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Biomedical subjects

Sanjay Kumar

Publications and source records attributed to Sanjay Kumar.

At least 19 recordsLinked to original sources
PubMed2026

Druggable genome CRISPRi screen in hydrogels reveals regulators of cortactin-driven actin remodeling promoting glioblastoma invasion.

To identify therapeutic targets limiting glioblastoma invasion, we applied druggable genome CRISPRi screens and multiomic analysis to patient-derived glioblastoma cells in micro-dissectible biomimetic 3D hydrogels that permitted separation and analysis of core versus invasive fractions. Of 2,550 genes screened, 12 encoded druggable targets whose suppression limited invasion, of which AURKB (encoding aurora kinase B) and ACP1 (encoding low molecular weight protein tyrosine phosphatase, LMW-PTP) were validated in neurosphere assays and in vivo. Proximity labeling identified cortactin as a link between LMW-PTP and aurora B, and we observed that cortactin underwent serine phosphorylation by aurora B and tyrosine dephosphorylation by LMW-PTP. Targeting ACP1 or AURKB via CRISPRi or inhibitors in culture and in vivo shifted the cortactin phosphorylation balance in glioblastoma, reducing levels of cortactin and the actin-related protein 2/3 (Arp2/3) complex that mediates cortactin-induced actin stabilization, thereby reducing actin-cortactin-Arp2/3 colocalization and subsequent actin polymerization. AURKB or ACP1 targeting shifted actin from cytoplasm to the nucleus, reducing mesenchymal gene expression. Biophysical analysis implicated AURKB in glioblastoma cell adhesion and stiffness needed for initial migration and ACP1 in mechanical stress resistance required for later migration. These findings revealed a targetable axis balancing kinase and phosphatase activities to regulate actin polymerization during glioblastoma invasion.

Humans↗
PubMed2025

Druggable genome CRISPRi screen in 3D hydrogels reveals regulators of cortactin-driven actin remodeling in invading glioblastoma cells.

To identify new therapeutic targets that limit glioblastoma (GBM) invasion, we applied druggable-genome CRISPR screens to patient-derived GBM cells in micro-dissectible biomimetic 3D hydrogel platforms that permit separation and independent analysis of core vs. invasive fractions. We identified 12 targets whose suppression limited invasion, of which ACP1 (LMW-PTP) and Aurora Kinase B (AURKB) were validated in neurosphere assays. Proximity labeling analysis identified cortactin as an ACP1-AURKB link, as cortactin undergoes serine phosphorylation by AURKB and tyrosine dephosphorylation by ACP1. Suppression of ACP1 or AURKB in culture and in vivo shifted the balance of cortactin phosphorylation in GBM and reduced actin polymerization and actin-cortactin co-localization. Additional biophysical analysis implicated AURKB in GBM cell adhesion and cortical stiffness, and ACP1 in resistance to mechanical stress and shape plasticity needed for 3D migration. These findings reveal a novel targetable axis that balances kinase and phosphatase activities to regulate actin polymerization during GBM invasion.

CRISPR↗
PubMed2004

Expression of Babesia equi EMA-1 and EMA-2 during merozoite developmental stages in erythrocyte and their interaction with erythrocytic membrane skeleton.

In the present study, we investigated the cellular localizations and expression patterns of equi merozoite antigens (EMA) -1 and -2 of Babesia equi during its asexual erythrocytic-developmental cycle using anti-EMA-1t or -2t mono-specific mouse serum. Indirect fluorescent antibody tests demonstrated that EMA-1 and EMA-2 were not expressed in all the erythrocytic-developmental stages of the merozoites and that these two antigens were co-expressed during the early developmental stages. Additionally, it was shown that EMA-1 and EMA-2 were mutually expressed on the surface of extra-erythrocytic merozoites and also that the intra-erythrocytic merozoites shed only EMA-2 antigen in the infected erythrocytic cytoplasm or inside the membrane surface. The specific binding of EMA-2 to a Triton X-100-insoluble horse erythrocyte membrane fraction was also demonstrated. These findings facilitate our understanding of the biological roles of merozoite surface proteins of B. equi and our investigation for new drug targets.

Animals↗
PubMed2004

Growth-inhibitory effect of heparin on Babesia parasites.

We examined the inhibitory effects of three heparins on the growth of Babesia parasites. The multiplication of Babesia bovis, B. bigemina, B. equi, and B. caballi in in vitro cultures and that of B. microti in vivo were significantly inhibited in the presence of heparins, as determined by light microscopy. Treatment with various concentrations of heparin showed complete clearance of the intracellular parasites. Interestingly, a higher percentage of abnormally multidividing B. bovis parasites was observed in the presence of low concentrations of heparin. Furthermore, fluorescein isothiocyanate-labeled heparin was preferably found on the surfaces of extracellular merozoites, as detected by confocal laser scanning microscopy. These findings indicate that the heparin covers the surfaces of babesial merozoites and inhibits their subsequent invasion of erythrocytes.

Animals↗
PubMed2003

Identification, characterization, and inhibition of Plasmodium falciparum beta-hydroxyacyl-acyl carrier protein dehydratase (FabZ).

The emergence of drug-resistant forms of Plasmodium falciparum emphasizes the need to develop new antimalarials. In this context, the fatty acid biosynthesis (FAS) pathway of the malarial parasite has recently received a lot of attention. Due to differences in the fatty acid biosynthesis systems of Plasmodium and man, this pathway is a good target for the development of new and selective therapeutic drugs directed against malaria. In continuation of these efforts we report cloning and overexpression of P. falciparum beta-hydroxyacyl-acyl carrier protein (ACP) dehydratase (PffabZ) gene that codes for a 17-kDa protein. The enzyme catalyzes the dehydration of beta-hydroxyacyl-ACP to trans-2-acyl-ACP, the third step in the elongation phase of the FAS cycle. It has a Km of 199 microM and kcat/Km of 80.4 m-1 s-1 for the substrate analog beta-hydroxybutyryl-CoA but utilizes crotonoyl-CoA, the product of the reaction, more efficiently (Km = 86 microM, kcat/Km = 220 m-1 s-1). More importantly, we also identify inhibitors (NAS-91 and NAS-21) for the enzyme. Both the inhibitors prevented the binding of crotonoyl-CoA to PfFabZ in a competitive fashion. Indeed these inhibitors compromised the growth of P. falciparum in cultures and inhibited the parasite fatty acid synthesis pathway both in cell-free extracts as well as in situ. We modeled the structure of PfFabZ using Escherichia coli beta-hydroxydecanoyl thioester dehydratase (EcFabA) as a template. We also modeled the inhibitor complexes of PfFabZ to elucidate the mode of binding of these compounds to FabZ. The discovery of the inhibitors of FabZ, reported for the first time against any member of this family of enzymes, essential to the type II FAS pathway opens up new avenues for treating a number of infectious diseases including malaria.

Amino Acid Sequence↗
PubMed2003

A rapid protocol for the prevention of contrast-induced renal dysfunction: the RAPPID study.

OBJECTIVES: This study was designed to test a rapid protocol of intravenous acetylcysteine for prevention of radiocontrast-induced nephropathy (RCIN). BACKGROUND: Oral acetylcysteine (NAC) may provide better prophylaxis against RCIN than intravenous (i.v.) hydration alone. Current protocols preclude prophylaxis of same-day or emergency patients owing to the need for prolonged pretreatment. METHODS: We prospectively randomized 80 patients with stable renal dysfunction undergoing cardiac catheterization/intervention to a rapid protocol of i.v. NAC (150 mg/kg in 500 ml N/saline over 30 min immediately before contrast followed by 50 mg/kg in 500 ml N/saline over 4 h, n = 41, 67 +/- 10 years, 90% men) or i.v. hydration (1 ml/kg/h N/saline for 12 h pre- and post-contrast, n = 39, 71 +/- 8.8 years, 85% men). RESULTS: Radiocontrast-induced nephropathy occurred in 2 of the 41 patients in the NAC group (5%) and in 8 of the 39 patients in the hydration group (21%; p = 0.045; relative risk: 0.28; 95% confidence interval 0.08 to 0.98). In the NAC group, mean serum creatinine fell from 1.85 +/- 0.59 to 1.77 +/- 0.73 and 1.79 +/- 0.73 mg/dl 48 h and four days post-contrast (p = 0.02 and 0.023 vs. baseline, respectively). In the hydration group, serum creatinine increased from 1.75 +/- 0.41 to 1.81 +/- 0.6 48 h and 1.80 +/- 0.50 mg/dl four days post-contrast (p = 0.99 and 0.23, respectively). NAC infusion was ceased after the bolus in three patients (7%) due to flushing, itching, or a transient rash. CONCLUSIONS: Administration of i.v. NAC should be considered in all patients at risk of RCIN before contrast exposure when time constraints preclude adequate oral prophylaxis, provided the patient is able to tolerate this degree of volume loading.

Acetylcysteine↗
PubMed2003

Innate immunity mediated by the cytokine IL-1 homologue 4 (IL-1H4/IL-1F7) induces IL-12-dependent adaptive and profound antitumor immunity.

Recently, several novel members of the IL-1 family have been identified. The possible therapeutic utility and the underlying biologic role of these new members remain unclear. In the present study we analyzed the anti-tumor activity of human IL-1 homologue 4(IL-1H4; renamed IL-F7) by adenovirus-mediated gene transfer (AdIL-1H4) directly into murine tumors. In vitro expression analysis showed that IL-1H4 was a secretory protein. Treatment of an established MCA205 mouse fibrosarcoma by single intratumoral injection of AdIL-1H4 resulted in significant growth suppression. Furthermore, complete inhibition of tumor growth was observed following multiple injections of AdIL-1H4. The anti-tumor activity of IL-1H4 was abrogated in nude and SCID mice and in IL-12-, IFN-gamma-, or Fas ligand-deficient mice. In contrast, IL-1H4 was able to confer substantial anti-tumor effects in NKT-deficient mice. These results suggest that IL-1H4 could play an important role in the link between innate and adaptive immunity and may be useful for tumor immunotherapy.

Adenoviridae↗