A single-cell RNA expression atlas of normal, preneoplastic and tumorigenic states in the human breast.

To examine global changes in breast heterogeneity across different states, we determined the single-cell transcriptomes of > 340,000 cells encompassing normal breast, preneoplastic BRCA1+/- tissue, the major breast cancer subtypes, and pairs of tumors and involved lymph nodes. Elucidation of the normal breast microenvironment revealed striking changes in the stroma of post-menopausal women. Single-cell profiling of 34 treatment-naive primary tumors, including estrogen receptor (ER)+ , HER2+ , and triple-negative breast cancers, revealed comparable diversity among cancer cells and a discrete subset of cycling cells. The transcriptomes of preneoplastic BRCA1+/- tissue versus tumors highlighted global changes in the immune microenvironment. Within the tumor immune landscape, proliferative CD8+ T cells characterized triple-negative and HER2+ cancers but not ER+ tumors, while all subtypes comprised cycling tumor-associated macrophages, thus invoking potentially different immunotherapy targets. Copy number analysis of paired ER+ tumors and lymph nodes indicated seeding by genetically distinct clones or mass migration of primary tumor cells into axillary lymph nodes. This large-scale integration of patient samples provides a high-resolution map of cell diversity in normal and cancerous human breast.

The role of podocytes in lupus nephritis: Insights and implications.

Lupus nephritis (LN) is a severe complication of systemic lupus erythematosus, with high mortality rates despite medical advancements. The complexity of its pathogenesis, including the pivotal role of podocytes - kidney-localized cells - remains a challenge, lacking effective treatments and biomarkers. Recent studies highlight the significant contribution of these cells to LN's development, particularly through their immune-related functions and interaction with other kidney cells. This new understanding opens possibilities for targeted therapies aimed at these cellular mechanisms. This review aims to summarize these recent developments, shedding light on the intricate involvement of podocytes in LN and potential avenues for innovative treatments.

A novel cryopreservation and biobanking strategy to study lymphoid tissue stromal cells in human disease.

Nonhematopoietic lymph node stromal cells (LNSCs) regulate lymphocyte trafficking, survival, and function for key roles in host defense, autoimmunity, alloimmunity, and lymphoproliferative disorders. However, the study of LNSCs in human diseases is complicated by a dependence on viable lymphoid tissues, which are most often excised prior to establishment of a specific diagnosis. Here, we demonstrate that cryopreservation can be used to bank lymphoid tissue for the study of LNSCs in human disease. Using human tonsils and lymph nodes (LN), lymphoid tissue fragments were cryopreserved for subsequent enzymatic digestion and recovery of viable nonhematopoietic cells. Flow cytometry and single-cell transcriptomics identified comparable proportions of LN stromal cell types in fresh and cryopreserved tissue. Moreover, cryopreservation had little effect on transcriptional profiles, which showed significant overlap between tonsils and LN. The presence and spatial distribution of transcriptionally defined cell types were confirmed by in situ analyses. Our broadly applicable approach promises to greatly enable research into the roles of LNSCs in human disease.

Flexible Anionic Groups-Activated Structure Dissymmetry for Strong Nonlinearity in Ln2 Ae3 MIV 3 S12 Family.

Structural dissymmetry and strong second-harmonic generation (SHG) responses are key conditions for nonlinear optical (NLO) crystals, and targeted combinatorial screening of suitable anionic groups has become extremely effective. Herein, optimal combination of flexible SnSn (n = 5, 6) groups and highly electropositive cations (lanthanides (Ln3+ ) and alkaline earth (Ae2+ : Sr, Ca) metals) affords the successful synthesis of 12 NLO thiostannates including Ln2 Sr3 Sn3 S12 (Pmc21 ) and Ln2 Ca3 Sn3 S12 (P-62m); whereas 17 rigid GeS4 or SiS4 tetrahedra-constructed Ln2 Ae3 Ge3 S12 and Ln2 Ae3 Si3 S12 crystallize in the centrosymmetric (CS) Pnma. This unprecedented CS to noncentrosymmetric (NCS) structural transformation (Pnma to P-62m to Pmc21 ) in the Ln2 Ae3 MIV 3 S12 family indicates that chemical substitution of the tetrahedral GeS4 /SiS4 units with SnSn breaks the original symmetry to form the requisite NCS structures. Remarkably, strong polarization anisotropy and hyperpolarizability of the Sn(4+) S5 unit afford huge performance improvement from the nonphase-matching (NPM) SHG response (1.4 × AgGaS2 and Δn = 0.008) of La2 Ca3 Sn3 S12 to the strong phase-matching (PM) SHG effect (3.0 × AgGaS2 and Δn = 0.086) of La2 Sr3 Sn3 S12 . Therefore, Sn(4+) S5 is proven to be a promising "NLO-active unit." This study verifies that the coupling of flexible SnSn building blocks into structures opens a feasible path for designing targeted NCS crystals with strong nonlinearity and optical anisotropy.

Can we predict kidney involvement in patients with systemic lupus erythematosus? A retrospective cohort study with independent validation.

OBJECTIVES: To discern predictive factors for incident kidney involvement in patients with systemic lupus erythematosus (SLE). METHODS: Patients with SLE from the 'Attikon' Lupus cohort were monitored for lupus nephritis (LN), defined by kidney histology and/or classification criteria. Demographic and clinical characteristics at baseline were compared against patients who did not develop LN. LN-free survival curves were generated by Kaplan-Meier. A multivariate Cox proportional hazards model was used to identify independent predictors of LN. Independent validation was performed in the University of Crete Lupus registry. RESULTS: Among the 570 patients in the derivation cohort, 59 exhibited LN as their initial presentation, while an additional 66 developed LN during the follow-up period (collectively, 21.9% incidence). In the latter group, baseline factors predictive of subsequent kidney involvement were male sex (multivariable-adjusted [a]HR 4.31, 95% CI: 1.82-10.2), age of SLE diagnosis below 26 years (aHR 3.71, 95% CI: 1.84-7.48), high anti-dsDNA titre (aHR 2.48, 95% CI: 1.03-5.97) and low C3 and/or C4 (although not statistically significant, aHR 2.24, 95% CI: 0.83-6.05, p= 0.11). A combination of these factors at time of diagnosis conferred an almost 90-fold risk compared with serologically inactive, older, female patients (aHR 88.77, 95% CI : 18.75-420.41), signifying a very high-risk group. Independent validation in the Crete Lupus registry showed concordant results with the original cohort. CONCLUSION: Male sex, younger age and serologic activity at SLE diagnosis are strongly associated with subsequent kidney involvement. Vigilant surveillance and consideration of early use of disease-modifying drugs is warranted in these subsets of patients.

Subtherapeutic hydroxychloroquine concentration is associated with increased disease activity and greater organ damage in lupus.

OBJECTIVE: To investigate the effects of the serum HCQ concentration on clinical manifestations, disease activity and organ damage in a longitudinal cohort of SLE patients. METHODS: The 338 SLE patients were assessed with respect to their demographic data, clinical and laboratory findings, Physician's Global Assessment (PGA), adjusted mean SLEDAI-2000 (AMS) and SLICC Damage Index (SDI) annually for 5 consecutive years. Patients were divided into two groups according to their serum HCQ concentration at baseline: subtherapeutic (<500 ng/ml) and therapeutic (≥500 ng/ml) groups. The impact of the HCQ concentration on the clinical outcomes was evaluated in a longitudinal analysis using a generalized estimating equation (GEE). RESULTS: Of the 338 patients, 287 (84.9%) were in the subtherapeutic group at baseline. This group had a higher incidence of newly developed LN (P = 0.036) and had been prescribed higher mean and cumulative doses of prednisolone (P = 0.003 and P = 0.013, respectively) than the therapeutic group. In multivariable analyses based on GEE, the subtherapeutic group had a higher AMS score (ß = 1.398, 95% CI 0.607, 2.189; P < 0.001), higher PGA score (ß = 0.328, 95% CI 0.215, 0.441; P < 0.001) and higher SDI score (ß = 0.366, 95% CI 0.061, 0.671; P = 0.019) across all 5 years. CONCLUSION: The subtherapeutic HCQ concentration was associated with the development of new-onset LN, and had significant associations with disease activity and cumulative organ damage in SLE patients over time.

Disruption of memory B-cell trafficking by belimumab in patients with systemic lupus erythematosus.

OBJECTIVES: Autoreactive memory B cells (MBCs) contribute to chronic and progressive courses in autoimmune diseases like SLE. The efficacy of belimumab (BEL), the first approved biologic treatment for SLE and LN, is generally attributed to depletion of activated naïve B cells and inhibition of B-cell activation. BEL's effect on MBCs is currently unexplained. We performed an in-depth cellular and transcriptomic analysis of BEL's impact on the blood MBC compartment in patients with SLE. METHODS: A retrospective meta-analysis was conducted, pooling flow cytometry data from four randomized trials involving 1245 patients with SLE treated with intravenous BEL or placebo. Then, extensive MBC phenotyping was performed using high-sensitivity flow cytometry in patients with mild/moderate SLE and severe SLE/LN treated with subcutaneous BEL. Finally, transcriptomic characterization of surging MBCs was performed by single-cell RNA sequencing. RESULTS: In BEL-treated patients, a significant increase in circulating MBCs, in a broad range of MBC subsets, was established at week 2, gradually returning to baseline by week 52. The increase was most prominent in patients with higher SLE disease activity, serologically active patients and patients aged ≤18 years. MBCs had a non-proliferating phenotype with a prominent decrease in activation status and downregulation of numerous migration genes. CONCLUSION: Upon BEL initiation, an increase of MBCs was firmly established. In the small cohort investigated, circulating MBCs were de-activated, non-proliferative and demonstrated characteristics of disrupted lymphocyte trafficking, expanding on our understanding of the therapeutic mechanism of B-cell-activating factor inhibition by BEL. TRIAL REGISTRATION: ClinicalTrials.gov, http://clinicaltrials.gov, NCT00071487, NCT00410384, NCT01632241, NCT01649765, NCT03312907, NCT03747159.

Interstitial fibrosis increases the risk of end-stage kidney disease in patients with lupus nephritis.

OBJECTIVE: To evaluate the risk of end-stage kidney disease (ESKD) in LN patients using tubulointerstitial lesion scores. METHODS: Clinical profiles and histopathological presentations of 151 biopsy-proven LN patients were retrospectively examined. Risk factors of ESKD based on characteristics and scoring of their tubulointerstitial lesions [e.g. interstitial inflammation (II), tubular atrophy (TA) and interstitial fibrosis (IF)] were analysed. RESULTS: The mean age of 151 LN patients was 36 years old, and 136 (90.1%) were female. The LN cases examined included: class I/II (n = 3, 2%), class III/IV (n = 119, 78.8%), class V (n = 23, 15.2%) and class VI (n = 6, 4.0%). The mean serum creatinine level was 1.4 mg/dl. Tubulointerstitial lesions were recorded in 120 (79.5%) patients. Prior to receiving renal biopsy, nine (6.0%) patients developed ESKD. During the follow-up period (mean, 58 months), an additional 47 patients (31.1%) progressed to ESKD. Multivariate analyses identified serum creatinine [hazard ratio (HR): 1.7, 95% CI: 1.42-2.03, P < 0.001] and IF (HR: 3.2, 95% CI: 1.58-6.49, P = 0.001) as independent risk factors of ESKD. Kaplan-Meier analysis further confirmed a heightened risk of ESKD associated with IF. CONCLUSION: Tubulointerstitial involvement is commonly observed in the histopathological presentation of LN. However, IF, rather than II or TA, was found to increase the risk of ESKD in our cohort. Therefore, to predict renal outcome in LN patients prior to adjusting immunosuppressive treatment, the degree of IF should be reviewed.

Belimumab and antimalarials combined against renal flares in patients treated for extra-renal systemic lupus erythematosus: results from 4 phase III clinical trials.

OBJECTIVES: To determine the effect of antimalarial agents (AMA) and different doses and pharmaceutical forms of belimumab on preventing renal flares in patients with SLE treated for extra-renal disease. METHODS: We pooled data from the BLISS-52, BLISS-76, BLISS-SC and BLISS-Northeast Asia trials of belimumab (n = 3225), that included patients with active SLE yet no severe ongoing nephritis. Participants were allocated to receive intravenous belimumab 1 mg/kg, intravenous belimumab 10 mg/kg, subcutaneous belimumab 200 mg, or placebo in addition to standard therapy. We estimated hazards of renal flare development throughout the study follow-up (52-76 weeks) using Cox regression analysis. RESULTS: In total, 192 patients developed a renal flare after a median of 197 days. Compared with placebo, the risk of renal flares was lower among patients receiving intravenous belimumab 10 mg/kg (HR: 0.62; 95% CI: 0.41, 0.92; P = 0.018) and intravenous belimumab 1 mg/kg (HR: 0.42; 95% CI: 0.22, 0.79; P = 0.007), while no significant association was found for subcutaneous belimumab 200 mg. AMA use yielded a lower hazard of renal flares (HR: 0.66; 95% CI: 0.55, 0.78; P < 0.001). The protection conferred was enhanced when belimumab and AMA were co-administered; the lowest flare rate was observed for the combination intravenous belimumab 1 mg/kg and AMA (18.5 cases per 1000 person-years). CONCLUSIONS: The protection conferred from belimumab against renal flare development in patients treated for extra-renal SLE appears enhanced when belimumab was administered along with AMA. The prominent effect of low-dose belimumab warrants investigation of the efficacy of intermediate belimumab doses. CLINICAL TRIAL IDENTIFICATION: BLISS-52: NCT00424476; BLISS-76: NCT00410384; BLISS-SC: NCT01484496; BLISS-NEA: NCT01345253.

Flow-cytometry Assessment of DNA content and Immunophenotyping of Immune-cells in Lymph-node-specimens as a Potential Diagnostic Signature of Aggressiveness in B-Non-Hodgkin Lymphomas.

Flow-cytometry (FC) is a powerful tool that can assist in lymphoma diagnosis in lymph node (LN) specimens. Although lymphoma diagnosis and classification are mainly based on tumor cell characteristics, surrounding cells are less employed in this process. We retrospectively investigated alterations in the ploidy status, proliferative cell fraction (PF) and the percentages of surrounding immune cells in 62 consecutive LN specimens with B-Cell Non-Hodgkin Lymphoma (B-NHL) that were submitted for FC evaluation between 2019-2022. Compared with indolent B-NHLs, aggressive B-NHLs show increased DNA aneuploidy and PF, increased monocytes, immature-granulocytes, mature granulocytes, CD8+ T-cells, Double-Negative-T-cells and Double-Positive-T-cells, and decreased total CD45+ cells, total lymphocytes, CD4+ T-cells and CD4/CD8 ratio. Receiver operating characteristic analysis determined PF > 6.8% and immature-granulocytes > 0.9% as optimal cutoffs with highest specificity and sensitivity in differentiating aggressive and indolent B-NHLs. These findings further strength the diagnostic value of DNA content analysis by FC and suggest the utilization of tumor surrounding immune cells in NHL diagnosis and classification.

Inhibitory effects of Gracilaria edulis and Gracilaria salicornia against the MGMT and VEGFA biomarkers involved in the onset and advancement of glioblastoma using in silico and in vitro approaches.

Glioblastoma (GBM), an aggressive primary brain tumor originating from glial cells, poses significant treatment challenges due to its rapid growth and invasiveness. The exact mechanisms of GBM's brain damage remain unclear. This study examines primary molecular markers commonly assessed in GBM patients, including brain-derived neurotrophic factor (BDNF), platelet-derived growth factor receptor A (PDGFRA), O6-methylguanine DNA methyltransferase (MGMT), epidermal growth factor receptor (EGFR), and vascular endothelial growth factor A (VEGFA) using computational approaches. The study revealed significant differences (p ≤ 0.05) in PDGFRA, EGFR, and VEGFA expression rates, which are particularly interesting. Additionally, MGMT and VEGFA showed higher hazard ratios. Expression levels of MGMT and VEGFA were visualized in immune and malignant cells using single-cell RNA datasets GSE103224 and GSE148842. From a total of 48 compounds in Gracilaria edulis and 86 in Gracilaria salicornia, we identified 15 compounds capable of crossing the blood-brain barrier. Notably, 2-tridecanone (binding affinity [BA] = -4.2 kcal/mol; root mean square deviation [RMSD] = 1.479 Å) and decanoic acid, ethyl ester (BA = -4.2 kcal/mol; RMSD = 1.702 Å) from G. edulis interacted with MGMT via hydrogen bonds. The compound alpha-terpineol interacted with MGMT (BA = -5.7 kcal/mol; RMSD = 0.501 Å) and VEGFA (BA = -4.7 kcal/mol; RMSD = 2.483 Å). Ethanolic and methanolic extracts from G. edulis and G. salicornia demonstrated mild anti-cell proliferation properties in the GBM LN-229 cell line, suggesting potential therapeutic benefits. This study highlights the significance of molecular markers and natural compounds in understanding and potentially treating GBM.

Metabolite characterisation of the peptide-drug conjugate LN005 in liver S9s by UHPLC-Orbitrap-HRMS.

LN005 is a peptide-drug conjugate (PDC) targeting glucose-regulated protein 78 (GRP78) to treat several types of cancer, such as breast, colon, and prostate cancer.As a new drug modality, understanding its metabolism and elimination pathways will help us to have a whole picture of it. Currently, there are no metabolic studies on LN005; therefore, this study aimed to investigate the metabolism of LN005, clarify its metabolic profile in the liver S9s of different species, and identify the major metabolic pathways and differences between species.The incubation samples were measured by ultra-high performance liquid chromatography combined with orbitrap tandem mass spectrometry (UHPLC-Orbitrap-HRMS).The results showed that LN005 was metabolised by liver S9s, and four metabolites were identified. The main metabolic pathway of LN005 in liver S9s was oxidative deamination to ketone or hydrolysis. Similar metabolic profiles were observed in mouse, rat, dog, monkey, and human liver S9s, indicating no differences between these four animal species and humans.This study provides information for the structural modification and optimisation of LN005 and affords a reference for subsequent animal experiments and human metabolism of other PDCs.

Cryopreserved leukapheresis material can be transferred from controlled rate freezers to ultracold storage at warmer temperatures without affecting downstream CAR-T cell culture performance and in-vitro functionality.

Chimeric antigen receptor (CAR) T-cell therapies are increasingly adopted as a commercially available treatment for hematologic and solid tumor cancers. As CAR-T therapies reach more patients globally, the cryopreservation and banking of patients' leukapheresis materials is becoming imperative to accommodate intra/inter-national shipping logistical delays and provide greater manufacturing flexibility. This study aims to determine the optimal temperature range for transferring cryopreserved leukapheresis materials from two distinct types of controlled rate freezing systems, Liquid Nitrogen (LN2)-based and LN2-free Conduction Cooling-based, to the ultracold LN2 storage freezer (≤-135 °C), and its impact on CAR T-cell production and functionality. Presented findings demonstrate that there is no significant influence on CAR T-cell expansion, differentiation, or downstream in-vitro function when employing a transfer temperature range spanning from -30 °C to -80 °C for the LN2-based controlled rate freezers as well as for conduction cooling controlled rate freezers. Notably, CAR T-cells generated from cryopreserved leukapheresis materials using the conduction cooling controlled rate freezer exhibited suboptimal performance in certain donors at transfer temperatures lower than -60 °C, possibly due to the reduced cooling rate of lower than 1 °C/min and extended dwelling time needed to reach the final temperatures within these systems. This cohort of data suggests that there is a low risk to transfer cryopreserved leukapheresis materials at higher temperatures (between -30 °C and -60 °C) with good functional recovery using either controlled cooling system, and the cryopreserved materials are suitable to use as the starting material for autologous CAR T-cell therapies.

Photophysical Characterization and In Vitro Evaluation of α-Mangostin-Loaded HDL Mimetic Nano-Complex in LN-229 Glioblastoma Spheroid Model.

Cytotoxic activity has been reported for the xanthone α-mangostin (AMN) against Glioblastoma multiforme (GBM), an aggressive malignant brain cancer with a poor prognosis. Recognizing that AMN's high degree of hydrophobicity is likely to limit its systemic administration, we formulated AMN using reconstituted high-density lipoprotein (rHDL) nanoparticles. The photophysical characteristics of the formulation, including fluorescence lifetime and steady-state anisotropy, indicated that AMN was successfully incorporated into the rHDL nanoparticles. To our knowledge, this is the first report on the fluorescent characteristics of AMN with an HDL-based drug carrier. Cytotoxicity studies in a 2D culture and 3D spheroid model of LN-229 GBM cells and normal human astrocytes showed an enhanced therapeutic index with the rHDL-AMN formulation compared to the unincorporated AMN and Temozolomide, a standard GBM chemotherapy agent. Furthermore, treatment with the rHDL-AMN facilitated a dose-dependent upregulation of autophagy and reactive oxygen species generation to a greater extent in LN-229 cells compared to astrocytes, indicating the reduced off-target toxicity of this novel formulation. These studies indicate the potential therapeutic benefits to GBM patients via selective targeting using the rHDL-AMN formulation.

Lupus Nephritis from Pathogenesis to New Therapies: An Update.

Lupus Nephritis (LN) still represents one of the most severe complications of Systemic Lupus Erythematosus (SLE) and a major risk factor for morbidity and mortality. However, over the last few years, several studies have paved the way for a deeper understanding of its pathogenetic mechanisms and more targeted treatments. This review aims to provide a comprehensive update on progress on several key aspects in this setting: pathogenetic mechanisms of LN, including new insight into the role of autoantibodies, complement, vitamin D deficiency, and interaction between infiltrating immune cells and kidney resident ones; the evolving role of renal biopsy and biomarkers, which may integrate information from renal histology; newly approved drugs such as voclosporin (VOC) and belimumab (BEL), allowing a more articulate strategy for induction therapy, and other promising phase III-immunosuppressive (IS) agents in the pipeline. Several adjunctive treatments aimed at reducing cardiovascular risk and progression of chronic renal damage, such as antiproteinuric agents, represent an important complement to IS therapy. Furthermore, non-pharmacological measures concerning general lifestyle and diet should also be adopted when managing LN. Integrating these therapeutic areas requires an effort towards a holistic and multidisciplinary approach. At the same time, the availability of an increasingly wider armamentarium may translate into improvements in patient's renal outcomes over the next decades.

Physicochemical Characterization and Antimicrobial Properties of Lanthanide Nitrates in Dilute Aqueous Solutions.

This work presents the results of studying dilute aqueous solutions of commercial Ln(NO3)3 · xH2O salts with Ln = Ce-Lu using X-ray diffraction (XRD), IR spectroscopy, X-ray absorption spectroscopy (XAS: EXAFS/XANES), and pH measurements. As a reference point, XRD and XAS measurements for characterized Ln(NO3)3 · xH2O microcrystalline powder samples were performed. The local structure of Ln-nitrate complexes in 20 mM Ln(NO3)3 · xH2O aqueous solution was studied under total external reflection conditions and EXAFS geometry was applied to obtain high-quality EXAFS data for solutions with low concentrations of Ln3+ ions. Results obtained by EXAFS spectroscopy showed significant contraction of the first coordination sphere during the dissolution process for metal ions located in the middle of the lanthanide series. It was established that in Ln(NO3)3 · xH2O solutions with Ln = Ce, Sm, Gd, Yb (c = 134, 100, 50 and 20 mM) there are coordinated and, to a greater extent, non-coordinated nitrate groups with bidentate and predominantly monodentate bonds with Ln ions, the number of which increases upon transition from cerium to ytterbium. For the first time, the antibacterial and antifungal activity of Ln(NO3)3 · xH2O Ln = Ce, Sm, Gd, Tb, Yb solutions with different concentrations and pH was presented. Cross-relationships between the concentration of solutions and antimicrobial activity with the type of Ln = Ce, Sm, Gd, Tb, Yb were established, as well as the absence of biocidal properties of solutions with a concentration of 20 mM, except for Ln = Yb. The important role of experimental conditions in obtaining and interpreting the results was noted.

Molecular architecture of proliferative lupus nephritis as elucidated using 50-plex imaging mass cytometry proteomics.

Due to unique advantages that allow high-dimensional tissue profiling, we postulated imaging mass cytometry (IMC) may shed novel insights on the molecular makeup of proliferative lupus nephritis (LN). This study interrogates the spatial expression profiles of 50 target proteins in LN and control kidneys. Proliferative LN glomeruli are marked by podocyte loss with immune infiltration dominated by CD45RO+, HLA-DR+ memory CD4 and CD8 T-cells, and CD163+ macrophages, with similar changes in tubulointerstitial regions. Macrophages are the predominant HLA-DR expressing antigen presenting cells with little expression elsewhere, while macrophages and T-cells predominate cellular crescents. End-stage sclerotic glomeruli are encircled by an acellular fibro-epithelial Bowman's space surrounded by immune infiltrates, all enmeshed in fibronectin. Proliferative LN also shows signs indicative of epithelial to mesenchymal plasticity of tubular cells and parietal epithelial cells. IMC enabled proteomics is a powerful tool to delineate the spatial architecture of LN at the protein level.

Efficacy and Safety of Belimumab in Patients With Lupus Nephritis: Subgroup Analyses of a Phase 3 Randomized Trial in the East Asian Population.

RATIONALE & OBJECTIVE: Belimumab improved kidney outcomes in patients with active lupus nephritis (LN) in BLISS-LN, leading to its approval in the United States and the European Union. As data on treatment of East Asian patients with LN are limited, we evaluated the efficacy and safety of belimumab in the BLISS-LN East Asian subgroup. STUDY DESIGN: Prespecified subgroup analysis of BLISS-LN, a phase 3, placebo-controlled, randomized 104-week trial. SETTING & PARTICIPANTS: Adults with biopsy-proven, active LN were randomized (1:1) to belimumab or placebo, plus standard therapy. INTERVENTION: Patients were administered intravenous belimumab 10mg/kg, or placebo, plus standard therapy (oral glucocorticoids and either cyclophosphamide for induction followed by azathioprine for maintenance, or mycophenolate mofetil for both induction and maintenance). At the investigator's discretion, 1-3 intravenous pulses of methylprednisolone, 500-1,000mg each, could be administered during induction. OUTCOMES: The primary end point was primary efficacy renal response (PERR; ie, urinary protein-creatinine ratio≤0.7g/g, estimated glomerular filtration rate no more than 20% below preflare value or≥60mL/min/1.73m2, and no treatment failure) at week 104. Key secondary end points included complete renal response (CRR; urinary protein-creatinine ratio<0.5g/g, estimated glomerular filtration rate no more than 10% below preflare value or≥90mL/min/1.73m2, and no treatment failure) at week 104; PERR at week 52; time to kidney-related event or death; and safety. ANALYTICAL APPROACH: PERR and CRR were analyzed using a logistic regression model, and time to a kidney-related event or death was analyzed using a Cox proportional hazards regression model. RESULTS: 142 patients from mainland China, Hong Kong, South Korea, and Taiwan were included (belimumab, n=74; placebo, n=68). At week 104, more belimumab than placebo patients achieved PERR (53% vs 37%; OR, 1.76 [95% CI, 0.88-3.51]) and CRR (35% vs 25%; OR, 1.73 [95% CI, 0.80-3.74]). At week 52, more belimumab than placebo patients achieved PERR (62% vs 37%; OR, 2.74 [95% CI, 1.33-5.64]). Belimumab reduced the risk of a kidney-related event or death compared with placebo at any time (HR, 0.37 [95% CI, 0.15-0.91]). Safety was similar across treatment groups. LIMITATIONS: Small sample size and lack of formal significance testing. CONCLUSIONS: Safety and efficacy profiles were consistent with BLISS-LN overall population, supporting benefits of belimumab treatment in the East Asian subgroup with LN. FUNDING: This study was funded by GSK (GSK study no. BEL114054). TRIAL REGISTRATION: Registered at ClinicalTrials.gov with study number NCT01639339.

TGF-ß3 in differentiation and function of Tph-like cells and its relevance to disease activity in patients with systemic lupus erythematosus.

OBJECTIVES: T peripheral helper (Tph) cells have major roles in pathological processes in SLE. We sought to clarify the mechanisms of Tph cell differentiation and their relevance to clinical features in patients with SLE. METHOD: Phenotypes and functions of Tph cell-related markers in human CD4+ T cells purified from volunteers or patients were analysed using flow cytometry and quantitative PCR. Renal biopsy specimens from patients with LN were probed by multicolour immunofluorescence staining. RESULTS: Among multiple cytokines, transforming growth factor (TGF)-ß3 characteristically induced programmed cell death protein 1 (PD-1)hi musculoaponeurotic fibrosarcoma (MAF)+, IL-21+IL-10+ Tph-like cells with a marked upregulation of related genes including PDCD-1, MAF, SOX4 and CXCL13. The induction of Tph-like cells by TGF-ß3 was suppressed by the neutralization of TGF-ß type II receptor (TGF-ßR2). TGF-ß3-induced Tph-like cells efficiently promoted the differentiation of class-switch memory B cells into plasmocytes, resulting in enhanced antibody production. The proportion of Tph cells in the peripheral blood was significantly increased in patients with SLE than in healthy volunteers in concordance with disease activity and severity of organ manifestations such as LN. TGF-ß3 was strongly expressed on macrophages, which was associated with the accumulation of CD4+ C-X-C chemokine receptor (CXCR5)-PD-1+ Tph cells, in the renal tissue of patients with active LN. CONCLUSION: The induction of Tph-like cells by TGF-ß3 mainly produced from tissue macrophages plays a pivotal role in the pathological processes of active LN by enhancing B-cell differentiation in patients with SLE.

Urinary CD163 is a marker of active kidney disease in childhood-onset lupus nephritis.

OBJECTIVE: The objective of this study was to evaluate the utility of urine CD163 for detecting disease activity in childhood-onset SLE (cSLE) patients. METHODS: Sixty consecutive pediatric patients fulfilling four or more ACR criteria for SLE and 20 healthy controls were recruited for testing of urinary CD163 using ELISA. SLE disease activity was assessed using the SLEDAI-2K. RESULTS: Urine CD163 was significantly higher in patients with active LN than inactive SLE patients and healthy controls, with receiver operating characteristics area under the curve values ranging from 0.93 to 0.96. LN was ascertained by kidney biopsy. Levels of CD163 significantly correlated with the SLEDAI, renal SLEDAI, urinary protein excretion and C3 complement levels. Urine CD163 was also associated with high renal pathology activity index and chronicity index, correlating strongly with interstitial inflammation and interstitial fibrosis based on the examination of concurrent kidney biopsies. CONCLUSION: Urine CD163 emerges as a promising marker for identifying cSLE patients with active kidney disease. Longitudinal studies are warranted to validate the clinical utility of urine CD163 in tracking kidney disease activity in children with lupus.