Orally bioavailable styryl derivative of rohitukine-N-oxide inhibits CDK9/T1 and the growth of pancreatic cancer cells.

The chromone alkaloid is one of the classical pharmacophores for cyclin-dependent kinases (CDKs) and represents the first CDK inhibitor to reach clinical trials. Rohitukine (1), a chromone alkaloid isolated from Dysoxylum binectariferum inspired the discovery of several clinical candidates. The N-oxide derivative of rohitukine occurs naturally, with no reports on its biological activity. Herein, we report isolation, biological evaluation, and synthetic modification of rohitukine N-oxide for CDK9/T1 inhibition and antiproliferative activity in cancer cells. Rohitukine N-oxide (2) inhibits CDK9/T1 (IC50 7.6 µM) and shows antiproliferative activity in the colon and pancreatic cancer cells. The chloro-substituted styryl derivatives, 2b, and 2l, inhibit CDK9/T1 with IC50 values of 0.17 and 0.15 µM, respectively. These derivatives display cellular antiproliferative activity in HCT 116 (colon) and MIA PaCa-2 (pancreatic) cancer cells with GI50 values of 2.5-9.7 µM with excellent selectivity over HEK293 (embryonic kidney) cells. Both analogs induce cell death in MIA PaCa-2 cells via inducing intracellular ROS production, reducing mitochondrial membrane potential, and inducing apoptosis. These analogs are metabolically stable in liver microsomes and have a decent oral pharmacokinetics in BALB/c mice. The molecular modeling studies indicated their strong binding at the ATP-binding site of CDK7/H and CDK9/T1.

Discovery of Pongol, the Furanoflavonoid, as an Inhibitor of CDK7/Cyclin H/MAT1 and Its Preliminary Structure-Activity Relationship.

Natural products have been a great source of leads for cancer drug discovery. The cyclin-dependent kinases (CDKs) play a vital role in the initiation and progression of cancer. The CDK-activating kinase, CDK7/cyclin H/MAT1, has recently gained tremendous attention in targeted cancer drug discovery. Herein, we screened a small library of pure natural products in an ADP-Glo CDK7/H kinase assay that yielded a series of furano- and naphthoflavonoids among actives. Pongol (SBN-88), the hydroxy-substituted furanoflavonoid, inhibits CDK7/H as well as CDK9/T1 with IC50 values of 0.93 and 0.83 µM, respectively, and >20-fold selectivity over CDK2/E1 (IC50 > 20 µM). The molecular docking and molecular dynamics simulation revealed that the presence of phenolic -OH in pongol is vital for kinase inhibition, as its absence resulted in a significant loss in activity (e.g., lanceolatin B). The prime MM-GBSA calculations revealed the presence of strong lipophilic and H-bonding interactions of pongol with CDKs.

The medicinal perspective of 2,4-thiazolidinediones based ligands as antimicrobial, antitumor and antidiabetic agents: A review.

2,4-Thiazolidinedione (2,4-TZD), commonly known as glitazone, is a ubiquitous heterocyclic pharmacophore possessing a plethora of pharmacological activities and offering a vast opportunity for structural modification. The diverse range of biological activities endowed with a novel mode of action, low cost, and easy synthesis has attracted the attention of medicinal chemists. Several researchers have integrated the TZD core with different structural fragments to develop a wide range of lead molecules against various clinical disorders. The most common sites for structural modifications at the 2,4-TZD nucleus are the N-3 and the active methylene at C-5. The review covers the recent development of TZD derivatives such as antimicrobial, anticancer, and antidiabetic agents. Various 2,4-TZD based agents or drugs, which are either under clinical development or in the market, are discussed in the study. Different synthetic methodologies for synthesizing the 2,4-TZD core are also included in the manuscript. The importance of various substitutions at N-3 and C-5 and the mechanisms of action and structure-activity relationships are also discussed. We hope this study will serve as a valuable tool for the scientific community engaged in the structural exploitation of the 2,4-TZD core for developing novel drug m\olecules for life-threatening ailments.

Cell-intrinsic Aryl Hydrocarbon Receptor signalling is required for the resolution of injury-induced colonic stem cells.

The aryl hydrocarbon receptor (AHR) is an environmental sensor that integrates microbial and dietary cues to influence physiological processes within the intestinal microenvironment, protecting against colitis and colitis-associated colorectal cancer development. Rapid tissue regeneration upon injury is important for the reinstatement of barrier integrity and its dysregulation promotes malignant transformation. Here we show that AHR is important for the termination of the regenerative response and the reacquisition of mature epithelial cell identity post injury in vivo and in organoid cultures in vitro. Using an integrative multi-omics approach in colon organoids, we show that AHR is required for timely termination of the regenerative response through direct regulation of transcription factors involved in epithelial cell differentiation as well as restriction of chromatin accessibility to regeneration-associated Yap/Tead transcriptional targets. Safeguarding a regulated regenerative response places AHR at a pivotal position in the delicate balance between controlled regeneration and malignant transformation.

Styryl Group, a Friend or Foe in Medicinal Chemistry.

The styryl (Ph-CH=CH-R) group is widely represented in medicinally important compounds, including drugs, clinical candidates, and molecular probes as it positively impacts the lipophilicity, oral absorption, and biological activity. The analysis of matched molecular pairs (styryl vs. phenethyl, phenyl, methyl, H) for the biological activity indicates the superiority aspect of styryl compounds. However, the Michael acceptor site in the styryl group makes it amenable to the nucleophilic attack by biological nucleophiles and transformation to the toxic metabolites. One of the downsides of styryl compounds is isomerization that impacts the molecular conformation and directly affects biological activity. The impact of cis-trans isomerism and isosteric replacements on biological activity is exemplified. We also discuss the styryl group-bearing drugs, clinical candidates, and fluorescent probes. Overall, the present review reveals the utility of the styryl group in medicinal chemistry and drug discovery.

Analyzing the scaffold diversity of cyclin-dependent kinase inhibitors and revisiting the clinical and preclinical pipeline.

Kinases have gained an important place in the list of vital therapeutic targets because of their overwhelming clinical success in the last two decades. Among various clinically validated kinases, the cyclin-dependent kinases (CDK) are one of the extensively studied drug targets for clinical development. Food and Drug Administration has approved three CDK inhibitors for therapeutic use, and at least 27 inhibitors are under active clinical development. In the last decade, research and development in this area took a rapid pace, and thus the analysis of scaffold diversity is essential for future drug design. Available reviews lack the systematic study and discussion on the scaffold diversity of CDK inhibitors. Herein we have reviewed and critically analyzed the chemical diversity present in the preclinical and clinical pipeline of CDK inhibitors. Our analysis has shown that although several scaffolds represent CDK inhibitors, only the amino-pyrimidine is a well-represented scaffold. The three-nitrogen framework of amino-pyrimidine is a fundamental hinge-binding unit. Further, we have discussed the selectivity aspects among CDKs, the clinical trial dose-limiting toxicities, and highlighted the most advanced clinical candidates. We also discuss the changing paradigm towards selective inhibitors and an overview of ATP-binding pockets of all druggable CDKs. We carefully analyzed the clinical pipeline to unravel the candidates that are currently under active clinical development. In addition to the plenty of dual CDK4/6 inhibitors, there are many selective CDK7, CDK9, and CDK8/19 inhibitors in the clinical pipeline.

A rohitukine derivative IIIM-290 induces p53 dependent mitochondrial apoptosis in acute lymphoblastic leukemia cells.

Rohitukine, a chromone alkaloid extracted from Dysoxylum binectariferum, has a propitious anticancer activity. Our previous study shows that a new Rohitukine derivative IIIM-290 restricts the growth of pancreatic cancer in vivo and in vitro. In the present findings, we report the mechanism of cell death induced by IIIM-290 in MOLT-4 cells (acute lymphoblastic leukemia) and its anticancer potential against various murine leukemic tumor models in vivo. We found that IIIM-290 induced apoptosis through upregulation of different apoptotic proteins like PUMA, BAX, cytochrome c, cleaved (active) caspase-3, and cleaved PARP in MOLT-4 cells. Moreover, IIIM-290 abated mitochondrial membrane potential, elevated calcium levels, reactive oxygen species, and arrested growth of MOLT-4 cells in the synthesis (S) phase of the cell cycle. Interestingly, the elevation in proapoptotic markers was p53 dependent-the silencing of p53 abrogated apoptosis (programmed cell death) triggered by IIIM-290 in MOLT-4 cells. Furthermore, IIIM-290 significantly enhanced the survival of animals with P388 and L1210 leukemia. Thus, our results put IIIM-290 as a potential candidate for the anticancer lead.

Evaluation of rohitukine-enriched fraction of Dysoxylum binectariferum Hook.f. (leaves) as anti-arthritic phytopharmaceutical candidate: Chemical standardization, in-vivo validation, formulation development and oral pharmacokinetics.

ETHNOPHARMACOLOGICAL RELEVANCE: Rheumatoid arthritis is a chronic inflammatory disease of joints. Dysoxylum binectariferum Hook.f (Family: Meliaceae) is a Indian medicinal plant which is traditionally being used to heal inflammation of joints. AIM OF THE STUDY: This work was aimed to carry out chemical standardization, in-vitro/in-vivo validation, oral pharmacokinetics and formulation development of anti-arthritic botanical lead, the rohitukine-enriched fraction of D. binectariferum. MATERIALS AND METHODS: The rohitukine-enriched fraction of D. binectariferum was standardized using four chemical markers and was checked for microbial load, heavy metal content, aflatoxins and pesticides. Its in-vitro inhibitory effect on the lipopolysaccharide (LPS) induced production of pro-inflammatory cytokines TNF-α and IL-6 was studied in THP-1 cells. The in-vivo anti-arthritic activity was investigated in collagen-induced arthritis model in DBA/1J mice. The sustained release capsule formulation was developed and characterized for physicochemical and pharmacokinetic properties. RESULTS: Rohitukine and schumaniofioside A were found to be major chemical constituents of the botanical lead. The rohitukine-enriched fraction of D. binectariferum significantly reduced the production of both pro-inflammatory cytokines TNF-α and IL-6 (>50% inhibition at 3.12 µg/mL) in THP-1 cells. In LPS-treated wild-type mice model, the rohitukine-enriched fraction at 200 mg/kg (PO, QD) completely reduced serum TNF-α levels. In transgenic mice model (collagen-induced arthritis in DBA/1J mice), rohitukine-enriched fraction at 100 mg/kg (PO, QD) dose has resulted in >75% reduction of TNF-α/IL-6 serum levels, 68% reduction in anti-mouse type II collagen IgG1 antibody levels, decreased joint proteoglycan loss and reduced paw edema in DBA/1J mice. The sustained release capsule formulation of rohitukine-enriched fraction showed sustained-release of rohitukine over the period of 24 h, and resulted in an improved plasma-exposure of rohitukine in SD rats. CONCLUSIONS: The data presented herein demonstrated anti-arthritic potential of rohitukine-enriched fraction of D. binectariferum and this study will serve as the benchmark for further research on this botanical lead and developed sustained release capsule formulation.

Impurity profiling of anticancer preclinical candidate, IIIM-290.

IIIM-290, an orally bioavailable preclinical candidate is effective in human xenograft models of leukemia, colon and pancreatic cancer. The promising preclinical data of this lead candidate has shown its potential for clinical development. As a part of its preclinical development, impurity profiling of pilot scale batches is one of the most important component of the CMC documentation. Herein, we report impurity profiling, its quantification in different scale-up batches and analytical method validation. Three impurities ranging from 0.09 to 1.25% in preclinical anticancer candidate, IIIM-290 were detected by validated HPLC method. The impurities (Imp-A, Imp-B and Imp-F) were isolated from the partially purified batch of IIIM-290 using semi-preparative HPLC. Isolated impurities were characterized by 1H, 13C NMR, FTIR and ESI-MS spectral data. Based on the characterization data, the sources of these impurities were identified as unreacted starting material (Imp-A), impurity from botanical raw material (Imp-B; impurity carried from starting material) and the chemically transformed product (Imp-F) of Imp-B, respectively.

Design, synthesis, biological evaluation and molecular modelling studies of novel diaryl substituted pyrazolyl thiazolidinediones as potent pancreatic lipase inhibitors.

A series of novel diaryl substituted pyrazolyl 2,4-thiazolidinediones were synthesized via reaction of appropriate pyrazolecarboxaldehydes with 2,4-thiazolidinedione (TZD) and nitrobenzyl substituted 2,4-thiazolidinedione. The resulting compounds were screened in vitro for pancreatic lipase (PL) inhibitory activity. Two assay protocols were performed viz., methods A and B using p-nitrophenyl butyrate and tributyrin as substrates, respectively. Compound 11e exhibited potent PL inhibitory activity (IC50=4.81µM and Xi50=10.01, respectively in method A and B), comparable to that of the standard drug, orlistat (IC50=0.99µM and Xi50=3.72). Presence of nitrobenzyl group at N-3 position of TZD and nature of substituent at para position of phenyl ring at C-3 position of pyrazole ring notably affected the PL inhibitory activity of the tested compounds. Enzyme inhibition kinetics of 11e revealed its reversible competitive inhibition, similar to that of orlistat. Molecular docking studies validated the rationale of pharmacophoric design and are in accordance to the in vitro results. Compound 11e exhibited a potential MolDock score of -153.349kcal/mol. Further, the diaryl pyrazolyl wing exhibited hydrophobic interactions with the amino acids of the hydrophobic lid domain. Moreover, the carbonyl group at 2nd position of the TZD ring existed adjacent to Ser 152 (≈3Å) similar to that of orlistat. A 10ns molecular dynamics simulation of 11e-PL complex revealed a stable binding conformation of 11e in the active site of PL (Maximum RMSD≈3Å). The present study identified novel thiazolidinedione based leads with promising PL inhibitory activity. Further development of the leads might result in potent PL inhibitors.