İnterlökin-2 içeren ilaç taşıyıcı sistemlerin inhalasyon formülasyonlarına yönelik çalışmalar

Bu calismanin amaci; akcigerlerdeki primer ve/veya metastatik tumor olgularinin immunoterapi yontemiyle tedavisi icin pulmoner yol ile uygulanmak uzere, rekombinant insan interlokin-2 (rhIL-2) iceren mikropartikuler sistemlerin gelistirilmesi ve degerlendirilmesidir.IL-2 aktive olmus T hucreleri tarafindan uretilen bir sitokindir ve bagisiklik yanitta onemli bir immunomodulator olarak rol oynamaktadir. Bu biyolojik fonksiyonu nedeniyle rhIL-2 klinik calismalarda antikanser ajan olarak test edilmis ve metastatik renal hucre karsinomasi ve metastatik melanoma tedavisi icin FDA'den onay almistir.Calismamizda rhIL-2 iceren mikropartikul formulasyonlari s/y/s emulsiyon cozucu ekstraksiyon yontemi kullanilarak hazirlanmistir. Mikropartikul formulasyonlarinin hazirlanmasinda polimer olarak biyolojik olarak parcalanabilir ve biyolojik olarak uyumlu olmalari nedeniyle poli(laktik-ko-glikolik asit) (PLGA) polimeri kullanilmistir.Hazirlanan mikropartikul formulasyonlarinda sekonder emulsiyon karistirma hizi, dis fazda dispersiyon ajani olarak kullanilan PVA konsantrasyonu, ortam sicakligi, organik cozucu hacmi ve tipi, kullanilan polimer tipi ve karisim oranlari, ic sulu faza eklenen yardimci maddeler ve mikropartikullerin yuzey ozelliklerini modifiye etmek icin kullanilan mukoadhezif polimer tipinin mikropartikullerin partikul buyuklugu ve dagilimi, morfolojik ozellikleri, enkapsulasyon etkinlikleri, imalat verimi, zeta potansiyel degerleri, etkin madde salim profilleri ve aerodinamik ozellikleri uzerine etkisi incelenmistir.Mikropartikullerin partikul buyuklugu ve dagilimi incelenmis ve inhalasyon icin uygun partikul buyuklugu araligina sahip olduklari belirlenmistir. Yapilan morfolojik incelemeler mikropartikullerin kuresel ve duzgun yuzeyli olduklarini gostermistir. Uygulanan formulasyon parametrelerine bagli olarak degismekle birlikte yuksek enkapsulasyon etkinligine sahip mikropartikuller hazirlanabilmistir. Yapilan zeta potansiyel olcum sonuclari mikropartikullerin mukoadhezif polimerler ile kaplandigini gostermistir. Mikropartikullerin mukoadhezif polimerler ile kaplanmasi akcigerlerdeki kalis surelerinin uzatilarak etkinligin artirilmasinda avantaj saglamasi acisindan onemlidir. Elde edilen cozunme hizi profilleri incelenmis ve baslangictaki hizli etkin madde salimini yavas etkin madde saliminin takip ettigi bifazik salim profili elde edildigi gorulmustur. Baslangictaki bu hizli etkin madde salimi lokal olarak uygulanan rhIL-2'nin T lenfositlerinin aktivasyonu icin avantaj saglamaktadir. Cozunme hizi sonuclarinin kinetik degerlendirilmesi yapilmis ve en yuksek uyumun Higuchi kinetigine oldugu gorulmustur. Bu da etkin madde salim mekanizmasinin difuzyon kontrollu oldugunu gostermektedir. Mikropartikullerde yapilan aerodinamik incelemeler sonucunda yuksek % puskuren doz, kucuk KOAC ve yuksek IPF degerleri elde edilmistir. Elde edilen bu sonuclar mikropartikul formulasyonlarinin kuru toz inhalasyon icin uygun olduklarini gostermektedir. Ayrica kuru toz inhalasyon formullerinde mikropartikullerle birlikte Pearlitol 50 C'nin kullanilmasi partikul agregasyonunu onleyerek aerodinamik ozelliklerinin iyilesmesini saglamistir.Yapilan biyoaktivite calismalari mikropartikullerden salinan biyolojik olarak aktif rhIL-2'nin T hucre proliferasyonunu saglayabildigini gostermistir.Sonuclarimiz biyolojik olarak aktif rhIL-2 iceren, inhalasyon icin uygun ozelliklere sahip mikropartikullerin elde edildigini gostermistir. AbstractThe purpose of this study is to develop and evaluate microparticular systems containing recombinant human interleukin-2 (rhIL-2) for immunotherapeutic treatment of primer or/and metastatic tumor cases of lung.IL-2 is a cytokine produced by activated T cells and plays an important immunomodulatory role in the immune response. Because of this biological function, rhIL-2 has been tested in clinical trials as an anticancer agent and has been approved by the FDA for the treatment of metastatic renal cell carcinoma and metastatic melanoma.Microparticular formulations containing rhIL-2 were prepared using w/o/w emulsion solvent extraction method. Poly(lactic-co-glycolic acid) (PLGA) polymer was used in microparticle formulations due to its biodegradable and biocompatible properties.The effects of secondary emulsion mixing rate, concentration of PVA which is used as dispersing agent in inner phase, medium temperature, volume and type of the organic solvent, polymer type and ratios, excipients added to inner aqueous phase and type of the mucoadhesive polymer used to modify surface properties, on particle size and distribution, morphological properties, encapsulation efficiencies, production yield, zeta potentials, drug release profiles and aerodynamic properties of microparticles were investigated.The particle size and distribution of microparticles was investigated and concluded that they are in the appropriate range for inhalation. The morphologic investigations revealed that microparticles were spherical and had smooth surface. Depending on the formulation parameters, microparticles with high loading efficiencies were prepared. The zeta potential measurements showed that microparticles were successfully coated with mucoadhesive polymers. This coating is important for prolonging the stay of microparticles in lungs and thus enhancing their efficiency. The release profiles showed that a biphasic release was obtained with initial rapid drug release followed by slow drug release. This rapid initial release is advantageous on activation of T lymphocytes when rhIL-2 is administered locally. The kinetic evaluation of drug release showed that best fit was obtained in Higuchi model. This shows that drug release mechanism is diffusion controlled. As a result of aerodinamic investigations on microparticles, high % emitted dose, low MMAD and high FPF values were obtained. These results revealed that microparticle formulations were appropriate for dry powder inhalation. Besides this usage of Pearlitol 50 C with microparticles in dry powder inhalations was beneficial on aerodynamic properties by preventing particle aggregation.Bioactivity studies demonstrated that biologically active rhIL-2 released microparticles could led to T cell proliferation.Our results showed that microparticles with appropriate properties for inhalation were prepared which contain biologically active rhIL-2.