Plasmodium knowlesi, a zoonotic malaria species, has become a significant public health concern in Southeast Asia. In regions such as Malaysia and southern Thailand, P. knowlesi incidence has risen, even as other human malaria parasites are nearing elimination. Like its close relative Plasmodium vivax, P. knowlesi relies on Duffy-Antigen Receptor for Chemokine (DARC) as a key receptor for erythrocyte invasion. Only Duffy-positive individuals are thought to be susceptible to clinical infection. Here, we reveal that P. knowlesi possesses greater invasion plasticity than previously recognized. This parasite can bypass the need for DARC, as shown by its in vitro adaptation to invade and replicate in Duffy-negative (Fy-) erythrocytes. This adaptation is stable and independent of DARC binding, enabling the adapted parasite line to be maintained in Fy-erythrocytes and to resist inhibition by a-DARC antibodies. Genomic analysis identified a genomic recombination event between the parasite's dbpα and dbpγ genes, resulting in a new chimeric gene dbpαγ. Using CRISPR-Cas9 targeted reversion, we were able to demonstrate that dbpαγ is essential for invasion of Fy- erythrocytes. These findings shed new light on the invasion plasticity of P. knowlesi, with implications for its potential spread beyond Southeast Asia and for understanding the complex host-cell specificity and atypical invasion pathways seen in P. vivax.

In this retrospective analysis of 584 patients who received CAR T-cell therapy for LBCL at six Australian centres we found no association between time of infusion and outcome, accounting for confounders, suggesting minimal clinical impact of chronobiology in this setting.

To elucidate the molecular basis underlying differential response and resistance to ibrutinib in Waldenström's macroglobulinemia (WM), we conducted a prospective phase II trial (ClinicalTrials.gov; NCT02604511) of ibrutinib monotherapy in treatment-naïve patients. Seventy-four sequential bone marrow (BM) aspirates from 17 patients, collected from baseline through 48 treatment cycles, were profiled using single-cell multi-omics. BM cells segregated primarily into B/plasma cell and T-cell compartments. Longitudinal clonal tracking of malignant B/plasma cells identified three distinct evolutionary patterns: "evolution" (early clone contraction with late clone expansion and increasing genomic complexity), "devolution" (early clone expansion with late clone contraction and genomic simplification), and "no-evolution" (stable clonal architecture). The "evolution" pattern was strongly associated with disease progression, whereas "devolution" correlated with durable clinical response. Transcriptomic profiling of resistant clones enabled development and validation of the Waldenström's Ibrutinib Prediction (WIP) score, which predicted treatment response at baseline. Within the WIP signature, LYN emerged as a key regulator; LYN knockdown or inhibition significantly increased WM cell sensitivity to ibrutinib, suggesting a rational combinatorial strategy. In parallel, GZMB⁺ CD8⁺ effector-memory (TEM) cells expanded post-treatment in progressing patients and co-existed with tumor "evolution". These cells exhibited persistently impaired cytotoxic programs (e.g., GNLY), a de-differentiated memory-like state, elevated PDCD1 expression, and reduced TCR diversity. Together, this study provides the first single-cell framework of tumor clonal evolution and T-cell dysfunction under ibrutinib in WM; introduces the WIP score as a predictive biomarker for treatment response; and identifies actionable tumor-intrinsic and immune mechanisms driving resistance.

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) can be caused by maternal IgG alloantibodies against the human platelet antigen 1a (HPA-1a), a Leu33Pro polymorphism at the PSI domain of the β3 subunit shared by integrins αIIbβ3 and αVβ3. Although serologic detection of anti-HPA-1a alloantibodies has long aided FNAIT diagnosis, the disease remains a major clinical challenge due to unpredictable severe outcomes, particularly intracranial hemorrhage and fetal loss. To define the molecular basis of HPA-1a immunogenicity and antibody pathogenicity, we determined crystal structures of the HPA-1a antigen bound to Fabs from four anti-HPA-1a monoclonal antibodies, including three clinically relevant maternal alloantibodies (26.4, D-204, and M-204) and one murine antibody (SZ21). The structures reveal that all antibodies recognize an HPA-1a epitope centered on Leu33, spanning both PSI and I-EGF1 domains of β3 integrin. Variations in antibody binding interfaces account for differential affinities to both αIIbβ3 and αVβ3, while distinct binding orientations determine whether antibody engagement stabilizes the bent inactive β3 conformation. Functional assays demonstrated antibody-dependent inhibition of αIIbβ3-mediated platelet aggregation, thrombus formation, and clot retraction, as well as αVβ3-mediated endothelial adhesion and spreading. These results establish the structural framework of HPA-1a alloantigenicity, explain the conformational selectivity of anti-HPA-1a antibodies, and elucidate how they block integrin activation. The findings provide mechanistic insight into FNAIT pathogenesis and suggest that antibody heterogeneity in affinity and function may contribute to clinical variability. Furthermore, these findings demonstrate antibody-mediated stabilization of the PSI/I-EGF1 domain as a potential strategy for allosteric modulation of integrin structure and function.

Hematopoietic stem cell (HSC) transplantation is a life-saving therapy for immune deficiencies and hematologic malignancies, but its efficacy is limited by poor engraftment. Therapeutic enhancement of HSC grafts requires deeper insight into the intrinsic determinants of their regenerative capacity. Here, we identify mechanical robustness as a critical feature distinguishing human HSCs from multipotent progenitors (MPPs). Through integrative biomechanical and transcriptomic profiling, we show that ZNF467 is a key regulator of HSC mechanical integrity. Loss of ZNF467 disrupts HSC mechanical fitness and abolishes long-term engraftment. Conversely, an engineered phase-separating ZNF467 variant enhanced mechanical strength and engraftment by activating a mechanoresponsive transcriptional program, including upregulation of ICAM1. ICAM1+ HSPCs exhibit superior biomechanical properties and improved engraftment efficiency. Furthermore, phase separation activity of nucleoplasmic ZNF467 (npZNF467) is crucial for its mechanical reprogramming function, and ectopic npZNF467 expression enhances the engraftment capacity of MPPs. Our findings establish biomechanical regulation as an important determinant of stem cell identity and reveal new strategies for engineering stem cells with enhanced regenerative capacity.

Immune thrombotic thrombocytopenic purpura (iTTP) is a rare, life-threatening condition. Caplacizumab substantially shortens the time to clinical response, yet delayed platelet count recovery is occasionally observed, raising concerns about iTTP refractoriness. This retrospective multicenter study analyzed 204 acute iTTP episodes reported to the German REACT-2020 and Austrian ATMAR registries, all treated with caplacizumab. Refractoriness was assessed using the 2017 International Working Group criteria and the more stringent definition by the French Reference Center for Thrombotic Microangiopathies. We evaluated time to platelet recovery and presence of confounding clinical conditions, potentially accounting for persistent thrombocytopenia, in all episodes. By day 5 after caplacizumab initiation, 83.8% of patients (171/204) achieved a clinical response, and the remaining 16.2% (33/204) showed at least a doubling of the platelet count. Only three patients (1.5%) met laboratory criteria for refractoriness. In all cases, plausible alternative causes were present (e.g., missed doses, infection). No patient was refractory without a confounding factor. In 8/204 patients (3.9%) we observed a markedly prolonged thrombocytopenia (≥10 days) and identified confounding conditions in all cases. In the stratified Cox model, the presence of alternative causes of thrombocytopenia was the only independent determinant of delayed platelet count normalization (HR 0.16, 95% CI 0.09-0.28, p < 0.001), whereas baseline parameters were not predictive. In the context of caplacizumab-based therapy, true refractoriness is rare. Delayed platelet count recovery is predominantly attributable to concomitant clinical conditions. Careful clinical assessment and context-sensitive interpretation of treatment response before escalating iTTP-specific therapy may avoid unnecessary treatment intensification and associated risks.

Acute graft-versus-host disease (GVHD) of the gastrointestinal (GI) tract is a primary cause of early transplant mortality after bone marrow transplantation (BMT), driven by local antigen presentation and T-cell expansion. We sought to clarify the role of various donor dendritic cell (DC) subsets in this process in order to define therapeutic strategies to prevent gut GVHD. Donor plasmacytoid DC (pDC) were prominent in the ileum of BMT recipients without GVHD but were largely absent in gut during GVHD. In contrast, donor XCR1+ conventional DC (cDC) were dramatically increased in the mesenteric lymph nodes (mLN) of BMT recipients with GVHD. We utilized Xcr1-DTR mice and Clec4c-DTR mice to enable highly efficient XCR1+ cDC versus pDC depletion. Donor XCR1+ cDC but not pDC deletion attenuated lethal GVHD, depleting most cDC presenting alloantigen, which inhibited α4β7 and the expansion of alloantigen-specific donor Th1 cells in the gut. Aldehyde dehydrogenase 1A (ALDH1A) expression by cDC is known to modulate GVHD and we thus examined the effect of a pan-ALDH1 inhibitor, WIN18,446, on this axis. WIN18,446 administration improved survival and these effects were ALDH1A1-specific but mediated by inhibition of CD11b+ rather than XCR1+ cDC, inhibiting alloantigen-specific Th17 cell differentiation in the gut. These findings highlight the limited role of pDC in the induction of gut GVHD and identify differential and dominant roles for XCR1+ and CD11b+ donor cDC in controlling Th1 and Th17 mediated gut aGVHD. The targeting of donor cDC subsets represents an effective strategy to prevent lethal gut GVHD.