In silico identification and biological evaluation of a selective MAP4K4 inhibitor against pancreatic cancer.

Inhibiting a specific target in cancer cells and reducing unwanted side effects has become a promising strategy in pancreatic cancer treatment. MAP4K4 is associated with pancreatic cancer development and correlates with poor clinical outcomes. By phosphorylating MKK4, proteins associated with cell apoptosis and survival are translated. Therefore, inhibiting MAP4K4 activity in pancreatic tumours is a new therapeutic strategy. Herein, we performed a structure-based virtual screening to identify MAP4K4 inhibitors and discovered the compound F389-0746 with a potent inhibition (IC50 120.7 nM). The results of kinase profiling revealed that F389-0746 was highly selective to MAP4K4 and less likely to cause side effects. Results of in vitro experiments showed that F389-0746 significantly suppressed cancer cell growth and viability. Results of in vivo experiments showed that F389-0746 displayed comparable tumour growth inhibition with the group treated with gemcitabine. These findings suggest that F389-0746 has promising potential to be further developed as a novel pancreatic cancer treatment.

Identification of a dual TAOK1 and MAP4K5 inhibitor using a structure-based virtual screening approach.

The STE20 kinase family is a complex signalling cascade that regulates cytoskeletal organisation and modulates the stress response. This signalling cascade includes various kinase mediators, such as TAOK1 and MAP4K5. The dysregulation of the STE20 kinase pathway is linked with cancer malignancy. A small-molecule inhibitor targeting the STE20 kinase pathway has therapeutic potential. In this study, a structure-based virtual screening (SBVS) approach was used to identify potential dual TAOK1 and MAP4K5 inhibitors. Enzymatic assays confirmed three potential dual inhibitors (>50% inhibition) from our virtual screening, and analysis of the TAOK1 and MAP4K5 binding sites indicated common interactions for dual inhibition. Compound 1 revealed potent inhibition of colorectal and lung cancer cell lines. Furthermore, compound 1 arrested cancer cells in the G0/G1 phase, which suggests the induction of apoptosis. Altogether, we show that the STE20 signalling mediators TAOK1 and MAP4K5 are promising targets for drug research.

Isoindoline scaffold-based dual inhibitors of HDAC6 and HSP90 suppressing the growth of lung cancer in vitro and in vivo.

This study reports the synthesis of a series of 2-aroylisoindoline hydroxamic acids employing N-benzyl, long alkyl chain and acrylamide units as diverse linkers. In-vitro studies led to the identification of N-benzyl linker-bearing compound (10) and long chain linker-containing compound (17) as dual selective HDAC6/HSP90 inhibitors. Compound 17 displays potent inhibition of HDAC6 isoform (IC50 = 4.3 nM) and HSP90a inhibition (IC50 = 46.8 nM) along with substantial cell growth inhibitory effects with GI50 = 0.76 µM (lung A549) and GI50 = 0.52 µM (lung EGFR resistant H1975). Compound 10 displays potent antiproliferative activity against lung A549 (GI50 = 0.37 µM) and lung H1975 cell lines (GI50 = 0.13 µM) mediated through selective HDAC6 inhibition (IC50 = 33.3 nM) and HSP90 inhibition (IC50 = 66 nM). In addition, compound 17 also modulated the expression of signatory biomarkers associated with HDAC6 and HSP90 inhibition. In the in vivo efficacy evaluation in human H1975 xenografts, 17 induced slightly remarkable suppression of tumor growth both in monotherapy as well as the combination therapy with afatinib (20 mg/kg). Moreover, compound 17 could effectively reduce programmed death-ligand 1 (PD-L1) expression in IFN-γ treated lung H1975 cells in a dose dependent manner suggesting that dual inhibition of HDAC6 and HSP90 can modulate immunosuppressive ability of tumor area.

Isolation of anti-VEGF monoclonal antibodies with neutralizing effects from an Astragalus-induced immune antibody library.

The Astragalus membranaceus polysaccharides (APS) can improve immunity and enhance treatment reactions. This study analyzed the effects of effective antivascular endothelial growth factor (anti-VEGF) antibody production in mice treated with APS. After APS treatment, the serum of mice produced the antibody reactions that can cross-validate VEGF. The isolated single-chain fragment variable (scFv) antibodies could neutralize VEGF and inhibit in vivo tumor growth. Of the scFvs, scFv 4E can significantly compete the interaction of bevacizumab with VEGF. In cell experiments, scFv 4E effectively inhibited human umbilical vein endothelial cells induced by VEGF in vitro. In a matrix gel-assisted angiogenesis model, scFv 4E significantly inhibited angiogenesis reactions. In addition, in a xenograft model established in the colorectal cancer cell strain HCT116, scFv 4E treatment inhibited tumor growth by up to 52.7%. Finally, molecule docking was performed to simulate the complex interactions of scFv 4E and VEGF, the main driving forces of which involve the hydrophobic interactions and hydrogen bonds of Tyr108 and Tyr 109 of the complementarity-determining region H3 loop with VEGF. The results help in establishing antibody library with high diversity for selecting antibodies with specificity. In addition, this study indirectly expounded the correlations of APS enhancing immunity regulation in vivo.

Combination treatment strategy for pancreatic cancer involving the novel HDAC inhibitor MPT0E028 with a MEK inhibitor beyond K-Ras status.

BACKGROUND: Oncogenic K-Ras signaling highly relies on the canonical Ras/MEK/ERK pathway to contribute to pancreatic cancer progression. However, numerous efforts of MEK inhibitors have failed to provide an optimal antitumor effect for pancreatic cancer in practice. The aim of the present work was to develop a more efficacious therapeutic intervention for MEK inhibitors through combination with histone deacetylase (HDAC) inhibitor MPT0E028. METHODS: The effects of combined therapy on cell viability, apoptosis, protein, and RNA expressions were determined by MTT assay, flow cytometry, western blotting, and quantitative PCR analysis. The AsPC-1 xenograft was used to assess antitumor effects in vivo. RESULTS: The co-administration of MPT0E028 and MEK inhibitor yielded synergistic effects on cell viability suppression both in K-Ras mutated and wild-type pancreatic cancer cells and also markedly triggered cell apoptosis. Surprisingly, ERK and epidermal growth factor receptor (EGFR) were activated by the long-term and low-concentration treatment of MPT0E028 or another HDAC inhibitor alone. Whereas, the pharmacological attenuation of ERK signaling dramatically abolished the MPTE028-induced p-ERK and EGFR expression. Overexpression of HDAC4, HDAC6, and MEK, respectively, reversed the cell death induced by the combined treatment. Finally, the combined treatment decreased the tumor volume in an AsPC-1 xenograft model compared to each individual treatment alone. CONCLUSIONS: The synergistic anti-survival effect of the combination was suggested to occur via compensation of the MEK inhibitor for activated ERK. Our results indicate that this combination strategy could benefit patients with pancreatic cancer beyond K-Ras status.

A Novel Selective JAK2 Inhibitor Identified Using Pharmacological Interactions.

The JAK2/STAT signaling pathway mediates cytokine receptor signals that are involved in cell growth, survival and homeostasis. JAK2 is a member of the Janus kinase (JAK) family and aberrant JAK2/STAT is involved with various diseases, making the pathway a therapeutic target. The similarity between the ATP binding site of protein kinases has made development of specific inhibitors difficult. Current JAK2 inhibitors are not selective and produce unwanted side effects. It is thought that increasing selectivity of kinase inhibitors may reduce the side effects seen with current treatment options. Thus, there is a great need for a selective JAK inhibitor. In this study, we identified a JAK2 specific inhibitor. We first identified key pharmacological interactions in the JAK2 binding site by analyzing known JAK2 inhibitors. Then, we performed structure-based virtual screening and filtered compounds based on their pharmacological interactions and identified compound NSC13626 as a potential JAK2 inhibitor. Results of enzymatic assays revealed that against a panel of kinases, compound NSC13626 is a JAK2 inhibitor and has high selectivity toward the JAK2 and JAK3 isozymes. Our cellular assays revealed that compound NSC13626 inhibits colorectal cancer cell (CRC) growth by downregulating phosphorylation of STAT3 and arresting the cell cycle in the S phase. Thus, we believe that compound NSC13626 has potential to be further optimized as a selective JAK2 drug.