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Lipopolymer mediated siRNA Therapy in Acute Lymphoblastic Leukemia

  • Author / Creator
    Mohseni, Mahsa
  • The nuclear transcription factor, Signal Transducer and Activator of Transcription 5 (STAT5) is one of the key drivers of Acute Lymphoblastic Leukemia (ALL), the most common childhood cancer, that promotes initiation, maintenance and progression to more aggressive stages of the malignancy by regulating the expression of oncogenes resulting in the aberrant proliferation of leukemic cells. Current therapeutic approaches for ALL including multi-agent chemotherapy and hematopoietic stem cell transplantation (HSCT), are associated with certain limitations, such as development of drug resistance leading to high rate of relapse. Therefore, it is outmost of importance to develop more effective and specific therapies for ALL including targeted therapeutic strategies. Small interfering RNA (siRNA) represents a promising tool to specifically target and inhibit the expression of genes that are involved in regulating fundamental cellular activities such as cell proliferation and migration. However, the properties of siRNA molecules including their biological instability, negative charge and large molecular weight prevent cellular delivery. Hence, potent carriers that can overcome extracellular and intracellular delivery hurdles of siRNA molecules and are effective in transfection of difficult-to-transfect cells, such as suspension ALL cells, are warranted for the progression of siRNA-based therapies towards clinical applications. Cationic polymers are one of the promising non-viral delivery systems for safe siRNA delivery due to their ability to bind and neutralize the anionic charges of siRNA molecules and package them into nano-scale complexes. In this thesis, we evaluated the potential of using lipid-modified polyethylenimines (PEIs) of low-molecular weights (0.6, 1.2, 2.0 kDa) in in vitro ALL models and patient-derived ALL cells and investigated the efficacy of lipopolymers for siRNA transfection with regard to cytotoxicity, siRNA uptake, gene silencing and biological effects (i.e., cell growth, and colony formation). The efficacy of siRNA transfection of lipid-substituted PEIs was explored in B-ALL RS4;11 and SUP-B15 cell lines grown in suspension and in ALL primary cells. The hypothesis of this thesis work was that lipid-modified PEIs would be able to effectively deliver siRNA molecules to ALL cells leading to the inhibition of target gene to achieve functional outcomes that could be potentially applied as an alternative therapeutic approach for ALL. Among the library of modified PEIs, linoleic acid (LA) and lauric acid (Lau) substituted PEIs (PEI-LA and PEI-Lau) have proven to be highly effective in delivering siRNAs to ALL cells, and higher uptake of siRNA/polymer complexes was observed compared to other polymer groups. In addition, STAT5A siRNA transfection by effective polymeric delivery systems caused a significant decrease in STAT5A mRNA levels in RS4;11 and SUP-B15 cells and some ALL primary cells which consequently led to marked increase in cell growth inhibition and decrease in colony formation ability in vitro compared to control groups. As promising outcomes were obtained in ALL cell lines and also primary cells by siRNA-mediated STAT5A silencing, we then explored the therapeutic effects of combining STAT5A siRNA and currently used chemotherapeutics for ALL including dexamethasone, doxorubicine, vincristine, and also TKIs such as nilotinib and dasatinib in RS4;11 and SUP-B15 cells. However, the results indicated that no synergistic effect was observed when cells treated with the combination of STAT5A siRNA and chemotherapeutics in comparison with individual siRNA or drug groups. Overall, the findings from this study showed the potential of siRNA-mediated STAT5A silencing as an individual therapy and are encouraging for the future design of non-viral delivery system with clinical translation capabilities for the treatment of ALL. This thesis work suggests opportunities for polymeric delivery systems that could be beneficial to inhibit other target genes in ALL and other leukemias (i.e., chronic and acute myeloid leukemias) for therapeutic purposes.

  • Subjects / Keywords
  • Graduation date
    Spring 2021
  • Type of Item
    Thesis
  • Degree
    Doctor of Philosophy
  • DOI
    https://doi.org/10.7939/r3-e54d-gy68
  • License
    This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.