Biology subjects

Wolfe, A. L.

Publications and source records attributed to Wolfe, A. L..

3 recordsLinked to original sources

bioRxiv2025

Selective targeting of oncogenic KRAS G12D using peptide nucleic acid oligomers attached to cell-penetrating peptides

KRAS is a proto-oncogene that contains activating mutations in up to 30% of tumors. Many conventional therapies inhibit both cancerous and normal cells, which may cause toxicity. Thus, programmable mutant-selective targeted inhibitors are needed. Peptide nucleic acids (PNAs) incorporate base sequences analogous to DNA, with modified peptide backbones instead of ribose-phosphate backbones, allowing PNAs to hybridize with DNA with high avidity to suppress transcription. Here, we developed KRAS G12D-selective PNA oligomers with novel cell-penetrating flanking regions. Fluorescein-labeled PNA oligomers displayed high uptake rates in cells and nuclei. Exposure to PNA-delivery peptide conjugates resulted in repression of KRAS G12D mRNA and protein expression within 2 hours and lasting up to 48 hours. Varying cell-penetrating peptide (CPP) compositions and lengths of complementary KRAS sequences were tested using dose-response cell viability assays. These experiments identified configurations that were effective at selectively preventing growth of on-target KRAS G12D cells, while relatively sparing off-target KRAS G12C cells. Electrophoretic mobility shift assays demonstrated in vitro binding and selectivity for KRAS G12D DNA sequences. CPP-PNA-G12D-1 was effective against a panel of pancreatic ductal adenocarcinoma cell lines and patient-derived xenografts in vivo. These results show promise for an enhanced PNA-delivery peptide conjugate strategy as both a tool for studying tumors driven by oncogenic point mutations and as a potential therapeutic strategy to selectively target mutant cancer cells.

cancer biology↗

bioRxiv2024

Effects of N361 Glycosylation on Epidermal Growth Factor Receptor Biological Function

Epidermal growth factor receptor (EGFR) is a transmembrane tyrosine kinase that is frequently modified by glycosylation post-translationally. In cancer, EGFR amplifications and hotspot mutations such as L858R that promote proliferation have been detected in a significant fraction of non-small cell lung carcinomas and breast adenocarcinomas. Molecular dynamic simulations suggested that glycosylation at asparagine residue 361 (N361) promotes dimerization and ligand binding. We stably expressed glycosylation-deficient mutant EGFR N361A, with or without the oncogenic mutation L858R. Immunofluorescence and flow cytometry demonstrated that the mutants were each well expressed at the cell membrane. N361A decreased proliferation relative to wild-type EGFR as well as decreased sensitivity to ligands. Proximity ligation assays measuring co-localization of EGFR with its binding partner HER2 in cells revealed that N361A mutations increased co-localization. N361A, located near the binding interface for the EGFR inhibitor necitumumab, desensitized cells expressing the oncogenic EGFR L858R to antibody-based inhibition. These findings underline the critical relevance of post-translational modifications on oncogene function. SIGNIFICANCEEGFR transduces signals from growth factors into cell proliferation and is frequently hyperactivated in tumors. Glycosylation of EGFR at N361 regulates EGFR dimerization, growth factor stimulation of proliferative signaling, and susceptibility to targeted inhibition. Insights into EGFR glycosylation may expand therapeutic opportunities to benefit cancer patients.

cancer biology↗

bioRxiv2020

UDP-glucose pyrophosphorylase 2, a regulator of glycosylation and glycogen, is essential for pancreatic cancer growth

Pancreatic ductal adenocarcinomas (PDACs) have enhanced nutrient uptake requirements and rapid metabolic processing. The enzyme UDP-glucose pyrophosphorylase 2 (UGP2) rests at the convergence of multiple metabolic pathways, however the role of UGP2 in tumor maintenance and cancer metabolism remains unclear. Here, we identify an essential role for UGP2 in the maintenance of PDAC growth in both in vitro and in vivo tumor models. Transcription of UGP2 is directly regulated by the YAP/TEAD complex. Loss of UGP2 leads to decreased intracellular glycogen and defects in N-glycosylation targets important for cell growth including epidermal growth factor receptor (EGFR). In murine xenograft models, knockdown of UGP2 halted tumor growth and repressed expression of EGFR. The critical roles of UGP2 in cancer maintenance, metabolism, and protein glycosylation may offer new avenues of therapy for otherwise intractable PDACs. Impact StatementConvergent findings reveal that UDP-glucose pyrophosphorylase 2 has a central role in growth and metabolism of pancreatic ductal adenocarcinomas, highlighting novel therapeutic possibilities for this deadly cancer.

cancer biology↗