Targeting CD25+ lymphoma cells with the antibody-drug conjugate camidanlumab tesirine as a single agent or in combination with targeted agents.

Camidanlumab tesirine (ADCT-301) is a CD25-specific antibody-drug conjugate (ADC) employing SG3199, a highly cytotoxic DNA minor groove cross-linking pyrrolobenzodiazepine dimer. The ADC has shown early clinical antitumour activity in various cancers, including B- and T-cell lymphomas. We assessed its preclinical activity as a single agent in 57 lymphoma cell lines and in combination with selected drugs in T-cell lymphoma-derived cell lines. Cells were exposed to increasing concentrations of the ADC or SG3199 for 96 h, followed by an MTT proliferation assay. CD25 expression was measured at cell surface and RNA levels. Experiments with PDX-derived cell lines were used for validation studies. Camidanlumab tesirine presented more potent single agent in vitro cytotoxic activity in T- than B-cell lymphomas. In vitro activity was correlated with CD25 cell surface and RNA expression. In vitro activity was correlated with CD25 cell surface and RNA expression. When camidanlumab tesirine-containing combinations were evaluated in four T-cell lymphoma models, the most active partners were everolimus, copanlisib, venetoclax, vorinostat, and pralatrexate, followed by bortezomib, romidepsin, bendamustine, and 5-azacytidine. The strong camidanlumab tesirine single-agent anti-lymphoma activity and the in vitro synergisms with targeted agents identify potential combination partners for future clinical studies.

Targeting CD19-positive lymphomas with the antibodydrug conjugate loncastuximab tesirine: preclinical evidence of activity as a single agent and in combination therapy.

Antibody-drug conjugates (ADC) represent one of the most successful therapeutic approaches introduced into clinical practice in the last few years. Loncastuximab tesirine (ADCT-402) is a CD19-targeting ADC in which the antibody is conjugated through a protease cleavable dipeptide linker to a pyrrolobenzodiazepine dimer warhead (SG3199). Based on the results of a phase II study, loncastuximab tesirine was recently approved for adult patients with relapsed/refractory large B-cell lymphoma. We assessed the activity of loncastuximab tesirine using in vitro and in vivo models of lymphomas, correlated its activity with levels of CD19 expression, and identified combination partners providing synergy with the ADC. Loncastuximab tesirine was tested across 60 lymphoma cell lines. It had strong cytotoxic activity in B-cell lymphoma cell lines. The in vitro activity was correlated with the level of CD19 expression and intrinsic sensitivity of cell lines to the ADC's warhead. Loncastuximab tesirine was more potent than other anti-CD19 ADC (coltuximab ravtansine, huB4-DGN462), although the pattern of activity across cell lines was correlated. The activity of loncastuximab tesirine was also largely correlated with cell line sensitivity to R-CHOP. Combinatorial in vitro and in vivo experiments identified the benefit of adding loncastuximab tesirine to other agents, especially BCL2 and PI3K inhibitors. Our data support the further development of loncastuximab tesirine for use as a single agent and in combination for patients affected by mature B-cell neoplasms. The results also highlight the importance of CD19 expression and the existence of lymphoma populations characterized by resistance to multiple therapies.

Characterization of GECPAR, a noncoding RNA that regulates the transcriptional program of diffuse large B-cell lymphoma.

Enhancers are regulatory regions of DNA, which play a key role in cell-type specific differentiation and development. Most active enhancers are transcribed into enhancer RNA (eRNA) that can regulate transcription of target genes by means of in cis as well as in trans action. eRNA stabilize contacts between distal genomic regions and mediate the interaction of DNA with master transcription factors. Here, we characterized an enhancer eRNA, GECPAR (germinal center proliferative adapter RNA), which is specifically transcribed in normal and neoplastic germinal center B cells from the super-enhancer of POU2AF1, a key regulatory gene of the germinal center reaction. Using diffuse large B-cell lymphoma cell line models, we demonstrated the tumor suppressor activity of GECPAR, which is mediated via its transcriptional regulation of proliferation and differentiation genes, particularly MYC and the Wnt pathway.

In Vivo and Ex Vivo Patient-Derived Tumor Xenograft Models of Lymphoma for Drug Discovery.

In the hemato-oncology field, remarkable scientific progress has been achieved, primarily propelled by the discovery of new technologies, improvement in genomics, and novel in vitro and in vivo models. The establishment of multiple cell line collections and the development of instrumental mouse models enhanced our ability to discover effective therapeutics. However, cancer models that faithfully mimic individual cancers are still imperfect. Patient-derived tumor xenografts (PDTXs) have emerged as a powerful tool for identifying the mechanisms which drive tumorigenesis and for testing potential therapeutic interventions. The recognition that PDTXs can maintain many of the donor samples' properties enabled the development of new strategies for discovering and implementing therapies. Described in this article are protocols for the generation and characterization of lymphoma PDTXs that may be used as the basis of shared procedures. Universal protocols will foster the model utilization, enable the integration of public and private repositories, and aid in the development of shared platforms. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Tissue handling and cryopreservation of primary and PDTX samples Basic Protocol 2: Performing tumor implant in immunocompromised mice PDTX models Alternate Protocol 1: Intra-medullary femoral injection Alternate Protocol 2: Intravenous injection Alternate Protocol 3: Intraperitoneal injection Support Protocol 1: Phenotypical characterization of PDTXs by flow cytometry Support Protocol 2: Biological and molecular characterization of PDTX tumors by PCR detection of IGK, IGH, and TCR rearrangements Basic Protocol 3: Harvesting PDTX-derived tumor cells for ex vivo experiments Basic Protocol 4: In vivo testing of multiple compounds in a PDTX mouse model.

Clinical and Biological Subtypes of B-cell Lymphoma Revealed by Microenvironmental Signatures.

Diffuse large B-cell lymphoma (DLBCL) is a biologically and clinically heterogeneous disease. Transcriptomic and genetic characterization of DLBCL has increased the understanding of its intrinsic pathogenesis and provided potential therapeutic targets. However, the role of the microenvironment in DLBCL biology remains less understood. Here, we performed a transcriptomic analysis of the microenvironment of 4,655 DLBCLs from multiple independent cohorts and described four major lymphoma microenvironment categories that associate with distinct biological aberrations and clinical behavior. We also found evidence of genetic and epigenetic mechanisms deployed by cancer cells to evade microenvironmental constraints of lymphoma growth, supporting the rationale for implementing DNA hypomethylating agents in selected patients with DLBCL. In addition, our work uncovered new therapeutic vulnerabilities in the biochemical composition of the extracellular matrix that were exploited to decrease DLBCL proliferation in preclinical models. This novel classification provides a road map for the biological characterization and therapeutic exploitation of the DLBCL microenvironment. SIGNIFICANCE: In a translational relevant transcriptomic-based classification, we characterized the microenvironment as a critical component of the B-cell lymphoma biology and associated it with the DLBCL clinical behavior establishing a novel opportunity for targeting therapies.This article is highlighted in the In This Issue feature, p. 1307.

Metabolic and Immune Markers for Precise Monitoring of COVID-19 Severity and Treatment.

Deep understanding of the SARS-CoV-2 effects on host molecular pathways is paramount for the discovery of early biomarkers of outcome of coronavirus disease 2019 (COVID-19) and the identification of novel therapeutic targets. In that light, we generated metabolomic data from COVID-19 patient blood using high-throughput targeted nuclear magnetic resonance (NMR) spectroscopy and high-dimensional flow cytometry. We find considerable changes in serum metabolome composition of COVID-19 patients associated with disease severity, and response to tocilizumab treatment. We built a clinically annotated, biologically-interpretable space for precise time-resolved disease monitoring and characterize the temporal dynamics of metabolomic change along the clinical course of COVID-19 patients and in response to therapy. Finally, we leverage joint immuno-metabolic measurements to provide a novel approach for patient stratification and early prediction of severe disease. Our results show that high-dimensional metabolomic and joint immune-metabolic readouts provide rich information content for elucidation of the host's response to infection and empower discovery of novel metabolic-driven therapies, as well as precise and efficient clinical action.

Profiling of immune dysfunction in COVID-19 patients allows early prediction of disease progression.

With a rising incidence of COVID-19-associated morbidity and mortality worldwide, it is critical to elucidate the innate and adaptive immune responses that drive disease severity. We performed longitudinal immune profiling of peripheral blood mononuclear cells from 45 patients and healthy donors. We observed a dynamic immune landscape of innate and adaptive immune cells in disease progression and absolute changes of lymphocyte and myeloid cells in severe versus mild cases or healthy controls. Intubation and death were coupled with selected natural killer cell KIR receptor usage and IgM+ B cells and associated with profound CD4 and CD8 T-cell exhaustion. Pseudo-temporal reconstruction of the hierarchy of disease progression revealed dynamic time changes in the global population recapitulating individual patients and the development of an eight-marker classifier of disease severity. Estimating the effect of clinical progression on the immune response and early assessment of disease progression risks may allow implementation of tailored therapies.