[Construction of a non-viral vector H1s-EGFc and a preliminary study on its function]

Area of Science:

• Biotechnology
• Molecular Biology
• Cancer Research

Background:

• Gene therapy offers a promising avenue for cancer treatment.
• Development of efficient and targeted non-viral vectors is crucial for successful gene delivery.
• Existing viral vectors often pose safety concerns, necessitating exploration of non-viral alternatives.

Purpose of the Study:

• To construct and characterize a novel non-viral vector for targeted cancer gene therapy.
• To evaluate the efficacy of the developed vector in delivering a therapeutic "killing gene" into cancer cells.
• To assess the targeting capability of the vector towards cancer cells expressing the epidermal growth factor receptor (EGFR).

Main Methods:

• The H1s-EGFc fusion protein was engineered and expressed in Pichia pastoris, followed by purification.
• The H1s-EGFc fusion protein was complexed with pKG plasmid DNA encoding a "killing gene".
• HeLa (EGFR-positive) and Jurkat (EGFR-negative) cells were transfected with the H1s-EGFc/pKG complex, and cell viability was assessed using trypan blue staining.

Main Results:

• The H1s-EGFc fusion protein was successfully constructed and purified to over 90% purity.
• Transfection with H1s-EGFc/pKG complex demonstrated dose-dependent cancer cell killing.
• Killing rates of 30.6%, 36.2%, and 58.1% were observed at complex concentrations of 3, 6, and 9 microg/ml, respectively.

Conclusions:

• The H1s-EGFc fusion protein serves as an effective non-viral vector for targeted gene delivery in cancer therapy.
• The vector demonstrates efficient binding and internalization into specific cells, particularly those overexpressing EGFR.
• This approach holds potential for advancing targeted cancer gene therapy strategies.