Cancer, once thought to be caused largely by genetic alterations, is now considered to be a mixed genetic and epigenetic disease. The epigenetic landscape, which is dictated by covalent DNA and ...histone modifications, is profoundly altered in transformed cells. These abnormalities may arise from mutations in, or altered expression of, chromatin modifiers. Recent reports on the interplay between cellular signalling pathways and chromatin modifications add another layer of complexity to the already complex regulation of the epigenome. In this Review, we discuss these new studies and how the insights they provide can contribute to a better understanding of the molecular pathogenesis of neoplasia.
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IJS, NUK, SBMB, UL, UM, UPUK
Progress in adoptive T-cell therapy for cancer and infectious diseases is hampered by the lack of readily available, antigen-specific, human T lymphocytes. Pluripotent stem cells could provide an ...unlimited source of T lymphocytes, but the therapeutic potential of human pluripotent stem cell-derived lymphoid cells generated to date remains uncertain. Here we combine induced pluripotent stem cell (iPSC) and chimeric antigen receptor (CAR) technologies to generate human T cells targeted to CD19, an antigen expressed by malignant B cells, in tissue culture. These iPSC-derived, CAR-expressing T cells display a phenotype resembling that of innate γδ T cells. Similar to CAR-transduced, peripheral blood γδ T cells, the iPSC-derived T cells potently inhibit tumor growth in a xenograft model. This approach of generating therapeutic human T cells 'in the dish' may be useful for cancer immunotherapy and other medical applications.
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DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
T cell engineering is a powerful means to rapidly generate anti-tumor T cells. The costimulatory properties of second-generation chimeric antigen receptors (CARs) determine the overall potency of ...adoptively transferred T cells. Using an in vivo “stress test” to challenge CD19-targeted T cells, we studied the functionality and persistence imparted by seven different CAR structures providing CD28 and/or 4-1BB costimulation. One configuration, which uses two signaling domains (CD28 and CD3ζ) and the 4-1BB ligand, provided the highest therapeutic efficacy, showing balanced tumoricidal function and increased T cell persistence accompanied by an elevated CD8/CD4 ratio and decreased exhaustion. Remarkably, induction of the IRF7/IFNβ pathway was required for optimal anti-tumor activity. Thus, 1928z-41BBL T cells possess strikingly potent intrinsic and immunomodulatory qualities.
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•CD28 CARs confer greater functionality and 4-1BB CARs greater persistence to T cells•Costimulatory ligands potentiate second generation CARs, most strikingly 1928z-41BBL•1928z-41BBL strongly induces IRF7/IFNβ pathway in T cells•IRF7/IFNβ activation enhances CAR T cell-mediated tumor elimination
Zhao et al. analyze seven chimeric antigen receptors structures and show that T cells receiving integrated CD28 and 4-1BB signals display superior tumoricidal activity and robust persistence. These T cells also activate the IFNβ/IRF7 pathway to support their anti-tumor activity.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
In the rapidly evolving landscape of medical research, the emergence of RNA-based therapeutics is paradigm shifting. It is mainly driven by the molecular adaptability and capacity to provide ...precision in targeting. The Covid-19 pandemic crisis underscored the effectiveness of the mRNA therapeutic development platform and brought it to the forefront of RNA-based interventions. These RNA-based therapeutic approaches can reshape gene expression, manipulate cellular functions, and correct aberrant molecular processes underlying various diseases. The new technologies hold the potential to engineer and deliver tailored therapeutic agents to tackle genetic disorders, cancers, infectious diseases in a highly personalized and precisely tuned manner. The review discusses most recent advancements in the field of mRNA therapeutics for cancer treatment, with a focus on the features of the most utilized RNA-based therapeutic interventions, current pre-clinical and clinical developments and the remaining challenges in delivery strategies, effectiveness, and safety considerations.
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Vu and colleagues offer an insightful overview of RNA-based therapy, a recently emerged as an exciting therapeutic approach. The review describes the rational design of synthetic mRNA and strategies to deliver mRNAs, and an extensive discussion of current developments as well as safety and regulatory consideration for clinical applications of the technologies.
Significant progress has been made in the treatment of multiple myeloma (MM) however, no strategy to date has proven curative and relapse invariably occurs. We recently developed an integrated ...pipeline to identify biologically and therapeutically relevant targets for MM immunotherapy by using Mass-Spectrometry analyses from seven MM cell lines and RNA-seq data from 900+ patients. Starting from 4,000+ candidates, ILT3 aka LILRB4, an immunoreceptor tyrosine-based inhibition motif-containing receptor emerged as a promising candidate target: it has absent/low expression in most normal tissues and high and frequent expression in many primary patient samples we used for validation, similarly to BCMA (Di Meo et al., Cell Reports Medicine 2023). We sought to examine its functional role in MM and develop Chimeric Antigen Receptor (CAR) T-cells targeting LILRB4. Interrogating RNA-seq data from MM patients, LILRB4 expression results higher in patients with high-risk cytogenetics such as t:(14;16), t:(14;20), and t:(8;14) translocations compared to patients with no genomic abnormalities. Investigating function, we find no difference between cell proliferation of U266/KMS11/OPM2 genetically depleted of LILRB4 (KO) compared to controls by measuring cell growth in vitro for 120 hours and MTS assays. Similarly, when KO and control cells are seeded in plastic chambers containing a porous membrane and serum-free medium, both KO and control cells are attracted to the lower chambers containing FBS, suggesting that LILRB4 is also not required for MM cell migration. Despite these initial observations, RNAseq data from LILRB4 KO and overexpressing (OE) U266 MM cells identify ~3,000 differentially expressed genes compared to controls. Thus, we next investigate potential extrinsic effects on T-cell proliferation by co-culturing T-cells from healthy donors with LILRB4 KO or OE U266 MM cells (T:E ratio 1:2). T cells are previously activated with anti-CD3/CD28-coated beads and IL-2. 5 days after activation, T cells and LILRB4 KO U266 MM are seeded in the same well plate. After 48h of co-culture, T-cell proliferation is decreased (-30%) in the presence of LILRB4 U266 OE MM cells, whereas it is increased (+50%) when the co-culture includes LILRB4 U266 KO cells. CD25 expression is also reduced by 20% on T cells co-cultured with LILRB4 OE cells and increased by the same extent when T cells are co-cultured with LILRB4 KO cells, suggesting a role as a negative immune receptor. This effect seems dependent on the direct cell-cell interaction given that when T and MM cells are separated by a transwell in co-culture, no effect on the T cell proliferation is observed. Finally, we develop multiple Chimeric Antigen Receptor (CAR) constructs targeting LILRB4, by using different single-chain variable fragments (scFVs) and rearranging the variable regions of the heavy (VH) and light (VL) regions. We used lentiviral LILRB4 scFV-41BB-CD3zeta vectors. LILRB4 CAR T-cells specifically kill U266, H929 and KMS11 MM cell lines in vitro with cytotoxicity ranging from 30 to 80% depending on the construct (T:E ratio 1:10). Equal efficacy is observed in U266 cells resistant to Bortezomib and BCMA KO U266cells. No effect on LILRB4 null cells is noted. These data suggest that LILRB4 is a negative immune receptor that is active on malignant plasma cells mediating T-cell suppression, similar to previously described data in leukemia (Deng et al., Nature 2018). Our work supports the rationale development of ILT3 CAR-T cell therapy as a viable therapeutic approach for MM patients especially high-risk patients. Ongoing studies are investigating the functional role of LILRB4 in mediating the cross-talk between MM ad immune cells.
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IJS, IMTLJ, KILJ, NLZOH, NUK, SAZU, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Chimeric antigen receptor (CAR) therapy targeting CD19 has yielded remarkable outcomes in patients with acute lymphoblastic leukemia. To identify potential CAR targets in acute myeloid leukemia ...(AML), we probed the AML surfaceome for overexpressed molecules with tolerable systemic expression. We integrated large transcriptomics and proteomics datasets from malignant and normal tissues, and developed an algorithm to identify potential targets expressed in leukemia stem cells, but not in normal CD34+CD38− hematopoietic cells, T cells, or vital tissues. As these investigations did not uncover candidate targets with a profile as favorable as CD19, we developed a generalizable combinatorial targeting strategy fulfilling stringent efficacy and safety criteria. Our findings indicate that several target pairings hold great promise for CAR therapy of AML.
•We generated and annotated an extensive dataset of AML cell surface proteins•We designed an algorithm to identify candidate CAR targets•We defined six criteria for combinatorial pairing of CAR targets•We identified target combinations fulfilling stringent criteria for CAR therapy
Perna et al. search for optimal chimeric antigen receptor targets in acute myeloid leukemia using extensive proteomics and transcriptomics. In the absence of a target as favorable as CD19, they develop a generalizable combinatorial targeting strategy identifying several promising target pairings.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Acquired aplastic anemia (AA) is the typical bone marrow failure syndrome characterized by an empty bone marrow; an immune-mediated pathophysiology has been demonstrated by experimental works as well ...as by clinical observations. Immunusuppressive therapy (IST) is a key treatment strategy for aplastic anemia; since 20 years the standard IST for AA patients has been anti-thymocyte globuline (ATG) plus cyclosporine A (CyA), which results in response rates ranging between 50% and 70%, and even higher overall survival. However, primary and secondary failures after IST remain frequent, and to date all attempts aiming to overcome this problem have been unfruitful. Here we review the state of the art of IST for AA in 2010, focusing on possible strategies to improve current treatments. We also discuss very recent data which question the equality of different ATG preparations, leading to a possible reconsideration of the current standards of care for AA patients.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative therapy for hematological malignancies. However, graft-versus-host disease (GVHD) and relapse after allo-HSCT ...remain major impediments to the success of allo-HSCT. Chimeric antigen receptors (CARs) direct tumor cell recognition of adoptively transferred T cells. CD19 is an attractive CAR target, which is expressed in most B cell malignancies, as well as in healthy B cells. Clinical trials using autologous CD19-targeted T cells have shown remarkable promise in various B cell malignancies. However, the use of allogeneic CAR T cells poses a concern in that it may increase risk of the occurrence of GVHD, although this has not been reported in selected patients infused with donor-derived CD19 CAR T cells after allo-HSCT. To understand the mechanism whereby allogeneic CD19 CAR T cells may mediate anti-lymphoma activity without causing a significant increase in the incidence of GVHD, we studied donor-derived CD19 CAR T cells in allo-HSCT and lymphoma models in mice. We demonstrate that alloreactive T cells expressing CD28-costimulated CD19 CARs experience enhanced stimulation, resulting in the progressive loss of both their effector function and proliferative potential, clonal deletion, and significantly decreased occurrence of GVHD. Concurrently, the other CAR T cells that were present in bulk donor T cell populations retained their anti-lymphoma activity in accordance with the requirement that both the T cell receptor (TCR) and CAR be engaged to accelerate T cell exhaustion. In contrast, first-generation and 4-1BB-costimulated CAR T cells increased the occurrence of GVHD. These findings could explain the reduced risk of GVHD occurring with cumulative TCR and CAR signaling.
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IJS, NUK, SBMB, UL, UM, UPUK
Epigenetic regulation of key transcriptional programs is a critical mechanism that controls hematopoietic development, and, thus, aberrant expression patterns or mutations in epigenetic regulators ...occur frequently in hematologic malignancies. We demonstrate that the Polycomb protein L3MBTL1, which is monoallelically deleted in 20q- myeloid malignancies, represses the ability of stem cells to drive hematopoietic-specific transcriptional programs by regulating the expression of SMAD5 and impairing its recruitment to target regulatory regions. Indeed, knockdown of L3MBTL1 promotes the development of hematopoiesis and impairs neural cell fate in human pluripotent stem cells. We also found a role for L3MBTL1 in regulating SMAD5 target gene expression in mature hematopoietic cell populations, thereby affecting erythroid differentiation. Taken together, we have identified epigenetic priming of hematopoietic-specific transcriptional networks, which may assist in the development of therapeutic approaches for patients with anemia.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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