University of Louisville

Poster Title

KRAS Quadruplex-Forming Oligonucleotide: a Novel Therapeutic Option for Pancreatic Ductal Adenocarcinoma

Institution

University of Louisville

Abstract

Pancreatic cancer is a malignant neoplasm originating from transformed cells within the pancreas. It is the fourth most common cause of cancer-related deaths in the United States and, even when diagnosed early, has one of the worst prognoses of all human malignancies, with a median survival time of less than six months. Surgery is the best option for patients with localized disease; however, most patients experience local or metastatic recurrence. Therefore, there is a dire need for new therapeutic options. The KRAS oncogene, which is mutated and over-expressed in more than 95% of pancreatic ductal adenocarcinomas (PDACs), is essential to the initiation and maintenance of PDAC cell growth. Therefore, inhibition of KRAS expression may provide an effective treatment approach. The use of oligonucleotides (ODNs) for gene targeting of KRAS may be effective for promoting regression; however, problems with uptake, stability, and specificity prevent them from being used clinically. A guanine-rich, quadruplexforming sequence upstream of the KRAS promoter has been implicated in the regulation of KRAS expression. It was hypothesized that treatment of PDAC with an ODN encoding the KRAS quadruplex-forming sequence (KRASq) may abrogate cell growth by inhibiting KRAS expression. The results demonstrate that the 32-bp KRAS quadruplex-forming ODN formed a parallel quadruplex in solution. Four PDAC cell lines treated with KRASq (ASPC1, Panc-1, S2PC9, and S2VP10) showed prominent cellular uptake, with cytoplasmic and nuclear localization, as well as a significant dose and time-dependent decrease in cell proliferation after 24-144 hours. Interestingly, the two most sensitive cell lines, ASPC1 and S2PC9, had the highest basal levels of KRAS protein expression. Inhibition of proliferation in these two cell lines corresponded with a decrease in KRAS expression after 72-144 hours of KRASq treatment, but no change in the cell cycle was noted. Preliminarily, an approximate two-fold decrease in KRAS mRNA expression has been observed in ASPC1 and S2PC9.For more than thirty years, NCI funded institutions have been studying KRAS; however, at the time of this study’s inception, no therapeutic solutions to KRAS mutations had been developed. These results are the first of their kind and introduce a unique therapeutic method for significantly decreasing pancreatic cancer growth that abrogates KRAS expression while also addressing current concerns associated with conventional ODN therapy.

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KRAS Quadruplex-Forming Oligonucleotide: a Novel Therapeutic Option for Pancreatic Ductal Adenocarcinoma

Pancreatic cancer is a malignant neoplasm originating from transformed cells within the pancreas. It is the fourth most common cause of cancer-related deaths in the United States and, even when diagnosed early, has one of the worst prognoses of all human malignancies, with a median survival time of less than six months. Surgery is the best option for patients with localized disease; however, most patients experience local or metastatic recurrence. Therefore, there is a dire need for new therapeutic options. The KRAS oncogene, which is mutated and over-expressed in more than 95% of pancreatic ductal adenocarcinomas (PDACs), is essential to the initiation and maintenance of PDAC cell growth. Therefore, inhibition of KRAS expression may provide an effective treatment approach. The use of oligonucleotides (ODNs) for gene targeting of KRAS may be effective for promoting regression; however, problems with uptake, stability, and specificity prevent them from being used clinically. A guanine-rich, quadruplexforming sequence upstream of the KRAS promoter has been implicated in the regulation of KRAS expression. It was hypothesized that treatment of PDAC with an ODN encoding the KRAS quadruplex-forming sequence (KRASq) may abrogate cell growth by inhibiting KRAS expression. The results demonstrate that the 32-bp KRAS quadruplex-forming ODN formed a parallel quadruplex in solution. Four PDAC cell lines treated with KRASq (ASPC1, Panc-1, S2PC9, and S2VP10) showed prominent cellular uptake, with cytoplasmic and nuclear localization, as well as a significant dose and time-dependent decrease in cell proliferation after 24-144 hours. Interestingly, the two most sensitive cell lines, ASPC1 and S2PC9, had the highest basal levels of KRAS protein expression. Inhibition of proliferation in these two cell lines corresponded with a decrease in KRAS expression after 72-144 hours of KRASq treatment, but no change in the cell cycle was noted. Preliminarily, an approximate two-fold decrease in KRAS mRNA expression has been observed in ASPC1 and S2PC9.For more than thirty years, NCI funded institutions have been studying KRAS; however, at the time of this study’s inception, no therapeutic solutions to KRAS mutations had been developed. These results are the first of their kind and introduce a unique therapeutic method for significantly decreasing pancreatic cancer growth that abrogates KRAS expression while also addressing current concerns associated with conventional ODN therapy.