Targeting the cancer stroma with a fibroblast activation protein-activated promelittin protoxin.
Fibroblast-Activation Protein-α (FAP) is a membrane-bound serine protease that is expressed on the surface of reactive stromal fibroblasts present within the majority of human epithelial tumors but is not expressed by normal tissues. FAP is a postprolyl peptidase that differs from other dipeptidyl prolyl peptidases such as diprolylpeptidase 4 in that it also has gelatinase and collagenase endopeptidase activity. Therefore, FAP represents a potential pan-tumor target whose enzymatic activity can be exploited for the intratumoral activation of prodrugs and protoxins. To evaluate FAP as a tumor-specific target, putative FAP-selective peptide protoxins were constructed through modification of the prodomain of melittin, a 26 amino acid amphipathic cytolytic peptide that is the main toxic component in the venom of the common European honeybee Apis milefera. Melittin is synthesized as promelittin, containing a 22 amino acid NH(2)-terminal prodomain rich in the amino acids proline and alanine. In this study, peptides containing truncated melittin prodomain sequences were tested on erythrocytes to determine the optimal prodomain length for inhibiting cytolytic activity. Once optimized, modified promelittin peptides were generated in which previously identified FAP substrate sequences were introduced into the prodomain. Peptide protoxins were identified that were efficiently activated by FAP and selectively toxic to FAP-expressing cell lines with an IC(50) value in the low micromolar range that is similar to melittin. Intratumoral injection of an FAP-activated protoxin produced significant lysis and growth inhibition of human breast and prostate cancer xenografts with minimal toxicity to the host animal.
Department of Pharmacology and Molecular Sciences, The Johns Hopkins University, Baltimore Maryland 21231, USA.
Aaron M
AM
LeBeau
W Nathaniel
WN
Brennen
Saurabh
S
Aggarwal
Samuel R
SR
Denmeade
D000595:Amino Acid Sequence; D000818:Animals; D000970:Antineoplastic Agents / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D010450:Endopeptidases; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D018093:Gelatinases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008297:Male; D008555:Melitten / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008819:Mice, Nude; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000188:drug therapy / Q000473:pathology*; D015202:Protein Engineering; D011498:Protein Precursors / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D023041:Xenograft Model Antitumor Assays; D000099167:Fibroblast Activation Protein Alpha
D000970:Antineoplastic Agents; D010450:Endopeptidases; D018093:Gelatinases; D008555:Melitten; D008565:Membrane Proteins; D011498:Protein Precursors; D012697:Serine Endopeptidases; D000099167:Fibroblast Activation Protein Alpha; C027736:promelittin