Biology subjects

Kamat, V.

Publications and source records attributed to Kamat, V..

6 recordsLinked to original sources

bioRxiv2023

A Compound that Inhibits Glycolysis in Prostate Cancer Controls Growth of Advanced Prostate Cancer

PurposeMetastatic castration-resistant prostate cancer remains incurable regardless of recent therapeutic advances. Prostate cancer tumors display highly glycolytic phenotypes as the cancer progresses. Non-specific inhibitors of glycolysis have not been utilized successfully for chemotherapy, because of their penchant to cause systemic toxicity. This study reports the preclinical activity, safety, and pharmacokinetics of a novel small molecule preclinical candidate, BKIDC-1553, with antiglycolytic activity. Experimental designWe tested a large battery of prostate cancer cell lines for inhibition of cell proliferation, in vitro. Cell cycle, metabolic and enzymatic assays were used to demonstrate their mechanism of action. A human PDX model implanted in mice and a human organoid were studied for sensitivity to our BKIDC preclinical candidate. A battery of pharmacokinetic experiments, absorption, distribution, metabolism, and excretion experiments, and in vitro and in vivo toxicology experiments were carried out to assess readiness for clinical trials. ResultsWe demonstrate a new class of small molecule inhibitors where antiglycolytic activity in prostate cancer cell lines is mediated through inhibition of hexokinase 2. These compounds display selective growth inhibition across multiple prostate cancer models. We describe a lead BKIDC-1553 that demonstrates promising activity in a preclinical xenograft model of advanced prostate cancer, equivalent to that of enzalutamide. BKIDC-1553 demonstrates safety and pharmacologic properties consistent with a compound that can be taken into human studies with expectations of a good safety margin and predicted dosing for efficacy. ConclusionThis work supports testing BKIDC-1553 and its derivatives in clinical trials for patients with advanced prostate cancer.

cancer biology↗

bioRxiv2023

Alpaca Single B Cell Interrogation and Heavy-Chain-Only Antibody Discovery on an Optofluidic Platform

In vivo discovery approaches for single domain antibodies such as VHH have been limited by the lack of methodologies available for camelid B cell interrogation. Here, we report a novel application of the Berkeley Lights Beacon(R) optofluidic platform to the discovery of heavy-chain-only antibodies by screening single B cells from alpacas immunized with two different targets. Custom methods for alpaca B cell enrichment, culture, on-Beacon IgG2/3 detection, and sequencing were developed and used to discover target-specific VHH candidates from an alpaca immunized with either human prostate specific membrane antigen (PSMA) or a second blinded target, in a proof-of-concept study. PSMA-specific VHH hits discovered on the Beacon were recombinantly expressed as VHH-Fc, purified, and characterized using label-free techniques. All but one VHH-Fc bound PSMA with a single-digit nanomolar affinity, and four candidates were successfully humanized in silico using a rapid bulk humanization approach. In addition, next-generation repertoire sequencing was performed at longitudinal timepoints after immunization, uncovering additional variants within the clonal lineages of the validated hits discovered on the Beacon platform. The establishment of this single B cell VHH discovery workflow extends the powerful Beacon technology to enable rapid discovery of VHH directly from natural camelid immune repertoires.

immunology↗

bioRxiv2023

Elamipretide Improves ADP Sensitivity in Aged Mitochondria by Increasing Uptake through the Adenine Nucleotide Translocator (ANT)

Aging muscle experiences functional decline in part mediated by impaired mitochondrial ADP sensitivity. Elamipretide (ELAM) rapidly improves physiological and mitochondrial function in aging and binds directly to the mitochondrial ADP transporter ANT. We hypothesized that ELAM improves ADP sensitivity in aging leading to rescued physiological function. We measured the response to ADP stimulation in young and old muscle mitochondria with ELAM treatment, in vivo heart and muscle function, and compared protein abundance, phosphorylation, and S-glutathionylation of ADP/ATP pathway proteins. ELAM treatment increased ADP sensitivity in old muscle mitochondria by increasing uptake of ADP through the ANT and rescued muscle force and heart systolic function. Protein abundance in the ADP/ATP transport and synthesis pathway was unchanged, but ELAM treatment decreased protein s-glutathionylation incuding of ANT. Mitochondrial ADP sensitivity is rapidly modifiable. This research supports the hypothesis that ELAM improves ANT function in aging and links mitochondrial ADP sensitivity to physiological function. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=79 SRC="FIGDIR/small/525989v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1980dc7org.highwire.dtl.DTLVardef@1b0c22eorg.highwire.dtl.DTLVardef@19c9244org.highwire.dtl.DTLVardef@46e79b_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical Abstract.C_FLOATNO ELAM Binds Directly to ANT and ATP Synthase and ELAM Treatment Improves ADP Sensitivity, Increases ATP Production, and Improves Physiological Function in Old Muscles. ADP (adenosine diphosphate), ATP (adenosine triphosphate), VDAC (voltage-dependent anion channel), ANT (Adenine nucleotide translocator), H+ (proton), ROS (reactive oxygen species), NADH (nicotinamide adenine dinucleotide), FADH2 (flavin adenine dinucleotide), O2 (oxygen), ELAM (elamipretide), -SH (free thiol), -SSG (glutathionylated protein). C_FIG

physiology↗

bioRxiv2022

A Peptide Triple Agonist of GLP-1, Neuropeptide Y1, and Neuropeptide Y2 Receptors Promotes Glycemic Control and Weight Loss

Mechanisms underlying long-term sustained weight loss and glycemic normalization after obesity surgery include changes in gut hormone levels, including glucagon-like peptide 1 (GLP-1) and peptide YY (PYY). We demonstrate that two peptide biased agonists (GEP44 and GEP12) of the GLP-1, neuropeptide Y1, and neuropeptide Y2 receptors (GLP-1R, Y1-R, and Y2-R, respectively) elicit Y1-R antagonist-controlled, GLP-1R-dependent stimulation of insulin secretion in both rat and human pancreatic islets, thus revealing the counteracting effects of Y1-R and GLP-1R agonism. These agonists also promote insulin-independent Y1-R-mediated glucose uptake in muscle tissue ex vivo and more profound reductions in food intake and body weight than liraglutide when administered to diet-induced obese rats. Our findings support a role for Y1-R signaling in glucoregulation and highlight the therapeutic potential of simultaneous receptor targeting to achieve long-term benefits for millions of patients.

pharmacology and toxicology↗

bioRxiv2021

ACVR1 antibodies exacerbate heterotopic ossification in fibrodysplasia ossificans progressiva (FOP) by activating FOP-mutant ACVR1

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disorder whose most debilitating pathology is progressive and cumulative heterotopic ossification (HO) of skeletal muscles, ligaments, tendons, and fascia. FOP is caused by mutations in the type I BMP receptor gene ACVR1, which enable ACVR1 to utilize its natural antagonist, Activin A, as an agonistic ligand. The physiological relevance of this property is underscored by the fact that HO in FOP is exquisitely dependent on activation of FOP-mutant ACVR1 by Activin A, an effect countered by inhibition of Activin A via monoclonal antibody treatment. Hence, we surmised that ACVR1 antibodies that block activation of ACVR1 by ligand should also inhibit HO in FOP and provide an additional therapeutic option for this condition. Therefore, we generated ACVR1 monoclonal antibodies that block ACVR1s activation by its ligands. Surprisingly, in vivo, these ACVR1 antibodies stimulate HO and activate signaling of FOP-mutant ACVR1. This property is restricted to FOP-mutant ACVR1 and results from ACVR1 antibody-mediated dimerization of ACVR1. Conversely, wild type ACVR1 is inhibited by ACVR1 antibodies. These results uncover an additional novel property of FOP-mutant ACVR1 and indicate that ACVR1 antibodies should not be considered as therapeutics for FOP.

biochemistry↗

bioRxiv2021

Fluidics System for Resolving Concentration-Dependent Effects of Dissolved Gases on Tissue Metabolism

Oxygen (O2) and other dissolved gases such as the gasotransmitters H2S, CO and NO affect cell metabolism and function. To evaluate effects of dissolved gases on processes in tissue, we developed a fluidics system that controls dissolved gases while simultaneously measuring parameters of electron transport, metabolism and secretory function. We use pancreatic islets, retina and liver to highlight its ability to assess effects of O2 and H2S. Protocols aimed at emulating hypoxia-reperfusion conditions resolved a previously unrecognized transient spike in O2 consumption rate (OCR) following replenishment of O2, and tissue-specific recovery of OCR following hypoxia. The system revealed both inhibitory and stimulatory effects of H2S on insulin secretion rate from isolated islets. The unique ability of this new system to quantify metabolic state and cell function in response to precise changes in dissolved gases provides a powerful platform for cell physiologists to study a wide range of disease states.

cell biology↗