Biochemical correlates of mTOR inhibition by the rapamycin ester CCI-779 and tumor growth inhibition

Citation
L. Dudkin et al., Biochemical correlates of mTOR inhibition by the rapamycin ester CCI-779 and tumor growth inhibition, CLIN CANC R, 7(6), 2001, pp. 1758-1764
Citations number
43
Categorie Soggetti
Oncology
Journal title
CLINICAL CANCER RESEARCH
ISSN journal
10780432 → ACNP
Volume
7
Issue
6
Year of publication
2001
Pages
1758 - 1764
Database
ISI
SICI code
1078-0432(200106)7:6<1758:BCOMIB>2.0.ZU;2-R
Abstract
The rapamycin ester, CCI-779, potently inhibits cell growth in vitro, inhib its tumor growth in vivo, and is currently in Phase I clinical trials. To f urther understand the relationship between plasma systemic exposure and inh ibition of the target Ser/Thr kinase, mTOR/FRAP, two assays have been devel oped. The first assay involves determination of the 4E suppressor protein ( 4E-BP1) bound to eukaryotic initiation factor 4E (eIF4E), and the second is direct Western analysis of phosphorylation of residue Thr(70) Of 4E-BP1. U nder normal growth conditions in vitro, rapamycin caused rapid association of 4E-BP1 with eIF4E within 1 h in Rh30 and GC(3) human tumor cells, Associ ation was persistent up to 16 h. In mice, administration of rapamycin (5 or 20 mg/kg) caused rapid association of 4E-BP1 with eIF4E within 4 h in both human colon adenocarcinoma GC, and rhabdomyosarcoma Rh30 xenografts, Using phosphospecific antibody against Thr(70) of 4E-BP1, rapid and persistent d ephosphorylation within 30 min of exposure to rapamycin was detected in Rh1 8 rhabdomyosarcoma cells. Evaluation of CCI-779 against Rh18 xenografts sho wed this tumor to be growth inhibited at daily dose levels of greater than or equal to8.7 mg/kg, Because immunoblotting may be more suitable for assay ing tumor biopsy tissue, a "blinded" comparison between the effect of CCI-7 79 on Thr(70) phosphorylation and growth inhibition of human tumor xenograf ts was undertaken. Mice were treated daily for 5 days with CCI-779 (20 mg/k g/day) or with drug vehicle, and tumor diameters were measured. Tumors were excised I h after the final administration and frozen, and phaspho Thr(70) was determined by Western blot analysis. The correlation coefficient for d ecreases in Thr(70) phosphorylation and growth inhibition was high (r(2), 0 .99). The results indicate that an assay of decreases in phosphorylation of Thr(70) Of 4E-BP1 may be a useful surrogate for determining the inhibition of mTOR activity in tumor specimens.