Development of a prolactin receptor-targeting fusion toxin using a prolactin antagonist and a recombinant form of Pseudomonas exotoxin A.
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.
Langenheim
John F
JF
Department of Biological Sciences, Clemson University, Clemson, SC, USA.
Chen
Wen Y
WY
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
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