Rationale: l-OHP showed p53-independent activity in neoplasm with acquired cisplatin resistance. PTX and DTX are active in NSCLC patients acting by stabilizing microtubule (MT) polymerization. MAP4 stabilizes MTs and is inhibited by p53 expression. DNA-damage induction of p53 can affect the sensitivity to anti-MT drugs. Materials and Methods: in vitro evaluation of l-OHP, PTX and DTX activity as single agents (S), concomitant (CS, l-OHP+PTX/DTX for 24h) and sequential association (SS, l-OHP/PTX/DTX and PTX/DTX/l-OHP for 24h+24h) in terms of cytotoxicity, recovery time (RT), combination index (CI), apoptosis and Western Blot (WB) in A549 (A) and H322 (H) (p53 wt and mt) NSCLC cell lines, were performed. Cytotoxicity was evaluated by a coulter counter; apoptosis with PI analyzed by FACS; p53 and MAP4 proteins levels by WB. Results: concentration inhibiting 50% of cells at 24h (IC50) was of 11.63±12 and 442.61±459.15 M, 24.6±6.1 and 10.5±1.3 nM and 7.6±3.8 and 3.7±0.3 nM for l-OHP, PTX and DTX in A and H respectively. CI showed synergism in all combinations in A (<1), resulting 6 fold stronger for l-OHP PTX/DTX and PTX/DTX l-OHP (0.116±0.0001 vs 0.67±0.35) SS; whereas antagonism (>1) for l-OHP+PTX/DTX (>2.38±0.219), additivity (=1)/antagonism for PTX/DTX l-OHP and l-OHP PTX (1.49±0.45) and modest synergism for l-OHP DTX (0.9±0.01), was found in H. RT, at 24 and 72h, showed a further growth inhibition as CS and SS in A and H. Apoptosis evaluation showed best activity for the PTX/DTX l-OHP sequence and for DTX alone in A; in H the best activity resulted to be the l-OHP PTX/DTX sequence and l-OHP alone after RT. WB analysis showed a direct correlation between p53 induction and MAP4 down-regulation (p=0.0018) more evident for the PTX/DTX l-OHP sequence in A, whereas it was not present in H. Conclusions: l-OHP shows a p53-independent activity with a 38 fold lower IC50 in A than in H. MAP4 p53-dependent inhibition, in terms of cytotoxicity, apoptosis induction and molecular interaction features, explains the better activity of PTX/DTX l-OHP and l-OHP PTX/DTX SS in A and H NSCLC cell lines respectively. Supported in part by ASCO onlus
Oxaliplatin (l-OHP) association with paclitaxel (PTX) or docetaxel (DTX) in NSCLC cell lines: Schedule-dependent activity based on p53/microtubule associated protein 4 (MAP4) interaction
FLATI, VINCENZO;RICEVUTO, Enrico;
2002-01-01
Abstract
Rationale: l-OHP showed p53-independent activity in neoplasm with acquired cisplatin resistance. PTX and DTX are active in NSCLC patients acting by stabilizing microtubule (MT) polymerization. MAP4 stabilizes MTs and is inhibited by p53 expression. DNA-damage induction of p53 can affect the sensitivity to anti-MT drugs. Materials and Methods: in vitro evaluation of l-OHP, PTX and DTX activity as single agents (S), concomitant (CS, l-OHP+PTX/DTX for 24h) and sequential association (SS, l-OHP/PTX/DTX and PTX/DTX/l-OHP for 24h+24h) in terms of cytotoxicity, recovery time (RT), combination index (CI), apoptosis and Western Blot (WB) in A549 (A) and H322 (H) (p53 wt and mt) NSCLC cell lines, were performed. Cytotoxicity was evaluated by a coulter counter; apoptosis with PI analyzed by FACS; p53 and MAP4 proteins levels by WB. Results: concentration inhibiting 50% of cells at 24h (IC50) was of 11.63±12 and 442.61±459.15 M, 24.6±6.1 and 10.5±1.3 nM and 7.6±3.8 and 3.7±0.3 nM for l-OHP, PTX and DTX in A and H respectively. CI showed synergism in all combinations in A (<1), resulting 6 fold stronger for l-OHP PTX/DTX and PTX/DTX l-OHP (0.116±0.0001 vs 0.67±0.35) SS; whereas antagonism (>1) for l-OHP+PTX/DTX (>2.38±0.219), additivity (=1)/antagonism for PTX/DTX l-OHP and l-OHP PTX (1.49±0.45) and modest synergism for l-OHP DTX (0.9±0.01), was found in H. RT, at 24 and 72h, showed a further growth inhibition as CS and SS in A and H. Apoptosis evaluation showed best activity for the PTX/DTX l-OHP sequence and for DTX alone in A; in H the best activity resulted to be the l-OHP PTX/DTX sequence and l-OHP alone after RT. WB analysis showed a direct correlation between p53 induction and MAP4 down-regulation (p=0.0018) more evident for the PTX/DTX l-OHP sequence in A, whereas it was not present in H. Conclusions: l-OHP shows a p53-independent activity with a 38 fold lower IC50 in A than in H. MAP4 p53-dependent inhibition, in terms of cytotoxicity, apoptosis induction and molecular interaction features, explains the better activity of PTX/DTX l-OHP and l-OHP PTX/DTX SS in A and H NSCLC cell lines respectively. Supported in part by ASCO onlusPubblicazioni consigliate
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