A PCNA-Derived Cell Permeable Peptide Selectively Inhibits Neuroblastoma Cell Growth

Abstract

Proliferating cell nuclear antigen (PCNA), through its interaction with various proteins involved in DNA synthesis, cell cycle regulation, and DNA repair, plays a central role in maintaining genome stability. We previously reported a novel cancer associated PCNA isoform (dubbed caPCNA), which was significantly expressed in a broad range of cancer cells and tumor tissues, but not in non-malignant cells. We found that the caPCNA-specific antigenic site lies between L126 and Y133, a region within the interconnector domain of PCNA that is known to be a major binding site for many of PCNA's interacting proteins. We hypothesized that therapeutic agents targeting protein-protein interactions mediated through this region may confer differential toxicity to normal and malignant cells. To test this hypothesis, we designed a cell permeable peptide containing the PCNA L126-Y133 sequence. Here, we report that this peptide selectively kills human neuroblastoma cells, especially those with MYCN gene amplification, with much less toxicity to non-malignant human cells. Mechanistically, the peptide is able to block PCNA interactions in cancer cells. It interferes with DNA synthesis and homologous recombination-mediated double-stranded DNA break repair, resulting in S-phase arrest, accumulation of DNA damage, and enhanced sensitivity to cisplatin. These results demonstrate conceptually the utility of this peptide for treating neuroblastomas, particularly, the unfavorable MYCN-amplified tumors.

Expression of a novel peptide derived from PCNA damages DNA and reverses cisplatin resistance

“…The presence of the R9 sequence facilitates the destabilization of the cell membrane and the formation of transient pores through which a R9-peptide is transported into the cell [33]. The results of these experiments showed that treatment with micro-molar levels of R9-cc-caPeptide was cytotoxic to a panel of breast cancer and neuroblastoma cell lines [Smith SJ et al in press] [21]. …”

“…Like the induced peptide, R9-cc-caPeptide predominantly gravitates to the nucleus [21], but the possibility remains that as it passes from outside the cell to the nucleus, it may have a distinct subset of interactions from those of the induced peptide.…”

“…6). Gu et al [21] illustrated that γH2AX levels in irradiated neuroblastoma cells treated with R9-cc-caPeptide remained elevated for at least 48 h compared with cells treated with a control peptide where γH2AX levels reverted to baseline levels. They also showed that the prolonged elevation of γH2AX levels in caPeptide-treated cells correlated with impaired DNA repair by homologous recombination (HR) and a reduction in the recruitment of the HR repair protein Rad51 to sites of DNA damage.…”

“…Because it is part of a binding domain, caPeptide might also have the potential to function as an in vivo blocking peptide and be used to interfere with binding partners of caPCNA, possibly mitigating the aberrant functions due to caPCNA. Preliminary work has found that when the membrane penetrating D-Arg 9 (R9) sequence is added to the N-termini of the caPeptide, the peptide is very toxic to breast cancer, lymphoma, pancreatic and neuroblastoma cell lines [21] [Smith et al in press].…”

Cytotoxic effect of extract from Dunaliella salina against SH-SY5Y neuroblastoma cells

“…Neuroblastoma is one of the most common childhood neoplasms originating from neural crest progenitor cells of the sympathetic nervous system (Gu et al 2014). In our study, SH-SY5Y human neuroblastoma cells were used to assess cytotoxic effects of Dunaliella salina extract.…”