Secreted and tumour targeted human carboxylesterase for activation of irinotecan.
Irinotecan (CPT-11) is an anticancer agent for the treatment of colon cancer. CPT-11 can be considered as a prodrug, since it needs to be activated into the toxic drug SN-38 by the enzyme carboxylesterase. An approach to achieve tumour specific activation of CPT-11 is to transduce the cDNA encoding carboxylesterase into tumour cells. A secreted form of carboxylesterase may diffuse through a tumour mass and may activate CPT-11 extracellularly. This could enhance the antitumour efficacy by exerting a bystander effect on untransduced cells. In addition a secreted tumour-targeted form of carboxylesterase should prevent leakage of the enzyme from the site of the tumour into the circulation. We have constructed a secreted form of human liver carboxylesterase-2 by deletion of the cellular retention signal and by cloning the cDNA downstream of an Ig kappa leader sequence. The protein was secreted by transfected cells and showed both enzyme activity and efficient CPT-11 activation. To obtain a secreted, tumour-targeted form of carboxylesterase-2 the cDNA encoding the human scFv antibody C28 directed against the epithelial cell adhesion molecule EpCAM, was inserted between the leader sequence and carboxylesterase-2. This fusion protein showed CPT-11 activation and specific binding to EpCAM expressing cells. Importantly, in combination with CPT-11 both recombinant carboxylesterase proteins exerted strong antiproliferative effects on human colon cancer cells. They are, therefore, promising new tools for gene directed enzyme prodrug therapy approaches for the treatment of colon carcinoma with CPT-11.
Oosterhoff
D
D
Division of Gene Therapy, Department of Medical Oncology, Vrije Universiteit Medical Center, PO Box 7057, 1007 MB, Amsterdam, The Netherlands. D.Oosterhoff.oncol@med.vu.nl
Pinedo
H M
HM
van der Meulen
I H
IH
de Graaf
M
M
Sone
T
T
Kruyt
F A
FA
van Beusechem
V W
VW
Haisma
H J
HJ
Gerritsen
W R
WR
D000818:Animals; D000951:Antigens, Neoplasm / Q000235:genetics / Q000378:metabolism; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D015153:Blotting, Western; D019556:COS Cells; D002166:Camptothecin / Q000031:analogs & derivatives / Q000494:pharmacology*; D043182:Carboxylesterase; D002265:Carboxylic Ester Hydrolases / Q000235:genetics / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002522:Chlorocebus aethiops; D003110:Colonic Neoplasms / Q000188:drug therapy* / Q000201:enzymology* / Q000473:pathology; D004354:Drug Screening Assays, Antitumor; D005786:Gene Expression Regulation; D006801:Humans; D007124:Immunoenzyme Techniques; D000077146:Irinotecan; D008222:Lymphokines / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011355:Prodrugs / Q000494:pharmacology*; D012795:Sialoglycoproteins / Q000235:genetics / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000187:drug effects
D000951:Antigens, Neoplasm; D000972:Antineoplastic Agents, Phytogenic; D015815:Cell Adhesion Molecules; C089422:Fv protein, human; D008222:Lymphokines; D011355:Prodrugs; D012795:Sialoglycoproteins; D000077146:Irinotecan; D002265:Carboxylic Ester Hydrolases; D043182:Carboxylesterase; D002166:Camptothecin