Synthesis and biological evaluation of pyrrolo[2,1-c][1,4]benzodiazepine (PBD) C8 cyclic amine conjugates

We report examples of a series of novel pyrrolo[2,1-c][1,4]benzodiazepine (PBD) analogues 12-15 prepared from a common functionalized building block 11 that can be conveniently synthesized on a large scale and in optically pure form. Isoindoline analogue 15 is the most cytotoxic agent in this series, has the highest DNA-binding affinity, and shows significant activity in the in vivo hollow fibre assay.     

Synthesis and biological evaluation of estradiol linked pyrrolo[2,1-c][1,4]benzodiazepine (PBD) conjugates as potential anticancer agents

A series of new estradiol linked pyrrolo[2,1-c][1,4]benzodiazepine (E(2)-PBD) conjugates (3a-f, 4a-f and 5a-f) with different linker architectures including a triazole moiety have been designed and synthesized. All the 18 compounds have been evaluated for their anticancer activity and it is observed that some of the compounds particularly 4c-e and 5c,d exhibited significant anticancer activity. The detailed biological aspects relating to the cell cycle effects and tubulin depolymerization activity have been examined with a view to understand the mechanism of action of these conjugates. Among all these conjugates, one of the compound 5c could be considered as the most effective compound particularly against MCF-7 breast cancer cell line.     

Quinazolinone linked pyrrolo[2,1-c][1,4]benzodiazepine (PBD) conjugates: Design,synthesis and biological evaluation as potential anticancer agents

A series of novel quinazolinone linked pyrrolobenzodiazepine (PBD) conjugates were synthesized. These compounds 4a–f and 5a–f were prepared in good yields by linking C-8 of DC-81 with quinazolinone moiety through different alkane spacers. These conjugates were tested for anticancer activity against 11 human cancer cell lines and found to be very potent anticancer agents with GI₅₀ values in the range of <0.1–26.2 μM. Among all the PBD conjugates, one of the conjugate 5c was tested against a panel of 60 human cancer cells. This compound showed activity for individual cancer cell lines with GI₅₀ values of <0.1 μM. The thermal denaturation studies exhibited effective DNA binding ability compared to DC-81 and these results are further supported by molecular modeling studies. The detailed biological aspects of these conjugates on A375 cell line were studied. It was observed that compounds 4b and 5c induced the release of cytochrome c, activation of caspase-3, cleavage of PARP and subsequent cell death. Further, these compounds when treated with A375 cells showed the characteristic features of apoptosis like enhancement in the levels of p53, p21 and p27 inhibition of cyclin dependent kinase-2 (CDK2) and suppression of NF-κB. Moreover, these two compounds 4b and 5c control the cell proliferation by regulating anti-apoptotic genes like (B-cell lymphoma 2) Bcl-2. Therefore, the data generated suggests that these PBD conjugates activate p53 and inhibit NF-κB and thereby these compounds could be promising anticancer agents with better therapeutic potential for the suppression of tumours.     

An asymmetric C8/C8'-tripyrrole-linked sequence-selective pyrrolo[2,1-c][1,4]benzodiazepine (PBD) dimer DNA interstrand cross-linking agent spanning 11 DNA base pairs

A novel sequence-selective extended PBD dimer 4 has been synthesized that binds with high affinity to an interstrand cross-linking site spanning 11 DNA base pairs. Despite its molecular weight (984.07) and length, the molecule has significant DNA interstrand cross-linking potency (approximately 100-fold greater than the clinically used agent melphalan) and sub-micromolar cytotoxicity in a number of tumour cell lines, suggesting that it readily penetrates cellular and nuclear membranes to reach its DNA target.     

Synthesis and biological activity of fluoroquinolone-pyrrolo[2,1-c][1,4]benzodiazepine conjugates

Fluoroquinolones have been synthesized and linked to DC-81 at C8 position through different alkyl chain spacers. These PBD conjugates have exhibited good DNA binding affinity, and a representative compound shows promising in vitro anticancer activity.     

Synthesis of a novel tetrahydroisoquinolino[2,1-c][1,4]benzodiazepine ring system with DNA recognition potential

We report the first stereospecific synthesis and reactivity of a novel tetrahydroisoquinolino[2,1-c][1,4]benzodiazepine ring system with DNA recognition potential.     

Spectroscopic and calorimetric studies on the DNA recognition of pyrrolo[2,1-c][1,4]benzodiazepine hybrids

DNA binding of two hybrid ligands composed of an alkylating pyrrolo[2,1-c][1,4]benzodiazepine (PBD) moiety tethered to either a naphthalimide or a phenyl benzimidazole chromophore was studied by DNA melting experiments, UV and fluorescence titrations, CD spectroscopy and isothermal titration calorimetry (ITC). Binding of both hybrids results in a remarkable thermal stabilization with an increase of DNA melting temperatures by up to 40 °C for duplexes that allow for a covalent attachment of the PBD moiety to guanine bases in their minor groove. CD spectroscopic measurements suggest that the naphthalimide moiety of the drug interacts through intercalation. In contrast, the PBD-benzimidazole hybrid binds in the DNA minor groove with a preference for (A,T)₄G sequences. Whereas the binding of both ligands is enthalpy-driven and associated with a negative entropy, the benzimidazole hybrid exhibits a less favourable binding enthalpy that is counterbalanced by a more favourable entropic term when compared to the naphthalimide hybrid.     

Pyrrolo[2,1-c][1,4]benzodiazepine-anthraquinone conjugates. Synthesis, DNA binding and cytotoxicity

New pyrrolobenzodiazepine-anthraquinone hybrids have been designed and synthesized, found to effectively bind to DNA and also exhibit cytotoxicity against many cancer cell lines     

Synthesis and antitumor activity of novel enediyne-linked pyrrolo[2,1-c][1,4]benzodiazepine hybrids

A series of novel pyrrolo[2,1-c][1,4]benzodiazepine (PBD) hybrids linked with enediyne is described. These compounds were prepared by linking C-8 of DC-81 (1) with an enediyne (10–16) through carbon chain linkers to afford PBD hybrid agents 17–23 in good yields. Most of the hybrids on human cancer cell lines exhibited higher cytotoxicity, and an increase in the sub-G1 population than 1. In a previous article, we have demonstrated that DC-81-indole conjugate agents (3–6) are potent inducers of cell apoptosis in melanoma. In the present article, we investigated whether DC-81-enediyne agents possess more cytotoxicity than 6 on human 293T cells. Our data revealed that treatment of 293T cells with DC-81-enediyne resulted in a significant increase of annexin V binding, caspase-3 degradation, and p53 arrest to identify apoptotic cells than 6. These results suggest that the DC-81-enediyne agents are more efficient in inducing apoptosis than DC-81-indole in 293T cells.     

Synthesis and DNA binding affinity of novel A-C8/C-C2-exo unsaturated alkoxyamido-linked pyrrolo[2,1-c][1,4]benzodiazepine dimers

The synthesis of novel A-C8/C-C2-exo unsaturated alkoxyamido-linked pyrrolo[2,1-c][1,4]benzodiazepine dimers is reported and these dimers show significant DNA binding affinity and they also exhibit moderate anticancer activity.     

Synthesis, DNA binding, and cytotoxicity studies of pyrrolo[2,1-c][1,4]benzodiazepine-anthraquinone conjugates

A series of pyrrolo[2,1-c][1,4]benzodiazepine-anthraquinone conjugates have been prepared and evaluated for their DNA binding ability as well as anticancer activity. Some of these molecules have shown significant anticancer activity in a number of cancer cell lines.     

Phosphonate-linked pyrrolo[2,1-c][1,4]benzodiazepine conjugates: synthesis, DNA-binding affinity and cytotoxicity

Pyrrolobenzodiazepine-diethylphosphonate conjugates have been designed and synthesized that link through two different types of spacers that are simple alkane chain and also a piperazine moiety side-armed with the alkane chains. These pyrrolobenzodiazepine conjugates have exhibited remarkable DNA-binding affinity and improved solubility in water, a representative compound 7d showing promising in vitro cytotoxicity.     

Novel vasopressin V2 receptor-selective antagonists, pyrrolo[2,1-a]quinoxaline and pyrrolo[2,1-c][1,4]benzodiazepine derivatives.

The intent of the work was to study the structure-activity relationships of AVP receptor antagonists bearing a chiral ring as a partial structure since such studies had been reported for only achiral compounds. In the present paper, we deal with compounds consisting of the chiral tricyclic hetero ring (1,2,3,3a,4,5-hexahydropyrrolo[1,2-a]quinoxaline and 1,2,3,10,11,11a-hexahydro-1H-pyrrolo[2,1-c][1,4]benzodiazepine) and 2-phenylbenzanilide analogues. These compounds exhibited a highly selective affinity for V2 receptor, and their stereochemical configuration had a great influence on V2 receptor binding. VP-343 (N-[4-[[(2S,3aR)-2-hydroxy-2,3,3a,4-tetrahydropyrrolo[1,2-a] quinoxalin-5(1H)-yl]carbonyl]phenyl]-4'-methyl[1,1'-biphenyl]-2-ca rboxamide), VP-365 (N-[4-[[(11aS)-2,3,11,11a-tetrahydro-1H-pyrrolo[2,1-c][1,4]benz odiazepin-10(5H)-yl]carbonyl]phenyl][1,1'-biphenyl-2-carboxamide) and VP-339 (N-[4-[[(11aS)-5-oxo-2,3,11,11a-tetrahydro-1H-pyrrolo[2,1-c][1,4]+ ++benzodiazepin-10(5H)-yl]carbonyl]phenyl][1,1'-biphenyl]-2-carboxami de) were the most potent compounds in vitro and in vivo. The IC50 values of VP-343, VP-365 and VP-339 against V2 receptor were 0.772, 1.18 and 0.216 nM, respectively. The ED300 values (dose required to increase three times the urine volume of the control rats; oral administration) of VP-343, VP-365 and VP-339 were 0.22, 0.31 and 0.78 mg/kg, respectively.     

Synthesis and biological evaluation of an N10-Psec substituted pyrrolo[2,1-c][1,4]benzodiazepine prodrug

The first example of an N10-protected (e.g., Psec, 15) pyrrolo[2,1-c][1,4]benzodiazepine (PBD) analogue that retains significant cytotoxicity in a number of tumour cell lines is reported. This prototype could lead to a new generation of clinically useful N10-protected PBD prodrugs.     

1,2,4-benzothiadiazine linked pyrrolo[2,1-c][1,4]benzodiazepine conjugates: synthesis, DNA-binding affinity and cytotoxicity

Benzothiadiazine-pyrrolobenzodiazepine conjugates linked through different alkane spacers have been prepared. These new classes of hybrid molecules exhibit cytotoxicity against many cancer cell lines. Their DNA thermal denaturation studies have been carried out and one of the compounds (4b) elevates the DNA helix melting temperature of the CT-DNA by 6.7 degrees C after incubation for 36 h.     

Solution structure of a 2:1 C2-(2-naphthyl) pyrrolo[2,1-c][1,4]benzodiazepine DNA adduct: molecular basis for unexpectedly high DNA helix stabilization

The naturally occurring pyrrolo[2,1- c][1,4]benzodiazepine (PBD) monomers such as sibiromycin, anthramycin, and tomaymycin form stable covalent adducts with duplex DNA at purine-guanine-purine sites. A correlative relationship between DNA-binding affinity, as measured by enhanced thermal denaturation temperature of calf thymus DNA ( T m), and cytotoxicity is well documented for these naturally occurring compounds and a range of synthetic analogues with sibiromycin having the highest Delta T m value (16.3 degrees C), reflecting favorable hydrogen-bonding interactions between the molecule and DNA bases. We report here that, surprisingly, the structurally simple synthetic C2-(2-naphthyl)-substituted pyrrolo[2,1- c][1,4]benzodiazepine monomer ( 5) has a Delta T m value (15.8 degrees C) similar to that of sibiromycin and significantly higher than the values for either anthramycin (13.0 degrees C) or tomaymycin (2.6 degrees C). 5 also has similar cytotoxic potency to sibiromycin which is widely regarded as the most potent naturally occurring PBD monomer. To investigate this, we have used NMR in conjunction with molecular dynamics to study the 2:1 adduct formed between 5 and the DNA duplex d(AATCTTTAAAGATT) 2. In contrast to the hydrogen-bonding interactions which predominate in the case of sibiromycin and anthramycin adducts, we have shown that the high binding affinity of 5 is due predominantly to hydrophobic (van der Waals) interactions. The high-resolution 2D NOESY, TOCSY, and COSY data obtained have also allowed unequivocal determination of the orientation of the PBD molecule (A-ring toward 3'-end of covalently bound strand), the stereochemistry at the C11 position of the PBD (C11 S), and the conformation of the C2-naphthyl ring which extends along the floor of the minor groove thus optimizing hydrophobic interactions with DNA. These results provide opportunities for future drug design in terms of extending planar hydrophobic groups at the C2 position of PBDs to maximize binding affinity.     

Synthesis and DNA-binding affinity of A-C8/C-C2 alkoxyamido-linked pyrrolo[2,1-c][1,4]benzodiazepine dimers

The synthesis of new A-C8/C-C2 alkoxyamido-linked pyrrolo[2,1-c][1,4]-benzodiazepine dimers have been described in this report. These dimers exhibit significant DNA-binding ability with moderate anticancer activity.     

Synthesis, anticancer activity and apoptosis inducing ability of bisindole linked pyrrolo[2,1-c][1,4]benzodiazepine conjugates

A series of bisindole-pyrrolobenzodiazepine conjugates (5a-f) linked through different alkane spacers was prepared and evaluated for their anticancer activity. All compounds exhibited significant anticancer potency and the most potent compounds 5b and 5e were taken up for detailed studies on MCF-7 cell line. Cell cycle effects were examined apart from investigating the inhibition of tubulin polymerization for compounds 2a, 2b, 5b and 5e at 2μM. FACS analysis showed that at higher concentrations (4 and 8μM) there was an increase of sub-G1 phase cells and decrease of G2/M phase cells, thus indicating that compounds 5b and 5e are effective in causing apoptosis in MCF-7 cells. It was also observed that compounds 5b and 5e showed the down regulation of histone deacetylase protein levels such as HDAC1, 2, 3, 8 and increase in the levels of p21, followed by apoptotic cell death. The apoptotic nature of these compounds was further evidenced by increased expression of cleaved-PARP and active caspase-7 in MCF-7 cells.     

Synthesis,DNA-binding abilities and anticancer activities of triazole-pyrrolo[2,1-c][1,4]benzodiazepines hybrid scaffolds

We synthesized a new series of PBD-hybrid derivatives having tethered triazoles and investigated for their cytotoxicity. The studies indicated that cis-olefin compounds induce higher cytotoxicity with increase in the G1 cell cycle phase compared with the trans-compounds. Quantitative RT-PCR assay indicated that compounds (16a–d) induced G1 phase arrest through down-regulation of cyclin D1 and up-regulation of p21, p27, and p53 mRNA expressions. Compounds 16a–d induced A375 early apoptosis as detected by flow cytometry after double-staining with annexin V and propidium iodide. Moreover, the Western blot analysis showed that A375 treated by compounds (16a–d) resulted in decreased levels of Bcl-2 and Bcl-xL, increased levels of Bax and Bad, and caspase/PARP degradation to identify apoptotic cells.