Evaluation of Synergistic Effect of Cytosine-Phosphodiesterbond-Guanosin-Oligodeoxynucleotide 7909, and Protamine on Transfection Process Mediated by Calcium Phosphate Nanoparticles

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Ghassaq T. Al-Ubaidi
Ahmed A. Abbas
Ali A. Taha3
Qasim S. Sharhan
Israa W. Ahmed
Ilham A. Jasim


Bacille Calmette-Guerin (BCG) still the only authenticated vaccine against tuberculosis. Due to its drawbacks, a need for a new formula has emerged. The implication of “Nanovaccinology” is one of the possible alternatives. The non-viral vectors have a low transfection ability. In the context, this work aims to add two adjuvants to a calcium phosphate nanoparticles (CPNPs) functionalized with early secreted antigenic target 6-kilo dalton ( ESAT-6) cloned pcDNA3.1(+) plasmid. ESAT-6 gene is specific to mycobacterium tuberculosis complex (MTC) and encodes a T-cell antigen. The adjuvants in practice are Herring protamine and cytosine-phosphodiester bond-guanosine-oligodeoxynucleotide 7909 ( CpG-ODN 7909). Each has a different strategy in enhancing immune response; protamine is particulate adjuvant while CpG is an immunopotentiator substance. Nano complex was transfected into THP-1 monocytic cell line after its activation to a macrophage via 100nM PMA. Cellular immune response, interleukin-12 (IL-12), and tumor necrosis factor –alfa (TNF-ɑ) also ESAT-6 protein production were assayed via the Sandwich ELISA technique. Results revealed that CPNPs offer only partial protection to the adsorbed plasmid against enzymatic degradation. Nano complex formula with two adjuvants resulted in significantly higher cellular immune response comparing to formula carrying one adjuvant. In conclusion, the implication of CPNPs in gene delivery accompanied with two adjuvants each possess different strategy, will result in partial protection to the delivered gene with upsurge cellular immune response.

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