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Mechanism of Action of Novel Anti-Cancer Peptide, PNC-27, that Selectively Kills Cancer Cells.
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Michl, Josef, Pincus, Matthew R.
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Spring 2011
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2011-02-03
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Doctoral Dissertation
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In vivo and in vitro studies have shown the efficiency of anti-cancer peptides PNC-27 and PNC-28 in killing cancer cells by necrosis without affecting normal cells (1, 2 and 5). These peptides were derived from the hdm2-binding domain of p53 (PNC-27: aa12-26 and PNC-28: aa17-26) and linked to a membrane residency peptide (MRP).
Here I provide morphological and biochemical evidence of the effect of PNC-27 on a wide range of cancer cells including primary cancer cells, and the absence of any effect on untransformed cells. I also demonstrate that PNC-27-induced cytotoxicity is time, dose and temperature dependent. Moreover, I demonstrate the role of HDM2 as a possible tumor marker and a target for PNC-27 in the tumor cell’s plasma membrane which is confirmed by: immunofluorescence, exogenous expression of HDM2 in normal cell plasma membrane, competition experiments and immunoblot of isolated plasma membrane. I also evaluate sequence of cellular events following the exposure of tumor cells to PNC-27 using live cell imaging spinning disc confocal microscopy.
Furthermore, using Immuno-gold TEM and SEM I show that gold-conjugated antibodies to HDM2 and PNC-27 decorated ring-like pore structures in the plasma membrane of tumor cells exposed to PNC-27, further confirming the co-localization of HDM2 and PNC-27 in tumor cell membrane. These results strongly suggest that the highly amphipatic PNC-27 inserts itself into the plasma membrane of only cancer cells upon binding to HDM2, where, by random movement, the HDM2-PNC-27 complexes assemble into oligomers and eventually into membrane pores. Influx of free PNC-27 molecules through the newly formed membrane pores into the cell’s cytoplasm leads to the assembly of pores in mitochondrial membranes and rapid cell death.
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Yazdi, E. (2011). Mechanism of Action of Novel Anti-Cancer Peptide, PNC-27, that Selectively Kills Cancer Cells. [Doctoral dissertation, SUNY Downstate Health Sciences University]. SUNY Open Access Repository. https://soar.suny.edu/handle/20.500.12648/16143
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Doctoral Dissertation