GATA-3-dependent Gene Transcription is Impaired upon HDAC Inhibition.

PURPOSE: Many peripheral and cutaneous T-cell lymphoma (CTCL) subtypes are poorly responsive to conventional chemotherapeutic agents and associated with dismal outcomes. The zinc finger transcription factor GATA-3 and the transcriptional program it instigates are oncogenic and highly expressed in various T-cell neoplasms. Posttranslational acetylation regulates GATA-3 DNA binding and target gene expression. Given the widespread use of histone deacetylase inhibitors (HDACi) in relapsed/refractory CTCL, we sought to examine the extent to which these agents attenuate the transcriptional landscape in these lymphomas. EXPERIMENTAL DESIGN: Integrated GATA-3 chromatin immunoprecipitation sequencing and RNA sequencing analyses were performed in complementary cell line models and primary CTCL specimens treated with clinically available HDACi. RESULTS: We observed that exposure to clinically available HDACi led to significant transcriptional reprogramming and increased GATA-3 acetylation. HDACi-dependent GATA-3 acetylation significantly impaired both its ability to bind DNA and transcriptionally regulate its target genes, thus leading to significant transcriptional reprogramming in HDACi-treated CTCL. CONCLUSIONS: Beyond shedding new light on the mechanism of action associated with HDACi in CTCL, these findings have significant implications for their use, both as single agents and in combination with other novel agents, in GATA-3-driven lymphoproliferative neoplasms.

ITK degradation to block T cell receptor signaling and overcome therapeutic resistance in T cell lymphomas.

Interleukin (IL)-2-inducible T cell kinase (ITK) is essential for T cell receptor (TCR) signaling and plays an integral role in T cell proliferation and differentiation. Unlike the ITK homolog BTK, no inhibitors of ITK are currently US Food and Drug Administration (FDA) approved. In addition, recent studies have identified mutations within BTK that confer resistance to both covalent and non-covalent inhibitors. Here, as an alternative strategy, we report the development of BSJ-05-037, a potent and selective heterobifunctional degrader of ITK. BSJ-05-037 displayed enhanced anti-proliferative effects relative to its parent inhibitor BMS-509744, blocked the activation of NF-kB/GATA-3 signaling, and increased the sensitivity of T cell lymphoma cells to cytotoxic chemotherapy both in vitro and in vivo. In summary, targeted degradation of ITK is a novel approach to modulate TCR signal strength that could have broad application for the investigation and treatment of T cell-mediated diseases.

GATA-3 is a proto-oncogene in T-cell lymphoproliferative neoplasms.

Neoplasms originating from thymic T-cell progenitors and post-thymic mature T-cell subsets account for a minority of lymphoproliferative neoplasms. These T-cell derived neoplasms, while molecularly and genetically heterogeneous, exploit transcription factors and signaling pathways that are critically important in normal T-cell biology, including those implicated in antigen-, costimulatory-, and cytokine-receptor signaling. The transcription factor GATA-3 regulates the growth and proliferation of both immature and mature T cells and has recently been implicated in T-cell neoplasms, including the most common mature T-cell lymphoma observed in much of the Western world. Here we show that GATA-3 is a proto-oncogene across the spectrum of T-cell neoplasms, including those derived from T-cell progenitors and their mature progeny, and further define the transcriptional programs that are GATA-3 dependent, which include therapeutically targetable gene products. The discovery that p300-dependent acetylation regulates GATA-3 mediated transcription by attenuating DNA binding has novel therapeutic implications. As most patients afflicted with GATA-3 driven T-cell neoplasms will succumb to their disease within a few years of diagnosis, these findings suggest opportunities to improve outcomes for these patients.

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.

Targeting Lymphoma-associated Macrophage Expansion via CSF1R/JAK Inhibition is a Therapeutic Vulnerability in Peripheral T-cell Lymphomas.

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.

Colony-Stimulating Factor 1 Receptor (CSF1R) Activates AKT/mTOR Signaling and Promotes T-Cell Lymphoma Viability.

PURPOSE: Peripheral T-cell lymphomas are clinically aggressive and usually fatal, as few complete or durable remissions are achieved with currently available therapies. Recent evidence supports a critical role for lymphoma-associated macrophages during T-cell lymphoma progression, but the specific signals involved in the cross-talk between malignant T cells and their microenvironment are poorly understood. Colony-stimulator factor 1 receptor (CSF1R, CD115) is required for the homeostatic survival of tissue-resident macrophages. Interestingly, its aberrant expression has been reported in a subset of tumors. In this article, we evaluated its expression and oncogenic role in T-cell lymphomas. EXPERIMENTAL DESIGN: Loss-of-function studies, including pharmacologic inhibition with a clinically available tyrosine kinase inhibitor, pexidartinib, were performed in multiple in vitro and in vivo models. In addition, proteomic and genomic screenings were performed to discover signaling pathways that are activated downstream of CSF1R signaling. RESULTS: We observed that CSF1R is aberrantly expressed in many T-cell lymphomas, including a significant number of peripheral and cutaneous T-cell lymphomas. Colony-stimulating factor 1 (CSF1), in an autocrine or paracrine-dependent manner, leads to CSF1R autophosphorylation and activation in malignant T cells. Furthermore, CSF1R signaling was associated with significant changes in gene expression and in the phosphoproteome, implicating PI3K/AKT/mTOR in CSF1R-mediated T-cell lymphoma growth. We also demonstrated that inhibition of CSF1R in vivo and in vitro models is associated with decreased T-cell lymphoma growth. CONCLUSIONS: Collectively, these findings implicate CSF1R in T-cell lymphomagenesis and have significant therapeutic implications.

Pharmacologic characterization of a kinetic in vitro human co-culture angiogenesis model using clinically relevant compounds.

Angiogenesis, the formation of new vessels from preexisting vessels, involves multiple cell types acting in concert to cause endothelial cell proliferation, migration, and differentiation into microvascular arrays. Under pathologic conditions, microenvironment changes result in altered blood vessel production. Historically, in vitro angiogenesis assays study individual aspects of the process and tend to be variable, difficult to quantify, and limited in clinical relevance. Here, we describe a kinetic, quantitative, co-culture angiogenesis model and demonstrate its relevance to in vivo pharmacology. Similar to in vivo angiogenesis, a co-culture of human umbilical vein endothelial cells with normal human dermal fibroblasts remains sensitive to multiple cytokines, resulting in a concentration-dependent stimulation of tube formation over time. Treatment with axitinib, a selective vascular endothelial growth factor (VEGF) antagonist, inhibited VEGF-mediated tube length and branch point formation and was selective for inhibiting VEGF over basic fibroblast growth factor (bFGF), similar to previous studies. Conversely, an FGFR-1 selective compound, PD-161570, was more potent at inhibiting bFGF-mediated angiogenesis. These results demonstrate the cytokine dynamics, selective pharmacology, and translational application of this model system. Finally, combining quantitative angiogenic biology with kinetic, live-content imaging highlights the importance of using validated in vitro models in drug discovery research.

Involving youth with disabilities in the development and evaluation of a new advocacy training: Project TEAM.

PURPOSE: This paper describes a participatory research process in which six youth with disabilities (Youth Panel) participated in the development and evaluation of a manualized advocacy training, Project TEAM (Teens making Environment and Activity Modifications). Project TEAM teaches youth with disabilities how to identify environmental barriers, generate solutions, and request accommodations. METHOD: The Youth Panel conducted their evaluation after the university researcher implemented Project TEAM with three groups of trainees. The Youth Panel designed and administered a survey and focus group to evaluate enjoyment and usefulness of Project TEAM with support from an advocate/researcher. Members of the Youth Panel analyzed survey response frequencies. The advocate/researcher conducted a content analysis of the open-ended responses. RESULTS: Sixteen of 21 Project TEAM trainees participated in the evaluation. The evaluation results suggest that the trainees found the interactive and individualized aspects of the Project TEAM most enjoyable and useful. Some instructional materials were difficult for trainees with cognitive disabilities to understand. CONCLUSIONS: The Youth Panel's involvement in the development of Project TEAM may explain the relatively positive experiences reported by trainees. Project TEAM should continue to provide trainees with the opportunity to apply concepts in real-life situations. Project TEAM requires revisions to ensure it is enjoyable and useful for youth with a variety of disabilities. IMPLICATIONS FOR REHABILITATION: • Group process strategies, picture-based data collection materials, peer teamwork, and mentorship from adults with disabilities can enable youth with disabilities to engage in research. • Collaborating with youth with disabilities in the development of new rehabilitation approaches may enhance the relevance of interventions for other youth with disabilities. • Youth with cognitive disabilities participating in advocacy and environment-focused interventions may prefer interactive and experiential learning activities over passive teaching approaches such as powerpoints and videos.

Choosing employment: factors that impact employment decisions for individuals with intellectual disability.

Little is known about the factors that shape the employment-related decisions of individuals with intellectual and/or developmental disabilities. Findings from qualitative interviews with individuals, their family members, and employment-support professionals from four community rehabilitation providers throughout Massachusetts were reported. Recognizing the value of participatory action research, we also included a co-researcher with intellectual disability who participated in all facets of the research process. Findings revealed a collection of people and factors considered influential in employment-related decision-making. The family in the formative years, school-based staff and early employment experiences, the culture of the community rehabilitation providers, the job developer, and personal preferences all influenced participants' decisions. Through understanding these persuasive elements, we offer recommendations to those in the intellectual and developmental disabilities field to optimize employment choices and outcomes.