Recipient endothelial cells (ECs) actively respond to inflammation during allogeneic hematopoietic cell transplant (allo-HCT), yet mechanisms by which ECs influence acute graft-versus-host-disease (GVHD) pathology remain incompletely defined. Single cell RNA-sequencing of ECs isolated from a GVHD target organ, the liver, showed rapid, subset-specific transcriptional reprogramming after allo-HCT, with induction of canonical interferon-γ (IFNγ)-inducible genes including interferon regulatory factor 1 (IRF1), MHC II, and PD-L1 in lymphatic ECs (LECs). In allo-HCT recipients, circulating IFNγ peaked on day 4 (early), decreased but remained elevated at day 14 (late), and declined by day 21, with a concordant induction of IRF1+ LECs in the liver and GI tract. IFNγ neutralization with anti-IFNγ monoclonal antibody at either early or late time points attenuated IRF1⁺MHCII⁺PD-L1⁺ LEC activation. Notably, early donor T cell expansion was IFNγ-independent, whereas late IFNγ blockade selectively impaired donor Treg, but not Th1, expansion, leading to accelerated GVHD. Using in vitro LEC-CD4 T cell co-cultures and allo-HCT in Irf1-/- bone marrow chimera recipients, we show that recipient ECs that cannot mediate IFNγ-IRF1 signaling exhibit reduced activation and apoptosis, but also have impaired Treg expansion, increased donor Th1/Treg ratios, and worsened GVHD severity. Finally, pharmacological JAK inhibition in allo-HCT recipient mice spares IRF1+ LECs and Tregs while reducing pathogenic Th1 cells, correlating with reduced GVHD severity and improved survival. Our findings identify a role for the lymphatic endothelial IFNγ-IRF1 axis in regulating early vascular remodeling, donor T cell mediated tolerance, underscoring a potential "goldilocks" level of pathway activation required to improve post-transplant outcomes.

The c-MYC (MYC) oncogene is a critical driver of multiple myeloma (MM), however, direct targeting of the MYC protein has proven challenging due to its intrinsic structural disorder. In this study, we evaluated the biological activity and molecular mechanism of a ribonuclease-targeting chimera (RiboTAC) designed to promote the degradation of MYC mRNA (MYC-RiboTAC) across a panel of primary patient MM samples and MM cell lines. This heterobifunctional molecule consists of a small molecule targeting the MYC internal ribosomal entry site (IRES) conjugated to a small-molecule recruiter of endogenous RNaseL. The MYC-RiboTAC reduces MYC mRNA and protein levels in an RNase L-dependent manner, selectively inhibits MYC-driven transcriptional programs, and exhibits potent anti-MM activity. It effectively suppresses cell growth in MM cells co-expressing MYC and RNase L, even in the presence of the protective bone marrow. Furthermore, it synergizes with clinically active agents such as carfilzomib, lenalidomide, and pomalidomide. Importantly, MYC-RiboTAC displays favorable safety and pharmacokinetic profiles in mice and significantly suppresses tumor growth in NOD SCID mice bearing MM xenografts, as shown in two different models. These findings highlight the potential of RNA degraders to target mRNAs encoding "undruggable" proteins such as MYC, offering a promising avenue for precision cancer therapy.

Sickle cell disease (SCD) is an inherited hemoglobinopathy characterized by chronic hemolytic anemia, painful vaso-occlusive episodes, and end-organ damage. Cellular therapies, including allogeneic hematopoietic cell transplant (allo-HCT) and gene therapy, are potentially curative treatments for SCD. Cases of myelodysplastic syndrome and acute myeloid leukemia (MDS/AML) occurring after cellular therapy for SCD and reports of an increased relative risk of AML in populations with SCD have raised questions about the association between SCD and clonal hematopoiesis (CH), a recognized precursor state for MDS/AML. We used error-corrected DNA sequencing to detect CH variants at a variant allele fraction > 0.0005 in blood samples from 98 individuals with SCD and 72 non-SCD donor controls who underwent HCT at the NIH between July 2004 and June 2023. Baseline CH prevalence was similar in recipients and donors; however, SCD recipients had 3.8-fold higher odds of DNA Damage Response (DDR)-mutant CH, involving TP53, PPM1D, ATM, and CHEK2, compared with non-SCD donors. Following HCT, the proportion of SCD recipients with CH increased, driven primarily by mutations in DNMT3A/TET2 and TP53. Post-HCT CH arose from newly emergent variants undetectable at baseline in both recipient and paired donor samples, engrafted donor-derived CH, and persistent recipient CH. All cases of donor-derived CH involved DNMT3A/TET2 mutations, while persistent-recipient CH involved DDR mutations. Hematologic malignancies were rare but included 3 cases of fatal TP53-mutant MDS/AML. Larger cohort studies are needed to identify risk factors for developing DDR-CH and MDS/AML in SCD and to optimize cellular therapy safety in this population.

Light chain (AL) amyloidosis is a fatal plasma cell dyscrasia characterized by the overproduction of misfolded l or k immunoglobulin light chains (LCs) produced by clonal plasma cells, which aggregate into amyloid fibrils that deposit in tissues and cause progressive organ damage. While current anti-plasma cell therapies reduce the production of new amyloidogenic LCs, pre-existing fibrils persist and continue to drive organ damage. Amyloid-targeting monoclonal antibodies birtamimab and anselamimab were developed and tested for active AL amyloid clearance; however, they failed to meet the primary endpoint in Phase 3 trials, presumably due to insufficient binding affinity for l LC amyloid, which occurs in ~80% of patients. Here, we reported the development of 1F10, a novel high-affinity l subtype-specific amyloid-binding monoclonal antibody that does not cross-react with soluble native l or k LCs. 1F10 exhibits superior binding affinity to l AL amyloid fibrils and significantly enhances antibody-dependent phagocytosis (ADP) of l AL amyloid compared with birtamimab and anselamimab. Specific binding of 1F10 to l amyloid is confirmed by immunohistochemical staining of patient-derived tissue biopsies. Importantly, 1F10 demonstrates robust in vivo activity, significantly accelerating the resolution of l amyloid fibrils in an AL amyloidoma mouse model. Together, these findings establish 1F10 antibody as a promising subtype-specific immunotherapeutic candidate for targeting l LC amyloid, addressing a critical unmet need for the majority of patients with AL amyloidosis.

Clonal cytopenia of undetermined significance (CCUS) is defined by unexplained cytopenias with myeloid-associated somatic mutations not meeting diagnostic criteria for myelodysplastic syndromes/neoplasms (MDS) yet carries a highly risk-stratified probability of progression to myeloid neoplasms. The clinical distinction between CCUS and lower-risk MDS (LR-MDS) is challenging because current criteria rely heavily on semi-quantitative morphologic thresholds, despite substantial clinical and molecular overlap. In this prospective study of 409 patients with CCUS and 241 with LR-MDS, we applied harmonized diagnostic and progression criteria, rigorous centralized pathology review, and uniform genomic profiling to compare clinical, molecular, and outcome data. Risk stratification was performed using two independent models-the Clonal Hematopoiesis Risk Score (CHRS) and the Clonal Cytopenia Risk Score (CCRS). Patients with high-risk CCUS, as defined by CHRS or CCRS, exhibited clinical features and event rates comparable to those with LR-MDS. In contrast, patients with low- or intermediate-risk CCUS had markedly improved outcomes, supporting conservative management. These findings underscore that CHRS and CCRS are clinically informative tools that extend beyond morphology-based classification and enable a risk-adapted approach to the management of CCUS. Importantly, a subset of patients with high-risk CCUS demonstrated substantial clinical and genomic convergence with LR-MDS, supporting their consideration for enrollment in prospective clinical trials designed for LR-MDS. These observations highlight the need for further study of risk-adapted therapeutic approaches in this population and underscore the importance of prospective clinical evaluation.

Current cell and gene therapies lack clinically practical mechanisms to selectively promote or suppress therapeutic cells in vivo, a limitation that is particularly acute in patients requiring ongoing immunosuppression. This includes gene therapy for immune dysregulation syndromes, and antigen-specific or chimeric antigen receptor (CAR) T-cell therapy for patients requiring immunosuppression (e.g., transplant recipients), where both pathogenic and therapeutic cells may be suppressed. Here, we develop a multiplex prime-editing platform that converts commonly used immunosuppressive drugs into tools for in vivo control of T-cell therapies via defined, pathway-specific drug resistance. Focusing initially on gene therapy, prime editing efficiently edited loci of multiple pathogenic variants associated with immune dysregulation in primary human T-cells and corrected the HAVCR2 driver mutation in T-cells from multiple patients with subcutaneous panniculitis-like T-cell lymphoma (SPTCL). Comprehensive genomic, transcriptional, immunophenotypic, and clonal analyses demonstrated minimal off-target perturbation. Multiplexed gene correction and drug-resistance editing of T-cells from patients with SPTCL enabled selective in vivo expansion of corrected cells under immunosuppressive pressure in humanized mouse models and exhibited retained sensitivity to alternative agents permitting rapid in vivo suppression. Extending this approach, prime edited, drug-resistant antigen-specific and CAR T-cells retained effector function despite pharmacologic immunosuppression, demonstrating the generalizability of this platform to diverse cellular therapies. Together, these findings establish multiplex prime editing as a promising preclinical framework for generating drug-controllable T-cell therapies, enabling selective in vivo modulation in settings where immunosuppression cannot be withdrawn.

Immunotherapy has revolutionized the treatment of solid cancers in recent years. However, T-cell lymphomas (T-NHLs) originate from immune cells themselves and are biologically heterogeneous, rendering investigations of immune checkpoint inhibitor (ICI) mechanisms of action complex. While case reports and individual Anaplastic Large Cell Lymphoma (ALCL) cases enrolled in T-NHL trials demonstrated favourable responses to ICI, hyperprogression was observed in other T-NHL subtypes. We therefore utilized a syngeneic mouse model of ALK+ ALCL to investigate immune surveillance and ICI-induced immune response. Transplantation experiments combined with depletion of relevant immune axes revealed that ALCL immune surveillance is mediated by CD4+ T cells and NK cells. Innate and adaptive immune cell infiltration was confirmed on a large series of primary human ALK+ ALCL samples. ICI monotherapy demonstrated robust efficacy in murine ALCL, inducing complete remissions in approximately 50% of treated animals. Mechanistically, PD-L1 blockade reversed Treg-mediated immunosuppression and increased the frequency of circulating effector CD8+ T lymphocytes, thereby prolonging survival significantly. Importantly, CD4+ T cells proved indispensable for driving and sustaining immunotherapy-induced anti-tumour responses in murine ALCL. CD4+ T cells of non-responder animals exhibited an exhausted phenotype and a transcriptomic Th22-like signature, implicating persistent T-cell exhaustion and polarization as a meaningful immune-evasion mechanism. Our findings uncover CD4+ T cells as key players in spontaneous and immunotherapy-mediated anti T-cell lymphoma immunity, which demonstrates the critically needed preclinical proof-of-concept for the safe and effective use of immunotherapy for ALCL.

Patients in the liso-cel plus ibrutinib cohort of the phase 1/2, open-label TRANSCEND CLL 004 study had relapsed/refractory chronic lymphocytic leukemia (CLL)/small lymphocytic lymphoma (SLL) and received liso-cel (50×106 [dose level (DL)1] or 100×106 [DL2] chimeric antigen receptor-positive T cells) with concurrent ibrutinib from enrollment through 90 days after liso-cel infusion or longer per investigator discretion. Primary end point was complete response/remission (CR)/CR with incomplete marrow recovery (CRi) by investigator assessment. Among 56 patients who received ibrutinib plus liso-cel (DL1, n=5; DL2, n=51), median (range) age was 64.5 years (44‒77), 98% had high-risk cytogenetics, 55% had progression on Bruton tyrosine kinase inhibitor and venetoclax failure, and median (range) number of prior therapies was 5 (1‒13). Median (range) follow-up was 24.8 months (3.1‒51.8). At DL2, CR/CRi rate was 45% (95% confidence interval [CI], 31‒60) and overall response rate was 86% (95% CI, 74‒94). Median (95% CI) duration of response was not reached (NR; 28.7‒NR) for patients with CR/CRi and 41.4 months (23.3‒NR) for all responders. Median (95% CI) progression-free survival was 31.4 months (20.1‒NR). In DL1+DL2, most common grade ≥3 treatment-emergent adverse events (TEAE) were neutropenia (52%) and anemia (41%). Cytokine release syndrome was reported in 80% of patients (grade 3, 4%; no grade 4/5), and neurological events in 41% (grade 3/4, 11%; no grade 5). Ibrutinib-related TEAEs were reported in 68% of patients (grade 3/4, 43%; no grade 5). Liso-cel plus ibrutinib demonstrated notable efficacy in patients with relapsed/refractory CLL/SLL with predictable and manageable safety. Clinicaltrials.gov: NCT03331198; NCT03435796.