Molecular docking approach for the design and synthesis of new pyrazolopyrimidine analogs of roscovitine as potential CDK2 inhibitors endowed with pronounced anticancer activity.

Cyclin-dependent kinase 2 (CDK2) is a vital protein for controlling cell cycle progression that is critically associated with various malignancies and its inhibition could offer a convenient therapeutic approach in designing anticancer remedies. Consequently, this study aimed to design and synthesize new CDK2 inhibitors featuring roscovitine as a template model. The purine ring of roscovitine was bioisosterically replaced with the pyrazolo[3,4-d]pyrimidine scaffold, in addition to some modifications in the side chains. A preliminary molecular docking study for the target chemotypes in the CDK2 binding domain revealed their ability to accomplish similar binding patterns and interactions to that of the lead compound roscovitine. Afterwards, synthesis of the new derivatives was accomplished. Then, the initial anticancer screening at a single dose by the NCI revealed that compounds 7a, 9c, 11c, 17a and 17b achieved the highest GI% values reaching up to 150 % indicating their remarkable activity. These derivatives were subsequently selected to undertake five-dose testing, where compounds 7a, 9c, 11c and 17a unveiled the most pronounced activity against almost the full panel with GI50 ranges; 1.41-28.2, 0.116-2.39, 0.578-60.6 and 1.75-42.4 µM, respectively and full panel GI50 (MG-MID); 8.24, 0.6, 2.46 and 6.84 µM, respectively. CDK2 inhibition assay presented compounds 7a and 9c as the most potent inhibitors with IC50 values of 0.262 and 0.281 µM, respectively which are nearly 2.4 folds higher than the reference ligand roscovitine (IC50 = 0.641 µM). Besides, flow cytometric analysis on the most susceptible and safe cell lines depicted that 7a caused cell cycle arrest at G1/S phase in renal cancer cell line (RXF393) while 9c led to cell growth arrest at S phase in breast cancer cell line (T-47D) along with pronounced apoptotic induction in the mentioned cell lines. These findings afforded new anticancer pyrazolo[3,4-d]pyrimidine, roscovitine analogs, acting via CDK2 inhibition.

Hamed OA ，Abou-Elmagd El-Sayed N ，Mahmoud WR ，F Elmasry G ... -  《-》

Novel purine derivatives as selective CDK2 inhibitors with potential anticancer activities: Design, synthesis and biological evaluation.

Purine analogues were discovered to be inhibitors of CDK2, suggesting a potential therapeutic scaffold. This paper addresses the design, synthesis, and anticancer evaluation of purine analogues as kinase inhibitors. In the early stages of the investigation, the designed compounds demonstrated a promising docking score and greater protein-ligand stability in MD simulation than the standard, indicating a higher affinity against CDK2. Thus, we synthesised new purine analogues under simple and optimised reaction conditions. Among the studies under NCI-60, 5g and 5i were the most effective, with a percentage GI of 98.09 and 90 against OVCAR-4 and SNB-75, respectively, at a dose of 10 µM. Additionally, 5g and 5i demonstrated 7.80-fold and 1.54-fold greater cytotoxicity against PA-1 and MCF-7, with IC50s of 1.08 µM and 3.54 µM, respectively, compared to seliciclib (8.43 µM and 5.46 µM). In addition, 5g and 5i showed selective cytotoxicity against PA-1 and MCF-7 than normal cells, with selectivity indexes of 26.40 and 15.45, respectively, as compared to the standard (SI=3.83 and 5.91). In the kinase selectivity assay, both compounds demonstrated greater affinity against CDK2 than other kinases, with IC50 of 0.21 µM and 0.59 µM, in contrast to the standard (IC50 = 0.63 µM). Furthermore, 5g confirmed kinase inhibition in the western blot by lowering CDK2, cyclin A2, and other downstream substrates. Moreover, it triggered cell death by apoptosis and cell cycle arrest in G2/M. Taken together, 5g merits further investigation in PKPD research to discover a potential therapeutic candidate against cancer.

Shah A ，Teraiya N ，Kamdar JH ，Juneja T ，Sangani CB ，Ahmed S ，Kapadiya K ... -  《-》

Synthesis, design, and antiproliferative evaluation of 6-(N-Substituted-methyl)pyrazolo[3,4-d]pyrimidines as the potent anti-leukemia agents.

Pyrazolopyrimidine derivatives, including pyrazolopyrimidines, 6-aminopyrazolopyrimidines, 6-[(formyloxy)methyl]pyrazolopyrimidines, 6-(hydroxymethyl)pyrazolopyrimidine, and 6-(aminomethyl)pyrazolopyrimidines have been successfully prepared and tested against NCI-H226, NPC-TW01, and Jurkat cancer cell lines. Among the tested pyrazolopyrimidine compounds, we found 6-aminopyrazolopyrimidines and 6-(aminomethyl)pyrazolopyrimidines with essential o-ClPh or p-ClPh substituted moieties on N-1 pyrazole ring exhibited the best IC50 inhibition activity for Jurkat cells. Furthermore, optimization of the SAR study on the C-6 position of pyrazolopyrimidine ring demonstrated that 6-(N-substituted-methyl)pyrazolopyrimidines 17b, 17d, and 19d possessed the significant IC50 inhibitory activity for the different leukemia cell lines, especially for Jurkat, K-562, and HL-60. On the other hand, further SAR inhibition and docking model studies revealed that compound 19d, which has a 3-(1H-imidazol-1-yl)propan-1-amino side-chain on the C-6 position, was able to form four hydrogen bonds with residues Ala226, Leu152, and Glu194 and specifically extended into the P1 pocket subsite with Aurora A, resulting in improved inhibitory activity almost similar to SNS-314. To explore the anti-cancer mechanism, compound 19d was measured by Western blot analysis in Jurkat T-cells, however, it showed non-responsibility to Aurora B. For the further structural modifications on the lateral chain of compound 19d, compounds 24 with longer lateral chain were designed and synthesized for testing leukemia cell lines. However, compounds 24 was significantly decrease inhibition potency against leukemia cell lines. Based on the in-vitro results, compounds 17b and 19d could be considered to be the best potential lead drug in our study for the development of new and effective therapies for leukemia treatment. On the other hand, the DHFR inhibition results indicated compound 19d possessed good inhibitory activity and better than the reported naphthalene derivative. Through further comparisons of the model superposition of three-dimensional (3D) conformations in DHFR, compound 19d presented a similar structural alignment to Methotrexate and the reported naphthalene derivative and led to similar drug-like functional relationships. As a results, compound 19d would be a potential DHFR inhibitor for anti-leukemia drug candidate.

Chung CY ，Li SM ，Zeng WZ ，Uramaru N ，Huang GJ ，Juang SH ，Wong FF ... -  《-》

Discovery of novel diaryl urea-oxindole hybrids as BRAF kinase inhibitors targeting BRAF and KRAS mutant cancers.

In the current study, a novel series of diaryl urea incorporating oxindole moiety was rationally designed as type II BRAF inhibitors targeting BRAF and KRAS mutant cancers. Molecular hybridization between the diaryl urea scaffold which binds to the inactive conformation of protein kinases on one side and the oxindole core which exhibit adenine mimic properties to be settled in the hinge region on the other side was performed. Studying the antiproliferative activity of the synthesized candidates 9a-t on NCI cancer cell lines showed that they exhibit potent and broad spectrum of antiproliferative activity on the tested cancer cell lines with compounds 9c, 9p, 9q, 9s, and 9t demonstrating potent GI50 reaching 0.01 µM. Noteworthy, compound 9s demonstrated a potent GI50 on cell lines expressing mutant KRAS and those express BRAFV600E with GI50 ranges of 1.79 and 7.94 µM and 1.68 to 2.0 µM, respectively. Further analysis on A375 and Mel501 cell lines expressing BRAFV600E revealed that compound 9s has a potent growth inhibitory activity with IC50 of 0.7 and 1.5 µM, respectively, in reference to sorafenib (IC50 = 8.7 and 0.3 µM, respectively). Additionally, nearly all the target candidates did not show any cytotoxic effect on the normal fibroblast cell line BJ-1 with compound 9s showing IC50 of 20.2 µM in reference to sorafenib (IC50 = 6.1 µM). Further cellular assays on A375 cell line, revealed the ability of compound 9s to halt the cell cycle progression at the G2 phase besides its ability to induce apoptosis. In parallel, all the synthesized candidates 9a-t were biochemically evaluated for their inhibitory activity on BRAFWT and compounds 9b, 9c, and 9n revealed a sub-micromolar IC50 of 0.11, 0.84 and 0.80 µM, respectively. Further investigation of selected compounds on BRAFV600E showed that compounds 9c, 9n, 9s, and 9t exhibit a sub-micromolar IC50 range of 0.17 to 0.89 µM. Noteworthy, the examined candidates demonstrated a higher selectively towards BRAFV600E over BRAFWT highlighting their promising optimization for treating BRAFV600E expressing cancers. Molecular docking and molecular dynamics simulations in the inactive DFG-out kinase domain of BRAFWT/V600E protein kinases confirmed the planned design strategy.

Ghannam IAY ，El Kerdawy AM ，Mounier MM ，Abo-Elfadl MT ，Abdel-Mohsen HT ... -  《-》

New molecular hybrids integrated with quinoxaline and pyrazole structural motifs: VGFR2 inhibitors and apoptosis inducers.

The vascular endothelial growth factor receptor is essential for the angiogenesis of cancer. Tumor propagation was effectively suppressed by inhibiting VEGFR-2 activity. As a result, the target quinoxaline-pyrazole hybrids were created in a way that closely resembled the structural characteristics of VEGFR-2 inhibitors. The synthesized compounds were assessed in vitro using the MTT assay with doxorubicin serving as a reference standard for their cytotoxic properties against the HCT-116 and MCF-7 cell lines. Additionally, when tested on human normal fibroblasts (WI38), the promising cytotoxic compounds 8, 11, 13, and 15 were shown to be selective to cancer cells. Using ELISA, they showed mechanistically inhibitory activities against VEGFR-2; compound 13 was the most effective inhibitor, with an IC50 of 0.045 ± 6.24 uM, surpassing sorafenib (IC50 of 0.049 ± 5.24 μM). Notably, it was discovered that our target compound, 13, was 1.1 times more potent than sorafenib and 3.19 times more potent than sunitinib as a VGFRA2 inhibitor. Furthermore, Western blot analysis revealed that its VEGFR2 protein levels were noticeably higher than the control. Compound 13's selectivity towards VEGFR2 was further confirmed by testing it against other kinases, such as PDGFRA (IC50 0.329 ± 0.014 μM) and EGFR (IC50 0.6 ± 0.019 μM). Furthermore, 13 demonstrated a 50 % decrease in VEGF-A secretion in comparison to the control group, demonstrating its anti-angiogenic quality. A scratch closure percentage of 57.78 %, which was much lower than the 97.04 percent of untreated control cells, showed 13's anti-migratory capability. According to the cell cycle study, compound 13 induces apoptosis at the sub-G1 phase and terminates the cell cycle at the G1 phase. Consequently,flow cytometric analysis revealed that it caused apoptosis; compound 13 increases the BAX/Bcl-2 ratio from control to 13.66, and it also activates caspase 3 to 422.48 ± 43.82 and induces p53 to 366.79 ± 40.21. Docking simulations revealed potential binding modes and crucial structural elements of active drugs, and they were almost in agreement with enzymatic examination. For every hybrid, in silico physicochemical attributes, drug likeness metrics, and ligand efficiency were plausible. It's interesting to note that 13 and 15 are plausible medication candidates.

Ismail MMF ，Shawer TZ ，Ibrahim RS ，Elnagar MR ，Ammar YA ... -  《-》