DEPARTMENT OF LIPIDS BIOCHEMISTRY
Head – Professor Nadiia M. Hula, Dr. Sc., Corresponding Member of the National Academy of Sciences of Ukraine and National Academy of Medical Sciences of Ukraine
The Department of Lipids Biochemistry was founded in 1980 as the Laboratory of Cell Cultures. In 1991, it was reorganized into the Department of Lipids Biochemistry. Its primary area of research was the functional role of lipids in cells. Since the foundation of the department, Nadiia M. Hula worked as its head.
Currently the Department includes 3 Doctors of sciences and 8 Candidates of sciences.
In the early 1980’s Nadiya Hula and her team discovered a new class of biologically active low-polar lipids N-acylethanolamines (NAE) in the cells of neuroblastoma С1300N18. The Department continued its research on the role and functioning of these compounds, demonstrating for the first time the membranotropic and membrane protective effect of NAE. The Department researchers have established that the effect of the different NAE is contingent upon the structure of acyl residue. This line of research, along with the research of other scholars, led to the discovery of the endocannabinoid system.
Main results (2007-2015)
The Department employees have established the antiallergic and anti-inflammatory effect of saturated NAE N-stearoylethanolamine (NSE): it modulates the histamine levels and inhibits the production of proinflammatory cytokines TNF-alpha, IL- 1, IL-6 at the early stages of pathologic processes. When studying the NSE effect on nonspecific inflammation in rats with experimental burns, it was discovered that the NSE could accelerate healing processes in thermal burn wounds by inhibiting the production of proinflammatory cytokines (TNF-alpha, IL-6, IL-1), normalizing the level of the stable metabolite of nitric oxide (nitrite-anion and activity of constitutive and inducible NO-synthases) and eliminating the imbalance between free-radical lipid oxidation and protective antioxidant enzyme activity in plasma, erythrocytes, liver and spleen.
It was demonstrated that unsaturated NAE N-oleoylethanolamide (OEA) and NSE could affect the proliferation of both intact and irradiated cells. The concentration impact of the NAE effect was demonstrated under these conditions. Antiproliferative effects of NAEs are not associated with the activation of apoptotic cell death. Failures of the reproductive function of cells are probably caused by the impact of NAE on the functioning of mitochondria. It was demonstrated that NAE can inhibit the proliferation of irradiated transformed cells that are associated with the activation of apoptosis. NSE and OEA act as modulators of electrical excitability of the plasma membrane of neonatal rat cardiomyocytes by affecting the action potential duration. The shortening of the action potential duration in response to NSE and OEA occurs as the result of both potassium currents potentiation and the inhibition of incoming calcium current through calcium channels of L-type. The reduced amplitude and initial rate of the action potential rise in the presence of NSE suggests probable depression of fast sodium current by the substance.
It has been shown on rat males treated with 17beta-estradiol that NSE, when co-administered with 17beta-estradiol, has a pronounced protective effect on testicular tissue of rats, preventing the deterioration in its structural and functional state, the decrease of antioxidant enzyme activity and the level of nitrite-anion, and a decline in testosterone production.
It has been demonstrated that the immobilization of NAE in new oligomeric carriers retains completely the cytoprotective activity of N-acylethanolamines in vitro, but at the same time improves significantly (50%) the antitumor activity and speeds up the delivery of substances (including doxorubicin) immobilized in these carriers to target cells. It has been demonstrated that composite NSE, when administered with polyvinylpirrolidon and doxorubicin (compNSE), results in the 100 % survival rate of mice with Lewis carcinoma.
It has been shown that NSE suspension and compNSE administered when cancer develops can reduce the level of urea in the blood plasma of mice. Combining cytoprotector and antitumor drug on one drug delivery platform solves the fundamental problem of cancer chemotherapy: it reduces the effective dose of chemotherapy agent while cytoprotector NAE molecules protect normal cells from the side effects of the drug.
It has been demonstrated that NSE can selectively affect the proliferation of normal, pseudonormal and transformed cells. The Department researchers have discovered its cytoprotective effect on normal (primary myogenic fibroblast-like cells in rats) and pseudonormal (pseudonormal cell culture 293T) cells, and a pronounced antiproliferative effect on transformed cell cultures (Jurkat line, L1210, L929, A-549) and hybridoma (SP2 / 0). It has been demonstrated that NSE has an antiproliferative effect on transformed cell cultures through several biochemical mechanisms.
It was established on Jurcat line transformed cells culture that the antiproliferative effect of NSE was based on apoptotic cell death. NSE destroys L929 and L1210 cells through interphase cell death, not through activating apoptosis.
Incubation of both transformed (L929) and primary cells (myogenic and fibroblast-like rat cells) with NSE has revealed changes in the activity of mitochondrial enzymes: succinate dehydrogenase and alpha-glycerophosphate dehydrogenase. This suggests that the lowered proliferation of certain cell types under the impact of NSE may occur due to the inhibition of energy metabolism in the cells.
It has been established that the incubation of transformed fibroblasts L929 on NSE in the presence of the known antiproliferative agent antimycin A, that severs the respiratory chain of mitochondria, may reduce the antiproliferative effects of antimicyn A, prevent the formation of giant cells and preserve the activity of succinate dehydrogenase. We assume that the NSE effect in this case is based on partial preventing the antimycin embedding in the membrane apparatus of mitochondria.
It has been established that the co-culturing of the transformed fibroblasts L929 with NSE in the presence of cyclodextrin prevents the removal of membrane cholesterol and precludes its effect on the phospholipid composition of transformed fibroblasts.
The obtained results are of practical significance. The researchers intend to develop fundamentally new drugs with anti-allergy, anti-inflammatory and antitumor effects based on N-acylethanolamines.
The research of 1980-2009 is summarized in the volume New Class of Bioactive Lipids N-Acylethanolamines. Discovery, Research and Practical Application, which was awarded the State Prize of Ukraine in Sciences and Technology in 2010.
As part of the project on researching the antivirus effects of NSE, which started in 2010, the Department explored its effect on the reproduction of the influenza subtype H1N1.
It has been established that N-stearoylethanolamine (NSE) inhibits the reproduction of influenza A virus subtype H1N1 in vitro at 3.0 lg 50% tissue cytopathic dose in 10-6 and 10-7 M concentration. Hence, the chemotherapeutic index of NSE as regards the influenza A virus subtype H1N1 is 100, that allows us to classify NSE as an active anti-influenza drug. In vivo experiments demonstrate that intranasal administering of NSE as a prophylactic scheme at the dose of 81.75 µg/kg and 0.8175 µg/kg increased the survival rates of infected animals by 40% and 100% respectively. At the 0.8175 µg/kg NSE dosage, a low virus titer was discovered in the lung tissue (<0.5 lg 50% tissue cytopathic dose). As a treatment scheme, the most effective dosage of NSE for one-time administering of the drug was 81.75 µg/kg and 0.08175 µg/kg, which significantly lowered the mortality rate from the influenza. NSE inhibits the virus reproduction in the lung tissue of the infected mice by 2.0 and 3.5 lg, respectively. It has been established that the anti-influenza effect of NSE is based on inhibiting the neuraminidase activity of influenza A virus subtype H1N1. When NSE is administered intraperitoneally at the dose of 81.75 µkg/kg, 8.175 µg/kg, 0.8175 µg/kg and 0.08175 µg/kg, it induces IFNγ in mice, while the 0.8175 µg/kg and 0.08175 µg/kg dosage induces IFNα, which might be yet another mechanism of the anti-influenza effect of NSE. It was also established that the intranasal administering of NSE in all the researched doses and schemes prevented the changes in cholesterol levels in lung and liver tissue of the mice infected with the influenza virus.
Rat models of the type 1 diabetes (Strepozotocin model) and type 2 diabetes (alimentary obesity-induced insulin resistance) demonstrated that NSE has a marked hypoglycemic effect and reestablishes insulin sensitivity in the tissues of insulin-resistant rats. Experiments demonstrated that NSE renews the phospholipid composition of the liver and inhibits the progression of steatosis in the pancreas of experimental insulin-resistance rats.
Four theses for the Candidate’s degree have been defended (2009, 2011, 2012, 2015). One doctoral student graduated in 2014.
In 2010, the Department signed a license agreement, giving Farmak the exclusive rights to objects of intellectual property for the mass production of pharmaceutical substances based on NAE.
In 2011, Farmak completed the development of the process procedure for the production of NAE and started developing various pharmaceutical forms of NAE-based drugs.
In 2014, Farmak completed the preclinical trials of the N-Acetylethanolamine-based drug and started preparing for clinical trials.