Selective induction of apoptosis by the cytotoxic analog AN-207 in cells expressing recombinant receptor for luteinizing hormone-releasing hormone.
The selectivity of ligands specific for certain cells can be used to preferentially target chemotherapeutic compounds to neoplastic cells. Human breast, ovarian, endometrial, and prostatic cancers express receptors that can mediate the delivery of targeted cytotoxic compounds to neoplastic cells. Recently, a potent derivative of 2-pyrrolinodoxorubicin (AN-201) conjugated to [D-Lys6] luteinizing hormone-releasing hormone (LH-RH) (AN-207), was demonstrated to be less toxic than the nonconjugated chemotherapeutic radical and significantly more active in slowing neoplastic cellular growth. In this study we investigate the molecular mechanisms underlying the cytotoxic action of AN-207. We stably transfected COS cells with a LH-RH receptor (LH-RH-Rc) mammalian expression vector and examined the effect of AN-207 on known markers of cellular apoptosis. Apoptotic induction by AN-207, as measured by Bax and Bcl-2 protein levels, was increased in stable cells that express LH-RH-Rc compared with parental cells. DNA fragmentation also was increased by AN-207 treatment when compared with AN-201. Clinically used LH-RH antagonists partially inhibited apoptotic Bax expression and DNA fragmentation induced by AN-207, and blocked AN-207 induced down-regulation of Bcl-2 steady-state protein levels. In cell proliferation studies, after 72 h AN-207 exhibited greater cytotoxicity than AN-201 at equivalent concentrations, in COS cells expressing LH-RH-Rc but not in parental COS cells. In addition, survival of LH-RH-Rc positive cells treated with AN-207 was partially restored by LH-RH antagonist. This study demonstrates the receptor-specific cytotoxic effect of 2-pyrrolinodoxorubicin conjugated to [D-Lys6] LH-RH, exerted through induction of apoptosis and modulation of Bax, Bcl-2, and DNA fragmentation.
Danila
D C
DC
Neuroendocrine Unit, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114, USA. Danila.Daniel@MGH.Harvard.edu
Schally
A V
AV
Nagy
A
A
Alexander
J M
JM
D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D019556:COS Cells; D002470:Cell Survival / Q000187:drug effects; D003603:Cytotoxins / Q000494:pharmacology*; D053938:DNA Fragmentation / Q000187:drug effects; D004317:Doxorubicin / Q000031:analogs & derivatives* / Q000494:pharmacology; D017930:Genes, Reporter; D006801:Humans; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011758:Pyrroles / Q000494:pharmacology; D011966:Receptors, LHRH / Q000378:metabolism*; D011994:Recombinant Proteins; D014162:Transfection / Q000235:genetics; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; D003603:Cytotoxins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011758:Pyrroles; D011966:Receptors, LHRH; D011994:Recombinant Proteins; D051028:bcl-2-Associated X Protein; C098751:AN 204; D004317:Doxorubicin