO-1 and claudin-5. Intravitreal delivery of AAV2-hELOVL4 reduced diabetes-induced increase in vascular permeability. Ultrastructure and lipidomic analysis revealed that ω-linked acyl-VLC ceramides colocalize with TJ complexes. Overall, normalization of retinal ELOVL4 expression could prevent blood-retinal barrier dysregulation in diabetic retinopathy through an increase in VLC ceramides and stabilization of TJs.

Partial duplication of chromosome 3q is a recognizable syndrome with characteristic facial features, microcephaly, digital anomalies, genitourinary and cardiac defects as well as growth retardation and developmental delays. While there is clinical overlap with the unrelated Cornelia de Lange syndrome (CDLS), there are distinguishing features and molecular etiologies. Most cases of 3q duplication appear to be the result of an unbalanced translocation or inversion and therefore accompanied by additional cytogenetic anomalies. Consequently, pure duplications of 3q are very rare; we are aware of only 12 such cases that have been reported previously. Here, we present a new case of pure, partial 3q duplication in a 3-month-old female who displayed a number of clinical signs consistent with previously reported phenotypes and the additional novel finding of a vascular ring.

The metabolic insults associated with diabetes lead to low-grade chronic inflammation, retinal endothelial cell damage, and inadequate vascular repair. This is partly due to the increased activation of bone marrow (BM)-derived proinflammatory monocytes infiltrating the retina, and the compromised function of BM-derived reparative circulating angiogenic cells (CACs), which home to sites of endothelial injury and foster vascular repair. We now propose that a metabolic link leading to activated monocytes and dysfunctional CACs in diabetes involves upregulation of a central enzyme of sphingolipid signaling, acid sphingomyelinase (ASM). Selective inhibition of ASM in the BM prevented diabetes-induced activation of BM-derived microglia-like cells and normalized proinflammatory cytokine levels in the retina. ASM upregulation in diabetic CACs caused accumulation of ceramide on their cell membrane, thereby reducing membrane fluidity and impairing CAC migration. Replacing sphingomyelin with ceramide in synthetic membrane vesicles caused a similar decrease in membrane fluidity. Inhibition of ASM in diabetic CACs improved membrane fluidity and homing of these cells to damaged retinal vessels. Collectively, these findings indicate that selective modulation of sphingolipid metabolism in BM-derived cell populations in diabetes normalizes the reparative/proinflammatory cell balance and can be explored as a novel therapeutic strategy for treating diabetic retinopathy.