Breast cancer is the most common malignancy among women worldwide, causing >650,000 deaths annually. Despite advances in personalized therapies, many patients fail to obtain durable benefits from treatments tailored to their disease subtype. For instance, only some patients with PD-L1⁺ triple-negative breast cancer (TNBC) respond to chemotherapy plus immune checkpoint inhibitors. Moreover, while tumor-infiltrating lymphocyte levels correlate with improved survival in TNBC and HER2⁺ breast cancer, this is not true for HR⁺HER2- disease. Thus, other immunobiological features of the breast cancer microenvironment may hold clinically relevant prognostic or predictive value, especially in HR+HER2- tumors. Here, we investigated transcriptional signatures of immune cell infiltration across three public transcriptomic datasets from patients with breast cancer. The relative abundance and activation status of tumor-infiltrating mast cells were consistently associated with shifts in survival indicators amongst patients with HR+HER2- breast cancer. Specifically, infiltration by resting (but not activated) mast cells was associated with prolonged survival, correlating inversely with tumor infiltration by immune cells and proliferation markers in cancer cells, but positively with stromal richness. While based on retrospective transcriptional analyses, these findings suggest that interactions between mast cells and non-malignant components of the breast cancer microenvironment, particularly fibroblasts, may shape disease progression and treatment sensitivity.
Tumor-infiltrating lymphocytes (TILs) shape melanoma behavior and response to immunotherapy, but the links between immune regulation, TIL patterning, and outcomes remain unclear. We retrospectively studied 32 primary melanoma samples from patients treated with anti-PD-1 therapy. Histopathologic TILs were classified as brisk or non-brisk, and primary tumors underwent targeted immune transcriptomic profiling (NanoString nCounter Human Immunology panel). External validation was performed with 96 TCGA-SKCM primary tumors. Brisk tumors displayed broad upregulation of immune transcripts, with enrichment of adhesion pathways (directed global significance scores - DGSS = 2.15) and MHC class II antigen presentation (DGSS = 2.128). Cross-cohort comparison identified 22 shared differentially expressed genes, with ZAP70 remaining significant in both datasets. Brisk tumors also showed higher total TIL (p = 0.036), cytotoxic-cell (p = 0.0095), and Th1 (p = 0.027) scores. Response-associated genes differed by TIL pattern, and subgroup-specific gene scores predicted anti-PD-1 benefit in brisk (4-gene, Area under curve - AUC = 1.000) and non-brisk (3-gene, AUC = 0.852) tumors; the non-brisk score remained independently associated with response (p = 0.029). Higher scores were also associated with prolonged survival. Integrating histopathological TIL patterning with immune transcriptomics may refine prognostication and support immunotherapy stratification in melanoma.
Interleukin-33 (IL-33) is a central regulator of immune responses and inflammation, and genetic variation in IL33 and its receptor IL1RL1 (ST2) is strongly linked to disease susceptibility, notably asthma. Emerging evidence suggests that IL-33 also influences hematopoietic processes and platelet biology, indicating functions beyond canonical immunity. We investigated two missense variants identified in a patient with unexplained thrombocytopenia: a rare novel IL33 variant, c.385T>C (p.Tyr129His) and the common IL1RL1 variant c.1501_1502CA>AG (p.Gln501Arg), previously associated with reduced IL-33 signaling. Structural modeling revealed that IL-33 Y129H disrupts a conserved hydrogen bond within the IL-33/ST2/IL-1RAcP ternary complex, destabilizing receptor engagement. Functional assays confirmed markedly reduced binding affinity and biological activity, establishing Y129H as a loss-of-function variant. The IL1RL1 Q501R variant affects the Toll/IL-1 receptor (TIR) domain, critical for recruiting adapter proteins such as MyD88. Modeling revealed pronounced perturbation in a peripheral helix of the TIR domain, potentially impacting adapter recruitment and downstream signaling, providing a mechanistic basis for its protective association with asthma. Together, these findings provide structural and functional insights into clinically relevant IL33 and IL1RL1 variants. Their simultaneous occurrence in a patient with thrombocytopenia further supports a potential role for this pathway in platelet homeostasis and stress-responsive hematopoiesis.
Brain-Derived Neurotrophic Factor (BDNF), highly enriched in platelets, contributes to vascular integrity and thrombotic responses. The rs11030119 variant, located in a BDNF intronic enhancer, has been linked to stroke recovery, but its influence on peripheral BDNF dynamics and hemostasis remains unexplored. In this study, we aimed to understand the impact of rs11030119 on circulating levels of BDNF, and its subsequent effects on thrombus formation. Twenty-six healthy individuals homozygous for either the GG or AA genotype of rs11030119 were matched for age and sex. BDNF and proBDNF levels were quantified in serum, plasma, and platelets. Platelet aggregation, ATP secretion, thrombin generation, and clot firmness were assessed. Washed platelets were also tested with recombinant BDNF. AA carriers displayed significantly lower plasma and serum BDNF levels and diminished BDNF release upon activation, despite comparable platelet BDNF content. The expression of its receptor, TrkB, on the platelet surface was also reduced in AA carriers. However, platelet reactivity, thrombin generation, and viscoelastic clot properties were preserved across genotypes. In conclusion, rs11030119 modulates circulating and platelet-releasable BDNF without impairing hemostatic function, suggesting that BDNF bioavailability may not directly correlate with thrombotic potential. These findings uncover a novel genetic mechanism controlling platelet-derived neurotrophin output.
Cyclic neutropenia (CyN) is a rare haematological disorder most often caused by mutations in the neutrophil elastase gene (ELANE). We describe three novel ELANE mutations in CyN patients and link them to impaired neutrophil extracellular trap (NET) formation. ELANE sequencing was performed using the Sanger method, and AlphaFold2 was used to visualize the impact of mutations on the structure of neutrophil elastase (NE). NET formation was assessed by colocalization of MPO, NE, and DNA using fluorescence microscopy, and free DNA was quantified fluorometrically. We identified two missense substitutions (M66K, V133G) and one deletion (G192-G196del). These mutations did not significantly alter the overall NE structure but exhibited functional effects: M66K may disrupt substrate binding, V133G increases protein mobility, and the Deletion of G192-G196 may limit the opening of the catalytic pocket. NETosis was impaired in CyN patients compared with controls. Their NETs appeared less expanded, and DNA release into supernatants was reduced, reaching the lowest levels during neutropenia, even in patients with normal neutrophil counts. These abnormalities were accompanied by reduced NE activity. In sum, novel ELANE variants may contribute to altered NE function and defective NET formation in CyN, suggesting that impaired NETosis is partly independent of cyclic changes in neutrophil numbers.
