Preclinical investigations of nanocrystal dispersions and lipid nanoparticles of deuterated pyrazoloquinolinone ligands – phyicochemical and biological aspects

[J. R. Mitrović]

119 str.

Farmacija - Farmaceutska tehnologija / Pharmacy - Pharmaceutical Technology

Trenutni trendovi u otkriću novih potencijalnih kandidata za lek doveli su do sinteze supstanci sa niskom rastvorljivošću, što uglavnom vodi ka niskoj bioraspoloživosti nakon primene i time može zaustaviti njihov dalji razvoj. Iz tog razloga, uključivanje savremenih pristupa formulaciji već u ranoj fazi razvoja leka može omogućiti prevazilaženje nepovoljnih fizičkohemijskih osobina novosintetisanih supstanci i sprovođenje farmakoloških i toksikoloških prekliničkih studija. Deuterisani pirazolohinolinoni predstavljaju nove, patentno zaštićene ligande za GABAA receptore koje odlikuje visoka selektivnost, efikasnost i metabolička stabilnost, ali i niska rastvorljivost u vodi, uljima i mnogim organskim rastvaračima. Stoga je osnovni cilj ove doktorske disertacije bio razvoj nanočestičnih formulacija dva pirazolohinolinonska liganda, DK-I-56-1 (7-metoksi-2-(4‑metoksi‑d3-fenil)−2,5-dihidro-3Hpirazolo[ 4,3-c]hinolin-3-on) i DK-I-60-3 (7-metoksi-d3-2-(4-metoksid3-fenil)-2,5-dihidro- 3Hpirazolo[4,3-c]hinolin-3-on), kao i procena mogućnosti primene razvijenih formulacija u prekliničkim ispitivanjima navedenih supstanci. Izbor formulacionih strategija za prevazilaženje slabe rastvorljivosti bio je vođen fizičkohemijskim karakteristikama liganada uzimajući u obzir planiran put primene. Razvijena su dva tipa formulacija: nanokristali i lipidne nanočestice. Njihova sveobuhvatna karakterizacija uključila je procenu kritičnih fizičkohemijskih osobina značajnih za stabilnost tokom čuvanja i nakon primene, kao i procenu njihovog ponašanja nakon oralne i pareneteralne primene u kontekstu sprovođenja farmakokinetičkih i farmakodinamskih studija. Dok se kao osnovna prednost nanokristalnih formulacija izdvojio jednostavan sastav i niska koncentracija stabilizatora, lipidne nanočestice kompleksnije strukture obezbeđuju dodatno povećanje oralne bioraspoloživosti zahvaljujući lecitinu i polisorbatu 80 kao stabilizatorima. Dodatno, liofilizacijom uspešno je povećana stabilnost nanokristalnih disperzija i načinjen prvi korak ka prevazilaženju osnovnog nedostatka ovih sistema. Pokazano je da odabrane formulacije mogu doprineti ispitivanju pirazolohinolinonskih liganada kroz omogućavanje njihove primene u sastavu biokompatibilnih formulacija koje dovode do povećanja biološke raspoloživosti, ali i zadržavanja osnovnih osobina liganada tokom distribucije kroz organizam, kao i obezbeđenja praćenja farmakoloških efekata bez uticaja ekscipijenasa tradicionalno korišćenih za rastvaranje slabo rastvornih kandidata za lekove tokom njihovog prekliničkog istraživanja.

Recent trends in the discovery of new potential drug canidates have led to the synthesis of substances with low solubility which usually leads to the low bioavailability after administration. This often results in the discontinuation of the drug development. Because of that, the usage of advanced formulation approaches already in early stages of drug development could help to overcome the unfavorable physicochemical properties of new chemical entities and enable farmacological and toxicological preclinical studies. Deuterated pyrazoloquinolinones represent the new, patent protected ligands for GABAA receptors, with high selectivnost, efficacy and metabolic stability but very low solubility in water, oils and many organic solvents. Therefore, the main aim of this doctoral thesis was the development of nanoparticle formulations of two pyrazoloquinolinone ligands, DK-I-56-1 (7-Methoxy-2-(4‑methoxy‑d3-phenyl)−2,5-dihydro- 3H-pyrazolo[4,3-c]quinolin-3-one) and DK-I-60-3 (7-Methoxy-d3-2-(4-methoxyd3-phenyl)-2,5- dihydro-3Hpyrazolo[4,3-c]quinolin-3-one), as well as the estimation of the application of developed formulations in preclinical studies of the mentioned substances. The selection of formulation strategies to overcome the low solubility was guided by the physicochemical properties of the mentioned substances taking into account the intended administration routes. Two types of formulations were developed: nanocrystals and lipid nanoparticles. Their comprehensive characterization included the assessment of the physicochemical properties significant for the stability during storage and after administration as well as the analysis of their behavior after oral and parenteral administration in context of the prospective pharmacokinetic and pharmacodynamic studies. While the main advantage of nanocrystals was the simple composition and low concentration of stabilizers, lipid nanoparticles with more complex structure provided the additional increase of bioavailability thanks to lecithin and polysorbate 80 as stabilizers. In addition, the stability of nanocrystal dispersion during storage was successfully enhanced by lyophilization and in that way the first step to overcome the main disadvantage of these systems was made. It was shown that the selected formulations could contribute to the investigation of pyrazoloquinolinone ligands through their delivery within the biocompatible formulations which enable bioavailability enhancement, at the same time keeping the basic characteristics of ligands during distribution in the body as well as providing pharmacological effects monitoring without the effects of excipients traditionally used to dissolve low solubility drug candidates during their preclinical studies.

nanokristali, lipidne nanočestice, liofilizacija, niska rastvorljivost, pirazolohinolinoni, parenteralna primena, oralna primena, nuklearna magnetna rezonanca, farmakokinetika, test spontane lokomotorne aktivnosti.

nanocrystals, lipid nanoparticles, lyophilization, poor solubility, pyrazoloquinolinones , parenteral administration, oral administration, nuclear magnetic resonance, pharmacokinetics, spontaneous locomotor activity assay

615.015:543.51(043.3)

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