PI3Kδ activation, IL6 over-expression, and CD37 loss cause resistance to the targeting of CD37-positive lymphomas with the antibody-drug conjugate naratuximab emtansine.

Purpose: The transmembrane protein CD37 is expressed almost exclusively in lymphoid tissues, with the highest abundance in mature B cells. CD37-directed antibody- and, more recently, cellular-based approaches have shown preclinical and promising early clinical activity. Naratuximab emtansine (Debio 1562, IMGN529) is an antibodydrug conjugate (ADC) that incorporates an anti-CD37 monoclonal antibody conjugated to the maytansinoid DM1 as payload. Naratuximab emtansine has shown activity as a single agent and in combination with the anti-CD20 monoclonal antibody rituximab in B cell lymphoma patients. Experimental Design: We assessed the activity of naratuximab emtansine using in vitro models of lymphomas, correlated its activity with CD37 expression levels, characterized two resistance mechanisms to the ADC, and identified combination partners providing synergy. Results: The anti-tumor activity of naratuximab emtansine was tested in 54 lymphoma cell lines alongside its free payload. The median IC 50 of naratuximab emtansine was 780 pM, and the activity, primarily cytotoxic, was more potent in B than in T cell lymphoma cell lines. In the subgroup of cell lines derived from B cell lymphoma, there was some correlation between sensitivity to DM1 and sensitivity to naratuximab emtansine (r=0.28, P = 0.06). After prolonged exposure to the ADC, one diffuse large B cell lymphoma (DLBCL) cell line developed resistance to the ADC due to the biallelic loss of the CD37 gene. After CD37 loss, we also observed upregulation of IL6 (IL-6) and other transcripts from MYD88/IL6-signaling. Recombinant IL6 led to resistance to naratuximab emtansine, while the anti-IL6 antibody tocilizumab improved the cytotoxic activity of the ADC in CD37-positive cells. In a second model, resistance was sustained by an activating mutation in the PIK3CD gene, associated with increased sensitivity to PI3K δ inhibition and a switch from functional dependence on the anti-apoptotic protein MCL1 to reliance on BCL2. The addition of idelalisib or venetoclax to naratuximab emtansine overcame resistance to the ADC in the resistant derivative while also improving the cytotoxic activity of the ADC in the parental cells. Conclusions: Targeting B cell lymphoma with the CD37 targeting ADC naratuximab emtansine showed vigorous anti-tumor activity as a single agent, which was also observed in models bearing genetic lesions associated with inferior outcomes, such as MYC translocations and TP53 inactivation or resistance to R-CHOP. Resistance DLBCL models identified active combinations of naratuximab emtansine with drugs targeting IL6, PI3K δ , and BCL2. Despite notable progress in recent decades, we still face challenges in achieving a cure for a substantial number of lymphoma patients (1,2). A pertinent example is diffuse large B cell lymphoma (DLBCL), the most prevalent type of lymphoma (3). More than half of DLBCL patients can achieve remission, but around 40% of them experience refractory disease or relapse following an initial positive response (3). Regrettably, the prognosis for many of these cases remains unsatisfactory despite introducing the most recent antibody-based or cellular therapies (3,4), underscoring the importance of innovating new therapeutic strategies and gaining insights into the mechanisms of therapy resistance. CD37 is a transmembrane glycoprotein belonging to the tetraspanin family, primarily expressed on the surface of immune cells, principally in mature B cells but also, at lower levels, in T cells, macrophages/monocytes, granulocytes and dendritic cells (5) (6-8). CD37 plays a crucial role in various immune functions, including B cell activation, proliferation, and signaling, although its precise role still needs to be fully elucidated. CD37 interacts with multiple molecules, including SYK, LYN, CD19, CD22, PI3K δ , PI3K γ , and different integrins, among others (6-8). In mice, the lack of CD37 is paired with reduced T cell-dependent antibody-secreting cells and memory B cells, apparently due to the loss of CD37-mediated clustering of α 4 ß 1 integrins (VLA-4) on germinal center B cells and decreased downstream activation of PI3K/AKT signaling and cell survival (5). Reflecting the expression pattern observed in normal lymphocytes, CD37 exhibits elevated expression in all mature B-cell lymphoid neoplasms, including most lymphoma subtypes, and absence in early progenitor cells or terminally differentiated plasma cells (6,8-14). In DLBCL, CD37 expression has been reported between 40% and 90% of cases across multiple studies performed using different antibodies (10,14-16). CD37-directed antibody- and, more recently, cellular-based approaches have shown preclinical (7,10-14,17-23) and early promising clinical activity (24-32). Among the CD37-targeting agents, naratuximab emtansine (Debio 1562, IMGN529) is an antibody-drug conjugate (ADC) that incorporates the anti-CD37 humanized IgG1 monoclonal antibody K7153A conjugated to the maytansinoid DM1, as payload, via the thioether linker, N-succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC) (10). Based on the initial in vitro and in vivo evidence of anti-tumor activity in lymphoma and chronic lymphocytic leukemia (CLL) (7,10), naratuximab emtansine entered the clinical evaluation as a single agent. The phase 1 study exploring naratuximab emtansine enrolled 39 patients with relapsed/refractory B cell lymphoma (27). The overall response rate (ORR) was 13% across all patients and 22% in DLBCL patients, including the only observed complete remission (CR) (27). In preliminary results of a phase 2 trial exploring the combination of naratuximab emtansine with the anti-CD20 monoclonal antibody rituximab (18), based on positive preclinical data (18), the ORR was 45% in 76 patients with DLBCL with 24 CRs (32%), 57% in 14 patients with follicular lymphoma (five CR), 50% in four MCL patients (2 CR) (31). Here, we studied the pattern of activity of naratuximab emtansine across a large panel of cell lines derived from DLBCL and other lymphoma subtypes and characterized two resistance mechanisms to the ADC.

Systems analyses of the Fabry kidney transcriptome and its response to enzyme replacement therapy identified and cross-validated enzyme replacement therapy-resistant targets amenable to drug repurposing.

Fabry disease is a rare disorder caused by variations in the alpha-galactosidase gene. To a degree, Fabry disease is manageable via enzyme replacement therapy (ERT). By understanding the molecular basis of Fabry nephropathy (FN) and ERT's long-term impact, here we aimed to provide a framework for selection of potential disease biomarkers and drug targets. We obtained biopsies from eight control individuals and two independent FN cohorts comprising 16 individuals taken prior to and after up to ten years of ERT, and performed RNAseq analysis. Combining pathway-centered analyses with network-science allowed computation of transcriptional landscapes from four nephron compartments and their integration with existing proteome and drug-target interactome data. Comparing these transcriptional landscapes revealed high inter-cohort heterogeneity. Kidney compartment transcriptional landscapes comprehensively reflected differences in FN cohort characteristics. With exception of a few aspects, in particular arteries, early ERT in patients with classical Fabry could lastingly revert FN gene expression patterns to closely match that of control individuals. Pathways nonetheless consistently altered in both FN cohorts pre-ERT were mostly in glomeruli and arteries and related to the same biological themes. While keratinization-related processes in glomeruli were sensitive to ERT, a majority of alterations, such as transporter activity and responses to stimuli, remained dysregulated or reemerged despite ERT. Inferring an ERT-resistant genetic module of expressed genes identified 69 drugs for potential repurposing matching the proteins encoded by 12 genes. Thus, we identified and cross-validated ERT-resistant gene product modules that, when leveraged with external data, allowed estimating their suitability as biomarkers to potentially track disease course or treatment efficacy and potential targets for adjunct pharmaceutical treatment.

Myc controls NK cell development, IL-15-driven expansion, and translational machinery.

MYC is a pleiotropic transcription factor involved in cancer, cell proliferation, and metabolism. Its regulation and function in NK cells, which are innate cytotoxic lymphocytes important to control viral infections and cancer, remain poorly defined. Here, we show that mice deficient for Myc in NK cells presented a severe reduction in these lymphocytes. Myc was required for NK cell development and expansion in response to the key cytokine IL-15, which induced Myc through transcriptional and posttranslational mechanisms. Mechanistically, Myc ablation in vivo largely impacted NK cells' ribosomagenesis, reducing their translation and expansion capacities. Similar results were obtained by inhibiting MYC in human NK cells. Impairing translation by pharmacological intervention phenocopied the consequences of deleting or blocking MYC in vitro. Notably, mice lacking Myc in NK cells exhibited defective anticancer immunity, which reflected their decreased numbers of mature NK cells exerting suboptimal cytotoxic functions. These results indicate that MYC is a central node in NK cells, connecting IL-15 to translational fitness, expansion, and anticancer immunity.

Subcapsular Sinus Macrophages Promote Melanoma Metastasis to the Sentinel Lymph Nodes via an IL1α-STAT3 Axis.

During melanoma metastasis, tumor cells originating in the skin migrate via lymphatic vessels to the sentinel lymph node (sLN). This process facilitates tumor cell spread across the body. Here, we characterized the innate inflammatory response to melanoma in the metastatic microenvironment of the sLN. We found that macrophages located in the subcapsular sinus (SS) produced protumoral IL1α after recognition of tumoral antigens. Moreover, we confirmed that the elimination of LN macrophages or the administration of an IL1α-specific blocking antibody reduced metastatic spread. To understand the mechanism of action of IL1α in the context of the sLN microenvironment, we applied single-cell RNA sequencing to microdissected metastases obtained from animals treated with the IL1α-specific blocking antibody. Among the different pathways affected, we identified STAT3 as one of the main targets of IL1α signaling in metastatic tumor cells. Moreover, we found that the antitumoral effect of the anti-IL1α was not mediated by lymphocytes because Il1r1 knockout mice did not show significant differences in metastasis growth. Finally, we found a synergistic antimetastatic effect of the combination of IL1α blockade and STAT3 inhibition with stattic, highlighting a new immunotherapy approach to preventing melanoma metastasis.

Apyrase-mediated amplification of secretory IgA promotes intestinal homeostasis.

Secretory immunoglobulin A (SIgA) interaction with commensal bacteria conditions microbiota composition and function. However, mechanisms regulating reciprocal control of microbiota and SIgA are not defined. Bacteria-derived adenosine triphosphate (ATP) limits T follicular helper (Tfh) cells in the Peyer's patches (PPs) via P2X7 receptor (P2X7R) and thereby SIgA generation. Here we show that hydrolysis of extracellular ATP (eATP) by apyrase results in amplification of the SIgA repertoire. The enhanced breadth of SIgA in mice colonized with apyrase-releasing Escherichia coli influences topographical distribution of bacteria and expression of genes involved in metabolic versus immune functions in the intestinal epithelium. SIgA-mediated conditioning of bacteria and enterocyte function is reflected by differences in nutrient absorption in mice colonized with apyrase-expressing bacteria. Apyrase-induced SIgA improves intestinal homeostasis and attenuates barrier impairment and susceptibility to infection by enteric pathogens in antibiotic-induced dysbiosis. Therefore, amplification of SIgA by apyrase can be leveraged to restore intestinal fitness in dysbiotic conditions.

Protein Expression of AEBP1, MCM4, and FABP4 Differentiate Osteogenic, Adipogenic, and Mesenchymal Stromal Stem Cells.

Mesenchymal stem cells (MSCs) gain an increasing focus in the field of regenerative medicine due to their differentiation abilities into chondrocytes, adipocytes, and osteoblastic cells. However, it is apparent that the transformation processes are extremely complex and cause cellular heterogeneity. The study aimed to characterize differences between MSCs and cells after adipogenic (AD) or osteoblastic (OB) differentiation at the proteome level. Comparative proteomic profiling was performed using tandem mass spectrometry in data-independent acquisition mode. Proteins were quantified by deep neural networks in library-free mode and correlated to the Molecular Signature Database (MSigDB) hallmark gene set collections for functional annotation. We analyzed 4108 proteins across all samples, which revealed a distinct clustering between MSCs and cell differentiation states. Protein expression profiling identified activation of the Peroxisome proliferator-activated receptors (PPARs) signaling pathway after AD. In addition, two distinct protein marker panels could be defined for osteoblastic and adipocytic cell lineages. Hereby, overexpression of AEBP1 and MCM4 for OB as well as of FABP4 for AD was detected as the most promising molecular markers. Combination of deep neural network and machine-learning algorithms with data-independent mass spectrometry distinguish MSCs and cell lineages after adipogenic or osteoblastic differentiation. We identified specific proteins as the molecular basis for bone formation, which could be used for regenerative medicine in the future.

NLRC5 promotes transcription of BTN3A1-3 genes and Vγ9Vδ2 T cell-mediated killing.

BTN3A molecules-BTN3A1 in particular-emerged as important mediators of Vγ9Vδ2 T cell activation by phosphoantigens. These metabolites can originate from infections, e.g. with Mycobacterium tuberculosis, or by alterations in cellular metabolism. Despite the growing interest in the BTN3A genes and their high expression in immune cells and various cancers, little is known about their transcriptional regulation. Here we show that these genes are induced by NLRC5, a regulator of MHC class I gene transcription, through an atypical regulatory motif found in their promoters. Accordingly, a robust correlation between NLRC5 and BTN3A gene expression was found in healthy, in M. tuberculosis-infected donors' blood cells, and in primary tumors. Moreover, forcing NLRC5 expression promoted Vγ9Vδ2 T-cell-mediated killing of tumor cells in a BTN3A-dependent manner. Altogether, these findings indicate that NLRC5 regulates the expression of BTN3A genes and hence open opportunities to modulate antimicrobial and anticancer immunity.

Role of ETS1 in the Transcriptional Network of Diffuse Large B Cell Lymphoma of the Activated B Cell-Like Type.

Diffuse large B cell lymphoma (DLBCL) is a heterogenous disease that has been distinguished into at least two major molecular entities, the germinal center-like B cell (GCB) DLBCL and activated-like B cell (ABC) DLBCL, based on transcriptome expression profiling. A recurrent ch11q24.3 gain is observed in roughly a fourth of DLBCL cases resulting in the overexpression of two ETS transcription factor family members, ETS1 and FLI1. Here, we knocked down ETS1 expression by siRNA and analyzed expression changes integrating them with ChIP-seq data to identify genes directly regulated by ETS1. ETS1 silencing affected expression of genes involved in B cell signaling activation, B cell differentiation, cell cycle, and immune processes. Integration of RNA-Seq (RNA sequencing) data and ChIP-Seq (chromatin immunoprecipitation sequencing) identified 97 genes as bona fide, positively regulated direct targets of ETS1 in ABC-DLBCL. Among these was the Fc receptor for IgM, FCMR (also known as FAIM3 or Toso), which showed higher expression in ABC- than GCB-DLBCL clinical specimens. These findings show that ETS1 is contributing to the lymphomagenesis in a subset of DLBCL and identifies FCMR as a novel target of ETS1, predominantly expressed in ABC-DLBCL.

Copanlisib synergizes with conventional and targeted agents including venetoclax in B- and T-cell lymphoma models.

Copanlisib is a pan-class I phosphoinositide 3-kinase (PI3K) inhibitor with preferred activity toward PI3Kα and PI3Kδ. Despite the clear overall clinical benefit, the number of patients achieving complete remissions with the single agent is relatively low, a problem shared by the vast majority of targeted agents. Here, we searched for novel copanlisib-based combinations. Copanlisib was tested as a single agent, in combination with an additional 17 drugs in 26 cell lines derived from mantle cell lymphoma (MCL), marginal zone lymphoma (MZL), and T-cell lymphomas. In vivo experiments, transcriptome analyses, and immunoblotting experiments were also performed. Copanlisib as a single agent showed in vitro dose-dependent antitumor activity in the vast majority of the models. Combination screening identified several compounds that synergized with copanlisib. The strongest combination was with the B-cell lymphoma 2 (BCL2) inhibitor venetoclax. The benefit of the combination over single agents was also validated in an MZL xenograft model and in MCL primary cells, and was due to increased induction of apoptosis, an effect likely sustained by the reduction of the antiapoptotic proteins myeloid cell leukemia 1 (MCL1) and BCL-XL, observed in MCL and MZL cell lines, respectively. These data supported the rationale for the design of the Swiss Group for Clinical Cancer Research (SAKK) 66/18 phase 1 study currently exploring the combination of copanlisib and venetoclax in relapsed/refractory lymphomas.

KMT2D mutations and TP53 disruptions are poor prognostic biomarkers in mantle cell lymphoma receiving high-dose therapy: a FIL study.

In recent years, the outcome of mantle cell lymphoma (MCL) has improved, especially in younger patients, receiving cytarabine-containing chemoimmunotherapy and autologous stem cell transplantation. Nevertheless, a proportion of MCL patients still experience early failure. To identify biomarkers anticipating failure of intensive chemotherapy in MCL, we performed target resequencing and DNA profiling of purified tumor samples collected from patients enrolled in the prospective FIL-MCL0208 phase 3 trial (high-dose chemoimmunotherapy followed by autologous transplantation and randomized lenalidomide maintenance). Mutations of KMT2D and disruption of TP53 by deletion or mutation associated with an increased risk of progression and death, both in univariate and multivariate analysis. By adding KMT2D mutations and TP53 disruption to the MIPI-c backbone, we derived a new prognostic index, the "MIPI-genetic" ("MIPI- g"). The "MIPI-g" improved the model discrimination ability compared to the MIPI-c alone, defining three risk groups: i) low-risk patients (4-year progression free survival and overall survival of 72.0% and 94.5%); ii) inter-mediate-risk patients (4-year progression free survival and overall survival of 42.2% and 65.8%) and iii) high-risk patients (4-year progression free survival and overall survival of 11.5% and 44.9%). Our results: i) confirm that TP53 disruption identifies a high-risk population characterized by poor sensitivity to conventional or intensified chemotherapy; ii) provide the pivotal evidence that patients harboring KMT2D mutations share the same poor outcome as patients harboring TP53 disruption; and iii) allow to develop a tool for the identification of high-risk MCL patients for whom novel therapeutic strategies need to be investigated. (Trial registered at clinicaltrials.gov identifier: NCT02354313).

The Novel TORC1/2 Kinase Inhibitor PQR620 Has Anti-Tumor Activity in Lymphomas as a Single Agent and in Combination with Venetoclax.

The phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) signaling cascade is an important therapeutic target for lymphomas. Rapamycin-derivates as allosteric mTOR complex 1 (TORC1) inhibitors have shown moderate preclinical and clinical anti-lymphoma activity. Here, we assessed the anti-tumor activity of PQR620, a novel brain penetrant dual TORC1/2 inhibitor, in 56 lymphoma cell lines. We observed anti-tumor activity across 56 lymphoma models with a median IC50 value of 250 nM after 72 h of exposure. PQR620 was largely cytostatic, but the combination with the BCL2 inhibitor venetoclax led to cytotoxicity. Both the single agent and the combination data were validated in xenograft models. The data support further evaluation of PQR620 as a single agent or in combination with venetoclax.

The ETS Inhibitors YK-4-279 and TK-216 Are Novel Antilymphoma Agents.

PURPOSE: Transcription factors are commonly deregulated in cancer, and they have been widely considered as difficult to target due to their nonenzymatic mechanism of action. Altered expression levels of members of the ETS-transcription factors are often observed in many different tumors, including lymphomas. Here, we characterized two small molecules, YK-4-279 and its clinical derivative, TK-216, targeting ETS factors via blocking the protein-protein interaction with RNA helicases, for their antilymphoma activity. EXPERIMENTAL DESIGN: The study included preclinical in vitro activity screening on a large panel of cell lines, both as single agent and in combination; validation experiments on in vivo models; and transcriptome and coimmunoprecipitation experiments. RESULTS: YK-4-279 and TK-216 demonstrated an antitumor activity across several lymphoma cell lines, which we validated in vivo. We observed synergistic activity when YK-4-279 and TK-216 were combined with the BCL2 inhibitor venetoclax and with the immunomodulatory drug lenalidomide. YK-4-279 and TK-216 interfere with protein interactions of ETS family members SPIB, in activated B-cell-like type diffuse large B-cell lymphomas, and SPI1, in germinal center B-cell-type diffuse large B-cell lymphomas. CONCLUSIONS: The ETS inhibitor YK-4-279 and its clinical derivative TK-216 represent a new class of agents with in vitro and in vivo antitumor activity in lymphomas. Although their detailed mechanism of action needs to be fully defined, in DLBCL they might act by targeting subtype-specific essential transcription factors.

Unraveling transformation of follicular lymphoma to diffuse large B-cell lymphoma.

Follicular lymphoma (FL) is an indolent but largely incurable disease. Some patients suffer histological transformation to a more aggressive subtype with poorer prognosis. This study aimed to improve our understanding of the genetics underlying FL histological transformation, and to identify genetic drivers or promoters of the transformation by elucidating the differences between FL samples from patients who did and did not transform. We conducted targeted massive parallel sequencing of 22 pre-transformed FL/transformed diffuse large B-cell lymphoma pairs and 20 diagnostic samples from non-transformed FL patients. Additionally, 22 matched samples from 11 transformed FL patients (pre-transformed FL and diffuse large B-cell lymphoma) and 9 non-transformed FLs were studied for copy number variation using SNP arrays. We identified recurrently mutated genes that were enriched at transformation, most notably LRP1B, GNA13 and POU2AF1, which have roles in B-cell differentiation, GC architecture and migration. Mutations in POU2AF1 might be associated with lower levels of expression, were more frequent in transformed FLs, and seemed to be specific to transformed- compared with de novo-diffuse large B-cell lymphomas. Pre-transformed FLs carried more mutations per sample and had greater subclonal heterogeneity than non-transformed FLs. Finally, we identified four mutated genes in FL samples that differed between patients who did and did not transform: NOTCH2, DTX1, UBE2A and HIST1H1E. The presence of mutations in these genes was associated with shorter time to transformation when mutated in the FL biopsies. This information might be useful for identifying patients at higher risk of transformation.

Genome-wide promoter methylation of hairy cell leukemia.

Classic hairy cell leukemia (HCL) is a tumor of mature clonal B cells with unique genetic, morphologic, and phenotypic features. DNA methylation profiling has provided a new tier of investigation to gain insight into the origin and behavior of B-cell malignancies; however, the methylation profile of HCL has not been specifically investigated. DNA methylation profiling was analyzed with the Infinium HumanMethylation27 array in 41 mature B-cell tumors, including 11 HCL, 7 splenic marginal zone lymphomas (SMZLs), and chronic lymphocytic leukemia with an unmutated (n = 7) or mutated (n = 6) immunoglobulin gene heavy chain variable (IGHV) region or using IGHV3-21 (n = 10). Methylation profiles of nontumor B-cell subsets and gene expression profiling data were obtained from public databases. HCL had a methylation signature distinct from each B-cell tumor entity, including the closest entity, SMZL. Comparison with normal B-cell subsets revealed the strongest similarity with postgerminal center (GC) B cells and a clear separation from pre-GC and GC cellular programs. Comparison of the integrated analysis with post-GC B cells revealed significant hypomethylation and overexpression of BCR-TLR-NF-κB and BRAF-MAPK signaling pathways and cell adhesion, as well as hypermethylation and underexpression of cell-differentiation markers and methylated genes in cancer, suggesting regulation of the transformed hairy cells through specific components of the B-cell receptor and the BRAF signaling pathways. Our data identify a specific methylation profile of HCL, which may help to distinguish it from other mature B-cell tumors.

BET bromodomain inhibitor birabresib in mantle cell lymphoma: in vivo activity and identification of novel combinations to overcome adaptive resistance.

BACKGROUND: The outcome of patients affected by mantle cell lymphoma (MCL) has improved in recent years, but there is still a need for novel treatment strategies for these patients. Human cancers, including MCL, present recurrent alterations in genes that encode transcription machinery proteins and of proteins involved in regulating chromatin structure, providing the rationale to pharmacologically target epigenetic proteins. The Bromodomain and Extra Terminal domain (BET) family proteins act as transcriptional regulators of key signalling pathways including those sustaining cell viability. Birabresib (MK-8628/OTX015) has shown antitumour activity in different preclinical models and has been the first BET inhibitor to successfully undergo early clinical trials. MATERIALS AND METHODS: The activity of birabresib as a single agent and in combination, as well as its mechanism of action was studied in MCL cell lines. RESULTS: Birabresib showed in vitro and in vivo activities, which appeared mediated via downregulation of MYC targets, cell cycle and NFKB pathway genes and were independent of direct downregulation of CCND1. Additionally, the combination of birabresib with other targeted agents (especially pomalidomide, or inhibitors of BTK, mTOR and ATR) was beneficial in MCL cell lines. CONCLUSION: Our data provide the rationale to evaluate birabresib in patients affected by MCL.

Bromodomain and extra-terminal domain inhibition modulates the expression of pathologically relevant microRNAs in diffuse large B-cell lymphoma.

Aberrant changes in microRNA expression contribute to lymphomagenesis. Bromodomain and extra-terminal domain inhibitors such as OTX015 (MK-8628, birabresib) have demonstrated preclinical and clinical activity in hematologic tumors. MicroRNA profiling of diffuse large B-cell lymphoma cells treated with OTX015 revealed changes in the expression levels of a limited number of microRNAs, including miR-92a-1-5p, miR-21-3p, miR-155-5p and miR-96-5p. Analysis of publicly available chromatin immunoprecipitation sequencing data of diffuse large B-cell lymphoma cells treated with bromodomain and extra-terminal domain (BET) inhibitors showed that the BET family member BRD4 bound to the upstream regulatory regions of multiple microRNA genes and that this binding decreased following BET inhibition. Alignment of our microRNA profiling data with the BRD4 chromatin immunoprecipitation sequencing data revealed that microRNAs downregulated by OTX015 also exhibited reduced BRD4 binding in their promoter regions following treatment with another bromodomain and extra-terminal domain inhibitor, JQ1, indicating that BRD4 contributes directly to microRNA expression in lymphoma. Treatment with bromodomain and extra-terminal domain inhibitors also decreased the expression of the arginine methyltransferase PRMT5, which plays a crucial role in B-cell transformation and negatively modulates the transcription of miR-96-5p. The data presented here indicate that in addition to previously observed effects on the expression of coding genes, bromodomain and extra-terminal domain inhibitors also modulate the expression of microRNAs involved in lymphomagenesis.

Population-based outcome analysis of diffuse large B-cell lymphoma in people living with HIV infection and competent individuals.

The prognostic factors and outcome of 58 acquired immunodeficiency syndrome-related diffuse large B-cell lymphoma (AR-DLBCL) patients from the Swiss HIV Cohort Study, diagnosed from 2004 to 2011, were compared with those of 326 immunocompetent (IC)-DLBCL from the Hematology Division of the Amedeo Avogadro University (Italy) and the Oncology Institute of Southern Switzerland. Median follow-up was 6 years; 5-year overall survival (OS) was 68% (95% CI: 63%-73%) in IC-DLBCL and 63% (95% CI: 49%-75%) in AR-DLBCL (P = .220). The acquired immunodeficiency syndrome-related lymphoma international prognostic index predicted OS in AR-DLBCL. Among 148 patients younger than 61 years (40 AR-DLBCL and 108 IC-DLBCL) treated with RCHOP/RCHOP-like regimens, 20 IC-DLBCL and 9 AR-DLBCL patients died and OS was not significantly different. A higher proportion of early deaths occurred in the AR-DLBCL: indeed, 1-year OS was 94% (95% CI: 87%-97%) in IC-DLBCL and 82% (95% CI: 66%-91%) in AR-DLBCL patients. After rituximab and active antiretroviral therapy introduction, AR-DLBCL and IC-DLBCL patients treated with curative intent have similar long-term survival.

PQR309 Is a Novel Dual PI3K/mTOR Inhibitor with Preclinical Antitumor Activity in Lymphomas as a Single Agent and in Combination Therapy.

Purpose: Activation of the PI3K/mTOR signaling pathway is recurrent in different lymphoma types, and pharmacologic inhibition of the PI3K/mTOR pathway has shown activity in lymphoma patients. Here, we extensively characterized the in vitro and in vivo activity and the mechanism of action of PQR309 (bimiralisib), a novel oral selective dual PI3K/mTOR inhibitor under clinical evaluation, in preclinical lymphoma models.Experimental Design: This study included preclinical in vitro activity screening on a large panel of cell lines, both as single agent and in combination, validation experiments on in vivo models and primary cells, proteomics and gene-expression profiling, and comparison with other signaling inhibitors.Results: PQR309 had in vitro antilymphoma activity as single agent and in combination with venetoclax, panobinostat, ibrutinib, lenalidomide, ARV-825, marizomib, and rituximab. Sensitivity to PQR309 was associated with specific baseline gene-expression features, such as high expression of transcripts coding for the BCR pathway. Combining proteomics and RNA profiling, we identified the different contribution of PQR309-induced protein phosphorylation and gene expression changes to the drug mechanism of action. Gene-expression signatures induced by PQR309 and by other signaling inhibitors largely overlapped. PQR309 showed activity in cells with primary or secondary resistance to idelalisib.Conclusions: On the basis of these results, PQR309 appeared as a novel and promising compound that is worth developing in the lymphoma setting. Clin Cancer Res; 24(1); 120-9. ©2017 AACR.