Please use this identifier to cite or link to this item: https://www.um.edu.mt/library/oar/handle/123456789/31944
Title: Investigating the effect of insect conditioned medium and its constituents on the terminal differentiation of leukaemia
Authors: Cassar, Analisse
Keywords: Leukemia -- Malta
Cancer -- Differentiation therapy -- Malta
Insect hormones -- Malta
Tretinoin
Issue Date: 2017
Citation: Cassar, A. (2017). Investigating the effect of insect conditioned medium and its constituents on the terminal differentiation of leukaemia (Doctoral dissertation).
Abstract: Acute myeloid leukaemia is characterised by a block in haematopoiesis, resulting in uncontrolled clonal proliferation of immature cells leading to bone marrow failure. Acute promyelocytic leukaemia with t(15;17) reciprocal translocation has proven that overcoming the block in differentiation with the use of all-trans retinoic acid (ATRA) will lead cancer cells to enter apoptotic pathways, thus providing treatment without the use of chemotherapy. This success is however dependent on the genetic mutation of this blood cancer and in this study the aim is to cause differentiation to several leukaemia sub-types with various genetic mutations. Metamorphosis is a site of differentiation and media conditioned from cells of an insect pupa has already been proven to effect the differentiation of the HL-60 cell line. Organic extract from the conditioned medium (ICM) was used to expand and improve results to other leukaemia cell lines, with the addition of chromatin-modifying agents (CMAs) if needed. A method was optimised for analysis of superoxide dismutase (SOD) activity per relative cell number, as a screening test for differentiation. This facilitated the testing of multiple combinations of CMAs with ATRA or ICM in order to find the best possible treatments against leukaemia. Successful results were analysed for toxicity, and their differentiation effect confirmed by Leishman's stain then followed by flow cytometry for analysis of cell differentiation markers. The use of lowe doses of CMAs with ATRA proved beneficial in improving differentiation results of leukaemia cell lines other than NB4. However, the ICM resulted in more efficient differentiation in all the other cell lines than the differentiating agents being currently used clinically. The extract, alone or in combination with 5' Azacytidine, was the only chemical that caused differentiation in two out of three leukaemia patient primary cells. Analysis of the extract by GC-MS proved unsuccessful, however HPLC-MS helped to expose the presence of a juvenile hormone and a hydroxyecdysone within the ICM. Tests on the commercially available juvenile hormone and hydroxyecdysone showed some effect on SOD activity, however their combined use elicited a better response. The ICM poses a real therapeutic advantage over known differentiating agents and provides a true possibility in formulating a novel drug for the treatment of leukaemia. Additional chemical analysis of the components within ICM may further improve the achieved results and allow for more studies on the underlying mechanism by which this natural extract causes differentiation. Keywords: leukaemia, differentiation therapy, retinoic acid, metamorphosis, insect hormones
Description: PH.D.
URI: https://www.um.edu.mt/library/oar//handle/123456789/31944
Appears in Collections:Dissertations - FacM&S - 2017
Dissertations - FacM&SAna - 2017

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