M.V.P.S. Vishnuvardhan

DNA-binding affinity and anticancer activity of β-carboline–chalcone conjugates as potential DNA intercalators: Molecular modelling and synthesis

A new series of DNA-interactive β-carboline-chalcone conjugates have been synthesized and evaluated for their in vitro cytotoxicity and DNA-binding affinity. It has been observed that most of these new hybrids have shown potent cytotoxic activities on A-549 (lung adenocarcinoma) cell lines with IC50 values lower than 10 μM. The hybrid 7b is more effective against some of the selected cancer cell lines with IC50 values less than 50 μM. In addition, compounds 7e, 7k, 7p-u has displayed significant elevation in ΔTm of DNA in comparison to Adriamycin, suggesting significant interaction and remarkable DNA stabilization. The DNA intercalation of these new hybrids has been investigated by fluorescence titration, DNA viscosity measurements, molecular docking as well as molecular dynamics and the results are in agreement with the thermal denaturation studies.

Design and synthesis of benzo(c,d)indolone-pyrrolobenzodiazepine conjugates as potential anticancer agents

A series of benzo[c,d]indol-2(1H)one-PBD conjugates (11a-l) have been designed and synthesized as potential anticancer agents. These compounds were prepared by linking the C8-position of DC-81 with a benzo[c,d]indol-2(1H)one moiety through different alkane spacers in good yields and confirmed by (1)H NMR, mass and HRMS data. The DNA binding ability of these conjugates was evaluated by thermal denaturation studies and interestingly, compound 11l showed enhanced DNA binding ability. These compounds were also evaluated for their anticancer activity in selected human cancer cell lines of lung, skin, colon and prostate by using MTT assay method. These new conjugates showed promising anticancer activity with IC(50) values ranging from 1.05 to 36.49 μM. Moreover, cell cycle arrest in SubG1 phase was observed upon treatment of A549 cells with 1 and 2 μM (IC(50)) concentrations of compound 11l and it induced apoptosis. This is confirmed by Annexin V-FITC, Hoechst staining, caspase-3 activity as well as DNA fragmentation analysis.

Synthesis and Biological Evaluation of Benzo[b]furans as Inhibitors of Tubulin Polymerization and Inducers of Apoptosis

A series of benzo[b]furans was synthesized with modification at the 5‐position of the benzene ring by introducing C‐linked substituents (aryl, alkenyl, alkynyl, etc.). These compounds were evaluated for their antiproliferative activities, inhibition of tubulin polymerization, and cell‐cycle effects. Some compounds in this series displayed excellent activity in the nanomolar range against lung cancer (A549) and renal cell carcinoma (ACHN) cancer cell lines. (6‐Methoxy‐5‐((4‐methoxyphenyl)ethynyl)‐3‐methylbenzofuran‐2‐yl)(3,4,5‐trimethoxyphenyl)methanone (26) and (E)‐3‐(6‐methoxy‐3‐methyl‐2‐(1‐(3,4,5‐trimethoxyphenyl)vinyl)benzofuran‐5‐yl)prop‐2‐en‐1‐ol (36) showed significant activity in the A549 cell line, with IC50 values of 0.08 and 0.06 μM, respectively. G2/M cell‐cycle arrest and subsequent apoptosis was observed in the A549 cell line after treatment with these compounds. The most active compound in this series, 36, also inhibited tubulin polymerization with a value similar to that of combretastatin A‐4 (1.95 and 1.86 μM, respectively). Furthermore, detailed biological studies such as Hoechst 33258 staining, DNA fragmentation and caspase‐3 assays, and western blot analyses with the pro‐apoptotic protein Bax and the anti‐apoptotic protein Bcl‐2 also suggested that these compounds induce cell death by apoptosis. Molecular docking studies indicated that compound 36 interacts and binds efficiently with the tubulin protein.

Synthesis and anticancer activity of heteroaromatic linked 4β-amido podophyllotoxins as apoptotic inducing agents

A series of different heteroaromatic linked 4β-amidopodophyllotoxin conjugates (16a-i, 17a-i and 18a-d) were synthesized and evaluated for anticancer activity against five human cancer cell lines. Among the series, one of the compound 17g showed significant antiproliferative activity in A549 (lung cancer) cell line. Flow cytometric analysis showed that 17g arrested the cell cycle in the G2/M phase leading to caspase-3 dependent apoptotic cell death. Further, Hoechst 33258 staining and DNA fragmentation assay also suggests that 17g induces cell death by apoptosis.

Synthesis of a terphenyl substituted 4-aza-2,3-didehydropodophyllotoxin analogues as inhibitors of tubulin polymerization and apoptosis inducers.

A series of terphenyl based 4-aza-2,3-didehydropodophyllotoxin conjugates (8a-r) were synthesized by a straightforward one-step multicomponent synthesis that demonstrated anticancer activity against five human cancer cell lines (lung, colon, renal, prostate and cervical). All the tested compounds showed potent anticancer activity with IC50 values ranging from 0.87 to 16.59 μM. Among them compounds 8n and 8p showed significant anticancer activity in lung cancer cells with IC50 values 0.91 and 0.87 μM, respectively. Flow cytometric analysis revealed that these compounds induced cell cycle arrest in G2/M phase in A549 cell line leading to caspase-3 dependent apoptotic cell death. The tubulin polymerization assay and immunofluorescence analysis showed that these compounds effectively inhibit microtubule assembly at both molecular and cellular levels in A549 cells. Further, Hoechst staining, DNA fragmentation analysis also suggested that these compounds induced cell death by apoptosis. Overall, the current study demonstrated that the synthesis of terphenyl based 4-aza-2,3-didehydropodophyllotoxin conjugates as promising anticancer agents with G2/M cell cycle arrest and apoptotic-inducing activities via targeting tubulin.

Synthesis and selective cytotoxic activity of novel hybrid chalcones against prostate cancer cells.

A new class of hybrid chalcones (17a-l &18a-l) was synthesized by Claisen-Schmidt condensation. All compounds were characterized by (1)H NMR, IR and mass spectral analysis and tested for their cytotoxic activity against PC-3 (prostate cancer), HT-29 (colon cancer), B-16 (mouse macrophages) and NCI-H460 (lung cancer) cell lines. Three compounds 18i, 18j and 18l (IC(50)=8.4, 7.9 & 5.9 μM) showed significant activity against PC-3 cell line.

Synthesis of 2-aryl-1,2,4-oxadiazolo-benzimidazoles: Tubulin polymerization inhibitors and apoptosis inducing agents

A new series of 2-aryl 1,2,4-oxadiazolo-benzimidazole conjugates have been synthesized and evaluated for their antiproliferative activity in the sixty cancer cell line panel of the National Cancer Institute (NCI). Compounds 5l (NSC: 761109/1) and 5x (NSC: 761814/1) exhibited remarkable cytotoxic activity against most of the cancer cell lines in the one dose assay and were further screened at five dose concentrations (0.01, 0.1, 1, 10 and 100 μM) which showed GI50 values in the range of 0.79-28.2 μM. Flow cytometric data of these compounds showed increased cells in G2/M phase, which is suggestive of G2/M cell cycle arrest. Further, compounds 5l and 5x showed inhibition of tubulin polymerization and disruption of the formation of microtubules. These compounds induce apoptosis by DNA fragmentation and chromatin condensation as well as by mitochondrial membrane depolarization. In addition, structure activity relationship studies within the series are also discussed. Molecular docking studies of compounds 5l and 5x into the colchicine-binding site of the tubulin, revealed the possible mode of interaction by these compounds.

Combretastatin linked 1,3,4-oxadiazole conjugates as a Potent Tubulin Polymerization inhibitors

A new class of combretastatin linked 1,3,4-oxadiazoles were designed, synthesized and screened for their cytotoxic activity against five human cancer cell lines such as HeLa, DU-145, A549, MDA-MB-231 and B16. These compounds showed significant cytotoxicity with IC50 values in the range 0.118-54.32μM. Conjugate 5m displayed potent antiproliferative activity against DU-145 cell line. Flow cytometric analysis revealed that these compounds arrested the cell cycle in G2/M phase. Moreover, the tubulin polymerization assay and immunofluorescence analysis indicate that 5m exhibits potent inhibitory effect on the tubulin assembly. Further, DNA fragmentation and Hoecst staining assays confirm that 5m induces apoptosis. Molecular docking studies and competitive binding assay indicated that 5m effectively bind at the colchicine binding site of the tubulin.

Investigation of the mechanism and apoptotic pathway induced by 4β cinnamido linked podophyllotoxins against human lung cancer cells A549

Apoptosis is essential for normal development and the maintenance of homeostasis. It plays a necessary role to protect against carcinogenesis by eliminating damaged cells. Many studies have demonstrated that the dysregulation of apoptosis results in cancer and this provides an approach to develop therapeutic agents via inducing apoptosis. In our previous studies 4β-cinnamido linked podophyllotoxin conjugates were synthesized and evaluated for their cytotoxic activity in a panel of five human cancer cell lines and the new molecules like 17a and 17f were considered as potential leads. The cytotoxic activity was comparable to etoposide. These observations prompted us to investigate the mechanism underplaying the cytotoxic activity and apoptotic pathway induced by these compounds in human lung cancer cells A459. The results of the present study revealed that these compounds exhibited DNA topoisomerase IIα inhibition and induced mitochondrial mediated apoptosis. It was further confirmed by Mitochondrial membrane potential, Cytochrome c release, cleavage of poly (ADP-ribose) polymerase (PARP), Reactive oxygen species (ROS) generation, regulation of antiapoptotic protein Bcl-2 and pro apoptotic protein Bax studied by Western blot analysis. Annexin V-FITC assay also suggested that these compounds induced cell death by apoptosis. Pretreatment with N-acetyl-l-cysteine (NAC) prevented the generation of ROS. Further, pretreatment with NAC significantly inhibited 17a and 17f induced apoptosis, suggesting that ROS are the key mediators for 17a and 17f induced apoptosis. These data indicate that these compounds might induce apoptosis in A549 cells through a ROS mediated mitochondrial dysfunction pathway. Moreover, these compounds did not significantly inhibit the noncancerous human embryonic kidney cells, HEK-293. Docking studies also elucidate the potential of these molecules to bind to the DNA topoisomerase II.Graphical AbstractPodophyllotoxin analogs were investigated for their mechanism and apoptotic pathway against lung cancer cell line, A549. These podophyllotoxin analogs inhibited DNA topoisomerase IIα and induced mitochondrial mediated apoptosis in lung cancer cell line, A549. Western blot analysis suggested that these compounds inhibited the DNA topoisomerase IIα. Studies like, Measurement of mitochondrial membrane potential (∆Ψm), Generation of intracellular reactive oxygen species (ROS) and Annexin V-FITC assay suggested that these compounds induced mitochondrial mediated apoptosis. Pretreatment with N-acetyl-l-cysteine (NAC) suggested that ROS plays a role in 17a and 17f induced apoptosis. Further the apoptotic effect of these compounds was confirmed by western blot analysis of pro apoptotic protein Bax and antiapoptotic protein Bcl-2, Cytochrome c release and cleavage of poly (ADP-ribose) polymerase (PARP). Moreover, these compounds did not significantly inhibit the noncancerous human embryonic kidney cells, HEK-293.

Synthesis and biological evaluation of cinnamido linked benzophenone hybrids as tubulin polymerization inhibitors and apoptosis inducing agents.

A new class of hybrid molecules containing cinnamide subunit linked to benzophenone as inhibitors of tubulin polymerization were synthesized and evaluated for their anticancer potential. These hybrids exhibit anticancer activity with IC50 values ranging from 0.06 to 16.3μM. Compounds 4f and 4g possessing fluoro and trifluoromethyl on the cinnamido subunit showed significant cytotoxic activity with IC50 values 0.06 and 0.09μM against HeLa cell line, respectively. These compounds showed cell cycle arrest at G2/M phase of the cell cycle and inhibited tubulin polymerization followed by activation of caspase-3 activity and apoptotic cell death. Further in vitro tubulin polymerization assay showed that the level of tubulin inhibition was comparable to that of 2a for the compounds 4f and 4g. Moreover, Hoechst 33258 staining and DNA fragmentation assay suggested that these compounds induce cell death by apoptosis. Overall, the current study demonstrates that the synthesis of benzophenone linked cinnamide subunit conjugates as promising anticancer agents with G2/M arrest and apoptotic-inducing ability via targeting tubulin.