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Annals of Oncology : Official Journal... May 2003The aim of this study was to assess the feasibility of administering bizelesin, a cyclopropylpyrroloindole with extraordinarily high potency as a bifunctional... (Clinical Trial)
Clinical Trial
BACKGROUND
The aim of this study was to assess the feasibility of administering bizelesin, a cyclopropylpyrroloindole with extraordinarily high potency as a bifunctional DNA-damaging agent and selectivity for specific AT-rich DNA sequences, as a single i.v. bolus injection every 4 weeks in patients with advanced solid malignancies. The study also sought to determine the maximum tolerated dose (MTD) of bizelesin, characterize its pharmacokinetic behavior, and seek preliminary evidence of anticancer activity.
PATIENTS AND METHODS
Patients with advanced solid malignancies were treated with escalating doses of bizelesin as an i.v. bolus injection every 4 weeks. The selection of the specific starting dose, 0.1 micro g/m(2), which was equivalent to one-tenth the toxic dose low in dogs, factored in large interspecies differences in myelotoxicity as gauged using an ex vivo hematopoietic colony-forming assay. Due to concerns about the high potency of bizelesin and the large interspecies differences in toxicity, a conservative dose-escalation scheme was used for dose-level assignment to determine the MTD levels for both minimally pretreated (MP) and heavily pretreated (HP) patients. A variety of analytical assays were assessed to reliably measure bizelesin concentrations in plasma.
RESULTS
Sixty-two patients were treated with 185 courses of bizelesin at eight dose levels ranging from 0.1 to 1.5 micro g/m(2). Myelosuppression, principally neutropenia that was always brief, was the most common toxicity observed. Thrombocytopenia and anemia were uncommon and severe non-hematological effects were not observed. Severe neutropenia alone and/or associated with fever was consistently experienced by HP and MP patients at doses exceeding 0.71 and 1.26 micro g/m(2), respectively. These doses also resulted in functionally non-cumulative myelosuppression as repetitive treatment was well-tolerated. A 40% reduction in measurable disease lasting 24 months was noted in a patient with advanced ovarian carcinoma. Various analytical methods were evaluated but none demonstrated the requisite sensitivity to reliably quantify the minute plasma concentrations of bizelesin and metabolites resulting from administering microgram quantities of drug.
CONCLUSIONS
The highly potent and unique cytotoxic agent, bizelesin can be feasibly administered to patients with advanced solid malignancies. The recommended doses for phase II studies of bizelesin as a bolus i.v. injection every 4 weeks are 0.71 and 1.26 micro g/m(2) in HP and MP patients, respectively. The characteristics of the myelosuppression, the paucity of severe toxicities with repetitive treatment, the preliminary antitumor activity noted, and, above all, its unique mechanism of action as a selective DNA-damaging agent and high potency, warrant disease-directed evaluations of bizelesin in solid and hematopoietic malignancies and consideration of its use as a cytotoxic in targeted conjugated therapeutics.
Topics: Adult; Aged; Aged, 80 and over; Antineoplastic Agents, Alkylating; DNA; Dose-Response Relationship, Drug; Drug Administration Schedule; Duocarmycins; Female; Humans; Indoles; Intercalating Agents; Male; Middle Aged; Neoplasms; Urea
PubMed: 12702533
DOI: 10.1093/annonc/mdg215 -
Biochimica Et Biophysica Acta Jul 2002Cellular DNA is not a uniform target for DNA-reactive drugs. At the nucleotide level, drugs recognize and bind short motifs of a few base pairs. The location of drug... (Review)
Review
Cellular DNA is not a uniform target for DNA-reactive drugs. At the nucleotide level, drugs recognize and bind short motifs of a few base pairs. The location of drug adducts at the genomic level depends on how these short motifs are distributed in larger domains. This aspect, referred to as region specificity, may be critical for the biological outcome of drug action. Recent studies demonstrated that certain minor groove binding (MGB) drugs, such as bizelesin, produce region-specific lesions in cellular DNA. Bizelesin binds mainly T(A/T)(4)A sites, which are on average scarce, but occasionally cluster in distinct minisatellite regions (200-1000 bp of approximately 85-100% AT), herein referred to as AT islands. Bizelesin-targeted AT islands are likely to function as strong matrix attachment regions (MARs), domains that organize DNA loops on the nuclear matrix. Distortion of MAR-like AT islands may be a basis for the observed inhibition of new replicon initiation and the extreme lethality of bizelesin adducts (<10 adducts/cell for cell growth inhibition). Hence, long AT-islands represent a novel class of critical targets for anticancer drugs. The AT island paradigm illustrates the potential of the concept of regional targeting as an essential component of the rational design of new sequence-specific DNA-reactive drugs.
Topics: Antineoplastic Agents; Base Sequence; Benzofurans; Binding Sites; Cyclohexanecarboxylic Acids; Cyclohexenes; DNA Damage; DNA, Neoplasm; Distamycins; Drug Design; Duocarmycins; Genome, Human; Humans; Indoles; Nitrogen Mustard Compounds; Tandem Repeat Sequences; Tumor Cells, Cultured; Urea
PubMed: 12084472
DOI: 10.1016/s0925-4439(02)00093-5 -
The Journal of Biological Chemistry Nov 2001Interstrand cross-links at T(A/T)4A sites in cellular DNA are associated with hypercytotoxicity of an anticancer drug, bizelesin. Here we evaluated whether these lethal...
Interstrand cross-links at T(A/T)4A sites in cellular DNA are associated with hypercytotoxicity of an anticancer drug, bizelesin. Here we evaluated whether these lethal effects reflect targeting critical genomic regions. An in silico analysis of human sequences showed that T(A/T)4A motifs are on average scarce and scattered. However, significantly higher local motif densities were identified in distinct minisatellite regions (200-1000 base pairs of approximately 85-100% AT), herein referred to as "AT islands." Experimentally detected bizelesin lesions agree with these in silico predictions. Actual bizelesin adducts clustered within the model AT island naked DNA, whereas motif-poor sequences were only sparsely adducted. In cancer cells, bizelesin produced high levels of lesions (approximately 4.7-7.1 lesions/kilobase pair/microM drug) in several prominent AT islands, compared with markedly lower lesion levels in several motif-poor loci and in bulk cellular DNA (approximately 0.8-1.3 and approximately 0.9 lesions/kilobase pair/microM drug, respectively). The identified AT islands exhibit sequence attributes of matrix attachment regions (MARs), domains that organize DNA loops on the nuclear matrix. The computed "MAR potential" and propensity for supercoiling-induced duplex destabilization (both predictive of strong MARs) correlate with the total number of bizelesin binding sites. Hence, MAR-like AT-rich non-coding domains can be regarded as a novel class of critical targets for anticancer drugs.
Topics: Adenine; Antineoplastic Agents; Base Sequence; DNA; Duocarmycins; Indoles; Thymine; Urea
PubMed: 11487576
DOI: 10.1074/jbc.M103390200 -
Nucleic Acids Research Nov 2003AT-rich minisatellites (AT islands) are sites of genomic instability in cancer cells and targets for extremely lethal AT-specific drugs, such as bizelesin. Here we...
AT-rich minisatellites (AT islands) are sites of genomic instability in cancer cells and targets for extremely lethal AT-specific drugs, such as bizelesin. Here we investigated the AT islands in the FRA16B fragile site region for their possible roles in the organization of DNA on the nuclear matrix. The FRA16B AT island nominally spans approximately 3 kb of mostly >90% A/T DNA. In silico analysis indicates that this domain exhibits characteristics of nuclear matrix attachment regions (MARs): an exceptionally intense computed 'MAR potential' and profound duplex destabilization and flexibility. FRA16B repeats specifically bind to isolated nuclear matrices, which indicates their in vitro MAR function. This binding is several-fold greater than that of a known MAR in the c-myc gene. AT islands in fragile sites FRA16B and FRA16D are significantly more abundant in CEM cells that are hypersensitive to bizelesin compared to normal WI-38 cells. FRA16B overabundance in CEM is due to an approximately 10-fold expansion of FRA16B repeats. The expanded FRA16B minisatellites in CEM cells preferentially localize to the nuclear matrix-associated DNA indicating their in vivo MAR function. The unexpanded repeats in WI-38 cells localize to the loop DNA. The c-myc MAR is also matrix-associated in CEM cells while localizing to loop DNA in WI-38 cells. These results are the first to demonstrate that AT islands in fragile sites can function as MARs both in vitro and in vivo. The ability of FRA16B-mediated MAR sites to rearrange depending on the repeat expansion status could be relevant to both genomic instability of cancer cells and their sensitivity to AT-island targeting drugs.
Topics: AT Rich Sequence; Binding Sites; Cell Line, Tumor; Chromosome Fragility; Chromosomes, Human, Pair 16; DNA; DNA Sequence, Unstable; Duocarmycins; Genes, myc; Globins; Humans; Indoles; Leukemia; Minisatellite Repeats; Molecular Sequence Data; Nuclear Matrix; Nucleic Acid Conformation; Polymerase Chain Reaction; Urea
PubMed: 14576323
DOI: 10.1093/nar/gkg832 -
PloS One 2012In Plasmodium, meiosis occurs in diploid zygotes as they develop into haploid motile ookinetes inside the mosquito. Further sporogonic development involves...
BACKGROUND
In Plasmodium, meiosis occurs in diploid zygotes as they develop into haploid motile ookinetes inside the mosquito. Further sporogonic development involves transformation of ookinetes into oocysts and formation of infective sporozoites.
METHODOLOGY/PRINCIPAL FINDINGS
Reverse genetics was employed to examine the role of the meiotic specific recombinase Dmc1, a bacterial RecA homolog during sporogony in Plasmodium berghei. PbDmc1 knockout (KO) parasites showed normal asexual growth kinetics compared to WT parasites; however oocyst formation in mosquitoes was reduced by 50 to 80%. Moreover, the majority of oocysts were retarded in their growth and were smaller in size compared to WT parasites. Only a few Dmc1 KO parasites completed maturation resulting in formation of fewer sporozoites which were incapable of infecting naive mice or hepatocytes in vitro. PbDmc1 KO parasites were shown to be approximately 18 times more sensitive to Bizelesin, a DNA alkylating drug compared to WT parasites as reflected by impairment of oocyst formation and sporogonic development in the mosquito vector.
CONCLUSIONS/SIGNIFICANCE
Our findings suggest that PbDmc1 plays a critical role in malaria transmission biology.
Topics: Alkylating Agents; Animals; Culicidae; DNA Damage; Duocarmycins; Female; Gene Knockout Techniques; Gene Targeting; Genes, Protozoan; Indoles; Kinetics; Life Cycle Stages; Male; Mice; Oocysts; Parasites; Plasmodium berghei; Protozoan Proteins; Recombinases; Reproduction, Asexual; Sporozoites; Urea
PubMed: 23285059
DOI: 10.1371/journal.pone.0052480