Chimeric antigen receptor (CAR) T cells represent an exciting therapeutic strategy with improved survival outcomes for patients with hematological malignancies. However, the efficacy of CAR T-cell therapy in the treatment of solid tumors remains suboptimal due to therapeutic barriers associated with the solid tumor microenvironment. We investigated whether ionizing radiation could improve vascular perfusion and CAR T-cell delivery in an EGFRvIII-expressing B16F10 melanoma model. Tumors received radiation doses of 2-12 Gy, and perfusion was evaluated at multiple time points using immunofluorescence detection of intravenously administered fluorescent dyes. We found that a single 8-Gy dose of ionizing radiation produced the most significant increase in B16F10 tumor perfusion 4 h after irradiation. Consistently, the irradiation of tumors 4 h prior to a systemic administration of EGFRvIII-targeting CAR T cells led to higher intratumoral CAR T-cell accumulation than in non-irradiated tumors. This approach also resulted in a significantly delayed tumor growth and improved survival relative to radiation or CAR T cells alone. Interestingly, the CD28ζ EGFRvIII-CAR T-cell levels substantially increased in irradiated tumors over time relative to 4-1BBζ EGFRvIII-CAR T cells and produced greater tumor growth delays and survival improvements in comparison to 4-1BBζ EGFRvIII-CAR T cells administered at a 10-fold higher concentration. Taken together, these data highlight the importance of co-stimulatory domains in CAR T-cell function in vivo and demonstrate that irradiating tumors prior to systemic CAR T-cell infusion can increase CAR T-cell infiltration and efficacy in solid tumors.
Clinical trials of adoptive cellular therapy demonstrate that a key characteristic associated with durable responses is in vivo expansion and persistence of transferred T cells. Strategies to develop a less differentiated, stem/memory population in the infusion product and peri-infusional regimens to promote the maintenance of desired T cell states following adoptive transfer would be desirable. Endogenous T cell therapy studies have routinely achieved memory T cells enriched for expression of interleukin (IL)-7 receptor; to eliminate the conventional requirement for immunosuppressive lymphodepletion and its attendant life-threatening toxicities, we performed the first-in-human use of IL-7 in combination with adoptively transferred antigen-specific memory CD8 T cells in a patient with refractory metastatic uveal melanoma. Single-cell immune repertoire profiling of serial peripheral blood sampling revealed substantial in vivo proliferation and expansion of a stem cell memory population in the endogenous T cell therapy product that achieved a >79% predominance of total circulating T cells by 3 weeks post-infusion in this non-lymphodepleted recipient. Although the patient's disease ultimately progressed, these findings demonstrate safety and proof of concept for an IL-7 treatment regimen for expansion of adoptively transferred T cells in vivo and induced memory differentiation in a heavily pretreated patient with refractory solid malignancy.
Gene regulatory networks (GRNs) at single-cell resolution provide a fundamental framework for understanding cellular functions and regulatory mechanisms. However, existing methods often focus on regulatory relationships among genes while overlooking intercellular heterogeneity and global expression organization across cell populations. Here, we present CSGRN, a supervised computational framework that integrates graph embedding and conditional cell-specific networks (CCSNs) to infer GRNs for individual cells from single-cell RNA sequencing (scRNA-seq) data. By incorporating causal regulatory structures and integrating local and global representations, CSGRN improves the accuracy and robustness of regulatory network inference. Benchmark analyses across three datasets demonstrated that CSGRN outperforms nine existing approaches. In addition, we developed two downstream analytical strategies-signal flow analysis and gene perturbation simulation-to quantify regulatory relationships and explore regulatory dynamics. These analyses reveal cell type-specific regulatory programs and key regulators involved in cellular differentiation and disease-related processes, providing a framework for investigating gene regulation in complex biological systems.
Spiral ganglion neurons (SGNs) relay auditory sensory information from the cochlea to the brain. Their loss results in permanent hearing impairment in humans due to their limited regenerative capacity. Progress in hearing restoration has been constrained by the inaccessibility of human inner ear tissue and challenges in generating functionally mature human SGN-like neurons from stem cells in vitro. To generate human SGN-like neurons from human induced pluripotent stem cells (hiPSCs), we recapitulated key signaling pathways involved in human inner ear development. On day (D) 11 of differentiation, nerve growth factor receptor-positive cells (precursors of pre-placodal ectoderm and neural crest) were isolated using magnetic sorting. From D18 to D25, cultures were treated with sonic hedgehogs to induce otic neural progenitors. Neuronal maturation was subsequently promoted by a cocktail of brain-derived neurotrophic factor, neurotrophin-3, and insulin-like growth factor-1, which supports SGN development. Cellular identity and functionality were assessed using single-cell RNA sequencing, immunocytochemistry, whole-cell patch-clamp electrophysiology, co-culture assays, and calcium ion (Ca²⁺) imaging. hiPSC-derived SGN-like neurons exhibited morphological, molecular, electrophysiological, and functional characteristics of SGNs in vivo. Neurons acquired bipolar morphology and were wrapped by glial cells. Transcriptomic analysis revealed that SGN-like neurons were distinct from other neuronal lineages and showed similarity to type I and type II SGNs based on expression of synaptic and intrinsic excitability-related genes. Electrophysiological recordings revealed progressive hyperpolarization of resting membrane potential and emergence of overshooting action potentials, consistent with neuronal maturation. In co-culture systems, human SGN-like neurons formed functional synaptic connections with mouse cochlear hair cells and cochlear nucleus neurons, evidenced by Ca2+ transients and induction of the immediate early gene c-Fos. This study reports a robust and reproducible protocol for generating human SGN-like neurons from hiPSCs, providing a versatile platform for studying human auditory development, disease modeling, drug screening, and cell-based therapies for hearing restoration.
Regenerating cementum remains a major unmet challenge in periodontal and peri-implant therapy, underscoring the need to understand how cementoblasts respond to engineered surface cues. This study examined the manner in which titanium nanosurfaces integrating anisotropic nanopatterns with three-dimensional (3D) nanospike architectures regulate mechanotransduction and matrix mineralization in human cementoblast-like cells (hCEM). Titanium surfaces with isotropic, anisotropic, and 3D anisotropic nanospike architectures were fabricated and characterized through quantitative analyses of nanoscale geometry and topographical organization. Surface chemistry and crystallinity were characterized using Fourier transform infrared spectroscopy, grazing-incidence X-ray diffraction, and X-ray photoelectron spectroscopy. hCEM cultures on each surface were evaluated for extracellular calcium (Ca) and phosphate (P) levels, Ca/P ratios, extracellular matrix crystallinity, cytomorphology, and phosphate metabolism-associated gene expression. Mechanotransduction activity was assessed through focal adhesion-Hippo pathway signaling. Relationships between nanoscale architecture, cell stimulation, morphology, and mineralization were examined using correlation and path analyses. Despite comparable wettability and oxide chemistry to that of other nanosurfaces, 3D anisotropic nanospike surfaces produced the highest mineralization and exhibited the highest Ca/P ratios, clear hydroxyapatite signatures, pronounced extracellular nodules, and coordinated activation of phosphate metabolism gene profiles. These surfaces induced prominent nanoscale vertex-cell interactions and distinct cytomorphological responses. Mineralization did not show association with vertical roughness, hydroxyl content, or crystallographic features but positively correlated (r = 0.94) with composite nanoscale architectural metrics capturing spatial heterogeneity and vertex density. The finding that anisotropic 3D nanospike architectures are associated with enhanced matrix mineralization in human cementoblast-like cells under osteogenic conditions provides mechanistic insight into how nanoscale architecture modulates mineralization responses and may inform the design of cementum-targeted bioactive titanium surfaces.
Can single-cell, mass spectrometry-based proteomics identify proteins associated with reduced developmental competence of Patl2-/- Metaphase II (MII) mouse oocytes and reveal therapeutic targets for Patl2-related infertility? Abnormal protein content is detected in Patl2-/- MII oocytes, which can be rescued by spindle transfer (ST). PATL2 is an RNA-binding protein that represses maternal mRNA translation during oocyte maturation. PATL2 mutations in humans often cause germinal vesicle (GV) arrest, although some affected patients produce MII oocytes with reduced fertilization and embryo developmental potential. Consequently, oocyte donation is required. The Patl2-/- knockout mouse model offers a unique opportunity to study Patl2-related infertility and evaluate potential treatments. Patl2 -/- mice (C57BL/6NTac-Patl2tm1a), with deletion of exon 7, were bred from April 2021 to October 2023, yielding 36 homozygous females from 271 pups. To investigate the role of Patl2 at the MII stage, in vivo MII oocytes from Patl2-/- and Patl2+/+ females were collected for analysis of key quality markers and single-cell proteomics. Based on these results, maternal ST was tested to rescue abnormal embryo development. At least three replicates were conducted per experiment. Four- to 12-week-old mice underwent superovulation and oocyte collection to assess in vitro and in vivo maturation. In vivo-matured MII oocytes were used to evaluate activation (AR) and blastocyst rates (BR) after PIEZO-ICSI, spindle configuration, and calcium oscillatory patterns following SrCl2 exposure. Vitrified-warmed oocytes were used for single-cell proteomics using a timsTOF ultra mass spectrometer operated in diaPASEF mode. ST involved transferring the Patl2-/- spindle to Patl2+/+ enucleated cytoplasm, followed by parthenogenetic activation (PA) via SrCl2 exposure. Patl2 -/- females exhibit lower in vivo MII rates (79.63%) than Patl2+/+ females (89.39%, P = 0.0123) but similar in vitro maturation rates (GV-MII = 48.74%) compared to Patl2+/+ females (52.85%, P = 0.5230). After PIEZO-ICSI with wild-type sperm, reduced AR (Patl2-/- = 31.71%, Patl2+/+ = 76.74%, P < 0.0001) and BR (Patl2-/- = 7.69%, Patl2+/+ = 42.42%, P = 0.0237) were observed in knockout oocytes. However, Patl2-/- oocytes exhibited normal spindle rates (78.57%) as seen in Patl2+/+ oocytes (86.00%, P = 0.3491), as well as a similar capacity to sustain long-lasting calcium oscillations (A×F = 6.15 ± 4.80) compared to Patl2+/+ oocytes (A×F = 4.59 ± 2.96, P = 0.1453). Single-cell proteomics identified 4882 proteins and confirmed the absence of Patl2 in knockout oocytes, from analyzing 25 Patl2+/+ and 27 Patl2-/- MII oocytes. After filtering, 3747 proteins were used for statistical analysis, revealing 1508 differentially expressed proteins (q-value < 0.05; 992 downregulated, 516 upregulated in Patl2-/- oocytes). The levels of multiple RNA-binding proteins, some of which are proposed Patl2 interactors (Cpeb1, Eif4e1b), were found to be significantly reduced in Patl2-/- oocytes. Additionally, the protein products of several maternal effect genes (MEGs) implicated in mRNA regulation (Zar1, Igf2bp2) and cell cycle division (Tcl1a, Cdk1, Mos) were downregulated, while MEGs participating in epigenetic modifications (Zfp57, Trim28) were upregulated in the knockout group. Consistent with these observations, ST-PA treatment significantly increased AR (100%) and BR (75%) in the Patl2-/- oocytes in comparison to PA alone (AR = 75.95%, P = 0.0078; BR = 45.00%, P = 0.0128), effectively rescuing development to wild-type levels. Lastly, ST-PA treatment did not alter embryonic development in Patl2+/+ oocytes and produced outcomes comparable to PA alone, supporting the technical safety and applicability of the technique. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE massIVE partner repository with the dataset identifier MSV000100606. Patl2 -/- mice exhibit a less severe phenotype compared to patients carrying PATL2 variants. Patl2-/- female mice display a high MII rate without significant spindle abnormalities, which contrasts with a previously published report. Additionally, ST treatment was conducted using parthenogenetically activated oocytes, rather than biparental embryos. ST represents a promising treatment for PATL2-related female infertility in patients with MII oocytes, as it appears to restore cytoplasmic defects linked to abnormal RNA-binding proteins and MEGs identified by single-cell proteomics. In contrast, other proposed treatments for poor embryo development, such as assisted oocyte activation, is unlikely to be effective since Patl2-/- oocytes show a normal calcium response. This study was supported by the Special Research Fund (BOF) (starting grant BOF.STG.2021.0042.01 awarded to B.H.) and the Research Foundation-Flanders (FWO) (fellowship 1177425 N awarded to E.A.). B.H. has been receiving unrestricted educational funding from Ferring Pharmaceuticals (Aalst, Belgium). A.C.B., E.A., A.C., M.F-I-A., J.G., A.R., M.B., A.B., C.A., K.C.P., D.S., K.G., and F.V.M. have nothing to disclose. B.H. reports being board member of the Belgian Ethical Committee on embryo research.
Monoclonal antibodies (mAbs) are powerful tools for elucidating disease mechanisms. Capturing heterogeneity of patient responses-including clonality, somatic mutations, isotypes, and pathogenic potential-requires building large libraries of mAbs using high-throughput approaches. However, current techniques for identifying and isolating patient-derived mAbs targeting conformational epitopes on membrane proteins remain labor-intensive and inefficient. To address this challenge, we evaluated a cell-based optofluidic antibody discovery pipeline to generate patient-derived mAbs against myelin oligodendrocyte glycoprotein (MOG), a transmembrane autoantigen targeted in MOG antibody-associated disease (MOGAD). An optofluidic-based workflow incorporated mammalian display of human MOG (hMOG) in a live cell-based assay (CBA) format. B-cell receptor sequences from individual hMOG-binding B cells were cloned and expressed to generate mAbs from 3 patients. The hMOG binding specificity of these mAbs was validated in off-platform hMOG-CBAs. hMOG-specific antibody-secreting cells in one patient represented 0.02% of all single B cells screened. From these low-frequency populations, one patient-derived IgG mAb was successfully generated and validated. This IgG mAb, characterized by a high frequency of V-region somatic mutations (5%-12.2%), bound hMOG at concentrations as low as 1 ng/mL. This workflow enables rapid discovery of rare, patient-derived mAbs targeting conformational epitopes on membrane antigens, offering a scalable approach for dissecting autoantibody repertoires.
The TEM8 receptor (coded by ANTXR1) plays several roles in oncogenesis and novel oncolytic therapies, such as the SVV-01 virus, uniquely bind this protein in neuroendocrine tumor (NET) histologies, such as small-cell lung cancer (SCLC). Emerging pre-clinical data suggest that TEM8-targeting therapies may convert immunologically "cold" tumor microenvironments (TME) into "hot" milieu with greater responses to immune checkpoint inhibitors (ICIs). NextGen sequencing of DNA (592 genes or whole exome)/RNA (whole transcriptome) was performed on SCLC (N = 1404) and other NET (N = 1668) samples submitted to Caris Life Sciences (Phoenix, AZ). Samples were stratified by ANTXR1 expression quartiles (Q1 low, Q4 high). TME cell fractions were estimated by RNA deconvolution using quanTIseq. Real-world overall survival (OS) was assessed from insurance claim data. The landscape of pathogenic gene mutations was similar among ANTXR1 Q1 vs Q4 tumors among SCLC and NET cohorts. The TME of SCLC and NET Q4 tumors comprised a greater fraction of B cells and M1/M2 macrophages and were more frequently classified as 'T cell-inflamed' based on a transcriptional signature predictive of response to ICI. However, OS from the start of ICI was similar between ANTXR1 Q1 and Q4 cohorts. Increased B cell and M1/M2 macrophage infiltrate, along with T-cell inflamed status, associated with ANTXR1 Q4 TMEs suggest these patients with SCLC and NET may respond preferentially to ICI. A Phase 1 trial incorporating SVV-01 along with ICI is underway. Prospective investigation of molecular associations and clinical outcomes related to ANTXR1 expression in SCLC is warranted.
The role of 4-hydroxyphenylpyruvate dioxygenase-like protein (HPDL) in endometrial cancer (EC) progression remains poorly understood, particularly its involvement in metabolic-epigenetic crosstalk via lactate-driven histone lactylation. This study aimed to investigate HPDL's mechanistic contribution to EC pathogenesis. Stable HPDL-overexpressing and knockdown EC cell lines (HEC-1-B and AN3CA) were generated using lentiviral vectors. Functional assays (proliferation, migration, invasion), subcutaneous xenograft models in BALB/c nude mice, and molecular analyses were conducted. Lactate levels, Pan-lysine lactylation (pan-kla), histone H3K18 lactylation (H3K18la), and effects of sodium oxamate (lactate modulator) were assessed. Lactate Dehydrogenase A/Lactate Dehydrogenase B (LDHA/LDHB) knockdown, promoter activity assays, and chromatin immunoprecipitation (ChIP) were performed to evaluate H3K18la occupancy at LDHA/LDHB promoters. HPDL knockdown reduced intracellular lactate, Pan-Kla, and H3K18la levels, while overexpression elevated these markers. Sodium oxamate amplified lactate and lactylation in HPDL-overexpressing cells but suppressed histone lactylation independently of HPDL. LDHA/LDHB knockdown diminished lactylation, repressed HPDL expression, and inhibited promoter activity. ChIP revealed H3K18la enrichment at LDHA/LDHB promoters in HPDL-overexpressing cells and reduced occupancy in knockdown models. HPDL enhanced EC cell proliferation, migration, and invasion in vitro. In vivo, HPDL-overexpressing xenografts exhibited accelerated tumor growth and larger volumes compared to controls. HPDL regulates histone lactylation via LDHA/LDHB and promotes the proliferation of EC cells.
Lupus nephritis (LN) is a common, potentially fatal manifestation of systemic lupus erythematosus. We aimed to gain new insights into the immune responses underlying LN and their relation to the histologic heterogeneity observed in this disease, focusing on myeloid cells. We used single-cell RNA-sequencing (scRNA-seq) data of dissociated kidney samples from 156 patients with LN and 30 healthy individuals. We applied spatial transcriptomics (ST), utilising a gene panel designed to capture all myeloid subsets identified in the scRNA-seq data, to profile kidney samples acquired from 6 patients with LN and 2 controls. We generated a catalogue of the myeloid subsets found in LN kidneys. Our analyses indicated that an increase in irreversible tissue damage, as measured by the National Institutes of Health chronicity index (CI), is associated with a gradual switch of the local immune response from one dominated by monocytes and macrophages to one featuring expanded CD4 T, GZMK+ CD8 T, B, and dendritic cells, with a parallel decrease in the interferon response. In proliferative/mixed LN only, the degree of active inflammation correlates with the expansion of disease-specific macrophage (DMac) subsets, which later contract as the CI increases. Trajectory analysis of the scRNA-seq data suggested that DMacs arise from both infiltrating monocytes and tissue-resident macrophages; this was supported by the ST data, as well as cell cultures. DMacs are implied to interact with parietal epithelial cells, promoting the development of glomerulosclerosis. We suggest a detailed picture of the changes in the kidney immune mechanisms in LN as this disease progresses.
Angioimmunoblastic T-cell lymphoma (AITL) is a rare peripheral T-cell lymphoma characterized by diverse and aggressive clinical manifestations that frequently mimic autoimmune disorders. Cutaneous presentations are common and may lead to diagnostic delay. A 67-year-old woman first presented with non-blanchable skin rash, with normal platelet count and negative autoimmune markers. The rash improved with corticosteroids but recurred two months later, accompanied by fever, night sweats, limb edema, diarrhea, and cervical lymphadenopathy. Skin biopsy reported vasculitis and panniculitis, as direct immunofluorescence was compatible with IgA vasculitis. Subsequent laboratory tests revealed atypical lymphocytes, Coombs-positive anemia, thrombocytopenia, and detected Epstein-Barr virus DNA. Computed tomography showed new splenomegaly and periaortic lymphadenopathy. Lymph node biopsy confirmed AITL. Although CHOP chemotherapy was planned after staging, the patient rapidly deteriorated and died of septic shock. Serial peripheral blood flow cytometry at admission, post-splenectomy, and follow-up showed dynamic immunophenotypic changes: reductions in exhaustion and senescence markers as well as activated regulatory T cells after splenectomy; and later upregulation of exhaustion markers on naïve T cells. This case illustrates the misleading presentation as immunoglobulin A (IgA) vasculitis and rapid progression of AITL. While vasculitis is accompanied by cytopenia, lymphadenopathy, or aggressive clinical course, early lymph node biopsy is essential for timely diagnosis.
To describe a case of cancer-associated retinopathy (CAR) after chimeric antigen receptor T (CART)-cell therapy. Retrospective case report. A 71-year-old man with follicular lymphoma refractory to multiple chemotherapies underwent CART-cell therapy. Two weeks after CART initiation, he presented with rapid-onset decreased peripheral vision in both eyes. Fluorescein angiography showed attenuation of peripheral vessel filling, and optical coherence tomography showed loss of ellipsoid zone with foveal sparing. Electroretinography revealed nondetectable rod responses and severely prolonged and diminished cone responses. Antiretinal antibody testing showed positive antienolase and antiheat shock protein 60 antibodies on Western blot with strong staining of the outer segments on immunohistochemistry. The patient was diagnosed with CAR and is currently on monthly IV immunoglobulin (IVIg) with stable central visual acuity. This patient developed CAR immediately after receiving CART-cell therapy, which depletes B cells. This suggests that CAR pathophysiology may not be purely autoantibody mediated as typically believed.
Type I interferon response, specifically, the cGAS-cGAMP-STING axis that results in IFN-β response, is well known for its complex roles early during viral infection. Previous reports suggest that HSV-1 DNA in Thp-1 cells and HIV-2 dsDNA in DCs and macrophages could be sensed by cGAS. The nuclear DNA sensor IFI16's viral DNA sensing leads to its acetylation, cytoplasmic translocation and STING activation and inflammasome activation. Although cGAS is known to be associated with IFI16 in the nucleus, however, during HSV-2 infection, the role of nuclear cGAS in viral DNA sensing, inflammasome formation and type I IFN response remains unknown. In the current study, extensive investigation of the complex IFN-β responses elicited early during de novo HSV-2 infections in HFF cells is undertaken. The SiIFI16 and SicGAS treated HFF cells infected with HSV-2 demonstrate that cGAS senses nuclear herpes-viral DNA in an IFI16 dependent manner leading to nuclear cGAMP production. These results unravel a novel nuclear cooperative role of cGAS and IFI16 and extend the cGAS DNA sensing and its enzymatic activity in the nucleus. IFI16 acetylation required for inflammasome complex formation is cGAS independent. The cGAS-pro-Caspase1 and cGAS-ASC interaction suggests plausible role of cGAS in inflammasome complex for Caspase-1 activation. The activated Caspase-1 interaction with cGAS was also observed. Further, the autophagy and DNA damage responses elicited during de novo HSV-2 infection are suggested. The crosstalk of the type I interferon pathway with the inflammasome, autophagy and DNA damage response pathways suggests an intricate mechanism of inter-regulation at different stages and time points during infection, that might orchestrate a balanced and efficient immune response or facilitate viral immune evasion. Unique and dynamic post translational modifications of cGAS, namely acetylation and K-63 poly-ubiquitination, are observed, and are plausibly involved in cGAS regulation during HSV-2 infection.
Granular cell tumor (GCT) of the neurohypophysis is a rare tumor originating from the posterior pituitary/infundibulum, classified as WHO grade I. Due to its imaging characteristics being highly similar to those of common sellar lesions, diagnosis typically relies on histopathology and immunohistochemistry. A 43-year-old male presented with a one-month history of pulsatile temporal headaches without significant visual complaints. Endocrine tests showed reduced levels of growth hormone and prolactin. CT scan of the head and MRI of the sellar region revealed a well-defined solid mass in the sellar/suprasellar region (approximately 20×19×21 mm), compressing the optic chiasm and closely associated with the pituitary stalk. The patient underwent endoscopic transnasal transsphenoidal tumor resection. Intraoperatively, the tumor was soft but highly vascular, and adhered to the pituitary stalk, requiring meticulous hemostasis and en bloc removal. Postoperative MRI confirmed total resection. Histologically, the tumor consisted of polygonal to spindle-shaped cells with abundant eosinophilic granules in the cytoplasm. Immunohistochemistry showed positivity for TTF-1, S100, and CD68, with a Ki-67 index of approximately 5%, while SOX10 was negative, supporting the diagnosis of neurohypophyseal GCT. We reviewed 88 published case reports to compare demographic characteristics, clinical presentations, endocrine abnormalities, treatment strategies, and recurrence rates. Neurohypophyseal GCT should be included in the differential diagnosis of solid sellar/suprasellar masses associated with the pituitary stalk. Endoscopic transnasal transsphenoidal resection is effective, but the rich vascular supply and adhesion to the pituitary stalk can increase the surgical difficulty. Due to the potential for late recurrence, long-term follow-up is recommended.
Lung cancer resection is curative but associated with postoperative morbidity and mortality. This study evaluated whether elevated blood eosinophil count (BEC) was associated with postoperative outcomes in early-stage lung cancer. This was a retrospective cohort study of consecutive adult patients undergoing lung resection for stage I and II non-small cell lung cancer in a large tertiary referral center from September 2017 to June 2021. Data were drawn from the institution's Data Warehouse. The primary outcome was 90-day healthcare utilization defined as emergency department visit or hospital readmission. Secondary outcomes were postoperative complications, index hospitalization length of stay, and 1-year survival. Preoperative 90-day BEC was categorized by a threshold of 200 cells/µL. Covariates were age, sex, smoking status, Charlson Comorbidity Index, chronic obstructive pulmonary disease (COPD), asthma, tumor size, nodal status, surgical approach, and blood results (white blood cells, hemoglobin, and creatinine). The main analyses were validated by a second international cohort. Log-Poisson with robust variance estimation and Cox proportional hazards regression models were fit for primary and secondary outcomes. Analyses were replicated for BEC thresholds of 150 and 300 cells/µL. Among 715 patients undergoing lung resection (median age = 69 years, 42% male, 29% with COPD), 146 patients (20%) had high preoperative BEC ≥ 200 cells/µL. BEC ≥ 200 cells/µL was associated with a higher rate of 90-day healthcare utilization:19% vs. 14% for BEC < 200 cells/µL. This association remained after adjustment (Risk Ratio [RR], 1.52; 95% Confidence Interval [CI], 1.02-2.25) and the validation cohort (RR, 2.23; 95% CI, 1.06-4.69). BEC as a continuous measure was also associated with the primary outcome in both cohorts: RR, 2.15 (95% CI, 1.49-3.12) and RR, 1.42 (95% CI, 1.10-1.94), respectively. BEC ≥ 200 cells/µL was associated with higher probability of death 1 year post-surgery (adjusted Hazard Ratio, 2.41; 95% CI, 1.08-5.35). There was no difference in the risk of postoperative pulmonary complications between high and low BEC (RR, 0.86; 95% CI, 0.58-1.27). Elevated preoperative BEC was associated with higher risk of postoperative healthcare utilization and lower 1-year survival after lung cancer resection among patients with or without respiratory disease.
Ovarian clear cell carcinoma (OCCC) frequently exhibits resistance to conventional chemotherapy with limited treatment options. This study reports a heavily pretreated advanced case with liver metastases to investigate the efficacy of gemcitabine combined with PD-1 inhibitor in such refractory patients. We present a case of stage IIIC OCCC that progressed after first-line platinum-based chemotherapy. The patient was treated with gemcitabine incombination with the PD-1 inhibitor toripalimab. Genetic alterations were analyzed using next-generation sequencing, and PD-L1 expression was assessed by immunohistochemistry (Combined Positive Score, CPS). Treatment response andsafety profiles were regularly monitored. The combination therapy induced a complete remission lasting over 24 months, with a manageable safety profile. Molecular profiling revealed a loss-of-function mutation in ARID1A, which may enhance tumor sensitivity to gemcitabine, while elevated PD-L1 expression (CPS = 30) may serve as a predictive biomarker for immunotherapy response in OCCC. Notably, the deep and durable clinical response observed suggests a potential synergistic effect between gemcitabine and immunotherapy, wherein ARID1A deficiency-induced chemosensitivity may prime the tumor microenvironment for enhanced immune checkpoint inhibition, offering a compelling rationale for this combination strategy in platinum-resistant OCCC. This study proposes a promising therapeutic strategy for advanced OCCC and supports the clinical value of molecular profiling-guided treatment. Both ARID1A status and PD-L1 expression warrant further validation as potential biomarkers inprospective clinical trials.
Pancreatic cystic lesions (PCLs) range from benign to malignant, creating diagnostic and therapeutic challenges. While most PCLs are serous or mucinous neoplasms, rare entities such as acinar cell cystadenoma (ACC) remain poorly characterized. This study reports a tertiary center cohort and a systematic literature review. We retrospectively analyzed 23 ACC patients at Karolinska University Hospital. A systematic review was performed following PRISMA guidelines. In our cohort (median age 67.0 years; 73.9% male), ACCs were most often incidentally detected, with abdominal pain being the most frequent indication for imaging. No patients underwent surgery, and no malignant transformation was observed during a median follow-up of 10.4 months. Most patients (87%) underwent both CT and MR imaging, and 65.2% fulfilled proposed imaging diagnostic criteria.The systematic review included 41 studies with 165 patients. Abdominal pain was the most common indication for imaging, and most patients underwent pancreatic surgery. Median follow-up was 1.3 years, with no malignant transformation observed. Imaging typically showed well-circumscribed cystic lesions, usually non-communicating with the main pancreatic duct and mimicking other cystic neoplasms. Histopathology consistently demonstrated an acinar phenotype with low proliferative activity. ACC appears to be an asymptomatic, incidentally discovered lesion, with imaging playing a central diagnostic role. Neither endocrine nor exocrine pancreatic insufficiency nor malignant transformation was observed. However, the relatively short overall follow-up limits conclusions regarding long-term outcomes.
In children affected by allergic proctocolitis, impaired weight gain and developmental failure to thrive can be a prominent manifestation. However, it remains unclear whether the association between White Blood Cell (WBC) excretion in these patients and the occurrence of food allergies is associated with subsequent impaired weight gain after the start of complementary feeding. In this prospective cohort study, 85 children with allergic proctocolitis were followed from birth to 1 year of age regarding continued WBC excretion, weight gain, and the presence or absence of allergic symptoms. Also, the weight gain of children was examined and compared between those with continued leukocyte excretion and those without. Data were analyzed using SPSS statistical software version 26. The age distribution of infants in the two groups with and without persistent WBC excretion did not differ significantly (p =0.35). However, the gender distribution was significantly different, with persistent excretion higher in boys (p =0.023). Examination of standardized weight changes showed no significant difference between the two groups, with and without persistent WBC excretion (p >0.05). Persistent leukocyte excretion did not affect weight gain, consistent with reports that inflammatory stool markers have limited predictive value in allergic proctocolitis. Future studies with larger samples and refined inflammatory biomarkers are needed to clarify the clinical significance and potential long-term allergic implications of persistent intestinal inflammation. Persistent WBC excretion in children with allergic proctocolitis was observed in some of the patients, but was not associated with impaired weight gain during the first year of life.
To determine whether baseline epigenomic and transcriptional profiles of human dental pulp stem cells (DPSCs) differ by sex, and to assess sex dependence of key markers related to growth and stemness. Primary DPSCs from nine young adult donors were isolated and cultured under standard growth conditions. Genome-wide DNA methylation was screened by Illumina Infinium Methylation EPIC v2.0. Bulk RNA sequencing was performed with poly A selected libraries on an Illumina NovaSeq X Plus sequencer. Additionally, a targeted RT-qPCR array covering canonical proliferation, stemness (multi- and pluri-potency, neural crest markers), MSC identity, and WNT and NOTCH signaling markers was tested. Global differences between male- and female-derived samples were predominantly confined to the sex chromosomes. DNA methylation profiling revealed sex-dependent patterns, with mild autosomal hypermethylation in males and marked hypermethylation of the X chromosome in females. This pattern was accompanied by female-specific expression of XIST, supporting X-chromosome inactivation as a major contributor to the observed sex-chromosome signal. Autosomal methylation differences were generally modest (|Δβ| < 0.2). At the transcriptomic level, only a small fraction of genes (52 of 17,204) was differentially expressed, with most mapping to the sex chromosomes. Consistently, the RT-qPCR array indicated minimal differences between male and female DPSCs, with no consistent sex dependence across targets except for CCNE1. Under basal culture conditions, DNA methylation and transcriptional profiles in DPSCs are largely similar between sexes. The limited differential signal is associated predominantly with sex chromosomes, while autosomal effects are few and modest. These findings provide an initial molecular baseline and motivate larger and context specific studies.
There is a lack of clinical data on hematopoietic stem cell transplantation (HSCT) in Ataxia-Telangiectasia (A-T) patients due to the underlying chromosomal instability that leads to low tolerance to chemotherapy. To effectively manage cancer and immune risks, there is a need for improved HSCT protocols, novel therapies, and long-term monitoring. This report describes a 16-year-old boy with A-T and T-ALL who achieved long-term leukemia-free survival after HSCT using a tailored, drug-monitored conditioning regimen. His results were analyzed in the context of a systematic review of the literature on HSCT outcomes in A-T patients. A thorough literature review was conducted using a comprehensive search of the PubMed, Scopus, and Google Scholar databases. The search was limited to studies published between September 1, 2000, and September 1, 2025. Eligible studies were required to involve human participants and to include at least one patient with a confirmed diagnosis of A-T, with transplantation interventions. The analysis included 16 A-T patients, including our patient, who underwent HSCT. The median age at transplantation was 48 months (interquartile range [IQR]: 22-142 months). Myeloablative conditioning (MAC) was administered to two patients, both of whom died. Reduced-intensity conditioning (RIC) was utilized for nine patients, with three deaths (33.3%). Reduced-toxicity conditioning (RTC) was administered in two patients, with one patient experiencing a fatal outcome. In total, eight patients (50%) experienced significant drug-related toxicities, eight (50%) had GvHD and only eight patients (50%) survived. Our patient underwent HSCT of a matched sibling donor after administration of adjusted treosulfan doses (cumulative AUC of 4671 mg/Lxh), and achieved leukemia-free survival with complete hematological and normalized thymic function without graft-versus-host disease (GvHD). Despite the historically poor survival outcomes observed in transplanted A-T patients, new HSCT strategies, such as treosulfan therapeutic drug monitoring and personalized drug profiles to select potent but less toxic agents, warrant reevaluation to achieve durable remission in leukemia and lymphoma. These findings underscore the necessity to persist in the development of innovative HSCT approaches with the objective of expanding therapeutic options for both malignancies and combined immunodeficiency.