Biology subjects

Patel, S.

Publications and source records attributed to Patel, S..

At least 109 records · Page 6Linked to original sources

bioRxiv2024

Synaptic Organization-Function Relationships of Amygdala Interneurons Supporting Associative Learning

Coordinated activity of basolateral amygdala (BLA) GABAergic interneurons (INs) and glutamatergic principal cells (PCs) is critical for associative learning, however the microcircuit organization-function relationships of distinct IN classes remain uncertain. Here, we show somatostatin (SOM) INs provide inhibition onto, and are excited by, local PCs, whereas vasoactive intestinal peptide (VIP) INs are driven by extrinsic afferents. Parvalbumin (PV) INs inhibit PCs and are activated by local and extrinsic inputs. Thus, SOM and VIP INs exhibit complementary roles in feedback and feedforward inhibition, respectively, while PV INs contribute to both microcircuit motifs. Functionally, each IN subtype reveals unique activity patterns across fear- and extinction learning with SOM and VIP INs showing most divergent characteristics, and PV INs display an intermediate phenotype parallelling synaptic data. Finally, SOM and PV INs dynamically track behavioral state transitions across learning. These data provide insight into the synaptic microcircuit organization-function relationships of distinct BLA IN classes.

neuroscience↗

bioRxiv2024

Targeted isolation of bacteria with potential to competitively exclude Staphylococcus aureus in the upper respiratory tract of pigs

Considering global antimicrobial resistance (AMR) prevalence, alternative or complementary strategies to antimicrobial use, are of interest. Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) is of particular interest as despite significant AMU reduction, LA-MRSA prevalence in pig husbandry has not decreased. We performed targeted isolation of bacterial species with potential antagonism against LA-MRSA in pig farms. Duplicate piglet nasal swabs from three European countries (Germany, Ireland and The Netherlands) were taken longitudinally from birth up to 10 weeks, one for amplicon sequencing and qPCR, and the other was cryopreserved for culturing. We identified potential probiotic species by anticorrelation analysis of bacterial abundance from amplicon sequencing data with quantitative S. aureus estimates from qPCR data from the samples. A literature-screen was performed on the species identified, to determine their probiotic potential. Following this, 1302 isolates were grown from selected cryopreserved swabs and identified using MALDI-TOF and additional 16S rRNA gene sequencing to isolate the anticorrelating species. Ninety-five isolates of interest were screened for absence of tetracycline resistance and hemolytic activity and whole genome sequencing was conducted to verify their taxonomy and to assess their AMR and virulence gene profile. Additional phenotypic antimicrobial resistance testing selected three different Lactococcus lactis strains. During an in vitro challenge using spent medium, all three strains demonstrated inhibition against two S. aureus strains. These L. lactis strains may have the ability to be used safely to reduce LA-MRSA carriage in the nasal passages of pigs but further in vivo testing is necessary to confirm this potential. ImportanceAn approach to tackle antimicrobial resistance in livestock is to use competitive exclusion agents. We employed a sequencing guided isolation approach to identify bacterial species with in silico antagonism to livestock-associated Staphylococcus aureus (LA-MRSA) in the nasal microbiome of piglets. Based on this, three Lactococcus lactis isolates were found to be suitable for further probiotic testing. This strategy can be used to effectively identify, isolate, and screen bacteria that live in antagonism with opportunistic pathogens such as LA-MRSA in a complex microbiome.

microbiology↗

bioRxiv2024

Stable, chronic in-vivo recordings from a fully wireless subdural-contained 65,536-electrode brain-computer interface device

Minimally invasive, high-bandwidth brain-computer-interface (BCI) devices can revolutionize human applications. With orders-of-magnitude improvements in volumetric efficiency over other BCI technologies, we developed a 50-m-thick, mechanically flexible micro-electrocorticography (ECoG) BCI, integrating a 256x256 array of electrodes, signal processing, data telemetry, and wireless powering on a single complementary metal-oxide-semiconductor (CMOS) substrate containing 65,536 recording channels, from which we can simultaneously record a selectable subset of up to 1024 channels at a given time. Fully implanted below the dura, our chip is wirelessly powered, communicating bi-directionally with an external relay station outside the body. We demonstrated chronic, reliable recordings for up to two weeks in pigs and up to two months in behaving non-human primates from somatosensory, motor, and visual cortices, decoding brain signals at high spatiotemporal resolution.

neuroscience↗

bioRxiv2024

Activation loop phosphorylation of Cdk11 is restrained by PNUTS-PP1 and regulates Cdk11 activity and function

Organisation of the transcription cycle is facilitated by the reversible phosphorylation of the C-terminal domain of RNA Polymerase II (RNAPII-CTD) and its accessory factors. The PNUTS-PP1 protein phosphatase is crucial for mRNA synthesis and processing, yet the complete spectrum of its physiological targets in these processes remain elusive. Here, using quantitative phosphoproteomics, we discover that Cdk11, in addition to various spliceosomal and RNA processing factors, associates with PNUTS, and that disruption of PP1-binding results in hyperphosphorylation of Cdk11 at an evolutionarily conserved Serine residue, seven amino acid residues C-terminal to DFG residues in the activation loop. In vitro experiments reveal a role for Ser DFG+7 in modulating Cdk11 kinase activity towards RNAPII-CTD Ser5. Making use of a novel technique to conditionally disrupt PP1 binding, we show that PNUTS-PP1 normally serves to restrain Cdk11 phosphorylation in vivo. Mutational analysis shows that cdk11 is not only essential for survival but also plays a widespread role in regulating normal mRNA expression and splicing. Notably, we find that a phosphomimetic mutation in cdk11 exhibits distinct biological effects compared to loss of cdk11 function, including defective processing of intronic small nucleolar RNAs, diminished intronic RNA Pol II velocity, and a decrease in intergenic transcription. These findings underscore physiologically significant roles of Cdk11 dephosphorylation by PNUTS-PP1 in the regulation of mRNA transcription and processing.

molecular biology↗

bioRxiv2024

Eco-evolutionary dynamics of anthelmintic resistance in soil-transmitted helminths

Anthelmintic resistance (AR) of helminth parasites against the most widely available drugs is an ongoing concern for both human and livestock-infecting species. Indeed, there has been substantial evidence of AR in livestock but less in humans, which may be due to a variety of reasons. In this paper, we develop an eco-evolutionary model that couples the life cycle of these parasites with their underlying evolution in a single biallelic genetic locus that confers resistance to treatment drugs. We determine the critical treatment frequency needed to effectively eliminate the population, for a fixed drug efficacy (without evolution) and use this to classify three qualitative distinct behaviors of the eco-evolutionary model. Then, we describe how aspects of the life cycle influence which qualitative outcome is achieved and the spread of the resistance allele, comparing across parameterized models of human- and livestock-infecting species. For all but one species, we find that lower fecundity rates and lower contact rates speed the spread of resistance, while lower larval death slows it down. The life cycle parameters of Ancylostoma duodenale and Ostertagia circumcincta are associated with the fastest and slowest spread of resistance, respectively. We discuss the mechanistic reason for these results.

ecology↗

bioRxiv2024

GeneSqueeze: A Novel Lossless, Reference-Free Compression Algorithm for FASTQ/A Files

As sequencing becomes more accessible, there is an acute need for novel compression methods to efficiently store this data. Omics technologies can enhance biomedical research and individualize patient care, but they demand immense storage capabilities, especially when applied to longitudinal studies. Addressing the storage challenges posed by these technologies is crucial for omics technologies to achieve their full potential. We present a novel lossless, reference-free compression algorithm, GeneSqueeze, that leverages the patterns inherent in the underlying components of FASTQ files (i.e., nucleotide sequences, quality scores and read identifiers). GeneSqueeze provides several benefits, including an auto-tuning compression protocol based on each samples distribution, lossless preservation of IUPAC nucleotides and read identifiers, and unrestricted FASTQ/A file attributes (i.e., read length, read depth, or read identifier format). We compared GeneSqueeze to the general-purpose compressor, gzip, and to the domain-specific compressor, SPRING. GeneSqueeze achieved up to three times higher compression ratios as compared to gzip, regardless of read length, read depth, or file size. GeneSqueeze achieved 100% lossless compression, with the original and decompressed files perfectly matching for all tested samples, preserving read identifiers, quality scores, and IUPAC nucleotides, in contrast to SPRING. Overall, GeneSqueeze represents a competitive and specialized compression method optimized for FASTQ/A files containing nucleotide sequences that has the potential to significantly reduce the storage and transmission costs associated with large omics datasets without sacrificing data integrity.

bioinformatics↗

bioRxiv2024

Forecasting Dengue Fever in Brazil Using Multimodal Data, Including Climate Data

Dengue fever is a major tropical disease transmitted by Aedes mosquitoes. Dengue affects more than 120 countries with highly variable year to year infection rates. Despite high variability, dengue has a clear relationship to climate factors and human demography. Global trends to higher temperatures and greater disorderly urban development are increasing the scale and scope of dengue risk. Dengue has complex human immunity with 4 known serotypes that make multiple infections possible. Accurate forecasting of dengue fever would allow for appropriate interventions and improved public health outcomes. We demonstrate, GeoSeeq Dengue, a forecasting model for dengue fever in Brazil. GeoSeeq Dengue predicts dengue outbreaks monthly in 5,570 Brazilian municipalities at 1, 3, and 6 months ahead of the outbreak. Model accuracy compares favorably to a historical baseline model, making it a promising model for informing public health response. We evaluate how different types of input variables effect model accuracy and explore how this model could be adapted to other countries. This model could inform public health responses to dengue including targeting vector control programs, public health education messaging, and the newly launched dengue vaccine rollout.

bioinformatics↗

bioRxiv2024

The bispectral EEG (BSEEG) method quantifies post-operative delirium-like states in young and aged mice after head mount implantation surgery

Delirium, a syndrome characterized by an acute change in attention, awareness, and cognition, is commonly observed in older adults and has multiple potential triggers, including illness, drug, trauma, and surgery. There are few quantitative monitoring methods in clinical settings. We developed the bispectral electroencephalography (BSEEG) method in clinical research that can detect the presence of and quantify the severity of delirium using a novel algorithm. In the pre-clinical model, we reported that the BSEEG method can capture a delirium-like state in mice following LPS administration. However, its application to post-operative delirium (POD) has not yet been validated in animal experiments. Therefore, this study aimed to create a POD model mouse with the BSEEG method by monitoring BSEEG scores after EEG head-mount implantation surgery throughout the recovery phase. We compared the BSEEG scores of C57BL/6J young (2-3 months old) with aged (18-19 months old) mice for quantitative evaluation of the delirium-like state after the surgery. Postoperatively, both groups showed increased BSEEG scores and a loss of regular diurnal changes in BSEEG scores every daytime and night. In young mice, BSEEG scores and regular diurnal changes recovered relatively quickly to baseline by around postoperative day 3. On the other hand, aged mice had prolonged increases in postoperative BSEEG scores and it reached steady state only after around postoperative day 8. This study suggests the BSEEG method can be utilized to quantitatively evaluate POD and also assess the effect of aging on recovery from POD in pre-clinical model.

neuroscience↗

bioRxiv2024

PREFRONTAL CORRELATES OF FEAR GENERALIZATION DURING ENDOCANNABINOID DEPLETION

Maladaptive fear generalization is one of the hallmarks of trauma-related disorders. The endocannabinoid 2-arachidonoylglycerol (2-AG) is crucial for modulating anxiety, fear, and stress adaptation but its role in balancing fear discrimination versus generalization is not known. To address this, we used a combination of plasma endocannabinoid measurement and neuroimaging from a childhood maltreatment exposed and non-exposed mixed population combined with human and rodent fear conditioning models. Here we show that 2-AG levels are inversely associated with fear generalization at the behavioral level in both mice and humans. In mice, 2-AG depletion increases the proportion of neurons, and the similarity between neuronal representations, of threat-predictive and neutral stimuli within prelimbic prefrontal cortex ensembles. In humans, increased dorsolateral prefrontal cortical-amygdala resting state connectivity is inversely correlated with fear generalization. These data provide convergent cross-species evidence that 2-AG is a key regulator of fear generalization and suggest 2-AG deficiency could represent a trauma-related disorder susceptibility endophenotype.

neuroscience↗

bioRxiv2024

NEUROG2 regulates a human-specific neurodevelopmental gene regulatory program

Unique hallmarks of human neocortical development include slower rates of neurogenesis and the establishment of an extracellular matrix-rich, outer-subventricular zone that supports basal neural progenitor cell expansion. How gene regulatory networks have evolved to support these human-specific neurodevelopmental features is poorly understood. Mining single cell data from cerebral organoids and human fetal cortices, we found that NEUROG2 expression is enriched in basal neural progenitor cells. To identify and purify NEUROG2-expressing cells and trace their short-term lineage, we engineered two NEUROG2-mCherry knock-in human embryonic stem cell lines to produce cerebral organoids. Transcriptomic profiling of mCherry-high organoid cells revealed elevated expression of PPP1R17, associated with a fast-evolving human-accelerated regulatory region, oligodendrocyte precursor cell and extracellular matrix-associated gene transcripts. Conversely, only neurogenic gene transcripts were enriched in mCherry-high cortical cells from Neurog2:mCherry knock-in mice. Finally, we show that Neurog2 is sufficient to induce Ppp1r17, which slows human neural progenitor cell division, and Col13a1, an extracellular matrix gene, in P19 cells. NEUROG2 thus regulates a human neurodevelopmental gene regulatory program implicated in supporting a pro-proliferative basal progenitor cell niche and tempering the neurogenic pace. SUMMARY STATEMENTTranscriptomic analyses of NEUROG2-mCherry knock-in human embryonic stem cell-derived cerebral organoids reveal a link between NEUROG2 and extracellular matrix remodeling during human cortical development.

developmental biology↗

bioRxiv2023

The developing pig respiratory microbiome harbours strains antagonistic to common respiratory pathogens

In the global efforts to combat antimicrobial resistance and reduce antimicrobial use in pig production, there is a continuous search for methods to prevent and/or treat infections. Within this scope, we explored the relationship between the developing piglet nasal microbiome and (zoonotic) bacterial pathogens from birth until ten weeks of life. The nasal microbiome of 54 pigs was longitudinally studied over 16 time-points on nine farms in three European countries (Germany, Ireland, and the Netherlands) using amplicon sequencing targeting the V3-V4 16S rRNA region as well as the tuf gene for its Staphylococcal discrimination power. The piglets age, the farm, and the litter affected the nasal microbiome, with piglets age explaining 19% of the variation in microbial composition between samples. Stabilization of the microbiome occurred around two weeks post-weaning. Notably, while opportunistic pathogens were ubiquitously present, they did not cause disease. The piglet nasal microbiome often carried species associated with gut, skin, or vagina, which suggests that contact with the vaginal and faecal microbiomes shape the piglet nasal microbiome. We identified bacterial Co-Abundance Groups (CAGs) of species that were present in the nasal microbiomes in all three countries over time. Anticorrelation between these species and known bacterial pathogens identified strains that might be exploited for pathogen reduction. Further experimental evidence is required to confirm these findings. Overall, this study advances our understanding of the longitudinal development and factors influencing the piglet nasal microbiome, providing insights into its role in health and disease. ImportanceOur study on longitudinal analysis of the developing nasal microbiota of piglets in farms in three European countries showed consistent microbiome compositions and that colonization of porcine pathogens occurred in relation with anticorrelating species. These findings enhance our knowledge of co-colonizing species in the nasal cavity, and the identified microbial interactions can be explored for the development of interventions to control pathogens in porcine husbandry.

microbiology↗

bioRxiv2023

A niche-adapted brain microbiome in salmonids at homeostasis

Ectotherms have long been known to have peculiar relationships with microorganisms. For instance, bacteria can be recovered from blood and internal organs of healthy teleost fish. However, until now, the presence of a microbial community in the healthy teleost brain has not been proposed. Here we report a living bacterial community in the brain of healthy salmonids. Brain bacterial loads in salmonids are comparable to those found in the spleen and 1000-fold lower than in the gut. Brain bacterial communities share >50% of their diversity with gut and blood bacterial communities. Using culturomics, we obtained 54 bacterial isolates from the brain of healthy rainbow trout. Comparative genomics uncovered unique niche adaptations associated with brain colonization and polyamine biosynthesis. In a natural system, salmonid brain microbiomes shift with the host life cycle, becoming dysbiotic in reproductively mature Chinook salmon, a species that undergoes reproductive death. Our study redefines the relationship between the teleost brain and bacterial microbiomes under physiological conditions. We posit that this symbiosis may endow salmonids with a direct mechanism to sense and respond to environmental microbes. One-Sentence SummarySalmonids have a brain-adapted, resident bacterial community

microbiology↗

bioRxiv2023

A mitochondria-regulated p53-CCF circuit integrates genome integrity with inflammation

Genomic instability and inflammation are distinct hallmarks of aging, but the connection between them is poorly understood. Understanding their interrelationship will help unravel new mechanisms and therapeutic targets of aging and age-associated diseases. Here we report a novel mechanism directly linking genomic instability and inflammation in senescent cells through a mitochondria-regulated molecular circuit driven by p53 and cytoplasmic chromatin fragments (CCF). We show, through activation or inactivation of p53 by genetic and pharmacologic approaches, that p53 suppresses CCF accumulation and the downstream inflammatory senescence-associated secretory phenotype (SASP), without affecting cell cycle arrest. p53 activation suppressed CCF formation by promoting DNA repair, and this is reflected in maintenance of genomic integrity, particularly in subtelomeric regions, as shown by single cell genome resequencing. Activation of p53 in aged mice by pharmacological inhibition of MDM2 reversed signatures of aging, including age- and senescence-associated transcriptomic signatures of inflammation and age-associated accumulation of monocytes and macrophages in liver. Remarkably, mitochondria in senescent cells suppressed p53 activity by promoting CCF formation and thereby restricting ATM-dependent nuclear DNA damage signaling. These data provide evidence for a mitochondria-regulated p53 signaling circuit in senescent cells that controls DNA repair, genome integrity, and senescence- and age-associated inflammation. This pathway is immunomodulatory in mice and a potential target for healthy aging interventions by small molecules already shown to activate p53.

cell biology↗

bioRxiv2023

Sexually dimorphic role of diet and stress on behavior, energy metabolism, and the ventromedial hypothalamus

Scientific evidence underscores the influence of biological sex on the interplay between stress and metabolic dysfunctions. However, there is limited understanding of how diet and stress jointly contribute to metabolic dysregulation in both males and females. To address this gap, our study aimed to investigate the combined effects of a high-fat diet (HFD) and repeated footshock stress on fear-related behaviors and metabolic outcomes in male and female mice. Using a robust rodent model that recapitulates key aspects of post-traumatic stress disorder (PTSD), we subjected mice to footshock stressor followed by weekly reminder footshock stressor or no stressor for 14 weeks while on either an HFD or chow diet. Our findings revealed that HFD impaired fear memory extinction in male mice that received initial stressor but not in female mice. Blood glucose levels were influenced by both diet and sex, with HFD-fed female mice displaying elevated levels that returned to baseline in the absence of stress, a pattern not observed in male mice. Male mice on HFD exhibited higher energy expenditure, while HFD-fed female mice showed a decreased respiratory exchange ratio (RER). Sex-specific alterations in pro-inflammatory markers and abundance of hematopoietic stem cells were observed in chronically stressed mice on an HFD in different peripheral tissues, indicating the manifestation of distinct comorbid disorders. Single-nuclei RNA sequencing of the ventromedial hypothalamus from stressed mice on an HFD provided insights into sex-specific glial cell activation and cell-type-specific transcriptomic changes. In conclusion, our study offers a comprehensive understanding of the intricate interactions between stress, diet, sex, and various physiological and behavioral outcomes, shedding light on a potential brain region coordinating these interactions.

neuroscience↗

bioRxiv2023

High temporal frequency light response in mouse retina is mediated by ON and OFF bipolar cells and requires FAT3 signaling

Vision is initiated by the reception of light by photoreceptors and subsequent processing via downstream retinal neurons. Proper cellular organization depends on the multi-functional tissue polarity protein FAT3, which is required for amacrine cell connectivity and retinal lamination. Here we investigated the retinal function of Fat3 mutant mice and found decreases in physiological and perceptual responses to high frequency flashes. These defects did not correlate with abnormal amacrine cell wiring, pointing instead to a role in bipolar cell subtypes that also express FAT3. The role of FAT3 in the response to high temporal frequency flashes depends upon its ability to transduce an intracellular signal. Mechanistically, FAT3 binds to the synaptic protein PTP{sigma}, intracellularly, and is required to localize GRIK1 to OFF-cone bipolar cell synapses with cone photoreceptors. These findings expand the repertoire of FAT3s functions and reveal its importance in bipolar cells for high frequency light response.

neuroscience↗

bioRxiv2023

OxPhos in adipose tissue macrophages regulated by BTK enhances their M2-like phenotype and confers a systemic immunometabolic benefit in obesity

Brutons tyrosine kinase (BTK) is a non-receptor bound kinase involved in pro-inflammatory signalling in activated macrophages, however, its role within adipose tissue macrophages remains unclear. We have demonstrated that BTK signalling regulates macrophage M2-like polarisation state by up-regulating subunits of mitochondrially encoded electron transport chain Complex I (ND4 and NDL4) and Complex IV (mt-CO1, mt-CO2 and mt-CO3) resulting in an enhanced rate of oxidative phosphorylation (OxPhos) in an NF-{kappa}B independent manner. Critically, BTK expression is elevated in adipose tissue macrophages from obese individuals with diabetes, while key mitochondrial genes (mtC01, mtC02 and mtC03) are decreased in inflammatory myeloid cells from obese individuals. Inhibition of BTK signalling either globally (Xid mice) or in myeloid cells (LysMCreBTK), or therapeutically (Acalabrutinib) protects HFD-fed mice from developing glycaemic dysregulation by improving signalling through the IRS1/Akt/GSK3b pathway. The beneficial effects of acalabrutinib treatment are lost in macrophage ablated mice. Inhibition of BTK signalling in myeloid cells but not B-cells, induced a phenotypic switch in adipose tissue macrophages from a pro-inflammatory M1-state to a pro-resolution M2-like phenotype, by shifting macrophage metabolism towards OxPhos. This reduces both local and systemic inflammation and protects mice from the immunometabolic consequences of obesity. Therefore, in BTK we have identified a macrophage specific, druggable target that can regulate adipose tissue polarisation and cellular metabolism that can confer systematic benefit in metabolic syndrome. Article high lightsObesity and diabetes are associated with inflammation, particularly within the adipose tissue. We have found a new druggable target called Brutons tyrosine kinase (BTK) that is highly expressed in adipose tissue macrophages. When BTK is inhibited in macrophages, it allows these cells to undergo a phenotypic switch towards an M2-like pro-resolution macrophage. This achieved by increasing expression of key mitochondrially encoded components of the electron transport chain allowing for enhanced OxPhos. Inhibition of BTK signalling in myeloid cells but not B-cells protects HFD-fed mice from developing glycaemic dysregulation.

pathology↗

bioRxiv2023

Colchicine promotes atherosclerotic plaque stability independently of inflammation

Atherosclerosis is a chronic inflammatory disease which is driven in part by the aberrant trans-differentiation of vascular smooth muscle cells (SMCs). No therapeutic drug has been shown to reverse detrimental SMC-derived cell phenotypes into protective phenotypes, a hypothesized enabler of plaque regression and improved patient outcome. Herein, we describe a novel function of colchicine in the beneficial modulation of SMC-derived cell phenotype, independent of its conventional anti-inflammatory effects. Using SMC fate mapping in an advanced atherosclerotic lesion model, colchicine induced plaque regression by converting pathogenic SMC-derived macrophage-like and osteoblast-like cells into protective myofibroblast-like cells which thickened, and thereby stabilized, the fibrous cap. This was dependent on Notch3 signaling in SMC-derived plaque cells. These findings may help explain the success of colchicine in clinical trials relative to other anti-inflammatory drugs. Thus, we demonstrate the potential of regulating SMC phenotype in advanced plaque regression through Notch3 signaling, in addition to the canonical anti-inflammatory actions of drugs to treat atherosclerosis.

cell biology↗

bioRxiv2023

Quantitative target engagement of RIPK1 in human whole blood via the cellular thermal shift assay for potential pre-clinical and clinical applications

The cellular thermal shift assay (CETSA(R)) is a target engagement method widely used for preclinical characterization of small molecule compounds. CETSA(R) has been used for semi-quantitative readouts in whole blood with PBMC isolation, and quantitative, plate-based readouts using cell lines. However, there has been no quantitative evaluation of CETSA(R) in unprocessed human whole blood, which is preferred for clinical applications. Here we report two separate assay formats - Alpha CETSA(R) and MSD CETSA(R) - that require less than 100 L of whole blood per sample without PBMC isolation. We chose RIPK1 as a proof-of-concept target and, by measuring engagement of seven different inhibitors, demonstrate high assay sensitivity and robustness. These quantitative CETSA(R) platforms enable possible applications in preclinical pharmacokinetic-pharmacodynamic studies, and direct target engagement with small molecules in clinical trials.

molecular biology↗