Characterization of the hollow fiber assay for the determination of microtubule disruption in vivo.
OBJECTIVE
The hollow fiber assay is used successfully as a routine in vivo screening model to quantitatively define anticancer activity by the National Cancer Institute. This study investigates whether the hollow fiber assay can be used as a short-term in vivo model to demonstrate specific pharmacodynamic end points, namely microtubule and cell cycle disruption.


METHODS
The growth of A549 cells was characterized within hollow fibers over 5 days in vivo at both subcutaneous (s.c.) and intraperitoneal (i.p.) sites. Drugs were administered on day 4 (i.p.).


RESULTS
At 24 hours, cells were retrieved from fibers at both i.p. and s.c. sites of paclitaxel-treated (20 mg/kg) and combretastatin A1 phosphate-treated (150 mg/kg) mice. Cell cycle analysis after paclitaxel treatment revealed a mean G(2)-M phase population of 48.04% (i.p.) and 25.76% (s.c.) compared with vehicle group mice (6.78 and 5.56%, respectively; P = <0.001 and 0.005, respectively). Tumor cells retrieved from combretastatin A1 phosphate-treated mice had a mean G2-M phase population of 36.3% (i.p.) and 29.36% (s.c.) compared with cells retrieved from vehicle group mice (5.58 and 5.49%, respectively; P = <0.001). Using fluorescence and laser-confocal microscopy, paclitaxel was revealed to induce the formation of spindle asters and tubulin polymerization. Combretastatin A1 phosphate was shown to hold cells in mitosis. Changes in nuclear morphology were also observed.


CONCLUSIONS
These data demonstrate that the hollow fiber assay can be used as a short-term in vivo model for studying the pharmacodynamic effects of both standard and novel compounds on microtubules. Evidence has also been provided to support the routine use of the in vivo hollow fiber assay for demonstrating the mechanism of action of a drug.
Tom Connors Cancer Research Centre, University of Bradford, Bradford, United Kingdom.
Marie
M
Suggitt
David J
DJ
Swaine
George R
GR
Pettit
Michael C
MC
Bibby
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004354:Drug Screening Assays, Antitumor / Q000295:instrumentation / Q000379:methods*; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D008567:Membranes, Artificial; D051379:Mice; D008815:Mice, Inbred Strains; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism*; D009368:Neoplasm Transplantation / Q000379:methods; D017239:Paclitaxel / Q000494:pharmacology; D011145:Polyvinyls; D008941:Spindle Apparatus / Q000187:drug effects / Q000378:metabolism; D013267:Stilbenes / Q000494:pharmacology; D014183:Transplantation, Heterologous; D014404:Tubulin / Q000032:analysis / Q000378:metabolism; D005465:Fluorocarbon Polymers
D000970:Antineoplastic Agents; D008567:Membranes, Artificial; D017239:Paclitaxel; D011145:Polyvinyls; D013267:Stilbenes; D014404:Tubulin; C063690:combretastatin A-1; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers