Lloyd R. Kelland

Mechanisms of tumor vascular shutdown induced by 5,6-dimethylxanthenone-4-acetic acid (DMXAA): Increased tumor vascular permeability

The novel vascular targeting agent 5,6‐dimethylxanthenone‐4‐acetic acid (DMXAA) has completed phase 1 clinical trial and has shown tumor antivascular activity in both mice and humans. We have investigated its ability to change tumor vascular permeability, relating it to tumor vascular perfusion and other responses. The murine colon 38 adenocarcinoma was grown in C57Bl wild‐type mice and mice lacking expression of either tumor necrosis factor receptor‐1 (TNFR1−/−) or TNF (TNF−/−). Tumor vascular permeability, as measured by extravasation of albumin‐Evans Blue complexes 4 hr after DMXAA treatment, was significantly increased in tumor tissue in C57Bl, TNFR1−/− and TNF−/− mice but not in normal (skin) tissue. Significant linear relationships were found between increased tumor vascular permeability, decreased functioning tumor blood vessels (measured by Hoechst 33342 staining at 4 hr), increased plasma 5‐hydroxyindole‐3‐acetic acid concentrations (as a measure of serotonin release by platelets) and the degree of induced tumor hemorrhagic necrosis. The results support the hypothesis that DMXAA increases tumor vascular permeability both directly and through the induction of other vasoactive mediators, including TNF. DMXAA might be useful clinically to potentiate the vascular permeability of other anticancer modalities such as cytotoxic drugs, antibodies, drug conjugates and gene therapy.

Farnesyl transferase inhibitors in the treatment of breast cancer

Until recently, the therapeutic treatment of breast cancer has been dominated by endocrine-based drugs (oestrogen receptor antagonists, aromatase inhibitors etc.) and conventional cytotoxics (doxorubicin, cyclophosphamide, 5fluorouracil etc.). However, the advent of new generation signal transduction inhibitor drugs targeted against the molecular abnormalities of breast cancer (e.g., the antibody trastuzumab, directed against the cERBB2 receptor) has the promise of providing a new era of more tumour selective therapy. Inhibitors of the enzyme farnesyl transferase (FTIs) are now undergoing early-stage clinical trials, including in patients with advanced breast cancer. Although originally developed as inhibitors of RAS signal transduction pathways, it is now apparent that these drugs are better described as prenylation inhibitors; the addition of a 15-carbon prenyl or farnesyl moiety by farnesyl transferase being critical to the function of a number of proteins, including RAS. At least three FTIs are currently undergoing clinical evaluation; R115777 (tipifarnib, Zarnestra®), SCH66336 (lonafarnib, Sarasar®) and BMS-214662. In terms of their potential use in the chemotherapeutic treatment of advanced breast cancer, a Phase II trial of R115777 (using either continuous or intermittent twice-daily oral dosing) has demonstrated promising activity (~ 10% partial response rate). Overall, however, the single agent activity of FTIs in various Phase II trials has been rather modest (as well as the above mentioned breast cancer trial, some responses have been seen in patients with acute and chronic myeloid leukaemias). The main dose-limiting toxicities that have been reported are myelosuppression and fatigue and neurotoxicity (with R115777). Two Phase III trials of R115777 in colorectal (versus placebo) and pancreatic (with gemcitabine versus placebo) cancer have failed to show a survival benefit. It is likely that the future clinical direction of FTIs will be as combination therapy, especially with the taxanes, where synergy has been seen in a variety of preclinical studies.

5,6-Dimethylxanthenone-4-Acetic Acid (DMXAA)

Currently, there is a great deal of interest in drugs that target tumor vasculature and their therapeutic potential in combination regimens for the treatment of cancer. This review focuses on one of the vascular disrupting agents, 5-6-dimethylxanthenone 4-acetic acid (DMXAA), and the rationale for its combination with standard taxane-based chemotherapy. DMXAA and taxanes have different mechanisms of action and, in combination, demonstrate at least additive activity against preclinical solid tumors. Their clinical adverse-effect and pharmacologic profiles as single agents appear to render these agents suitable for use in combination. Phase I studies of DMXAA have identified a range of doses for combination clinical trials. In addition, the clinical indications and chemotherapy doses for combination clinical studies have been selected from positive randomized controlled trials of taxanes in advanced cancers. A phase II clinical trials program of combination studies with DMXAA is now underway; paclitaxel and carboplatin in patients with NSCLC and ovarian cancer and docetaxel in patients with hormone-refractory prostate cancer are being evaluated.

Evidence for the involvement of p38 MAP kinase in the action of the vascular disrupting agent 5,6-dimethylxanthenone-4-acetic acid (DMXAA)

SummaryAims: DMXAA (AS1404), a small-molecule vascular disrupting agent that has now completed Phase II clinical trial, induces endothelial cell apoptosis, increased vascular permeability and decreased tumour blood flow in vivo. Its action is incompletely understood and we wished to develop an in vitro system to study its effects.n Methods: Human tumour cell lines developed from aggressive tumours were grown on Matrigel to simulate a tumour microenvironment. Cells were analysed by light microscopy and by gene expression profiling.n Results: Several cell lines formed networks when grown on Matrigel and the NZM7 melanoma cell line was chosen for further study. Addition of DMXAA at a clinically achievable concentration (30xa0μg/mL) prevented network formation, but co-addition of SB203580 (10xa0μM), a selective inhibitor of p38 MAP kinase, reversed the effect of DMXAA and restored network formation. Analysis of expression genes for endothelial and related functions showed that cells growing on Matrigel expressed a pattern similar to that of NZM7 cells growing as xenografts in vivo but different from that of cells grown on standard tissue culture plates. Addition of DMXAA resulted in the inhibition of expression of several genes including the transcriptional activator Ets1 and matrix metalloproteinase-2 (MMP2), but co-addition of SB203580 did not reverse these effects of DMXAA on gene expression.n Conclusion: The results suggest that p38 MAP kinase plays an important role in the action of DMXAA and that growth of tumour cells on Matrigel provides a promising model for further studies on the action of this drug.

Overcoming Resistance to Platinum Therapy in Patients with Advanced Cancer

The platinum-containing compounds cisplatin and carboplatin represent the mainstay of chemotherapeutic treatment for a variety of solid tumors occurring in adults, especially testicular and ovarian cancers. These drugs confer their antitumor efficacy by binding to DNA, especially to guanine bases, either adjacent on the same DNA strand or across strands. Tumour resistance, either present at the onset of therapy or acquired during successive courses of therapy, represents the significant limiting factor to long-term patient survival. From numerous studies of cancer cell lines in vitro, it is apparent that tumor resistance to cisplatin may arise through two general mechanisms: one preventing sufficient platinum from binding to DNA (reduced membrane transport and cytoplasmic inactivation by thiol-containing species such as glutathione or metallothioneins); and a second whereby platinum-DNA damage does not result in cell death (enhanced DNA nucleotide excision repair, increased tolerance, loss of DNA mismatch repair, failure to undergo apoptotic programmed cell death). It remains largely unclear as to the relative importance of these mechanisms in the clinic or whether additional mechanisms also exist.Overcoming clinical resistance has proven particularly difficult. Two broad approaches have been investigated: (i) using cisplatin/carboplatin with new drugs in combination or second-line with agents also possessing single agent antitumor activity (e.g. taxanes, topotecan); and (ii) the synthesis of platinum-containing analogues capable of circumventing resistance (e.g. oxaliplatin, AMD473, BBR3464). To date, these new platinum compounds (e.g. oxaliplatin in combination with fluorouracil to treat patients with advanced colorectal cancer) have made some relatively minor inroads into broadening the clinical utility of cisplatin/carboplatin. Much remains to be achieved.

New Platinum Drugs

Following the well-documented serendipitous discovery of the antitumour properties of cis-diamminedichloro platinum(II) (cisplatin) in the mid 1960s (see Rosenberg, 1985, for a review), the drug was introduced into clinical practice in 1971. While the introduction of cisplatin has undoubtedly made a dramatic impact on the response rates (and long-term survival) obtained for patients presenting with some tumour types (notably testicular teratoma and ovarian carcinoma), much effort has been, and continues to be, expended towards the discovery and development of additional platinum-based anticancer drugs. Platinum drug development has proceeded in two broad directions concomitant with the two main limitations of cisplatin itself: namely its severe side-effects (especially on the kidneys, gastrointestinal tract and peripheral nerves) and its poor activity against some common tumours (e.g. colorectal and non-small-cell lung cancers) combined with its inability to confer lasting remissions in responding tumour types (especially ovarian) due to the emergence of drug resistance.

Chemical synthesis and cytotoxicity of dihydroxylated cyclopentenone analogues of neocarzinostatin chromophore.

Compounds containing the naphthoate moiety of Neocarzinostatin chromophore or 2-hydroxynaphthoate have been synthesized and evaluated for cytotoxic activity against a leukemia cell line and a small panel of human-tumor cell lines. Those compounds containing a cyclopentenone moiety were active, with the carbonyl group being essential for biological activity.

PRECLINICAL TESTING AND VALIDATION OF NOVEL ANTICANCER AGENTS

Publisher Summary This chapter discusses preclinical testing and validation of novel anticancer agents. Target validation is an essential component prior to initiating a drug discovery program. The amalgamation of combinatorial chemistry and robotic high-throughput screens allows for the rapid identification of hits, which can then be converted into leads and further optimized by medicinal chemistry. There is an increased emphasis on pharmacokinetics and pharmacodynamics throughout the process. The use of mice in in vivo antitumor studies should be restricted to molecules possessing good pharmaceutical properties where plasma levels above those known to be required for in vitro anticancer effects are achievable. It is found that for some types of compounds, testing using orthotopically grown tumor models rather than sc xenografts may be more appropriate. The more pharmacological-based approach combined with techniques to increase throughput (robotic cell-free assays, high-throughput whole cell growth inhibition and ELISA-based PD methods, cassette-dosing, hollow fiber assay) should result in the more rapid progression of carefully selected molecules for proof of principle phase I trials. It is suggested that adequately sensitive and validated assays for determining drug pharmacokinetics and pharmacodynamics should be in place to allow the possibility of dosing to a biological mechanism-based PD end point.