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Abstract: We report a first-in-human clinical trial using chimeric antigen receptor (CAR) T cells targeting CD37, an antigen highly expressed in B- and T-cell malignancies. Five patients with relapsed or refractory CD37+ lymphoid malignancies were enrolled and infused with autologous CAR-37 T cells. CAR-37 T cells expanded in the peripheral blood of all patients and, at peak, comprised >94% of the total lymphocytes in 4 of 5 patients. Tumor responses were observed in 4 of 5 patients with 3 complete responses, 1 mixed response, and 1 patient whose disease progressed rapidly and with relative loss of CD37 expression. Three patients experienced prolonged and severe pancytopenia, and in 2 of these patients, efforts to ablate CAR-37 T cells, which were engineered to coexpress truncated epidermal growth factor receptor, with cetuximab were unsuccessful. Hematopoiesis was restored in these 2 patients after allogeneic hematopoietic stem cell transplantation. No other severe, nonhematopoietic toxicities occurred. We investigated the mechanisms of profound pancytopenia and did not observe activation of CAR-37 T cells in response to hematopoietic stem cells in vitro or hematotoxicity in humanized models. Patients with pancytopenia had sustained high levels of interleukin-18 (IL-18) with low levels of IL-18 binding protein in their peripheral blood. IL-18 levels were significantly higher in CAR-37-treated patients than in both cytopenic and noncytopenic cohorts of CAR-19-treated patients. In conclusion, CAR-37 T cells exhibited antitumor activity, with significant CAR expansion and cytokine production. CAR-37 T cells may be an effective therapy in hematologic malignancies as a bridge to hematopoietic stem cell transplant. This trial was registered at www.ClinicalTrials.gov as #NCT04136275., (© 2024 American Society of Hematology. Published by Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

Despite the clear indications and worldwide application of specific guidelines, the recognition of Infective Endocarditis (IE) may be challenging in day-to-day clinical practice. Significant changes in the epidemiological and clinical profile of IE have been observed, including variations in the populations at risk and an increased incidence in subjects without at-risk cardiac disease. Emergent at-risk populations for IE particularly include immunocompromised patients with a comorbidity burden (e.g., cancer, diabetes, dialysis), requiring long-term central venous catheters or recurrent healthcare interventions. In addition, healthy subjects, such as skin-contact athletes or those with piercing implants, may be exposed to the transmission of highly virulent bacteria (through the skin or mucous), determining endothelial lesions and subsequent IE, despite the absence of pre-existing at-risk cardiac disease. Emergent at-risk populations and clinical presentation changes may subvert the conventional paradigm of IE toward an unexpected clinical scenario. Owing to its unusual clinical context, IE might be overlooked, resulting in a challenging diagnosis and delayed treatment. This review, supported by a series of clinical cases, analyzed the subtle and deceptive phenotypes subtending the complex syndrome of unexpected IE. The awareness of an unexpected clinical course should alert clinicians to also consider IE diagnosis in patients with atypical features, enhancing vigilance for preventive measures in an emergent at-risk population untargeted by conventional workflows., Competing Interests: The authors declare no conflicts of interest.

Purpose: Targeting solid tumors with chimeric antigen receptor (CAR) T cells remains challenging due to heterogenous target antigen expression, antigen escape, and the immunosuppressive tumor microenvironment (TME). Pancreatic cancer is characterized by a thick stroma generated by cancer-associated fibroblasts (CAF), which may contribute to the limited efficacy of mesothelin-directed CAR T cells in early-phase clinical trials. To provide a more favorable TME for CAR T cells to target pancreatic ductal adenocarcinoma (PDAC), we generated T cells with an antimesothelin CAR and a secreted T-cell-engaging molecule (TEAM) that targets CAF through fibroblast activation protein (FAP) and engages T cells through CD3 (termed mesoFAP CAR-TEAM cells)., Experimental Design: Using a suite of in vitro, in vivo, and ex vivo patient-derived models containing cancer cells and CAF, we examined the ability of mesoFAP CAR-TEAM cells to target PDAC cells and CAF within the TME. We developed and used patient-derived ex vivo models, including patient-derived organoids with patient-matched CAF and patient-derived organotypic tumor spheroids., Results: We demonstrated specific and significant binding of the TEAM to its respective antigens (CD3 and FAP) when released from mesothelin-targeting CAR T cells, leading to T-cell activation and cytotoxicity of the target cell. MesoFAP CAR-TEAM cells were superior in eliminating PDAC and CAF compared with T cells engineered to target either antigen alone in our ex vivo patient-derived models and in mouse models of PDAC with primary or metastatic liver tumors., Conclusions: CAR-TEAM cells enable modification of tumor stroma, leading to increased elimination of PDAC tumors. This approach represents a promising treatment option for pancreatic cancer., (©2024 American Association for Cancer Research.)

Infective endocarditis (IE) is still a life-threatening disease with frequent lethal outcomes despite the profound changes in its clinical, microbiological, imaging, and therapeutic profiles. Nowadays, the scenario for IE has changed since rheumatic fever has declined, but on the other hand, multiple aspects, such as elderly populations, cardiovascular device implantation procedures, and better use of multiple imaging modalities and multidisciplinary care, have increased, leading to escalations in diagnosis. Since the ESC and AHA Guidelines have been released, specific aspects of diagnostic and therapeutic management have been clarified to provide better and faster diagnosis and prognosis. Surgical treatment is required in approximately half of patients with IE in order to avoid progressive heart failure, irreversible structural damage in the case of uncontrolled infection, and the prevention of embolism. The timing of surgery has been one of the main aspects discussed, identifying cases in which surgery needs to be performed on an emergency (within 24 h) or urgent (within 7 days) basis, irrespective of the duration of antibiotic treatment, or cases where surgery can be postponed to allow a brief period of antibiotic treatment under careful clinical and echocardiographic observation. Mainly, guidelines put emphasis on the importance of an endocarditis team in the handling of systemic complications and how they affect the timing of surgery and perioperative management. Neurological complications, acute renal failure, splenic or musculoskeletal manifestations, or infections determined by multiresistant microorganisms or fungi can affect long-term prognosis and survival. Not to be outdone, anatomical and surgical factors, such as the presence of native or prosthetic valve endocarditis, a repair strategy when feasible, anatomical extension and disruption in the case of an annular abscess (mitral valve annulus, aortic mitral curtain, aortic root, and annulus), and the choice of prosthesis and conduits, can be equally crucial. It can be hard for surgeons to maneuver between correct pre-operative planning and facing unexpected obstacles during intraoperative management. The aim of this review is to provide an overview and analysis of a broad spectrum of specific surgical scenarios and how their challenging management can be essential to ensure better outcomes and prognoses.

Nonbacterial thrombotic endocarditis (NBTE) is a form of endocarditis that occurs in patients with predisposing conditions, including malignancies, autoimmune diseases (particularly antiphospholipid antibody syndrome, which accounts for the majority of lupus-associated cases), and coagulation disturbances for which the correlation with classical determinants is unclear. The condition is commonly referred to as "marantic", "verrucous", or Libman-Sacks endocarditis, although these are not synonymous, representing clinical-pathological nuances. The clinical presentation of NBTE involves embolic events, while local valvular complications, generally regurgitation, are typically less frequent and milder compared to infective forms of endocarditis. In the past, the diagnosis of NBTE relied on post mortem examinations, while at present, the diagnosis is primarily based on echocardiography, with the priority of excluding infective endocarditis through comprehensive microbiological and serological tests. As in other forms of endocarditis, besides pathology, transesophageal echocardiography remains the diagnostic standard, while other imaging techniques hold promise as adjunctive tools for early diagnosis and differentiation from infective vegetations. These include cardiac MRI and 18 FDG-PET/CT, which already represents a major diagnostic criterion of infective endocarditis in specific settings. We will herein provide a comprehensive review of the current knowledge on the clinics and therapeutics of NBTE, with a specific focus on the diagnostic tools.

Background: Chimeric antigen receptor (CAR) T cells have achieved remarkable responses in patients with hematological malignancies; however, the potential of this therapeutic platform for solid tumors like glioblastoma (GBM) has been limited, due in large part to the targeting of single antigens in a heterogeneous disease. Strategies that allow CAR T cells to engage multiple antigens concomitantly may broaden therapeutic responses and mitigate the effects of immune escape., Methods: Here we have developed a novel, dual-specific, tandem CAR T (TanCART) cell with the ability to simultaneously target both EGFRvIII and IL-13Rα2, two well-characterized tumor antigens that are frequently found on the surface of GBM cells but completely absent from normal brain tissues. We employed both standard immunological assays and multiple orthotopic preclinical models including patient-derived xenograft to demonstrate efficacy of this approach against heterogeneous tumors., Results: Tandem CAR T cells displayed enhanced cytotoxicity in vitro against heterogeneous GBM populations, including patient-derived brain tumor cultures ( P < .05). Compared to CAR T cells targeting single antigens, dual antigen engagement through the tandem construct was necessary to achieve long-term, complete, and durable responses in orthotopic murine models of heterogeneous GBM, including patient-derived xenografts ( P < .05)., Conclusions: We demonstrate that TanCART is effective against heterogeneous tumors in the brain. These data lend further credence to the development of multi-specific CAR T cells in the treatment of GBM and other cancers., (© The Author(s) 2022. Published by Oxford University Press, the Society for Neuro-Oncology and the European Association of Neuro-Oncology.)

Chimeric antigen receptor (CAR) T cell therapy is effective in lymphoid malignancies, but there has been limited data in myeloid cancers. Here, we start with a CD27-based CAR to target CD70 ("native") in acute myeloid leukemia (AML), and we find modest efficacy in vivo, consistent with prior reports. We then use orthogonal approaches to increase binding on both the tumor and CAR-T cell sides of the immune synapse: a pharmacologic approach (azacitidine) to increase antigen density of CD70 in myeloid tumors, and an engineering approach to stabilize binding of the CAR to CD70. To accomplish the latter, we design a panel of hinge-modified regions to mitigate cleavage of the extracellular portion of CD27. Our CD8 hinge and transmembrane-modified CD70 CAR-T cells are less prone to cleavage, have enhanced binding avidity, and increased expansion, leading to more potent in vivo activity. This enhanced CD70-targeted CAR is a promising candidate for further clinical development., Competing Interests: Declaration of interests M.V.M. is an inventor on patents related to adoptive cell therapies, held by Massachusetts General Hospital (some licensed to Promab) and University of Pennsylvania (some licensed to Novartis). M.V.M. holds equity in 2SeventyBio, Century Therapeutics, Genocea, Neximmune, Oncternal, and TCR2 and has served as a consultant for multiple companies involved in cell therapies. M.V.M. is on the Board of Directors of 2Seventy Bio. M.V.M. has grant/research support from CRISPR therapeutics, Kite Pharma, Servier, and Novartis. M.J.F. has served as a consultant for several cell therapy companies. No other authors report conflicts of interest. M.B.L. and M.V.M. have filed patents on the constructs described herein., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Background: Myxomatous mitral valve prolapse (MVP) and mitral-annular disjunction (Barlow disease) are at-risk for ventricular arrhythmias (VA). Fibrosis involving the papillary muscles and/or the infero-basal left ventricular (LV) wall was reported at autopsy in sudden cardiac death (SCD) patients with MVP., Objectives: We investigated the electrophysiological substrate subtending VA in MVP patients with Barlow disease phenotype., Methods: Twenty-three patients with VA were enrolled, including five with syncope and four with a history of SCD. Unipolar (Uni < 8.3 mV) and bipolar (Bi < 1.5 mV) low-voltage areas were analyzed with electro-anatomical mapping (EAM), and VA inducibility was evaluated with programmed ventricular stimulation (PES). Electrophysiological parameters were correlated with VA patterns, electrocardiogram (ECG) inferior negative T wave (nTW), and late gadolinium enhancement (LGE) assessed by cardiac magnetic resonance., Results: Premature ventricular complex (PVC) burden was 12 061.9 ± 12 994.6/24 h with a papillary-muscle type (PM-PVC) in 18 patients (68%). Twelve-lead ECG showed nTW in 12 patients (43.5%). A large Uni less than 8.3 mV area (62.4 ± 45.5 cm 2 ) was detected in the basal infero-lateral LV region in 12 (73%) patients, and in the papillary muscles (2.2 ± 2.9 cm 2 ) in 5 (30%) of 15 patients undergoing EAM. A concomitant Bi less than 1.5 mV area (5.0 ± 1.0 cm 2 ) was identified in two patients. A history of SCD, and the presence of nTW, and LGE were associated with a greater Uni less than 8.3 mV extension: (32.8 ± 3.1 cm 2 vs. 9.2 ± 8.7 cm 2 ), nTW (20.1 ± 11.0 vs. 4.1 ± 3.8 cm 2 ), and LGE (19.2 ± 11.7 cm 2 vs. 1.0 ± 2.0 cm 2 , p = .013), respectively. All patients with PM-PVC had a Uni less than 8.3 mV area. Sustained VA (ventricular tachycardia 2 and VF 2) were induced by PES only in four patients (one with resuscitated SCD)., Conclusions: Low unipolar low voltage areas can be identified with EAM in the basal inferolateral LV region and in the papillary muscles as a potential electrophysiological substrate for VA and SCD in patients with MVP and Barlow disease phenotype., (© 2021 Wiley Periodicals LLC.)

T cell immunotherapies have revolutionized treatment for a subset of cancers. Yet, a major hurdle has been the lack of facile and predicative preclinical animal models that permit dynamic visualization of T cell immune responses at single-cell resolution in vivo. Here, optically clear immunocompromised zebrafish were engrafted with fluorescent-labeled human cancers along with chimeric antigen receptor T (CAR T) cells, bispecific T cell engagers (BiTEs), and antibody peptide epitope conjugates (APECs), allowing real-time single-cell visualization of T cell-based immunotherapies in vivo. This work uncovered important differences in the kinetics of T cell infiltration, tumor cell engagement, and killing between these immunotherapies and established early endpoint analysis to predict therapy responses. We also established EGFR-targeted immunotherapies as a powerful approach to kill rhabdomyosarcoma muscle cancers, providing strong preclinical rationale for assessing a wider array of T cell immunotherapies in this disease., Competing Interests: Disclosures: D.G. Millar reported personal fees from Revitope Inc. during the conduct of the study and personal fees from Revitope Inc. outside the submitted work; in addition, D.G. Millar had a patent to WO2012123755A1 licensed "Revitope Inc." and owns stock in Revitope Inc. B.J. Drapkin reported personal fees from AstraZeneca outside the submitted work. M. Cobbold reported "other" from Revitope Oncology outside the submitted work; in addition, M. Cobbold had a patent to WO 2012/123755 issued "Revitope Oncology"; and is currently an employee of Astrazeneca. M.V. Maus is an inventor on patents related to adoptive cell therapies, held by Massachusetts General Hospital and University of Pennsylvania (some licensed to Novartis). M.V. Maus holds equity in TCR2, Century Therapeutics, and Neximmune, and has served as a consultant for multiple companies involved in cell therapies. In addition, M.V. Maus is a consultant for Adaptimmune, Agenus, Allogene, Arcellx, Astellas, AstraZeneca, Atara, Bayer, BMS, Cabaletta Bio (SAB), Cellectis (SAB), CRISPR therapeutics, In8bio (SAB), Innovakine, Intellia, GSK, Kite Pharma, Micromedicine, Neximmune, Novartis, TCR2 (SAB), Tmunity, Torque, and WindMIL (SAB); has grant/research support from CRISPR therapeutics, Kite Pharma, Servier, and Novartis; is a stockholder of Century Therapeutics, TCR2, and Ichnos; and is a member of board of directors for Ichnos Sciences. D.M. Langenau reported a patent pending on use of immune deficient zebrafish for xenograft transplantation studies. No other disclosures were reported., (© 2021 Yan et al.)

Background: Myocardial contrast two-dimensional echocardiography (MC-2DE) is widely used to address alcohol septal ablation (ASA) in obstructive hypertrophic cardiomyopathy (HCM). Owing to its limited cut-planes, MC-2DE may inaccurately identify the contrast misplacement associated with an unsuccessful or complicated ASA outcome., Objective: The aim of this study was to assess the added value of myocardial contrast three-dimensional echocardiography (MC-3DE) compared with MC-2DE to identify the appropriate matching between the target septal zone (TSZ) and coronary artery branch for safe and long-term effective ASA in HCM patients., Methods: A consecutive series of 52 symptomatic obstructive HCM patients referred for isolated surgical myectomy (SM) was analyzed with MC-2DE and MC-3DE following injection of echocontrast into one or more septal branches. MC-2DE and MC-3DE patterns were categorized according to complete ( Type 1 ) or incomplete ( Type 2 ) TSZ covering, high-risk ( Type 3 ) exceeding TSZ, or life-threatening outside TSZ distribution ( Type 4 )., Results: MC-2DE per patient analysis showed a Type 1 pattern in 32 patients and Types 2-4 in the remaining 20 patients; subsequent MC-3DE analysis provided a re-phenotyping of MC-2DE findings in 22 of the 52 patients (42%), showing a high-risk Type 2 pattern in 17 of the 32 patients with Type 1 , and a new life-threatening Type 4 in three patients with Type 2, respectively. All patients with MC-3DE Type 1 pattern underwent safe and effective ASA with a long-term uneventful follow-up, while the remaining patients underwent SM., Conclusions: Refining high risk or life-threatening contrast misplacement, MC-3DE is more accurate than conventional MC-2DE to target safe and long-term effective septal reduction with ASA in obstructive HCM patients referred for isolated SM.

Cell-based therapies are emerging as effective agents against cancer and other diseases. As autonomous "living drugs," these therapies lack precise control. Chimeric antigen receptor (CAR) T cells effectively target hematologic malignancies but can proliferate rapidly and cause toxicity. We developed ON and OFF switches for CAR T cells using the clinically approved drug lenalidomide, which mediates the proteasomal degradation of several target proteins by inducing interactions between the CRL4 CRBN E3 ubiquitin ligase and a C2H2 zinc finger degron motif. We performed a systematic screen to identify "super-degron" tags with enhanced sensitivity to lenalidomide-induced degradation and used these degradable tags to generate OFF-switch degradable CARs. To create an ON switch, we engineered a lenalidomide-inducible dimerization system and developed split CARs that required both lenalidomide and target antigen for activation. Subtherapeutic lenalidomide concentrations controlled the effector functions of ON- and OFF-switch CAR T cells. In vivo, ON-switch split CARs demonstrated lenalidomide-dependent antitumor activity, and OFF-switch degradable CARs were depleted by drug treatment to limit inflammatory cytokine production while retaining antitumor efficacy. Together, the data showed that these lenalidomide-gated switches are rapid, reversible, and clinically suitable systems to control transgene function in diverse gene- and cell-based therapies., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Background: Adoptive cell therapy with chimeric antigen receptor T cells (CAR-T) has become a standard treatment for patients with certain aggressive B cell malignancies and holds promise to improve the care of patients suffering from numerous other cancers in the future. However, the high manufacturing cost of CAR-T cell therapies poses a major barrier to their broader clinical application. Among the key cost drivers of CAR-T production are single-use reagents for T cell activation and clinical-grade viral vector. The presence of variable amounts of contaminating monocytes in the starting material poses an additional challenge to CAR-T manufacturing, since they can impede T cell stimulation and transduction, resulting in manufacturing failure., Methods: We created K562-based artificial antigen-presenting cells (aAPC) with genetically encoded T cell stimulation and costimulation that represent an inexhaustible source for T cell activation. We additionally disrupted endogenous expression of the low-density lipoprotein receptor (LDLR) on these aAPC (aAPC-ΔLDLR) using CRISPR-Cas9 gene editing nucleases to prevent inadvertent lentiviral transduction and avoid the sink effect on viral vector during transduction. Using various T cell sources, we produced CD19-directed CAR-T cells via aAPC-ΔLDLR-based activation and tested their in vitro and in vivo antitumor potency against B cell malignancies., Results: We found that lack of LDLR expression on our aAPC-ΔLDLR conferred resistance to lentiviral transduction during CAR-T production. Using aAPC-ΔLDLR, we achieved efficient expansion of CAR-T cells even from unpurified starting material like peripheral blood mononuclear cells or unmanipulated leukapheresis product, containing substantial proportions of monocytes. CD19-directed CAR-T cells that we produced via aAPC-ΔLDLR-based expansion demonstrated potent antitumor responses in preclinical models of acute lymphoblastic leukemia and B-cell lymphoma., Conclusions: Our aAPC-ΔLDLR represent an attractive approach for manufacturing of lentivirally transduced T cells that may be simpler and more cost efficient than currently available methods., Competing Interests: Competing interests: Some of the authors have filed provisional patent applications related to the use of the aAPC’s described herein. BPK is an inventor on various patents and patent applications that describe gene editing and epigenetic editing technologies. BPK is a consultant for Avectas and an advisor for Acrigen Biosciences. JKJ has financial interests in Beam Therapeutics, Editas Medicine, Excelsior Genomics, Pairwise Plants, Poseida Therapeutics, Transposagen Biopharmaceuticals, and Verve Therapeutics (f/k/a Endcadia). JKJ’s interests were reviewed and are managed by Massachusetts General Hospital and Partners HealthCare in accordance with their conflict of interest policies. JKJ is a member of the Board of Directors of the American Society of Gene and Cell Therapy. JKJ is a coinventor on various patents and patent applications that describe gene editing and epigenetic editing technologies., (© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Dynamic left ventricular (LV) obstruction has important clinical and therapeutic implications in patients with hypertrophic cardiomyopathy (HC). Although absent at rest, LV obstruction may be elicited using varying stressors. Meal-related hemodynamic changes may favor LV obstruction and support postprandial (PP) symptoms in HC patients. The aim of this study was to evaluate PP-LV obstruction inducibility in HC patients in comparison with fasting Valsalva maneuver and exercise test. Ninety-two HC patients without LV obstruction underwent at-rest Transthoracic Echocardiography (TTE) during Valsalva maneuver and exercise test under fasting condition followed by at-rest re-test PP-TTE 30 to 60 minutes after a standardized midday meal. LV obstruction was noted and classified as being related to systolic anterior motion (SAM) of the mitral valve (SAM-related) and/or non-SAM-related (mid-cavity or apical), and intraventricular gradient was measured. At-rest re-test PP-TTE showed significant intraventricular gradient (>30 mm Hg) in 68 patients (60 SAM-related, 8 non-SAM related, 30 combined) with a higher prevalence (74%) of HC phenotype re-classified as obstructive compared with the fasting Valsalva maneuver (23%) or exercise test (33%) (p < 0.001). At multivariate analysis, a clinical history of PP symptoms and mitral anterior leaflet length and/or LV outflow ratio >2 were independently correlated with PP-TTE obstruction. In conclusion, PP TTE re-test is a simple and effective approach to unmask latent LV dynamic obstruction in daily clinical practice over fasting Valsalva maneuver or exercise test. PP clinical phenotype refinement may be relevant in targeting and evaluating HC therapy., Competing Interests: Disclosures The authors have no conflicts of interest to disclose., (Copyright © 2020 Elsevier Inc. All rights reserved.)

CAR T cells with different costimulation domains have proven clinical efficacy in leukemia and lymphoma but have different kinetics of activation, antigen sensitivity, and susceptibility to exhaustion. Two recent studies identified that these functions are shaped by a balance among opposing signaling complexes and transcription factors competing for binding motifs., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Purpose: T cells engineered to express a chimeric antigen receptor (CAR) against CD19 have recently been FDA approved for the treatment of relapsed or refractory large B-cell lymphoma. Despite the success and curative potential of CD19 CAR T cells, several reports describing disease relapse due to antigen loss are now emerging., Experimental Design: We developed a novel CAR construct directed against CD79b, a critical receptor for successful B-cell development that remains highly expressed in several subtypes of B-cell lymphoma, including mantle cell lymphoma (MCL). We tested CAR T cells directed against CD79b alone or in combination with CD19 targeting in a single construct, against cell line- and patient-derived xenograft models., Results: We demonstrate CAR79b antigen-specific recognition and cytotoxicity against a panel of cell lines and patient-derived xenograft models of MCL. Importantly, we show that downregulation of CD19 does not influence surface expression of CD79b and that anti-CD79b CAR T cells alone or arranged in a dual-targeting format with a CD19 single-chain variable fragment (scFv) are able to recognize and eliminate CD19 + , CD19 - , and mixed CD19 + /CD19 - B-cell lymphoma., Conclusions: Our findings demonstrate that CAR T cells targeting CD79b alone or in combination have promise for treating and preventing CD19 antigen escape in B-cell lymphomas., (©2019 American Association for Cancer Research.)

Despite remarkable success in the treatment of hematological malignancies, CAR T-cell therapies for solid tumors have floundered, in large part due to local immune suppression and the effects of prolonged stimulation leading to T-cell dysfunction and exhaustion. One mechanism by which gliomas and other cancers can hamper CAR T cells is through surface expression of inhibitory ligands such as programmed cell death ligand 1 (PD-L1). Using the CRIPSR-Cas9 system, we created universal CAR T cells resistant to PD-1 inhibition through multiplexed gene disruption of endogenous T-cell receptor (TRAC), beta-2 microglobulin (B2M) and PD-1 (PDCD1). Triple gene-edited CAR T cells demonstrated enhanced activity in preclinical glioma models. Prolonged survival in mice bearing intracranial tumors was achieved after intracerebral, but not intravenous administration. CRISPR-Cas9 gene-editing not only provides a potential source of allogeneic, universal donor cells, but also enables simultaneous disruption of checkpoint signaling that otherwise impedes maximal antitumor functionality.

Chimeric antigen receptor (CAR) T cells (CARTs) have shown tremendous potential for the treatment of certain B-cell malignancies, including patients with relapsed/refractory multiple myeloma (MM). Targeting the B-cell maturation antigen (BCMA) has produced the most promising results for CART therapy of MM to date, but not all remissions are sustained. Emergence of BCMA escape variants has been reported under the selective pressure of monospecific anti-BCMA CART treatment. Thus, there is a clinical need for continuous improvement of CART therapies for MM. Here, we show that a novel trimeric APRIL (a proliferation-inducing ligand)-based CAR efficiently targets both BCMA+ and BCMA- MM. Modeled after the natural ligand-receptor pair, APRIL-based CARs allow for bispecific targeting of the MM-associated antigens BCMA and transmembrane activator and CAML interactor (TACI). However, natural ligands as CAR antigen-binding domains may require further engineering to promote optimal binding and multimerization to adequately trigger T-cell activation. We found that using a trimeric rather than a monomeric APRIL format as the antigen-binding domain enhanced binding to BCMA and TACI and CART activity against MM in vitro and in vivo. Dual-specific, trimeric APRIL-based CAR are a promising therapeutic approach for MM with potential for preventing and treating BCMA escape., (© 2019 by The American Society of Hematology.)

Cancer immunotherapy based on the engineering of chimeric antigen receptors (CAR) on T cells has emerged as one of the most promising new therapies for patients with B-cell malignancies. Preclinical assessments of essential CAR T cell functions such as trafficking and cytotoxicity are critical for accelerating the development of highly effective therapeutic candidates. However, current tools for evaluating CAR-T functions lack sufficient precision. Here, a micropatterned tumor array (MiTA) is described that enables detailed and dynamic characterization of CAR T cell trafficking toward tumor-cell islands and subsequent killing of tumor cells. It is shown that CAR T cells often merge into large clusters that envelop and kill the tumor cells with high efficiency. Significant differences are also measured between CAR T cells from different donors and between various CAR T cell constructs. Overall, the assay allows for multifaceted, dynamic, high-content evaluation of CAR T trafficking, clustering, and killing and could eventually become a useful tool for immune-oncology research and preclinical assessments of cell-based immunotherapies., Competing Interests: The authors declare no conflict of interest., (© 2019 The Authors. Published by WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim.)

Chimeric antigen receptor (CAR)-T-cell therapy for solid tumors is limited due to heterogeneous target antigen expression and outgrowth of tumors lacking the antigen targeted by CAR-T cells directed against single antigens. Here, we developed a bicistronic construct to drive expression of a CAR specific for EGFRvIII, a glioblastoma-specific tumor antigen, and a bispecific T-cell engager (BiTE) against EGFR, an antigen frequently overexpressed in glioblastoma but also expressed in normal tissues. CART.BiTE cells secreted EGFR-specific BiTEs that redirect CAR-T cells and recruit untransduced bystander T cells against wild-type EGFR. EGFRvIII-specific CAR-T cells were unable to completely treat tumors with heterogenous EGFRvIII expression, leading to outgrowth of EGFRvIII-negative, EGFR-positive glioblastoma. However, CART.BiTE cells eliminated heterogenous tumors in mouse models of glioblastoma. BiTE-EGFR was locally effective but was not detected systemically after intracranial delivery of CART.BiTE cells. Unlike EGFR-specific CAR-T cells, CART.BiTE cells did not result in toxicity against human skin grafts in vivo.

Objective: Owing to mediastinal and cardiac damage burden, the surgical treatment of radiotherapy-related mitral regurgitation (MR) may be associated with high operative risk or might even contraindicated. We evaluated the feasibility and outcome of MitraClip therapy in patients with radiotherapy-related MR as an alternative to surgery., Methods: Based on Doppler Echocardiography, 15 of 33 screened patients underwent MitraClip implantation., Results: Following MitraClip MR improved (residual MR ≤2+) without significant mitral valve stenosis (planimetric area 2.83 ± 0.8 cm 2 , mean gradient 4.6 ± 1.8 mm Hg). All patients completed a 6-month follow-up, while 14 of 15 patients achieved a longer follow-up, ranging from 12 to 72 months (median 24 months, IQR 42 months). At 6-month follow-up we observed NYHA improvement in 13 patients with an increase of 6-min walking covered distance (from 260 ± 34 to 367 ± 70, p < 0.001), sustained moderate or less MR, mild mitral stenosis in 3 patients, and significant systolic Pulmonary Artery Pressure (PAPs) reduction (from 52.5 ± 14 to 42 ± 9, p < 0.01). Sustained clinical improvement and ≤2+ MR was observed in 13 of 14 patients who completed the 12-month follow-up. Two patients died of acute pneumonia (11 months and 60 months, respectively). One patient developed moderate MV stenosis (MVA 1.4 cm 2 ) at last follow-up (48 months) without related clinical instability. Tricuspid regurgitation improved in 12 patients with further improvement at late follow-up in 2 of 3 patient with 3+., Conclusion: MitraClip may be an effective treatment for RT-induced MR, although unexpected late stenosis may occur in the context of sustained reactive mitral apparatus damage following mediastinal radiation., (Copyright © 2018 Hellenic Society of Cardiology. Published by Elsevier B.V. All rights reserved.)

Cutaneous T cell lymphomas (CTCL) are a heterogeneous group of malignancies characterized by the expansion of a malignant T cell clone. Chimeric Antigen Receptor (CAR) T cell therapy has shown impressive results for the treatment of B-cell tumors, but several challenges have prevented this approach in the context of T cell lymphoma. These challenges include the possibilities of fratricide due to shared T-cell antigens, T cell immunodeficiency, and CAR transduction of malignant cells if CAR T are manufactured in the autologous setting. In this review, we discuss these and other challenges in detail and summarize the approaches currently in development to overcome these challenges and offer cellular targeting of T cell lymphomas.

Broad use of CRISPR-Cas12a (formerly Cpf1) nucleases 1 has been hindered by the requirement for an extended TTTV protospacer adjacent motif (PAM) 2 . To address this limitation, we engineered an enhanced Acidaminococcus sp. Cas12a variant (enAsCas12a) that has a substantially expanded targeting range, enabling targeting of many previously inaccessible PAMs. On average, enAsCas12a exhibits a twofold higher genome editing activity on sites with canonical TTTV PAMs compared to wild-type AsCas12a, and we successfully grafted a subset of mutations from enAsCas12a onto other previously described AsCas12a variants 3 to enhance their activities. enAsCas12a improves the efficiency of multiplex gene editing, endogenous gene activation and C-to-T base editing, and we engineered a high-fidelity version of enAsCas12a (enAsCas12a-HF1) to reduce off-target effects. Both enAsCas12a and enAsCas12a-HF1 function in HEK293T and primary human T cells when delivered as ribonucleoprotein (RNP) complexes. Collectively, enAsCas12a provides an optimized version of Cas12a that should enable wider application of Cas12a enzymes for gene and epigenetic editing.

Chimeric antigen receptor (CAR) T cells have emerged as a novel form of treatment of patients with B-cell malignancies. In particular, anti-CD19 CAR T-cell therapy has effected impressive clinical responses in B-cell acute lymphoblastic leukemia and diffuse large B-cell lymphoma. However, not all patients respond, and relapse with antigen loss has been observed in all patient subsets. Here, we report on the design and optimization of a novel CAR directed to the surface antigen CD37, which is expressed in B-cell non-Hodgkin lymphomas, in chronic lymphocytic leukemia, and in some cases of cutaneous and peripheral T-cell lymphomas. We found that CAR-37 T cells demonstrated antigen-specific activation, cytokine production, and cytotoxic activity in models of B- and T-cell lymphomas in vitro and in vivo, including patient-derived xenografts. Taken together, these results are the first showing that T cells expressing anti-CD37 CAR have substantial activity against 2 different lymphoid lineages, without evidence of significant T-cell fratricide. Furthermore, anti-CD37 CARs were readily combined with anti-CD19 CARs to generate dual-specific CAR T cells capable of recognizing CD19 and CD37 alone or in combination. Our findings indicate that CD37-CAR T cells represent a novel therapeutic agent for the treatment of patients with CD37-expressing lymphoid malignancies., (© 2018 by The American Society of Hematology.)

Chimeric antigen receptor (CAR) T-cell therapy represents a revolutionary treatment for haematological malignancies (i.e. B-ALL). However, the success of this type of treatment has not yet been achieved in solid tumors. One hypothesis is that the immunosuppressive nature of the tumor microenvironment (TME) influences and affects the efficacy of adoptive immunotherapy. Understanding the role of the TME and its interaction with CAR T-cells is crucial to improve the potency of adoptive immunotherapy. In this review, we discuss the strategies and potential combinatorial approaches recently developed in mouse models to enhance the efficacy of CAR T-cells, with particular emphasis on the translational potential of these approaches.

Anaplastic Large Cell Lymphoma (ALCL) is a clinical and biological heterogeneous disease including systemic ALK positive and ALK negative entities. Whereas ALK positive ALCLs are molecularly characterized and readily diagnosed, specific immunophenotypic or genetic features to define ALK negative ALCL are missing, and their distinction from other T-cell non-Hodgkin lymphomas (T-NHLs) can be controversial. In recent years, great advances have been made in dissecting the heterogeneity of ALK negative ALCLs and in providing new diagnostic and treatment options for these patients. A new revision of the World Health Organization (WHO) classification promoted ALK negative ALCL to a definite entity that includes cytogenetic subsets with prognostic implications. However, a further understanding of the genetic landscape of ALK negative ALCL is required to dictate more effective therapeutic strategies specifically tailored for each subgroup of patients.

Epithelial splicing regulatory protein 1 (ESRP1) is an epithelial cell-specific RNA binding protein that controls several key cellular processes, like alternative splicing and translation. Previous studies have demonstrated a tumor suppressor role for this protein. Recently, however, a pro-metastatic function of ESRP1 has been reported. We thus aimed at clarifying the role of ESRP1 in Colorectal Cancer (CRC) by performing loss- and gain-of-function studies, and evaluating tumorigenesis and malignancy with in vitro and in vivo approaches. We found that ESRP1 plays a role in anchorage-independent growth of CRC cells. ESRP1-overexpressing cells grown in suspension showed enhanced fibroblast growth factor receptor (FGFR1/2) signalling, Akt activation, and Snail upregulation. Moreover, ESRP1 promoted the ability of CRC cells to generate macrometastases in mice livers. High ESRP1 expression may thus stimulate growth of cancer epithelial cells and promote colorectal cancer progression. Our findings provide mechanistic insights into a previously unreported, pro-oncogenic role for ESRP1 in CRC, and suggest that fine-tuning the level of this RNA-binding protein could be relevant in modulating tumor growth in a subset of CRC patients.

Understanding transformation mechanisms other than genetic aberrations has recently captured the attention of cancer researchers. To date, the role of transposable elements (TEs) in tumor development remains largely undefined. However, an increasing number of studies have reported that loss of epigenetic control causes TE reactivation and consequent oncogenic transcription. Here, we discuss principal examples of TEs-driven oncogenesis. Available data suggest that long terminal repeats and long interspersed nuclear elements play a pivotal role as alternative promoters. These findings provide definitive experimental evidence that repetitive elements are a powerful underestimated force toward oncogenesis and open the possibility to new therapeutic treatments., (Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.)

Background: The neurally mediated syncope (NMS) is sustained by complex cardiac and vascular reflexes, acting on and amplified by central autonomic loops, resulting in bradycardia and hypotension., Hypothesis: Our aim was to assess whether the pathophysiology of NMS is also related to an abnormal peripheral vasoreactivity., Methods: We evaluated by ultrasound the flow-mediated vasodilation (FMD) and the nitrate-mediated dilation (NMD) in 17 patients with NMS, induced by drug-free tilt test in 6 subjects and by nitrate-potentiated tilt test in the other 11 cases; the syncope was classified as vasodepressive (VD) in 8 cases, cardioinhibitory (CI) in 7, and mixed in 2., Results: The FMD was not different from controls (10.2 ± 4.5 vs 11.4 ± 3.9, P = ns), with normal recovery times; the NMD was greater in fainting subjects than in controls (26.7 ± 7.3 vs 19.0 ± 3.6, P < 0.05), with higher values in VD than in CI syncope (31.1 ± 7.0 vs 23.1 ± 5.0, P = ns); compared to controls, subjects with NMS showed normal recovery times after FMD but longer recovery times after nitrate administration (13.0 ± 5.6 vs 6.3 ± 0.7 minutes, P < 0.05)., Conclusions: The evaluation of endothelial function supports evidence that NMS is characterized by a marked and sustained endothelial-independent vasodilation, in the presence of a normal FMD; vascular hyperreactivity in response to nitrate administration is particularly overt in vasodepressive syncope and can explain the high rate of responses to nitrate administration during tilt test., (© 2011 Wiley Periodicals, Inc.)