Target engagement approaches for pharmacological evaluation in animal models

2019 ◽  
Vol 55 (63) ◽  
pp. 9241-9250 ◽  
Author(s):  
James E. Kath ◽  
Aleksandra Baranczak
Keyword(s):  
Animal Models ◽  
Small Molecules ◽  
Chemical Biology ◽  
Target Engagement ◽  
Pharmacological Evaluation

We highlight recent applications of chemical biology approaches to measure target engagement of small molecules in animal models to support the nomination of clinical candidates.

scholarly journals P013 The JAK-3 and TYK-2 / STAT pathways are activated in moderate to severe ulcerative colitis

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S136-S137
Author(s):  
M Loza ◽  
J M Brea ◽  
C Calviño-Suarez ◽  
I Baston-Rey ◽  
R Ferreiro-Iglesias ◽  
...  
Keyword(s):  
Ulcerative Colitis ◽  
Animal Models ◽  
Inflammatory Response ◽  
Western Blot ◽  
Signalling Pathways ◽  
Target Engagement ◽  
Inclusion Criteria ◽  
Phosphorylated Proteins ◽  
Single Centre Study ◽  
Stat Pathway

Abstract Background Ulcerative colitis (UC) is a chronic, progressive and disabling disease with a complex pathology of unknown aetiology influenced by genetic, environmental and microbiota factors that lead to an immunological and inflammatory response in the colon. Janus Activated Kinase (JAK) family plays a key role in modulating the adaptive and innate inflammatory response. The JAK/STAT pathway involvement in UC has been demonstrated in both animal models and human studies. Thus, overexpressed JAK-3 has been detected in the intestine of patients with UC, suggesting a key role in their pathophysiology and the inhibition of TYK-2 in animal models resulted in an improvement of the disease, which would explain its implication in the inflammatory process. We hypothesise here that there could be an activation of JAK-3 and TYK-2 signalling pathways in UC patients. Thus, we aimed to detect the activation of both signalling pathways by means of western-blot studies in UC patient samples Methods A prospective, observational single-centre study was designed. Inclusion criteria were adult patients with endoscopic active UC (more than Mayo-0) confirmed in a programmed colonoscopy. All patients signed informed consent. Samples were obtained from overstock of routine biopsies in the more severe segment affected of the large bowel. Tissues were homogenised and processed in order to obtain cell lysates by employing RIPA buffer and ultrasounds. The degree of activation of the JAK-3 and TYK-2 pathways was measured by detecting the phosphorylation of both targets as well as of STAT1, STAT3, STAT4, STAT5 and STAT6 through western blot by employing specific antibodies for total and phosphorylated proteins. Results 19 UC patients were consecutively included. Mean age was 46 years old. 53% were female, 47% were extensive colitis (E3) and 53% left-side colitis (E2). Regarding endoscopic activity, 26% had Mayo-1, 53% Mayo-2, and 21% Mayo-3. Immunoreactive bands for both phosphorylated JAK-3 and TYK-2 were detected in the biopsies from UC patients, evidencing that colonic inflammation leads to an activation of both targets. The study of STATs phosphorylation showed immunoreactive bands for phosphorylated forms of STAT1, STAT3, STAT4, STAT5 and STAT6 confirming the activation of both signalling-pathways in these patients (Figure 1). Conclusion The developed translational workflows involving basic/clinical research confirm the activation of both JAK-3 and TYK-2-dependent signalling pathways in UC patients, validating both kinases as targets for treating UC. The developed methodology allows studying the target engagement for future JAK-3/ TYK-2 inhibitors employed in clinical trials.

scholarly journals Improving Precise CRISPR Genome Editing by Small Molecules: Is there a Magic Potion?

Cells ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 1318 ◽  
Author(s):  
Nadja Bischoff ◽  
Sandra Wimberger ◽  
Marcello Maresca ◽  
Cord Brakebusch
Keyword(s):  
Gene Therapy ◽  
Dna Repair ◽  
Animal Models ◽  
Molecular Biology ◽  
Small Molecules ◽  
Genome Editing ◽  
Drug Targets ◽  
Targeted Mutagenesis ◽  
Dna Repair Mechanisms ◽  
Repair Mechanisms

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) genome editing has become a standard method in molecular biology, for the establishment of genetically modified cellular and animal models, for the identification and validation of drug targets in animals, and is heavily tested for use in gene therapy of humans. While the efficiency of CRISPR mediated gene targeting is much higher than of classical targeted mutagenesis, the efficiency of CRISPR genome editing to introduce defined changes into the genome is still low. Overcoming this problem will have a great impact on the use of CRISPR genome editing in academic and industrial research and the clinic. This review will present efforts to achieve this goal by small molecules, which modify the DNA repair mechanisms to facilitate the precise alteration of the genome.

scholarly journals Small molecules with big roles in microRNA chemical biology and microRNA-targeted therapeutics

RNA Biology ◽  
2019 ◽  
Vol 16 (6) ◽  
pp. 707-718 ◽  
Author(s):  
Rengen Fan ◽  
Chaocheng Xiao ◽  
Xinqiang Wan ◽  
Wenzhang Cha ◽  
Yufeng Miao ◽  
...  
Keyword(s):  
Small Molecules ◽  
Chemical Biology ◽  
Targeted Therapeutics

scholarly journals CETSA MS Profiling for a Comparative Assessment of FDA-Approved Antivirals Repurposed for COVID-19 Therapy Identifies TRIP13 as a Remdesivir Off-Target

2020 ◽  
pp. 247255522097359
Author(s):  
Tomas Friman ◽  
Alexey Chernobrovkin ◽  
Daniel Martinez Molina ◽  
Laurence Arnold
Keyword(s):  
Small Molecules ◽  
Antiviral Drugs ◽  
Biologically Active ◽  
Target Engagement ◽  
Intact Cells ◽  
Protein Targets ◽  
Antiviral Compounds ◽  
Binding Partners ◽  
Cellular Thermal Shift Assay ◽  
Target Effects

The reuse of preexisting small molecules for a novel emerging disease threat is a rapid measure to discover unknown applications for previously validated therapies. A pertinent and recent example where such a strategy could be employed is in the fight against coronavirus disease 2019 (COVID-19). Therapies designed or discovered to target viral proteins also have off-target effects on the host proteome when employed in a complex physiological environment. This study aims to assess these host cell targets for a panel of FDA-approved antiviral compounds including remdesivir, using the cellular thermal shift assay (CETSA) coupled with mass spectrometry (CETSA MS) in noninfected cells. CETSA MS is a powerful method to delineate direct and indirect interactions between small molecules and protein targets in intact cells. Biologically active compounds can induce changes in thermal stability, in their primary binding partners, and in proteins that in turn interact with the direct targets. Such engagement of host targets by antiviral drugs may contribute to the clinical effect against the virus but can also constitute a liability. We present here a comparative study of CETSA molecular target engagement fingerprints of antiviral drugs to better understand the link between off-targets and efficacy.

scholarly journals From Phenotypic Hit to Chemical Probe: Chemical Biology Approaches to Elucidate Small Molecule Action in Complex Biological Systems

Molecules ◽  
2020 ◽  
Vol 25 (23) ◽  
pp. 5702
Author(s):  
Quentin T. L. Pasquer ◽  
Ioannis A. Tsakoumagkos ◽  
Sascha Hoogendoorn
Keyword(s):  
Small Molecules ◽  
Chemical Biology ◽  
Target Identification ◽  
Biologically Active ◽  
Phenotypic Screening ◽  
Biological Processes ◽  
Chemical Probes ◽  
Target Deconvolution ◽  
Cellular Target ◽  
Screening Approaches

Biologically active small molecules have a central role in drug development, and as chemical probes and tool compounds to perturb and elucidate biological processes. Small molecules can be rationally designed for a given target, or a library of molecules can be screened against a target or phenotype of interest. Especially in the case of phenotypic screening approaches, a major challenge is to translate the compound-induced phenotype into a well-defined cellular target and mode of action of the hit compound. There is no “one size fits all” approach, and recent years have seen an increase in available target deconvolution strategies, rooted in organic chemistry, proteomics, and genetics. This review provides an overview of advances in target identification and mechanism of action studies, describes the strengths and weaknesses of the different approaches, and illustrates the need for chemical biologists to integrate and expand the existing tools to increase the probability of evolving screen hits to robust chemical probes.

Protein degradation: a validated therapeutic strategy with exciting prospects

2017 ◽  
Vol 61 (5) ◽  
pp. 517-527 ◽  
Author(s):  
Honorine Lebraud ◽  
Tom D. Heightman
Keyword(s):  
Animal Models ◽  
Protein Degradation ◽  
Small Molecules ◽  
Cancer Patients ◽  
Clinical Efficacy ◽  
Therapeutic Strategy ◽  
Substrate Recognition ◽  
Pharmacological Targets ◽  
Preclinical Animal Models

In a time of unprecedented challenges in developing potent, selective and well-tolerated protein inhibitors as therapeutics, drug hunters are increasingly seeking alternative modalities to modulate pharmacological targets. Selective inhibitors are achievable for only a fraction of the proteome, and are not guaranteed to elicit the desired response in patients, especially when pursuing targets identified through genetic knockdown. Targeted protein degradation holds the potential to expand the range of proteins that can be effectively modulated. Drugs inducing protein degradation through misfolding or by modulating cereblon (CRBN) substrate recognition are already approved for treatment of cancer patients. The last decade has seen the development of proteolysis targeting chimeras (PROTACs), small molecules that elicit proteasomal degradation by causing protein polyubiquitination. These have been used to degrade a range of disease-relevant proteins in cells, and some show promising efficacy in preclinical animal models, although their clinical efficacy and tolerability is yet to be proven. This review introduces current strategies for protein degradation with an emphasis on PROTACs and the role of click chemistry in PROTAC research through the formation of libraries of preclicked PROTACs or in-cell click-formed PROTACs (CLIPTACs).

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