title: Development of a prolactin receptor-targeting fusion toxin using a prolactin antagonist and a recombinant form of Pseudomonas exotoxin A.
abstract: Human prolactin (hPRL) promotes the proliferation and differentiation of mammary epithelial cells during mammary gland development and has been linked to breast tumor development. The receptor for hPRL (hPRL-R) is elevated in a majority of human breast tumors, suggesting the overexpression of hPRL-R makes cancer cells highly sensitive to the mitogenic and anti-apoptotic activity of hPRL. These findings provide the rationale for the development of hPRL-R targeting breast cancer therapeutics. Previously, an hPRL-R antagonist, G129R, was developed that competitively binds to the hPRL-R resulting in growth inhibition and the induction of apoptosis in certain types of breast cancer cells. To further increase the potency of G129R, we fused G129R to a truncated form of Pseudomonas exotoxin A (PE(40)) that lacks the cell recognition domain of the toxin but retains the domains necessary for PE(40)_ to translocate into the cytosol and inhibit protein synthesis. We postulated that the fusion of G129R with PE(40)-KDEL would (1) deliver the recombinant toxin to breast cancer cells where hPRL-R is overexpressed; (2) block hPRL signaling via its G129R moiety; and (3) inhibit protein synthesis via its PE(40)-KDEL moiety. We demonstrate that the fusion toxin can competitively bind to hPRL-Rs on T-47D human breast cancer cells and inhibit STAT5 phosphorylation induced by hPRL. In addition, we show that G129R-PE(40)-KDEL is selectively cytotoxic to breast cancer cell lines expressing the hPRL-R and that cell death is associated with the inhibition of protein synthesis and does not involve caspase mediated apoptosis.
authors.affiliation: Department of Biological Sciences, Clemson University, Clemson, SC, USA.
authors.forename: John F
authors.initials: JF
authors.lastname: Langenheim
authors.forename: Wen Y
authors.initials: WY
authors.lastname: Chen
mesh_terms: D001426:Bacterial Proteins / Q000494:pharmacology*; D001427:Bacterial Toxins; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology*; D016923:Cell Death; D004268:DNA-Binding Proteins / Q000378:metabolism; D005260:Female; D006801:Humans; D008894:Milk Proteins / Q000378:metabolism; D010766:Phosphorylation; D011388:Prolactin / Q000494:pharmacology*; D011981:Receptors, Prolactin / Q000037:antagonists & inhibitors* / Q000502:physiology*; D011993:Recombinant Fusion Proteins; D050799:STAT5 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004268:DNA-Binding Proteins; D008894:Milk Proteins; D011981:Receptors, Prolactin; D011993:Recombinant Fusion Proteins; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; C402398:prolactin, Arg(129)-; C111749:pseudomonas exoprotein A protein, Pseudomonas aeruginosa; D011388:Prolactin