Folate Receptor Expression and Folate Receptor Targeted Chemotherapy in Cisplatin-sensitive and – resistant Human cancer cell lines Marverti G.a, Pirondi S.a, MarracciniC.a, Frassineti C.a, Helleman J.b, Berns E.M.J.J. b, and Costi, M.P. c aDept. Biomedical Sciences and cDept. Pharmaceutical Sciences, Via Campi 287-183, University of Modena and Reggio Emilia, 41125, Modena, Italy; Dept. of Medical Oncology, bErasmus University Medical Center - Daniel den Hoed Cancer Center, PO Box 2040, 3000 CA, Rotterdam,The Netherlands. Targeted drug delivery systems promise to expand the therapeutic windows of drugs by increasing delivery to the target tissue as well as the target–non-target tissue ratio. Interest in exploiting the folate receptor (FR) for drug targeting applications has rapidly increased because it is a tumor-associated antigen that is overex- pressed in greater than 90% of human ovarian carcinomas and associated with increased biological aggressive- ness of this tumor . The most significant advantage to emerge from studies of FR-mediated delivery has been the surprisingly low level of toxicity to normal tissues which constitute one of the most significant merits of the FR-targeting strategy. Therefore, FR presents an attractive target for tumor-selective drug delivery. The final goal of this research is to analyze the effects of oligopeptides, designed to inhibit thymidylate syn- thase (TS) activity by interfering with its dimerization, without causing TS over-expression and the develop- ment of cellular drug resistance against the traditional TS-targeted compounds. The over-expression of TS and the others folate cycle enzymes, is one of the major mechanisms of resistance to cisplatin (cDDP), encountered in most of resistant human ovarian cancer cell lines , accounting for the more efficient DNA repair and syn- thesis. To this aim, we have chosen two cisplatin-sensitive human ovarian cancer cell lines, 2008 and A2780, and their –resistant counterparts, C13* and A2780/CP cell lines, respectively, in order to display and study possi- ble different responses modulated by cDDP-resistance. At first, the cell lines have been tested for their total levels of FR and for functionally active receptors (FR or also other receptors?). The quantification of functional FR by the microfiltration method  and by an alterna- tive method, which deduces FR amount from the folic acid (FA) binding to FR  indicate that 2008 and C13* cells present a 3-4 fold higher expression of FR than A2780 and A2780/CP cells. However, time-dependent and concentration-dependent studies revealed that despite their higher expression of FR and the higher [3H]folic acid binding capacity, 2008 and C13* cell lines appeared to saturate earlier than A2780 and A2780/CP cell lines since they accumulated less [3H]folic acid at concentrations higher than 150 nM. Substrate specificity studies of the saturable uptake process, revealed that the 30 nM [3H]Folic acid uptake in A2780 cells was more affected than in the resistant line A2780/CP to the presence of 20μM of unlabeled folic acid (FA) and methotrexate (MTX), respectively (I do not understand this sentence, A2780 is more sensitive to unlabeled folic acid and mtx after labeled FA treatment? How do you measure this sensitivity? Do you mean affected/decreased uptake?). On the contrary, 30 nM [3H]Folic acid uptake was unaffected by unlabeled FA and MTX in both 2008 and C13* cell lines. ￼ To evaluate the presence of ATP-dependent process affecting FA uptake, experiments were also performed at 4°C and compared with those at 37°C. The accumulation of [3H]folic acid was greatly reduced at 4°C in comparison to 37°C both in 2008 and C13* cells, whereas only about 6- and 4-fold reductions were detected in A2780 and A2780/CP cells, respectively . These results are, at least partly, in accordance with the competitive uptake studies, since they suggest that in the resistant A2780/CP cells the accumulation is about 50% energy-dependent, carrier mediated and partly or slightly inhibited by competitive compounds. On the contrary, the sensitive A2780 cells are more sensitive to both temperature and folate analogs. However, the second cell couple (2008 and C13* cells), confirming to accumulate less [3H]folic acid, show an uptake more affected by low temperature and thus mostly energy- dependent, but not significantly affected by competitive compounds. To confirm the observed differences in FR expression and [3H]folic acid uptake, immunoblot analysis of FRα are in progress in a larger panel of human ovarian cancer cells including OAW28, COV504, IGROV1 and TOV112D cell lines. 1. Leamon CP and Reddy JA. Advanced Drug Delivery Review 2004, 56, 1127-41. 2. Scanlon KJ et al. Cancer Commun 1990, 2(10), 339-43. 3. Parker N, et al. Anal Biochem 2005;338: 284–93. 4. Mauritz, R. et al. Cancer Chemother Pharmacol 2008, 62:937–48. Acknowledgement This work has been supported by AIRC-DROC 10474 project to MPC.
Folate Receptor Expression and Folate Receptor Targeted Chemotherapy in Cisplatin-sensitive and – resistant Human cancer cell lines Marverti G.a, Pirondi S.a, MarracciniC.a, Frassineti C.a, Helleman J.b, Berns E.M.J.J. b, and Costi, M.P. cThe 4th International Symposium on Folate Receptors and Transporters Cozumel, Mexico October 7 - 11, 2012 / Marverti, Gaetano; Pirondi, Silvia; Marraccini, Chiara; Frassineti, Chiara; Costi, Maria Paola. - ELETTRONICO. - (2012), pp. 11-11.
|Data di pubblicazione:||2012|
|Titolo:||Folate Receptor Expression and Folate Receptor Targeted Chemotherapy in Cisplatin-sensitive and – resistant Human cancer cell lines Marverti G.a, Pirondi S.a, MarracciniC.a, Frassineti C.a, Helleman J.b, Berns E.M.J.J. b, and Costi, M.P. cThe 4th International Symposium on Folate Receptors and Transporters Cozumel, Mexico October 7 - 11, 2012|
|Autore/i:||Marverti, Gaetano; Pirondi, Silvia; Marraccini, Chiara; Frassineti, Chiara; Costi, Maria Paola|
|Data del convegno:||October 7 - 11, 2012|
|Luogo del convegno:||Cozumel-Messico|
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