Proteome-wide drug screening using mass spectrometric imaging of bead-arrays

Zhou, Ying; Liu, Ziying; Rothschild, Kenneth J.; Lim, Mark J.

doi:10.1038/srep26125

Cited by 11 publications

(8 citation statements)

References 33 publications

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“…It should also be noted that we, like others, have mostly considered CNS targets as being responsible for the clinical trial accident and serine hydrolases in particular. Although our study in SHRSP was carried out to explore possible mechanisms outside the CNS (albeit related to CNS vasculature), further studies outside the CNS and looking more broadly at protein targets using proteome‐wide screening techniques, such as surface plasmon resonance or MS imaging (Yang et al, ; Zhou, Liu, Rothschild, & Lim, ) could be of relevance. One other aspect that needs to be considered is that in this study and most of the non‐clinical studies with BIA 10‐2474, the rats were relatively young, although at the end of the 26‐week toxicology study, they would correspond to 25‐ to 30‐year‐old humans according to Sengupta () and no pathology of the type seen in the clinic was observed.…”

Section: Discussionmentioning

confidence: 99%

Preclinical pharmacological evaluation of the fatty acid amide hydrolase inhibitor BIA 10‐2474

Bonifácio¹,

Sousa²,

Aires³

et al. 2020

British J Pharmacology

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Background and Purpose: In 2016, one person died and four others had mild-tosevere neurological symptoms during a phase I trial of the fatty acid amide hydrolase (FAAH) inhibitor BIA 10-2474.Experimental Approach: Pharmacodynamic and pharmacokinetic studies were performed with BIA 10-2474, PF-04457845 and JNJ-42165279 using mice, rats and human FAAH expressed in COS cells. Selectivity was evaluated by activity-based protein profiling (APBB) in rats. BIA 10-2474 effect in stroke-prone spontaneously hypertensive rats (SHRSP) was investigated.Key Results: BIA 10-2474 was 10-fold less potent than PF-04457845 in inhibiting human FAAH in situ but inhibited mouse brain and liver FAAH with ED 50 values of 13.5 and 6.2 μgÁkg −1 , respectively. Plasma and brain BIA 10-2474 levels were consistent with in situ potency and neither BIA 10-2474 nor its metabolites accumulated following repeat administration. FAAH and α/β-hydrolase domain containing 6 were the primary targets of BIA 10-2474 and, at higher exposure levels, ABHD11, PNPLA6, PLA2G15, PLA2G6 and androgen-induced protein 1. At 100 mgÁkg −1 for 28 days, the level of several lipid species containing arachidonic acid increased. Daily treatment of SHRSP with BIA 10-2474 did not affect mortality rate or increased the incidence of haemorrhage or oedema in surviving animals.Conclusions and Implications: BIA 10-2474 potently inhibits FAAH in vivo, similarly to PF-04457845 and interacts with a number of lipid processing enzymes, some previously identified in human cells as off-targets particularly at high levels of exposure.These interactions occurred at doses used in toxicology studies, but the implication of these off-targets in the clinical trial accident remains unclear.Abbreviations: 2-AG, 2-arachidonoylglycerol; AA, arachidonic acid; ABHD6, α/β-hydrolase domain containing 6; ABPP, activity-based protein profiling; AEA, anandamide; AIG1, androgeninduced protein 1; BCRP, breast cancer resistance protein; CES, carboxyl esterase; CYP, cytochrome P450; DAG, diacylglycerol; FAAH, fatty acid amide hydrolase; LIPE, lipase E; MATE, multidrug and toxin extrusion; MDR, multidrug resistance; OAT, organic anion transporter; OCT, organic cation transporter; OEA, oleoylethanolamide; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PEA, palmitoylethanolamide; P-gp, P-glycoprotein; PLA2G15, PLA 2 Group 15; PLA2G6, PLA 2 Group 6; PNPLA6, patatin-like phospholipase domain containing 6; SHRSP, Spontaneously hypertensive rats stroke prone; TAMRA-FP, tetramethylrhodamine-fluorophosphonate.

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Section: Discussionmentioning

confidence: 99%

Preclinical pharmacological evaluation of the fatty acid amide hydrolase inhibitor BIA 10‐2474

Bonifácio¹,

Sousa²,

Aires³

et al. 2020

British J Pharmacology

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“…In our approach, we apply the advantage of the high-throughput scanning capability of MALDI MS to measure Affi-BAMS' microarrays containing thousands of spots of bead-captured analytes. The microarrays are produced on standard-size microscope slides that fit the majority of current MALDI MS instruments and have flat surfaces, thereby eliminating the need for orthogonal laser beam geometry, which is a prerequisite for probing arrays of microwells [74].…”

Section: Discussionmentioning

confidence: 99%

Affinity-Bead Assisted Mass Spectrometry (Affi-BAMS): A Multiplexed Microarray Platform for Targeted Proteomics

Hamza

Bergo

Mamaev

et al. 2020

IJMS

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The ability to quantitatively probe diverse panels of proteins and their post-translational modifications (PTMs) across multiple samples would aid a broad spectrum of biological, biochemical and pharmacological studies. We report a novel, microarray analytical technology that combines immuno-affinity capture with Matrix Assisted Laser Desorption Ionization Mass Spectrometry (MALDI MS), which is capable of supporting highly multiplexed, targeted proteomic assays. Termed “Affinity-Bead Assisted Mass Spectrometry” (Affi-BAMS), this LC-free technology enables development of highly specific and customizable assay panels for simultaneous profiling of multiple proteins and PTMs. While affinity beads have been used previously in combination with MS, the Affi-BAMS workflow uses enrichment on a single bead that contains one type of antibody, generally capturing a single analyte (protein or PTM) while having enough binding capacity to enable quantification within approximately 3 orders of magnitude. The multiplexing capability is achieved by combining Affi-BAMS beads with different protein specificities. To enable screening of bead-captured analytes by MS, we further developed a novel method of performing spatially localized elution of targets from individual beads arrayed on a microscope slide. The resulting arrays of micro spots contain highly concentrated analytes localized within 0.5 mm diameter spots that can be directly measured using MALDI MS. While both intact proteins and protein fragments can be monitored by Affi-BAMS, we initially focused on applying this technology for bottom-up proteomics to enable screening of hundreds of samples per day by combining the robust magnetic bead-based workflow with the high throughput nature of MALDI MS acquisition. To demonstrate the variety of applications and robustness of Affi-BAMS, several studies are presented that focus on the response of 4EBP1, RPS6, ERK1/ERK2, mTOR, Histone H3 and C-MET to stimuli including rapamycin, H2O2, EPO, SU11274, Staurosporine and Vorinostat.

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“…In addition, brigatinib inhibits various fibroblast growth factor receptors (FGFRs), several EPH receptors, and non-RTKs at low micromolar concentrations that have been achieved clinically by many other RTK inhibitors. Using a proteome-wide drug screening approach, it was confirmed that brigatinib strongly interacts with ALK, PTK2/FAK1, and PTK2B/FAK2 and has a medium affinity to IGF-1R and EGFR [20]. These results suggest that the non-RTKs FAK/PTK2 and FER, which are strongly inhibited by brigatinib in NF2-deficient meningioma and schwannoma cells, are important for the growth and survival of these tumors.…”

Section: Plos Onementioning

confidence: 85%

Brigatinib causes tumor shrinkage in both NF2-deficient meningioma and schwannoma through inhibition of multiple tyrosine kinases but not ALK

et al. 2021

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Neurofibromatosis Type 2 (NF2) is an autosomal dominant genetic syndrome caused by mutations in the NF2 tumor suppressor gene resulting in multiple schwannomas and meningiomas. There are no FDA approved therapies for these tumors and their relentless progression results in high rates of morbidity and mortality. Through a combination of high throughput screens, preclinical in vivo modeling, and evaluation of the kinome en masse, we identified actionable drug targets and efficacious experimental therapeutics for the treatment of NF2 related schwannomas and meningiomas. These efforts identified brigatinib (ALUNBRIG®), an FDA-approved inhibitor of multiple tyrosine kinases including ALK, to be a potent inhibitor of tumor growth in established NF2 deficient xenograft meningiomas and a genetically engineered murine model of spontaneous NF2 schwannomas. Surprisingly, neither meningioma nor schwannoma cells express ALK. Instead, we demonstrate that brigatinib inhibited multiple tyrosine kinases, including EphA2, Fer and focal adhesion kinase 1 (FAK1). These data demonstrate the power of the de novo unbiased approach for drug discovery and represents a major step forward in the advancement of therapeutics for the treatment of NF2 related malignancies.

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Proteome-wide drug screening using mass spectrometric imaging of bead-arrays

Cited by 11 publications

References 33 publications

Preclinical pharmacological evaluation of the fatty acid amide hydrolase inhibitor BIA 10‐2474

Preclinical pharmacological evaluation of the fatty acid amide hydrolase inhibitor BIA 10‐2474

Affinity-Bead Assisted Mass Spectrometry (Affi-BAMS): A Multiplexed Microarray Platform for Targeted Proteomics

Brigatinib causes tumor shrinkage in both NF2-deficient meningioma and schwannoma through inhibition of multiple tyrosine kinases but not ALK

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