An improved human carboxylesterase for enzyme/prodrug therapy with CPT-11.
CPT-11 is a potent antitumor agent that is activated by carboxylesterases (CE) and intracellular expression of CEs that can activate the drug results in increased cytotoxicity to the drug. As activation of CPT-11 (irinotecan-7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin) by human CEs is relatively inefficient, we have developed enzyme/prodrug therapy approaches based on the CE/CPT-11 combination using a rabbit liver CE (rCE). However, the in vivo application of this technology may be hampered by the development of an immune response to rCE. Therefore, we have developed a mutant human CE (hCE1m6), based on the human liver CE hCE1, that can activate CPT-11 approximately 70-fold more efficiently than the wild-type protein and can be expressed at high levels in mammalian cells. Indeed, adenoviral-mediated delivery of hCE1m6 with human tumor cells resulted in up to a 670-fold reduction in the IC(50) value for CPT-11, as compared to cells transduced with vector control virus. Furthermore, xenograft studies with human tumors expressing hCE1m6 confirm the ability of this enzyme to activate CPT-11 in vivo and induce antitumor activity. We propose that this enzyme should likely be less immunogenic than rCE and would be suitable for the in vivo application of CE/CPT-11 enzyme/prodrug therapy.
Wierdl
M
M
Department of Molecular Pharmacology, St Jude Children's Research Hospital, Memphis, TN 38105, USA.
Tsurkan
L
L
Hyatt
J L
JL
Edwards
C C
CC
Hatfield
M J
MJ
Morton
C L
CL
Houghton
P J
PJ
Danks
M K
MK
Redinbo
M R
MR
Potter
P M
PM
D000256:Adenoviridae / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000378:metabolism / Q000627:therapeutic use; D015153:Blotting, Western; D019556:COS Cells; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000378:metabolism / Q000627:therapeutic use; D043182:Carboxylesterase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002522:Chlorocebus aethiops; D003131:Combined Modality Therapy; D018360:Crystallography, X-Ray; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D000077146:Irinotecan; D051379:Mice; D016513:Mice, SCID; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011355:Prodrugs / Q000378:metabolism / Q000627:therapeutic use*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D023041:Xenograft Model Antitumor Assays
D000972:Antineoplastic Agents, Phytogenic; D011355:Prodrugs; D000077146:Irinotecan; D043182:Carboxylesterase; D002166:Camptothecin