C8-linked Pyrrolobenzodiazepine Research Articles

Mechanistic insight into the repair of C8-linked pyrrolobenzodiazepine monomer-mediated DNA damage.

Pyrrolobenzodiazepines (PBDs) are naturally occurring DNA binding compounds that possess anti-tumor and anti-bacterial activity. Chemical modifications of PBDs can result in improved DNA binding, sequence specificity and enhanced efficacy. More recently, synthetic PBD monomers have shown promise as payloads for antibody drug conjugates and anti-bacterial agents. The precise mechanism of action of these PBD monomers and their role in causing DNA damage remains to be elucidated. Here we characterized the damage-inducing potential of two C8-linked PBD bi-aryl monomers in Caulobacter crescentus and investigated the strategies employed by cells to repair the same. We show that these compounds cause DNA damage and efficiently kill bacteria, in a manner comparable to the extensively used DNA cross-linking agent mitomycin-C (MMC). However, in stark contrast to MMC which employs a mutagenic lesion tolerance pathway, we implicate essential functions for error-free mechanisms in repairing PBD monomer-mediated damage. We find that survival is severely compromised in cells lacking nucleotide excision repair and to a lesser extent, in cells with impaired recombination-based repair. Loss of nucleotide excision repair leads to significant increase in double-strand breaks, underscoring the critical role of this pathway in mediating repair of PBD-induced DNA lesions. Together, our study provides comprehensive insights into how mono-alkylating DNA-targeting therapeutic compounds like PBD monomers challenge cell growth, and identifies the specific mechanisms employed by the cell to counter the same.

Effects of Systematic Shortening of Noncovalent C8 Side Chain on the Cytotoxicity and NF-κB Inhibitory Capacity of Pyrrolobenzodiazepines (PBDs).

The systematic shortening of the noncovalent element of a C8-linked pyrrolobenzodiazepine (PBD) conjugate (13) led to the synthesis of a 19-member library of C8-PBD monomers. The critical elements of 13, which were required to render the molecule cytotoxic, were elucidated by an annexin V assay. The effects of shortening the noncovalent element of the molecule on transcription factor inhibitory capacity were also explored through an enzyme-linked immunosorbent assay-based measurement of nuclear NF-κB upon exposure of JJN-3 cells to the synthesized molecules. Although shortening the noncovalent interactive element of 13 had a less than expected effect upon compound cytotoxicity due to reduced DNA interaction, the transcription factor inhibitory capacity of the molecule was notably altered. This study suggests that a relatively short noncovalent side chain at the C8 position of PBD is sufficient to confer cytotoxicity. The shortened PBD monomers provide a new ADC payload scaffold because of their potent cytotoxicity and drug-like properties.

Activity of DNA-targeted C8-linked pyrrolobenzodiazepine–heterocyclic polyamide conjugates against aerobically and hypoxically grown Mycobacterium tuberculosis under acidic and neutral conditions

Mycobacterium tuberculosis (Mtb) is the aetiological agent of tuberculosis, the leading cause of death worldwide from a single infectious agent. Mtb is a highly adaptable human pathogen that might enter a dormant non-replicating (NR), drug-tolerant stage. Reactivation of dormant Mtb can lead to active disease. Antibiotic treatments of active and latent tuberculosis are long, complex and may fail to fully eradicate the infection. Therefore, it is imperative to identify novel compounds with new mechanisms of action active against NR bacilli. Dormant Mtb habitat is mostly thought to be the pH-neutral and hypoxic caseous granuloma. We have used the Wayne culture model to reproduce this environment and tested the activities of two DNA-targeted agents, C8-linked-pyrrolobenzodiazepine(PBD)-polyamide conjugates 1 and 2, against Mtb grown in aerobic and hypoxic conditions in both acidic and pH-neutral media. PBD 2 showed growth inhibitory activity at 5.1 µg/ml against 19-day-old hypoxic NR Mtb cultures with 1.8 log10 CFU reduction on day 21 at pH 7.3. PBD 2 was particularly effective against 5-day-old aerobic cells at pH 7.3, with CFU reduction (>6.8 log10) on day 21 at 5.1 µg/ml being identical to that of rifampin at 8 µg/ml. PBD 2 qualifies as a promising lead against aerobic and NR Mtb.

C8-Linked Pyrrolobenzodiazepine Monomers with Inverted Building Blocks Show Selective Activity against Multidrug Resistant Gram-Positive Bacteria.

Antimicrobial resistance has become a major global concern. Development of novel antimicrobial agents for the treatment of infections caused by multidrug resistant (MDR) pathogens is an urgent priority. Pyrrolobenzodiazepines (PBDs) are a promising class of antibacterial agents initially discovered and isolated from natural sources. Recently, C8-linked PBD biaryl conjugates have been shown to be active against some MDR Gram-positive strains. To explore the role of building block orientations on antibacterial activity and obtain structure activity relationship (SAR) information, four novel structures were synthesized in which the building blocks of previously reported compounds were inverted, and their antibacterial activity was studied. The compounds showed minimum inhibitory concentrations (MICs) in the range of 0.125-32 μg/mL against MDR Gram-positive strains with a bactericidal mode of action. The results showed that a single inversion of amide bonds reduces the activity while the double inversion restores the activity against MDR pathogens. All inverted compounds did not stabilize DNA and lacked eukaryotic toxicity. The compounds inhibit DNA gyrase in vitro, and the most potent compound was equally active against both wild-type and mutant DNA gyrase in a biochemical assay. The observed activity of the compounds against methicillin resistant S.aureus (MRSA) strains with equivalent gyrase mutations is consistent with gyrase inhibition being the mechanism of action in vivo, although this has not been definitively confirmed in whole cells. This conclusion is supported by a molecular modeling study showing interaction of the compounds with wild-type and mutant gyrases. This study provides important SAR information about this new class of antibacterial agents.

DNA sequence-selective C8-linked pyrrolobenzodiazepine-heterocyclic polyamide conjugates show anti-tubercular-specific activities.

New chemotherapeutic agents with novel mechanisms of action are in urgent need to combat the tuberculosis pandemic. A library of 12 C8-linked pyrrolo[2,1-c][1,4]benzodiazepine (PBD)-heterocyclic polyamide conjugates (1-12) was evaluated for anti-tubercular activity and DNA sequence selectivity. The PBD conjugates were screened against slow-growing Mycobacterium bovis Bacillus Calmette-Guérin and M. tuberculosis H37Rv, and fast-growing Escherichia coli, Pseudomonas putida and Rhodococcus sp. RHA1 bacteria. DNase I footprinting and DNA thermal denaturation experiments were used to determine the molecules' DNA recognition properties. The PBD conjugates were highly selective for the mycobacterial strains and exhibited significant growth inhibitory activity against the pathogenic M. tuberculosis H37Rv, with compound 4 showing MIC values (MIC=0.08 mg l-1) similar to those of rifampin and isoniazid. DNase I footprinting results showed that the PBD conjugates with three heterocyclic moieties had enhanced sequence selectivity and produced larger footprints, with distinct cleavage patterns compared with the two-heterocyclic chain PBD conjugates. DNA melting experiments indicated a covalent binding of the PBD conjugates to two AT-rich DNA-duplexes containing either a central GGATCC or GTATAC sequence, and showed that the polyamide chains affect the interactions of the molecules with DNA. The PBD-C8 conjugates tested in this study have a remarkable anti-mycobacterial activity and can be further developed as DNA-targeted anti-tubercular drugs.

Abstract 4555: C8-linked pyrrolobenzodiazepine (PBD)-benzofused hybrids as transcription factor inhibitors

Abstract The pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) have been extensively investigated as DNA-interactive agents since their discovery in 1963. The significant cytotoxicity of the naturally derived PBDs has led to the search for synthetic analogues that retain the cytotoxicity of the parent structure while providing an enhanced therapeutic index. There is growing evidence that PBD monomers inhibit the binding of transcription factors to gene promoter regions, blocking their effects on gene expression. For example, GWL-78, a C8-linked PBD-Py-Py (Py = pyrrole) conjugate, has been shown to block interaction of the transcription factor NF-Y, and KMR-28-39, a C8-linked PBD-MPB conjugate inhibits the activity of the transcription factor NF-κB. As up-regulation of NF-κB has long been implicated in the development and progression of several cancer types, this may account for the femtomolar potency observed for PBD monomers such as KMR-28-39. The initial synthesis of a small library of C8-PBD conjugates with benzofused C8-substituents provided notable activity against a panel of leukaemia cell lines. In addition to exhibiting low picomolar IC50 values in a number of cell lines, these compounds also showed selectivity for binding to NF-κB consensus sequences. This small library was used as the starting point for the synthesis of three analogous libraries of PBD C8-conjugates designed to explore the SARs of such structures, with a view to selecting a candidate for pre-clinical development. The design rationale for the novel conjugates involved modification of the shape of the C8-linked substituent with the introduction of specific functional groups in an effort to induce sequence-selective DNA interactivity. The sequence selectivity of the novel benzofused PBD conjugates was investigated using a FRET-based assay in which ligand binding to several different transcription factor binding sequences implicated in cancer was explored (i.e., AP-1, NF-κB1, NF-κB2, STAT3 and EGR). Differential DNA stabilisation compared to control was observed for all analogues, with ΔTm values between 23.4 and 3.4oC at 1 μM ligand concentration (Ligand:DNA = 1:5) for these sequences, indicating the molecules had preferential binding to certain transcription factor binding sequences. Stabilisation was especially pronounced for the AP-1 consensus sequence with two lead compounds (DC-3-28-33 and DC-1-113) providing a high degree of stabilisation (23.9oC and 26oC, respectively). Preliminary cytotoxicity screening studies were carried out in Primary CLL and a Myeloma cell line (JJN3), and low nanomolar IC50 values were observed in the CLL cells for several of the synthesised compounds. These included DC-1-92 (9.7nM), DC-1-115 (14nM), DC-1-36 (16nM), DC-1-170 (17nM), and DC-1-113 (18nM). Control screening in normal lymphocytes is currently underway in order to establish the selective toxicity profiles of these compounds. Citation Format: David B. Corcoran, Paul J. M. Jackson, Ambereen Ajaz, Thomas Lewis, Chris Pepper, David E. Thurston, Khondaker M. Rahman. C8-linked pyrrolobenzodiazepine (PBD)-benzofused hybrids as transcription factor inhibitors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4555. doi:10.1158/1538-7445.AM2015-4555

Abstract 3696: Molecular dynamics simulations of C7/C7′-linked pyrrolobenzodiazepine (PBD) dimers

Abstract The pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) are a group of naturally occurring and synthetic DNA minor-groove binding agents (MGBAs) characterised by their ability to covalently bind to the C2-amino groups of guanine bases. Although natural PBD monomers have low micromolar cytotoxicity, synthetic PBD dimers have garnered significant attention due to their potent cytotoxicity and use in antibody-drug conjugates (ADCs). The cytotoxic nature of PBD dimers is thought to relate to their ability to form inter- and intrastrand cross-links in DNA. The first known PBD dimer had its two PBD units linked through their C7/C7′-positions, and was reported by Suggs and co-workers. This molecule was shown to reversibly cross-link DNA, a feature that may account for its poor cytotoxicity relative to C8-linked dimers which are used as ADC payloads. To investigate this, we have undertaken Molecular Dynamics (MD) studies on the interaction of C7-dimers with DNA and compared results to those for C8-linked PBD dimers. This has led us to propose an alternative mechanism to explain the discrepancy in potencies as described below. MD simulations were conducted over 10ns in explicit solvent. In this study, Suggs’ published C7/C7′-linked dimer and an equivalent C8/C8′ linked structure were first compared, and this was followed by comparison of the known C8-linked PBD dimer SG-2202 and an equivalent C7/C7′-linked version. SG-2202 is known to preferentially bind to the sequence 5′-pu-G-A-A/T-A/T-C-py-3′. As each of the PBD dimers analysed have an equivalent central linker (i.e., five methylenes), each of the structures were simulated in the sequence 5′-GCGATTCTCGC-3′. The results suggested that C8-linked dimers interact with DNA to a far greater extent than C7-linked dimers, a feature attributable to the level of curvature (“isohelicity”) of C8-linked dimers, and this was supported by free energy of binding calculations. Crucially, the simulations showed that the C7-linked dimer family does not form stabilising non-covalent interactions with the DNA minor-groove as previously thought. Furthermore, due to its poor isohelicity, simulations predict that only a single alkylation event takes place initially, and this is followed by the second alkylation which is dependent on DNA breathing to align the second PBD unit with an appropriate guanine. Simulations suggested that the C7-position of any PBD has two potential orientations: (1) bulky groups such as a sugar (i.e., as in sibiromycin) point out of the minor groove, and (2) smaller substituents (e.g., a methylene linker) point along the minor groove floor. The latter explains why it is possible for C7-methylene-linked PBD dimers to cross-link DNA albeit with less efficiency compared to C8-linked dimers. Studies are now underway to analyse the curvature of PBD-type molecules to develop tools (e.g., QSAR) for the correlation of shape and cytotoxicity to facilitate the discovery of MGBAs with more potent cytotoxicity. Citation Format: Paul J. M. Jackson, Khondaker Miraz Rahman, David E. Thurston. Molecular dynamics simulations of C7/C7′-linked pyrrolobenzodiazepine (PBD) dimers. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3696. doi:10.1158/1538-7445.AM2015-3696

Abstract 1630: C8-linked pyrrolobenzodiazepine (pbd)-benzofused conjugates with low-picomolar in vitro cytotoxicity

Abstract There is significant interest in developing small-molecule ligands that can locate and bind to specific sequences of DNA as potential antitumor agents. Such molecules could be used, for example, to inhibit transcription factors and other control proteins from interacting with DNA at their consensus recognition sites. A set of DNA-interactive benzofused building blocks based on phenyl-substituted heterocycles have been developed with sufficient length to span two DNA base pairs. These building blocks have been conjugated to DNA minor-groove covalent-binding pyrrolobenzodiazepine (PBD) molecules via a four-carbon linker to produce C8-linked PBD-Benzofused hybrid molecules. These focused libraries contained structural variations where the benzofused building blocks were either directly linked to the PBD core or separated by pyrrole/imidazole heterocycles or GC-recognizing methylpyrrolebenzenamine (MPB) biaryl building blocks. The design rationale was based on the unique longitudinal curvature of these PBD conjugates which produces greater isohelicity with the DNA minor groove, and also on predicted hydrogen-bonding interactions with individual DNA bases. These molecules showed significant enhancement of melting temperatures of designed oligonucleotides in a FRET-based assay compared to the PBD core alone, highlighting the synergistic effect of joining non-covalent benzofused rings to the covalent-binding PBD unit. The cytotoxicity of these compounds was assessed in primary CLL cells and a panel of other tumor cell lines (A431, A2780, A549, MIA PaCa2, HeLa and MDA-MB-231) after 48h and 96h exposure using the Annexin V apoptosis assay and the MTT colorimetric assay, respectively. Normal lymphocytes and the non-tumor fibroblast cell line WI38 were used as controls. Some of these compounds were highly potent (i.e., low picomolar) in the primary CLL and other cell lines with at least 3-orders of magnitude selectivity compared to the normal lymphocytes or WI38 cells. Importantly, molecules with different distances of separation between the benzofused ring moieties had significant differences in cytotoxicity, thus not only providing useful SAR information but also suggesting that there was scope for optimizing activity through a medicinal chemistry approach. The effect of these conjugates on regulation of the NF-κB-associated regulatory proteins IKKα and IKKβ was studied in the CLL and breast cancer cell lines in which NF-κB is known to be active and closely correlated with the initiation and progression of malignancy. The lead PBD-Benzofused conjugate KMR-28-32 exhibited marked inhibition of both IKKα and IKKβ proteins at 4h and 24h time points at concentrations of 0.1nM and 1nM, respectively, with negligible effect on tubulin expression which was used as a housekeeping control. Citation Format: Julia Mantaj, Paul JM Jackson, Chris Pepper, David E. Thurston, Khondaker M. Rahman. C8-linked pyrrolobenzodiazepine (pbd)-benzofused conjugates with low-picomolar in vitro cytotoxicity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1630. doi:10.1158/1538-7445.AM2014-1630

GC-Targeted C8-Linked Pyrrolobenzodiazepine–Biaryl Conjugates with Femtomolar in Vitro Cytotoxicity and in Vivo Antitumor Activity in Mouse Models

DNA binding 4-(1-methyl-1H-pyrrol-3-yl)benzenamine (MPB) building blocks have been developed that span two DNA base pairs with a strong preference for GC-rich DNA. They have been conjugated to a pyrrolo[2,1-c][1,4]benzodiazepine (PBD) molecule to produce C8-linked PBD-MPB hybrids that can stabilize GC-rich DNA by up to 13-fold compared to AT-rich DNA. Some have subpicomolar IC50 values in human tumor cell lines and in primary chronic lymphocytic leukemia cells, while being up to 6 orders less cytotoxic in the non-tumor cell line WI38, suggesting that key DNA sequences may be relevant targets in these ultrasensitive cancer cell lines. One conjugate, 7h (KMR-28-39), which has femtomolar activity in the breast cancer cell line MDA-MB-231, has significant dose-dependent antitumor activity in MDA-MB-231 (breast) and MIA PaCa-2 (pancreatic) human tumor xenograft mouse models with insignificant toxicity at therapeutic doses. Preliminary studies suggest that 7h may sterically inhibit interaction of the transcription factor NF-κB with its cognate DNA binding sequence.

Novel C8-linked pyrrolobenzodiazepine (PBD)–heterocycle conjugates that recognize DNA sequences containing an inverted CCAAT box

A series of novel DNA-interactive C8-linked pyrrolobenzodiazepine (PBD)-heterocycle polyamide conjugates has been synthesised to explore structure/sequence-selectivity relationships. One conjugate (2d) has a greater selectivity and DNA binding affinity for inverted CCAAT sequences within the Topoisomerase IIα promoter than the known C8-bis-pyrrole PBD conjugate GWL-78 (1b).

Abstract 2321: Pyrrolobenzodiazepine (PBD) antitumour agents as chemical probes to determine the number of base pairs required for DNA minor groove structure

Abstract The pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) are sequence-selective covalent-binding DNA-interactive agents that fit perfectly in the minor groove of double-stranded DNA due to their isohelical 3-dimensional shape. They possess an electrophilic N10-C11 imine (or the equivalent) that can form a covalent aminal bond between their C11-position and the C2-NH2 group of a guanine. One synthetic PBD dimer, SJG-136, has just entered Phase II clinical trials. Previous studies using HPLC/MS, CD spectroscopy, high-field NMR, X-ray crystallography and gel-based assays such as DNA footprinting and in vitro transcription stop are all consistent with the requirement for duplex DNA for covalent binding. Conversely, it is well-documented that PBDs will not interact with single stranded DNA which lacks minor groove structure. In this study, these observations served as a basis to use PBDs as chemical probes to determine the minimum number of bases required to create a minor groove environment within a DNA duplex. Thus we studied reaction of the naturally-occurring anthramycin (3 base pair span) and the synthetic C8-linked PBD bis-pyrrole conjugate GWL-78 (6 base pair span) with a range of designed duplex oligonucleotides of increasing length (3 to 11 base pairs), each containing an AGA binding triplet. Reaction was monitored by HPLC/MS which provided data on both the rate and extent of reaction, and the structure of DNA adducts formed. For both anthramycin and GWL-78, we found that a minimum of 7 DNA base pairs were required for reaction to occur, suggesting that this may represent the minimum necessary to provide a suitable minor groove environment. Furthermore, we found that the rate of reaction for both PBDs increased with the length of the oligonucleotide between 7 to 10 base pairs. It is noteworthy that GWL-78 is approximately twice the length of anthramycin but both require at least 7 base pairs in order to covalently bind. This suggests that both may be responding to the formation of a minor groove environment in an oligonucleotide that reaches 7 base pairs in length, rather than a direct relationship between the length of the PBD and the number of base pairs in the oligonucleotide. These observations were supported by molecular modeling which confirmed that 7 base pairs are required to provide minor groove structure. This is the first study to use a chemical probe of this type to monitor formation of a minor groove. This information should be useful in the design of future generations of PBDs and similar DNA-interacting anticancer agents. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2321. doi:10.1158/1538-7445.AM2011-2321

Design, Synthesis, and Biophysical and Biological Evaluation of a Series of Pyrrolobenzodiazepine−Poly(N-methylpyrrole) Conjugates

A novel series of methyl ester-terminated C8-linked pyrrolobenzodiazepine (PBD)-poly(N-methylpyrrole) conjugates (50a-f) has been synthesized and their DNA interaction evaluated by thermal denaturation, DNA footprinting, and in vitro transcription stop assays. The synergistic effect of attaching a PBD unit to a polypyrrole fragment is illustrated by the large increase in DNA binding affinity (up to 50-fold) compared to the individual PBD and pyrrole components. 50a-f were found to bind mainly to identical DNA sequences but with apparent binding site widths increasing with molecular length and the majority of sites conforming to the consensus motif 5'-XGXWz (z = 3 +/- 1; W = A or T; X = any base but preferably a purine). They also provided robust sequence-selective blockade of transcription at sites corresponding approximately to their DNA footprints. 50a-f were shown to have good cellular/nuclear penetration properties, and a degree of correlation between cytotoxicity and DNA-binding affinity was observed.

Synthesis of Sequence-Selective C8-Linked Pyrrolo[2,1-c][1,4]benzodiazepine DNA Interstrand Cross-Linking Agents.

An efficient convergent synthesis of a homologous series of C8-linked pyrrolobenzodiazepine dimers with remarkable DNA interstrand cross-linking activity and potent in vitro cytotoxicity is reported. The "amino thioacetal" cyclization procedure was used to produce the electrophilic DNA-interactive N10-C11 imine moiety during the final synthetic step. In order to construct the key A-ring fragments (9a-d), a versatile convergent approach has been developed to join two units of vanillic acid with alpha,omega-dihaloalkanes of varying length to provide the required bis(4-carboxy-2-methoxyphenoxy)alkanes while avoiding the formation of mixtures of monoalkylated and bisalkylated products.

Cellular pharmacology of novel C8-linked anthramycin-based sequence-selective DNA minor groove cross-linking agents

The cellular pharmacology of a series of C8-linked pyrrolobenzodiazepine dimers with polymethylene linkers of n = 3-6 (compounds 1-4) has been studied in a range of human tumour cell lines. The four compounds showed the same pattern of relative activity in five ovarian carcinoma cell lines and one cervical carcinoma cell line with the order of IC50 values of 1 < or = 3 < 4 < 2, which correlated with the previously demonstrated DNA interstrand cross-linking ability of the compounds in plasmid DNA. In human leukaemic K562 cells the agents produced a block in the G2/M phase of the cell cycle characteristic of cross-linking drugs, and extensive interstrand cross-linking was observed in cells by alkaline elution with no evidence of single-strand breaks. Cross-links continued to increase up to 24 h following a 1 h exposure to drug, and no repair was evident by 48 h. In a series of ovarian and cervical carcinoma cell lines with acquired resistance to cisplatin no cross-resistance to the most potent compound 1 was observed in two lines whose major mechanism of resistance to cisplatin was reduced platinum transport. Cross-resistance to 1 was observed in a cell line (A2780cisR) possessing elevated glutathione, and depletion of intracellular glutathione using D,L-buthionine-S,R-sulphoximine (BSO) from 10.25 nmol to 2.8 nmol 10(-6) cells reduced the level of resistance from 11-fold to 2-fold compared with sensitive cells. Cross-linking in the resistant cells was restored to 80% of the level in the parent line by BSO pretreatment. There was also a correlation between glutathione levels and sensitivity to 1 measured in several other ovarian cell lines. Compound 1 also showed cross-resistance in the doxorubicin-resistant cell line 41MdoxR and partial cross-resistance in CH1doxR cells. Both these lines possess elevated levels of p170 glycoprotein. Following treatment with 6 microM verapamil, the resistance in these lines decreased almost 2-fold and 8-fold respectively.

Effect of linker length on DNA-binding affinity, cross-linking efficiency and cytotoxicity of C8-linked pyrrolobenzodiazepine dimers

An efficient synthesis of a homologous series of C8-linked pyrrolobenzodiazepine dimers is reported; compounds with an odd number of methylenes (n= 3 or 5) in the linker show a higher affinity for DNA, enhanced cross-linking efficiency, and are more cytotoxic compared with compounds with either n= 4 or 6.