Pathogenesis Of Peripheral T-cell Lymphomas Research Articles

Precise diagnosis and treatment for peripheral T-cell lymphomas: From pathogenic mechanisms to innovative approaches

<p>Peripheral T-cell lymphomas (PTCLs) encompass a diverse group of aggressive non-Hodgkin��s lymphomas originating from mature T lymphocytes. Despite their relatively low prevalence compared to B-cell lymphomas, PTCLs pose significant clinical challenges due to their heterogeneity, lack of specific biomarkers, and often poor response to conventional chemotherapies. Unraveling the intricate pathogenic mechanisms underlying PTCLs is pivotal for the development of targeted therapies, which has become a major focus of PTCL research in recent years. It was previously postulated that genomic alterations play a pivotal role in T-cell transformation and lymphomagenesis. Recent multi-omics investigations and functional studies focusing on pathogenic mechanisms have unveiled that in addition to genetic abnormalities, diverse intrinsic and extrinsic mechanisms are implicated in the pathogenesis of PTCLs. In this Review, we provide an overview of the recent advances in the pathogenesis and management of PTCLs, focusing on the current and potential precise diagnosis and treatment based on intrinsic and extrinsic mechanisms. Bioengineering and drug development have enabled the effective modulation of many previously ��undruggable�� targets, which offers the potential to target new biomarkers associated with PTCL pathogenesis. The integration of artificial intelligence (AI) in clinical practice enables a comprehensive framework, merging diverse data sources and clinical context to discover precise biomarkers for personalized medicine. Finally, we present pressing questions and challenges that demand attention and discuss emerging solutions.</p>

Precise diagnosis and treatment for peripheral T-cell lymphomas: From pathogenic mechanisms to innovative approaches

<p>Peripheral T-cell lymphomas (PTCLs) encompass a diverse group of aggressive non-Hodgkin��s lymphomas originating from mature T lymphocytes. Despite their relatively low prevalence compared to B-cell lymphomas, PTCLs pose significant clinical challenges due to their heterogeneity, lack of specific biomarkers, and often poor response to conventional chemotherapies. Unraveling the intricate pathogenic mechanisms underlying PTCLs is pivotal for the development of targeted therapies, which has become a major focus of PTCL research in recent years. It was previously postulated that genomic alterations play a pivotal role in T-cell transformation and lymphomagenesis. Recent multi-omics investigations and functional studies focusing on pathogenic mechanisms have unveiled that in addition to genetic abnormalities, diverse intrinsic and extrinsic mechanisms are implicated in the pathogenesis of PTCLs. In this Review, we provide an overview of the recent advances in the pathogenesis and management of PTCLs, focusing on the current and potential precise diagnosis and treatment based on intrinsic and extrinsic mechanisms. Bioengineering and drug development have enabled the effective modulation of many previously ��undruggable�� targets, which offers the potential to target new biomarkers associated with PTCL pathogenesis. The integration of artificial intelligence (AI) in clinical practice enables a comprehensive framework, merging diverse data sources and clinical context to discover precise biomarkers for personalized medicine. Finally, we present pressing questions and challenges that demand attention and discuss emerging solutions.</p>

Diagnosis, management and follow up of peripheral T-cell lymphomas: a consensus practice statement from the Australasian Lymphoma Alliance.

Peripheral T-cell lymphomas (PTCL) represent a heterogeneous disease group accounting for 10% of non-Hodgkin lymphomas. PTCL patients have typically poorer outcomes compared with aggressive B-cell lymphomas. However, such outcomes are heavily dependent on subtype. Although anthracycline-based regimens such as cyclophosphamide, doxorubicin, vincristine and prednisone remain the standard first-line treatment for most aggressive PTCL, there are important variations including incorporation of novel agents, use of radiotherapy and judicious consideration of stem cell transplantation. Relapsed or refractory disease represents a significant area of unmet need where chemotherapy intensification has limited efficacy and novel agents such as brentuximab vedotin and pralatrexate provide additional opportunities for attainment of remission and potential stem cell transplant. In the future, pre-therapy prognostic biomarkers including genomic characterisation, may aid in risk stratification and help guide initial patient management to improve survival. There is an urgent need to understand better the pathogenesis of PTCL to facilitate novel drug combinatorial approaches to improve survival. This position statement represents an evidence-based synthesis of the literature for application in Australian and New Zealand practice.

The Role of NOTCH1, GATA3, and c-MYC in T Cell Non-Hodgkin Lymphomas.

Simple SummaryPeripheral T cell lymphomas (PTCLs) comprise 10–15% of all non-Hodgkin lymphomas, and are more aggressive with worse prognosis. Due to their rarity and significant molecular abnormalities, the pathogenesis of PTCLs is not well understood. The expressions of NOTCH1, GATA3, and c-MYC have been linked to a poorer prognosis in PTCL, and are implicated in downstream processes. The aims of this review are to elucidate the role of NOTCH1, GATA3, and c-MYC in patients with PTCLs to provide additional information about the disease’s pathogenesis and to investigate the master regulator between these genes that will provide a basis for new therapeutic strategies and improve PTCL patients’ survival rates. NOTCH1 greatly influences the progression, pathogenicity, and development of PTCLs. Because NOTCH signalling does not rely on enzymatic signal amplification, and all Notch ligands and receptors include extracellular domains that are essential for receptor binding and activation, making them accessible to circulating therapies and targeting for different therapeutic strategies.Lymphomas are heterogeneous malignant tumours of white blood cells characterised by the aberrant proliferation of mature lymphoid cells or their precursors. Lymphomas are classified into main types depending on the histopathologic evidence of biopsy taken from an enlarged lymph node, progress stages, treatment strategies, and outcomes: Hodgkin and non-Hodgkin lymphoma (NHL). Moreover, lymphomas can be further divided into subtypes depending on the cell origin, and immunophenotypic and genetic aberrations. Many factors play vital roles in the progression, pathogenicity, incidence, and mortality rate of lymphomas. Among NHLs, peripheral T cell lymphomas (PTCLs) are rare lymphoid malignancies, that have various cellular morphology and genetic mutations. The clinical presentations are usually observed at the advanced stage of the disease. Many recent studies have reported that the expressions of NOTCH1, GATA3, and c-MYC are associated with poorer prognosis in PTCL and are involved in downstream activities. However, questions have been raised about the pathological relationship between these factors in PTCLs. Therefore, in this review, we investigate the role and relationship of the NOTCH1 pathway, transcriptional factor GATA3 and proto-oncogene c-MYC in normal T cell development and malignant PTCL subtypes.

A phase I/II study of golidocitinib, a selective JAK1 inhibitor, in refractory or relapsed peripheral T-cell lymphoma.

7563 Background: Peripheral T cell lymphoma (PTCL) is a group of heterogeneous T cell lymphomas. Patients who relapse from or are refractory to 1st line therapy face dismal prognosis. The response rates to commonly used 2nd line agents such as histone deacetylase inhibitors are below 30%. Immunotherapies, such as anti-PD1 antibodies, may induce hyperprogression in certain PTCL subtypes. Hence, r/r PTCL patients urgently need better therapies. Preclinical data shows JAK/STAT pathway may mediate the pathogenesis of PTCL, making it a promising target. Golidocitinib (DZD4205) is an orally available, potent, JAK1 specific inhibitor, demonstrating profound anti-tumor activities in T lymphoma cells in vitro and tumor xenograft in vivo. Here we report the preliminary data from an ongoing phase I/II study (NCT04105010) of Golidocitinib in r/r PTCL. Methods: The study included two parts: Part A (dose escalation) and Part B (dose expansion). In Part A, patients with r/r PTCL were enrolled and received Golidocitinib at different doses (150 mg or 250 mg, QD) to determine the recommended phase II dose (RP2D). Evaluation of safety and efficacy were performed by investigators per CTCAE and Lugano criteria, respectively. Part B is a single-arm, pivotal study, where patients with r/r PTCL will receive Golidocitinib at the RP2D till disease progression or intolerance. Results: As of May 31, 2021, a total of 51 patients enrolled in Part A and received Golidocitinib at 150 mg (n = 35) or 250 mg (n = 16). Patient characteristics: median age (range): 61.0 years (29-79); median prior systemic therapies (range): 2 lines (1-8). Ten patients (19.6%) had undergone hematopoietic stem cell transplantation. Fifteen patients (29.4%) had bone marrow involvement at baseline. Histological subtypes included PTCL-NOS (41.2%), AITL (39.2%), ALCL ALK- (7.8%), NKTCL (7.8%), and MEITL (3.9%). At the data cut-off (DCO), 49 patients completed at least one post-treatment Lugano assessment, of whom 21 (42.9%) achieved tumor response, including 11 complete responses (CRs, 22.4%) and 10 partial responses (20.4%). Tumor response was observed in various subtypes, including AITL (13/20), PTCL-NOS (5/19), ALCL ALK- (2/4) and NKTCL (1/4). At the DCO, the median duration of response (DoR) was not reached, and the longest DoR was > 14 months. Forty-eight patients (94.1%) experienced treatment emergent adverse events (TEAEs), of whom 30 (58.8%) experienced ≥ grade 3 TEAEs. Per investigators’ assessment, 20 patients (39.2%) experienced ≥ grade 3 TEAEs possibly related to the drug. The most common (≥ 10%) ≥ grade 3 TEAEs were neutropenia (29.4%), thrombocytopenia (15.7%) and pneumonia (11.8%). The majority of TEAEs were reversible or clinically manageable with dose modifications. Conclusions: Golidocitinib shows good safety and promising anti-tumor efficacy in r/r PTCL, indicating its potential as a therapeutic option for this unmet medical need. Clinical trial information: NCT04105010.

Expression of Vav1-Myo1F Fusion Affects T-Cell Differentiation and Induces T-Cell Lymphoma

Peripheral T-cell lymphomas (PTCL) are highly aggressive, malignant hematologic tumors that arise from clonal proliferation of mature T-cells. Among these, angioimmunoblastic T-cell lymphoma (AITL), and peripheral T cell lymphomas not otherwise specified (PTCL, NOS) account for >45% diagnoses, show limited response to intensified chemotherapy treatment and are associated with dismal survival. Genomic studies from our group have uncovered recurrent mutations and novel cancer-associated gene fusions involving the guanine nucleotide exchange factor VAV1 in AITL and PTCL, NOS. Interestingly, mutation co-occurrence analysis in AITL and PTCL, NOS showed significant mutual exclusivity of VAV1 genomic alterations and the highly prevalent RHOA mutations (p-value 0.0142), supporting a common mechanism of action. Gene fusions involving VAV1 are characterized by the substitution of their auto-inhibitory C-terminal SH3 domains by different domains from their fusion partners, leading to increased activation of VAV1-dependent signaling pathways. Among them, the VAV1-MYO1F fusion shows the strongest increase in VAV1 activity and activation of the mitogen-activated protein kinase (ERK1/2), c-Jun N-terminal kinase (JNK) and nuclear factor of activated T-cells (NFAT) pathways. To study the role of VAV1-MYO1F, we engineered a conditional knockin mouse that expresses the Vav1-myo1f fusion in CD4+ T-cells. Expression of Vav1-myo1f in CD4 T cells induces cell activation and alterations in T-cell specification, associated with up-regulation of master transcription factors involved in helper T-cell cell differentiation. Moreover, Vav1-Myo1f increased CD4+ T-cell survival upon cytokine withdrawal and enhanced Vav1 phosphorylation and activation of the MAPK pathway, resulting in increased cell activation and proliferation both in vivo and in vitro in response to TCR engagement. Notably, expression of Vav1-Myo1f fusion in CD4+ T-cells is sufficient to induce development of fatal malignant lymphomas with a latency of 6-14 months. Histological examination showed disrupted splenic architecture associated with clonal expansion of CD4+ cells indicative of T-cell lymphoma with PTCL, NOS phenotype. Similar results were obtained in a genetic model that combined the expression of Vav1-myo1f with the deletion of the Tet2 epigenetic regulator. In both genetic models, tumor cells specifically present a memory cell-associated immunophenotype (CD44+ CD62L-) and characteristic Th2-like features including increased expression of the transcription factors Gata3 and c-Maf and the IL4 and Il10 cytokines. Overall, these results demonstrate a direct oncogenic role for Vav1-Myo1f in the pathogenesis of PTCL, associated with deregulation of T-cell specification and of signaling programs critical for the control of T-cell proliferation. Disclosures Palomero: Kura Onclology: Research Funding.

VAV1-GFP Fusion Protein Generated By Intron-Based Genome Editing through CRISPR/Cas9 Leads to Spontaneous Activation in TCR Signaling

Recurrent VAV1 mutations and gene fusions (VAV1-THAP4, VAV1-MYO1F, and VAV1-S100A7) have been identified in peripheral T-cell lymphoma (PTCL) including angioimmunoblastic T-cell lymphoma (AITL) patients. A common theme of these genetic aberrations is the loss of the auto-inhibitory C-terminal SH3 domain of VAV1 resulting in aberrant activation of VAV1 independent of normal activation events. Although mouse models support VAV1 mutation/fusion as having a driver oncogenic role in the pathogenesis of PTCL, investigations on VAV1 activity in human cells were performed mainly on the Jurkat cell line with exogenous expression of VAV1 fusion proteins. This approach has un-physiological expression of VAV1 and the functions of VAV1 fusion/mutation under normal endogenous regulation need to be explored. In this study, we introduced a fusion gene, similar to what has been observed in PTCL, into the endogenous VAV1 locus. The fusion gene was under normal regulatory controls instead of being over-expressed by a viral vector, thus providing a more accurate assessment of its function in vivo. To simulate VAV1 fusion, we knocked in a green fluorescence protein (GFP) sequence followed by a simian virus 40 (SV40) poly(A) signal into intron 25 of VAV1 locus by clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) technology. A homologous DNA repair (HDR) template with tandem homologous sequences of VAV1 gene, GFP gene, and a SV40 transcription poly(A) signal was electroporated into Jurkat cells together with the Cas9/sgRNA ribonucleoprotein (RNP) complex. This knock-in disrupted the transcription of exon 26 and exon 27, resulting in an in-frame fusion protein with GFP fused to the C-terminal of SH2 of VAV1 (VAV1SH2-GFP)(Figure A). Because our guide RNA targeted the intron 25 sequence by CRISPR/Cas9 system, any possible indels caused by non-homologous end joining will occur within the intron and will not change the protein sequence of the wild type VAV1. The GFP expressing cells were isolated from the edited cell population by FACS. The fusion of GFP with VAV1 in the sorted cells was confirmed by western blot (Figure B) and these cells displayed a heterozygous VAV1SH2-GFP fusion/wild type (WT) phenotype that mimicked the VAV1 translocations observed in PTCL patients. Jurkat cells with VAV1SH2-GFP showed spontaneous activation of the T-cell receptor (TCR) signaling pathway. Analysis of signaling events downstream of VAV1 demonstrated increased phosphorylation in ITK, LCK, and subsequent ERK in Jurkat cells with VAV1SH2-GFP compared with WT Jurkat cells by western blot (Figure B). We also observed consistently elevated pERK in Jurkat cells with VAV1SH2-GFP by flow cytometry (Figure C). Notably, this elevation in pERK was spontaneous and independent of TCR stimulation with anti-CD3 antibody. VAV1SH2-GFP fusion protein also led to marked activation of downstream NFAT and NF-κB pathways as shown by Luciferase reporter assays (Figure D). Similarly, the enhanced NFAT and NF-κB pathway activation in Jurkat with the fusion protein was independent of TCR stimulation. Interestingly, with antiCD3 stimulation, Jurkat cells with VAV1SH2-GFP showed significantly lower pERK, NFAT and NF-κB activity compared to WT Jurkat cells with anti-CD3 stimulation. In conclusion, in Jurkat cells genetically edited with VAV1SH2-GFP, spontaneous activation of TCR signaling and subsequently increased NFAT and NF-κB activity were observed. Our findings further support that VAV1 C-terminal SH3 domain plays an important regulatory role in blocking VAV1 activity in the absence of proper activation. Removing C-terminal SH3 domain or replacing it with GFP or other protein relieves this inhibition, allowing spontaneous activation independent of TCR stimulation. Our study also indicates the sensitivity of the TCR signaling pathway to the level of activation and hyperactivation is detrimental. To validate the function of VAV1SH2-GFP in normal T cells, we have also successfully edited primary CD4+ T cells with VAV1SH2-GFP and the TCR signaling pathway in edited primary CD4+ T cell is currently being evaluated with and without anti-CD3/CD28 stimulation. Figure Disclosures No relevant conflicts of interest to declare.

A Phase I/II Study (JACKPOT8) of DZD4205, a Selective JAK1 Inhibitor, in Refractory or Relapsed Peripheral T- Cell Lymphoma

Background: Peripheral T-Cell Lymphoma (PTCL) is an aggressive malignancy which lacks effective treatment. Emerging data suggests that the JAK-STAT pathway may play an important role in mediating pathogenesis of PTCL. DZD4205 (AZD4205) is an orally available, potent and highly selective JAK1 inhibitor. In T-cell lymphoma cell lines, DZD4205 modulates pSTAT3 pathway and suppresses cell proliferation, and in tumor xenograft models, DZD4205 exhibits dose-dependent anti-tumor activities, with good correlation with drug exposure and modulations of pSTAT3 in tumor tissues. A phase I/II study (JACKPOT8) was initiated to assess the safety, tolerability, pharmacokinetics and anti-tumor efficacy of DZD4205 in patients with refractory/relapsed (r/r) PTCL. Methods: The JACKPOT8 study (ClinicalTrials.gov Identifier: NCT04105010) included two parts: Part A, dose escalation and Part B, dose expansion. In Part A, patients with r/r PTCL who have progressed on or were refractory to systemic therapy will be enrolled and treated with DZD4205 at two different dose levels, 150 mg or 250 mg once daily. In Part B, patients will receive DZD4205 treatment at a defined dose. The primary objective is to assess the safety and tolerability of DZD4205, and the secondary objectives include anti-tumor efficacy and pharmacokinetics. Evaluation of safety/tolerability and tumor response will be based on the CTCAE version 5.0 and 2014 Lugano classification, respectively. Patients will be treated until disease progression, intolerance to adverse events, or withdrawal of consent. Results: As of June 30, 2020, a total of 23 patients with r/r PTCL were enrolled and received DZD4205 at 150 mg (n = 19) or 250 mg (n = 4) once daily. Patient characteristics: median age (range): 65.0 years (34-79); median prior systemic therapies (range): 2 lines (1-8). Four patients had undergone prior hematopoietic stem cell transplantation (HSCT). Six patients had bone marrow involvement at the study entry. Histological subtypes included peripheral T-cell lymphoma not otherwise specified (PTCL-NOS) (n = 10, 43.5%), angioimmunoblastic T-cell lymphoma (AITL) (n = 10, 43.5%), ALK-negative anaplastic large cell lymphoma (ALCL ALK-) (n = 2, 8.7%) and extra-nodal nasal NK/T-cell lymphoma (NKTCL) (n = 1, 4.3%) . Preliminary data showed that 20 patients experienced treatment emergent adverse events (TEAEs), among whom 13 (56.5%) had ≥ grade 3 TEAEs. Based on local investigators' assessment, eight patients (34.8%) had ≥ grade 3 TEAEs which were considered to be related to DZD4205. The most common (> 5% incidence) DZD4205-related TEAEs (any grade) included thrombocytopenia (n = 6, 26.1%), neutropenia (n = 4, 17.3%), decreased appetite (n = 3, 13.0%), nausea (n = 2, 8.7%), interstitial lung disease (n = 2, 8.7%) and pneumonia (n = 2, 8.7%). Most TEAEs were manageable with dose interruption and reduction. Preliminary PK data is available in 20 patients (n = 19 at 150 mg, n = 1 at 250 mg). Exposure at 250 mg is higher than 150 mg following a single and multiple dose. As expected from long t1/2 of DZD4205, accumulation of about 3 folds in AUC was observed. DZD4205 had flat PK profile with small difference between Css,max and Css,min after 22 days of once daily dosing, which is an optimal PK profile to maintain DZD4205 concentrations above effective levels throughout the dosing interval. As of June 30, 2020, 22 patients (n = 19 at 150 mg, n = 3 at 250 mg) had at least one post-treatment Lugano assessment. In 150 mg cohort, 8 out of 19 patients showed tumor response, with objective response rate (ORR) of 42%, among whom 4 (21%) had complete response (CR) and another 4 (21%) partial response (PR). In 250 mg cohort, 1 out of 3 patients showed tumor response, with ORR of 33%. Tumor response was observed in subtypes including AITL, PTCL-NOS, ALCL (ALK-) and NKTCL. At the data cut-off date, the longest duration on treatment was > 8 months. Patient enrolment is ongoing. Updated clinical data will be shared at the meeting. Conclusion: DZD4205 shows good tolerability, pharmacokinetic profiles and promising anti-tumor efficacy in patients with r/r PTCL, indicating its potential as a therapeutic option for this unmet medical need. Disclosures Kim: Donga: Research Funding; Kyowa-Kirin: Research Funding; Roche: Research Funding; Johnson&Johnson: Research Funding; Celltrion: Research Funding; Mundipharma: Research Funding. Yoon:Celltrion: Honoraria; Samyang: Research Funding; Amgen, Chongkundang, Celgene, Astrazeneca: Consultancy. Wang:Dizal Pharmaceuticals: Current Employment. Li:Dizal Pharmaceuticals: Current Employment. Huang:Dizal Pharmaceuticals: Current Employment. Deng:Dizal Pharmaceuticals: Current Employment. Chen:Dizal Pharmaceuticals: Current Employment. Yang:Dizal Pharmaceuticals: Current Employment.

Identification of Hub Genes and Key Pathways Associated with Peripheral T-cell Lymphoma.

Peripheral T-cell lymphoma (PTCL) is a very aggressive and heterogeneous hematological malignancy and has no effective targeted therapy. The molecular pathogenesis of PTCL remains unknown. In this study, we chose the gene expression profile of GSE6338 from the Gene Expression Omnibus (GEO) database to identify hub genes and key pathways and explore possible molecular pathogenesis of PTCL by bioinformatic analysis. Differentially expressed genes (DEGs) between PTCL and normal T cells were selected using GEO2R tool. Gene ontology (GO) analysis and Kyoto Encyclopedia of Gene and Genome (KEGG) pathway analysis were performed using Database for Annotation, Visualization and Integrated Discovery (DAVID). Moreover, the Search Tool for the Retrieval of Interacting Genes (STRING) and Molecular Complex Detection (MCODE) were utilized to construct protein-protein interaction (PPI) network and perform module analysis of these DEGs. A total of 518 DEGs were identified, including 413 down-regulated and 105 up-regulated genes. The down-regulated genes were enriched in osteoclast differentiation, Chagas disease and mitogen-activated protein kinase (MAPK) signaling pathway. The up-regulated genes were mainly associated with extracellular matrix (ECM)-receptor interaction, focal adhesion and pertussis. Four important modules were detected from the PPI network by using MCODE software. Fifteen hub genes with a high degree of connectivity were selected. Our study identified DEGs, hub genes and pathways associated with PTCL by bioinformatic analysis. Results provide a basis for further study on the pathogenesis of PTCL.

Pathology and Pathogenesis of T-Cell Lymphoma

Peripheral T cell lymphomas (PTCLs) are a heterogeneous and often clinically aggressive group of neoplasms derived from mature post-thymic T lymphocytes. These neoplasms are rare and usually diagnostically challenging. Our understanding of the pathogenesis of PTCL is increasing and this improved knowledge is leading us to better molecular characterization, more objective diagnostic criteria, more effective risk assessment, and potentially to better treatments for these neoplasms.

Role of Vav1 Genomic Alterations in T-Cell Differentiation and Transformation in Peripheral T-Cell Lymphoma

Peripheral T-cell lymphomas (PTCL) are highly aggressive, malignant hematologic tumors that arise from clonal proliferation of mature post-thymic T-cells. Among these, angioimmunoblastic T-cell lymphoma (AITL), and peripheral T-cell lymphomas not otherwise specified (PTCL, NOS) account for >45% diagnoses, show limited response to intensified chemotherapy treatment with dismal overall survival rate. Our genomic studies have uncovered recurrent mutations and novel cancer-associated gene fusions involving the guanine nucleotide exchange factor VAV1 in AITL and PTCL, NOS, supporting a role for the VAV1 oncogene in the pathogenesis of PTCL. Genetic alterations in VAV1 found in PTCL included recurrent deletions resulting in the in-frame deletion of 9 codons (DV778_T786) in the linker region connecting the C-terminal SH2 domain to the downstream SH3 domain on VAV1 as well as recurrent different gene-fusion transcripts encoding proteins in which the C-terminal SH3 domain of VAV1 is replaced by different domains of the fusion partner. Consistent with the auto-inhibitory role of the C-SH3 domain, the PTCL-associated VAV1 alterations resulted in increased VAV1 activity and strong activation of the mitogen-activated protein kinase (ERK1/2), c-Jun N-terminal kinase (JNK) and nuclear factor of activated T-cells (NFAT) pathways. Interestingly, mutation co-occurrence analysis shows significant mutual exclusivity of the highly prevalent AITL RHOA G17V mutation and VAV1 genomic alterations (p-value 0.0142) in AITL and PTCL, NOS. This observation supports that VAV1 genetic alterations and RHOA G17V could share a common mechanism of action. To study the role and mechanisms of VAV1 alterations in PTCL in vivo we have engineered a C57/BL6 mouse lines with conditional expression of the recurrent Vav1-Myo1f fusion. By crossing Vav1-myo1f conditional knockin with CD4-CreERT2 mouse deletor line, we have generated animals with CD4+ T cell-specific and tamoxifen-inducible expression of Vav1-myo1f. Preliminary studies indicate that expression of Vav1-myo1f in CD4+ cells induces expansion of TFH cells. This phenotype partially resembles that of animals with CD4 specific expression of Rhoa G17V, indicating a potential convergent effect and mechanisms for VAV1 alterations and RHOA G17V in T-cell development and transformation. However, and more interestingly, Vav1-myo1f expressing cells specifically present a memory cell-associated immunophenotype (CD44+ CD62L-) and characteristic Th2-like features that had not been observed in the Rhoa G17V model. Analysis of signaling pathways activated in control and Vav1-myo1f expressing CD4+ cells showed increased phosphorylation of Vav1 and increased activation of the MAPK pathway upon expression of Vav1-myo1f, supporting the role of Vav1 alterations as regulators of T-cell activation. Notably, single cell transcriptome analysis of bulk splenic cells isolated from CD4-CreERT2 control and CD4-CreERT2Vav1co-Vav1-myo1f/WT mice indicate that expression of Vav1-myo1f in CD4+ cells leads to a significant expansion of monocytes and TFH cells and a decrease in B-cell, both naïve and memory. These results support that expression of Vav1-myo1f in CD4+ cells can deeply modify the lymphoid microenvironment. Finally, preliminary analysis of a limited number of mice with targeted expression of the Vav1-Myo1f fusion in CD4+ cells (CD4-Cre Vav1co-Vav1-Myo1f /WT) showed development of a fatal CD4+ malignant lymphoproliferation with a latency of 6-14 months. Histological examination showed disrupted splenic architecture accompanied with clonal expansion of CD4+ cells indicative of T-cell lymphoma with PTCL, NOS characteristics and verified expression of both wild type Vav1 and Vav1-myo1f alleles. Interestingly, Vav1-myo1f lymphoma cells showed Th2 associated features including expression of Gata3, c-Maf and IL4 and Il10. These results support an important role for VAV1 alterations in T-cell differentiation, remodeling of the tumor microenvironment and T-cell lymphoma development and highlight the importance of our model as a new experimental system to study the evolution of PTCL and develop novel targeted therapies for this disease. Disclosures Palomero: Kura Oncology: Research Funding.

A Phase 2 Study of the Dual SYK/JAK Inhibitor Cerdulatinib Demonstrates Good Tolerability and Clinical Response in Relapsed/Refractory Peripheral T-Cell Lymphoma and Cutaneous T-Cell Lymphoma

A Phase 2 Study of the Dual SYK/JAK Inhibitor Cerdulatinib Demonstrates Good Tolerability and Clinical Response in Relapsed/Refractory Peripheral T-Cell Lymphoma and Cutaneous T-Cell Lymphoma

Survival outcomes of patients with peripheral T-cell lymphomas (PTCL) treated with chemotherapy and/or novel agents: The Columbia University experience.

e19049 Background: CHOP-based regimens have been considered the standard of care for PTCL despite disappointing results. Several lines of evidence, including the regulatory approval of novel drugs since 2009, raise the question as to whether these drugs are changing the natural history of relapsed PTCL. Recent molecular insights into the pathogenesis of PTCL, especially in angioimmunoblastic T-cell lymphoma (AITL), have revealed a number of genetic lesions that might portend a unique vulnerability to select agents, such as histone deacetylase inhibitors (HDACi). Methods: This is a retrospective study of patients with PTCL who were seen and treated between 1994 and 2018. Kaplan-Meier curves for overall survival were generated and compared based on the log-rank test. Cox proportional hazard models were used to investigate the association by adjusting for age, gender, subtype, prognostic score and type of treatment. The analysis was done in SAS version 9.4. A p-value < 0.05 was considered statistically significant. Results: At Columbia University Medical Center, a total of 186 patients with PTCL were identified. Excluding patients with incomplete data, 168 patients were analyzed. The median overall survival was 7.5 months (95% CI 5.6-8.9) for the whole study group. Cox modeling adjusted for the aforementioned variables found little impact from the novel agents on the natural history of the disease. In particular, among patients with AITL, no survival benefit was associated with any HDACi. In addition, these data demonstrated that patients with adult T-cell leukemia lymphoma (ATLL) had a better overall survival compared to other subgroups, p = 0.0097. Conclusions: These data are concordant with other large experiences noting the poor prognosis of patients with PTCL, and in particular, those with relapsed disease. The lack of impact of HDACi on the natural history of AITL raises issues about the role of these drugs in this disease. We will present a variety of subanalyses to identify prognostic factors in PTCL. [Table: see text]

The Novel Tumor Suppressor SAMHD1 Is Differentially Expressed and Partly Regulated By MYC in Peripheral T-Cell Lymphomas (PTCL)

Introduction: The SAM domain and HD domain 1 (SAMHD1) protein is a deoxynucleoside triphosphate (dNTP) triphosphohydrolase, which has been initially described to restrict human immunodeficiency virus type 1 (HIV-1) in certain cell types through depletion of intracellular dNTP substrates required for HIV-1 reverse transcription. Mutations of SAMHD1 gene have been linked to Aicardi-Goutières syndrome (AGS) and have been identified as putative drivers of chronic lymphocytic leukemia resulting in decreased SAMHD1 mRNA and protein levels. More recently, SAMHD1 mutations have been reported in T-prolymphocytic leukemia (T-PLL). Based on these findings and the fact that SAMHD1 limits the dNTP pool in the cell, it may play a role in oncogenesis as a tumor suppressor. In addition, SAMHD1 may confer resistance to nucleoside-based chemotherapies by hydrolysing their active triphosphate metabolites, with cytarabine in acute myeloid leukemia being an example (Herold et al, Nat Med 2017; 23(2):256-263). The expression patterns and the potential role of SAMHD1 in the pathogenesis of peripheral T-cell lymphomas (PTCL) are not yet known.Methods: The patient cohort included 64 PTCLs of various histologic types which were diagnosed and treated at Karolinska University Hospital (Sweden). A control group of 4 reactive lymph nodes and 2 reactive tonsils was included in the study for comparison. All tissue samples were obtained prior to therapy. SAMHD1 expression was assessed by immunohistochemistry performed on a PTCL tissue microarray (TMA) with duplicate tumor cores from each case or full tissue sections using dual immunostaining (SAMHD1 / CD68) and a monoclonal antibody against SAMHD1 (Bethyl Laboratories, San Antonio, TX). At least 500 lymphoma cells were counted to calculate the percentage of SAMHD1-positive tumor cells. Overall survival (OS) was defined as time from diagnosis to death or last follow-up. Event-free survival (EFS) was defined as time from diagnosis to relapse, death, or last follow-up. Survival analyses were performed using the Kaplan-Meier method (log-rank test) and Cox regression models. Two T-cell lymphomas cell lines (Mac1, Mac2A) were used as an in vitro system. As our preliminary findings from in silico analysis revealed potential binding sites for MYC on the SAMHD1 gene promoter, we hypothesized that MYC might regulate SAMHD1 expression. Therefore, the T-cell lymphoma cell lines were treated with the selective BET / MYC inhibitor JQ-1 or transiently transfected with a MYC-overexpressing plasmid or MYC gene-specific siRNA constructs, respectively. Western blot analysis was used to assess the protein levels.Results: SAMHD1 protein was expressed in reactive T-cells and histiocytes (CD68+) in all reactive lymphoid tissues (lymph nodes and tonsils) with strong staining intensity. SAMHD was differentially expressed among PTCL subtypes generally with weaker staining intensity as compared to normal T-cells and histiocytes, thus being positive in all (100%) angioimmunoblastic T-cell lymphomas (AILT), 67% PTCL-NOS, 45% ALK+ ALCL, 20% of ALK+ ALCL, and none (0%) of T-lymphoblastic lymphomas (p=0.0017, chi-square test). Among the SAMHD1- positive cases, the percentage of positive lymphoma cells ranged from 0 to 100% and its highest median was observed in AILT. SAMHD1 expression inversely correlated with CD30 expression (% CD30+ positive lymphoma cells) (p=0.0025, Mann-Whitney test). No significant associations between SAMHD1 levels and other clinicopathologic parameters or clinical outcome (EFS or OS) were found, however, the number of patients analyzed in each histologic subtype was limited. Inhibition of MYC activity by JQ-1 or MYC gene silencing with specific siRNA resulted in a substantial increase in the SAMHD1 protein level in T-cell lymphoma cell lines. Inversely, transient transfection of the cell lines with a MYC overexpressing plasmid resulted in decreased levels of SAMHD1. Taken together, the in vitro data suggest a possible MYC-associated regulation (repression) of SAMHD1 gene expression in T-cell lymphoma.Conclusions: SAMHD1 is shown for the first time to be differentially expressed among PTCL types and its regulation may involve MYC. Preliminary survival analysis shows no significant associations of SAMHD1 expression with EFS and OS in this cohort of PTCL, however, analysis of a larger PTCL study group is underway to draw definite conclusions. DisclosuresÖsterborg:Gilead: Consultancy, Research Funding; Beigene: Research Funding; Pharmacyclics: Research Funding; Janssen: Research Funding; Abbvie: Research Funding.

Recent Advances in the Treatment of Peripheral T-Cell Lymphoma.

Recent progress in understanding of the biology and pathogenesis of peripheral T-cell lymphoma has led to the emergence of new drugs. Unfortunately, this has not been met with similar advances in outcome improvement. Anthracycline-containing regimens, mostly cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP), are considered the standard of care, although the best first-line approach remains to be defined. In the relapsed and refractory settings, several new agents achieved response rates between 10% and 30%, although these drugs do not significantly affect survival rates. Therapeutic options based on better molecular characterization of various histological types and combinations with the CHOP regimen or synergic combinations of new drugs may lead to better outcomes.

Activating mutations and translocations in the guanine exchange factor VAV1 in peripheral T-cell lymphomas

Peripheral T-cell lymphomas (PTCLs) are a heterogeneous group of non-Hodgkin lymphomas frequently associated with poor prognosis and for which genetic mechanisms of transformation remain incompletely understood. Using RNA sequencing and targeted sequencing, here we identify a recurrent in-frame deletion (VAV1 Δ778-786) generated by a focal deletion-driven alternative splicing mechanism as well as novel VAV1 gene fusions (VAV1-THAP4, VAV1-MYO1F, and VAV1-S100A7) in PTCL. Mechanistically these genetic lesions result in increased activation of VAV1 catalytic-dependent (MAPK, JNK) and non-catalytic-dependent (nuclear factor of activated T cells, NFAT) VAV1 effector pathways. These results support a driver oncogenic role for VAV1 signaling in the pathogenesis of PTCL.

Histone-Modifying Gene Mutations Are Related to Tumor Progression and Response to Histone Deacetylase Inhibitors in Patients with Peripheral T-Cell Lymphoma Not Otherwise Specified

ObjectiveHistone modifications are critically involved in pathogenesis of peripheral T-cell lymphoma not otherwise specified (PTCL-NOS).Histonedeacetylaseinhibitors (HDACIs) exhibited potential anti-lymphoma activity through restoring normal chromatin modification patterns. Here we detected histone-modifying gene mutations in PTCL-NOS and correlated them with tumor progression and therapeutic response to different HDACIs.MethodsMutations of important lymphoma-associated histone-modifying genes (MLL2, TET2, CREBBP, EP300, SETD2 and EZH2) were detected by targeted sequencing in tumor samples of 105 newly diagnosed patients with PTCL-NOS and compared with clinical parameters. Moreover, T-lymphoma cell lines bearing representative mutations were treated with four clinically available HDACIs (valproic acid, VPA;suberoylanilidehydroxamic acid, SAHA;Romidepsin,Romi; andChidamide,Chid).ResultsA total of 65 histone-modifying gene mutations were identified in 39 of 105 PTCL patients (Figure 1A). As illustrated in Figure 1B, somatic mutations in MLL2 (encoding H3K4-specific methyltransferase) were most frequently observed (20.0%), followed by mutations of TET2 (13.3%), EP300 and CREBBP (encoding H3K18 acetyltransferase, 7.6% and 6.7%, respectively), and SETD2 (encoding H3K36 methyltransferase, 7.6%). No EZH2 mutation was detected. Clinically, mutation-positive PTCL patients presented with shorteroverall survival than patients without mutations (P=0.0038, Figure 1C).Based on the crystal structure of each gene (Figure 1D), most mutations were located on SET domain of MLL2 (p.5397-5513), HAT domain of EP300 (p.1287-1663) and SET domain of SETD2 (p.1550-1667), which could lead to loss of H3K18ac, H3K4me3 and H3K36me3 expression, respectively.Meanwhile, Jurkatcells were treated with VPA, SAHA, Romiand Chidfor 48h and the results showed that all the HDACIs increased the expression of EP300/CREBBP, as compared to the untreated group. Interestingly, VPA and SAHA specifically upregulated SETD2 expression, while Romi and Chid upregulated MLL2 expression (Figure 2A). No obvious change of TET2 expression was found during HDACI treatment. Thereafter,Jurkat cells were transfected with MLL2 mutant (p.V5389M), EP300 mutant (p.H1377R) and SETD2 mutant (p.R1598_). Compared with the WT, mutant reduced the level of H3K18ac, H3K4me3 and H3K36me3 expression, respectively. All the HDACIs restored H3K18 hypo-acetylation induced by EP300 mutant. VPA and SAHA restored H3K36me3 expression inhibited by SETD2 mutant, whileRomi andChid restored H3K4me3 expression inhibited by MLL2 mutant (Figure 2B-2D).ConclusionOur study provided the mutational spectrum of histone-modifying genes in PTCL-NOS. HDACIs targeted differently histone H3 acetylation or methylation modulated by the mutations, suggestive their distinct therapeutic efficiency in PTCL-NOS. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Molecular Subgroups of Peripheral T-Cell Lymphoma Evolve By Distinct Genetic Pathways

Peripheral T-cell lymphoma (PTCL) is a group of clinically and pathologically heterogeneous non-Hodgkin lymphomas (NHL). Using gene expression profiling (GEP), we have defined molecular classifiers for PTCL subtypes reflecting their pathobiology and oncogenic pathways (Iqbal et al. 2014). We have also shown associations of specific mutations with the molecular subgroups (Wang et al. 2015). Although genomic information is increasing, the pathogenetic mechanisms of PTCLs remain largely unknown. Therefore, we analyzed copy number variation (CNV) and GEP to identify unique genetic abnormalities in the defined PTCL molecular subgroups.CNV data were generated on fresh frozen or formalin-fixed paraffin-embedded genomic DNA (n=114) on 3 Affymetrix platforms (SNP 6.0, 250K SNP, and OncoScan). Two published cohorts (PTCL-NOS, Hartmann et al. 2010; ALCL, Boi et al. 2013) were included for validation. The gene expression analysis, morphological review and clinical characteristics of these cases have been included in previous studies (Iqbal et al. 2010, 2014).Angioimmunoblastic T-cell lymphoma (AITL) represents 20% of all PTCL cases. The most recurrent CNV in AITL was chromosome (chr) 5 gain (39%), followed by chr 21 gain (21%). Interestingly, chr 21 gain co-occurred with chr 5 gain (p=0.003). No recurrent losses (≥20%) were identified among these cases. Molecularly re-classified AITL cases from morphologically classified PTCL-NOS cases showed concordant results with bonafide AITL cases. Of the commonly mutated genes, DNMT3A, IDH2, RHOA and TET2, only IDH2R172Kshowed a significant association (p=0.012) with chr 5 gain. GEP showed enrichment of gene signatures associated with oxidative phosphorylation (PGC-1α target genes) in cases with chr 5 gain.PTCL, not otherwise specified (PTCL-NOS) is the most common PTCL subtype and cannot be further sub-classified using conventional approaches; however, we have identified 2 molecular subgroups within PTCL-NOS, the GATA3 and TBX21 subgroups which are related to 2 distinct T-helper subsets (Iqbal et al. 2014), by employing GEP. Consistent with earlier observations (Hartmann et al. 2010), PTCL-NOS showed remarkably varied CNVs with nearly 50% of cases showing high CNV frequencies. When correlated with molecular subgroups, distinctive CNVs were observed in the molecular GATA3 and TBX21 subgroups. The GATA3 subgroup displayed a large assortment of CNVs. Complete or partial gain of chr 7 (57%) was the most recurrent gain in these cases. Losses affecting 17p, 10q and 9p21, encompassing tumor suppressors such as TP53 (57%), PTEN (43%) and CDKN2A (43%), were frequent in the GATA3 subgroup. The TBX21 subgroup had significantly fewer CNVs, as none were recurring (≥20%); but gains of 5p or 11p were observed in 14%. Additionally, PTCL-NOS cases with ≥10% abnormal genome had significantly poorer overall survival (p=0.012) compared to those with fewer abnormalities. This finding validates the GEP molecularly defined subgroups, as the GATA3 subgroup displayed more CNVs and has been associated with a worse prognosis compared to the TBX21 subgroup (Iqbal et al. 2014).We were able to distinguish CNVs characteristic of the different entities, including the co-occurrence of chr 5 and 21 gains specific in AITL. Gain of 1q (complete or partial) was identified in the GATA3 subgroup of PTCL-NOS and anaplastic lymphoma kinase (ALK) (-) ALCL with equal frequencies (~ 36%), but only 16% in ALK(+) ALCL. Complete or partial gain of chr 7 was also observed in ALCL, but at a considerably lower frequency than in the GATA3 subgroup. Additionally, gain of chr 18 or regions of 17q, and loss of 5q or regions on both arms of chr 9, were more frequent in the GATA3 subgroup compared to other entities. The TBX21 subgroup was primarily differentiated from the GATA3 subgroup by presence of fewer CNVs.Our analysis provides a framework for future investigations into the molecular pathogenesis of PTCL, and highlights potential candidate oncogenes and tumor suppressors deregulated by copy number aberrations. Comparative analysis revealed that certain chromosomal abnormalities are entity-specific. AITL cases with IDH2R172K also had trisomy 5 suggesting that these oncogenic events cooperate in malignant transformation. Thus, the complexity of PTCL is finally becoming clearer with the integration of high resolution molecular techniques for global genomic analysis. DisclosuresNo relevant conflicts of interest to declare.

GATA3 expression correlates with poor prognosis and tumor-associated macrophage infiltration in peripheral T cell lymphoma.

Peripheral T cell lymphoma (PTCL) is an aggressive form of non-Hodgkin's lymphoma characterized by a poor prognosis. In this study, we examined the prognostic value of two T-cell-specific transcription factors, GATA3 and T-bet, in PTCL, uncovered the pathogenesis of PTCL, and investigated new PTCL therapeutic targets. Samples from 109 PTCL patients were examined for expression of GATA3, T-bet and CD68. High GATA3 expression correlated with poor survival in PTCL patients and with tumor-associated macrophage (TAM) infiltration, as indicated by the presence of CD68-positive cells. Multivariate analysis further confirmed that high GATA3 expression and Eastern Cooperative Oncology Group (ECOG) scores higher than 2 were independent predictors of patient survival. Using lentiviral transfection to induce stable GATA3 knockdown in a PTCL cell line, we observed that GATA-3 knockdown in Hut78 cells decreased levels of IL4, IL5, IL13 and VEGF mRNA and reduced the number of co-cultured U937 cells that differentiated towards the M2 phenotype. These results suggest that high GATA3 expression is a predictor of a poor prognosis in PTCL, and that T lymphoma cells promote M2-type macrophage differentiation through a GATA3-dependent mechanism.

Molecular mechanisms of angioimmunoblastic T-cell lymphoma development.

The molecular pathogenesis of peripheral T-cell lymphoma (PTCL) has gradually been clarified in terms of genomic abnormalities. Insights into these genomic abnormalities have provided clues to understanding the pathogenesis of PTCL. Furthermore, the origins of lymphoma cells have been clarified by investigating the distribution of genomic abnormalities in tumor cells and non-tumor blood cells. Multistep tumorigenesis has been suggested to be a fundamental mechanism underlying the development of angioimmunoblastic T-cell lymphoma (AITL), a distinct subtype of PTCL: premalignant cells evolve from hematopoietic progenitors via mutations in epigenetic regulators. These cells then further differentiate into tumor cells via the addition of tumor-specific G17V RHOA mutations. Meanwhile, AITL are composed of various infiltrating cells as well as tumor cells. Most notably, AITL tissues are characterized by massive infiltration of B cells partially infected by Epstein-Barr virus, follicular dendritic cells, and high endothelial venules. Infiltration of these cell types has been thought to be a reactive process, promoted by cytokines and chemokines released from tumor cells. Considering the multistep mechanisms of AITL allows us to analyze whether these infiltrating cells are also derived from premalignant cells. Indeed, the mechanisms underlying massive infiltration of bystander cells might be more complicated than previously imagined.