Effect of HTI–286 and CCI–779 alone and in combination on hepatic tumour cell proliferation

Overall survival in general is poor in HCC patients as most present with condition of advanced disease when surgical resection or transplantation is not possible anymore. Lack of conventional systemic chemotherapy is a non-acceptable circumstance. HTI–286 (HTI) is a new synthetic analogue of the tripeptide Hemiasterlin with antimicrotubule properties. It circumvents the P-glycoprotein mediated resistance in tumour cells. CCI–779 is known to inhibit tumour growth by antiangiogenesis. Here we studied the inhibitory effect of HTI and CCI–779 alone or in combination on hepatic tumour cell proliferation in vitro and in vivo.

Morris Hepatoma (MH); Hep3B, HepG2, primary human hepatocytes and HUVEC cells were used. MTT assay was done to determine the HTI and CCI–779 effect on each cell line. 5- Bromodeoxyuridine (BrdU) incorporation assay was performed to measure DNA-synthesis. Immunofluoresence assay with alpha-tubulin antibody was done to analyse the microtubule structure. For the in vivo experiments female American cancer institute rats were used. Subcutaneously grown MH cells were implanted in the liver. Rats were randomised into four groups (Control/ HTI/ CCI–779 and HTI+CCI–779). HTI was applied i.v. and CCI–779 was administered orally. Tumour growth was followed over a month with MR-imaging.

MH, HepG2 and Hep3B cell proliferation was inhibited highly significant by HTI whereas CCI–779 did not show any effect. Primary human hepatocytes were not affected by HTI. HUVEC proliferation was inhibited by CCI–779. Immunofluoresence revealed disruption of the microtubule structure under HTI treatment and multipolar spindles were seen in mitotic cells. BrdU incorporation showed cell accumulation into the G2-M phase. In vivo HTI treatment inhibited highly significant tumour growth compared to the control. CCI–779 had also significant inhibitory effect vs. control. Interestingly time pattern of growth inhibition under HTI and CCI–779 were different. In combination the inhibitory effect was highly significant compared to control and single treatments alone.

HTI inhibits hepatic tumour cell proliferation in vitro and tumour growth in vivo while not affecting primary human hepatocytes. CCI–779 seems primarily to target the vascular compartment. Combination of HTI and CCI–779 has an additive effect on tumour growth inhibition. HTI is to be considered a potent promising antiproliferative drug in liver oncology.

Literatur: HTI-286, a synthetic analogue of the tripeptide hemiasterlin, is a potent antimicrotubule agent that circumvents P-glycoprotein-mediated resistance in vitro and in vivo. Loganzo et al.; Oncology Research, Wyeth Research, Pearl River, New York 10965, USA. [e-mail] Rapamycin inhibits primary and metastatic tumor growth by antiangiogenesis: involvement of vascular endothelial growth factor. Guba et al.;Department of Surgery, University of Regensburg, Regensburg, Germany. [e-mail]

Key words

CCI-779 tumour proliferation - HCC - HTI-286