以微球及奈米劑型遞送鲁拉西酮之研究

Abstract

鹽酸魯拉西酮(LSD-HCl)是一種帶鹽類的結晶化合物，在2010年由FDA核准上市為第二代非典型抗精神病藥，其溶解度不佳(0.224 mg/mL at 20℃)，生體可用率只有約9-19%，因此如何提升LSD-HCl之生體可用率是需要被解決的問題。本研究利用三種不同的高分子，分別為聚乳酸-甘醇酸(Poly(lactide-co-glycolide), PLGA)、聚己內酯(poly(caprolactone), PCL)及聚丙烯酸樹脂(Eudragit® RS100, ERS)作為載體材料，希望開發具有胃滯留特性，可延長藥物釋放的微球劑型，及因奈米尺寸的優勢，促使藥物更容易被胃壁細胞吸收的奈米顆粒劑型，來遞送鹽酸鲁拉西酮，進而達到提升其生體可用率的目的。 在第一部分中，使用均質機以油/水溶媒乳化揮發法的方法製作微球，由實驗結果顯示有機相的聚合物濃度與微球的粒徑大小及包覆率有依存性，當有機相聚合物的濃度越高時，因有機相黏度提高使其製備出來的微球粒徑越大，而藥物的包覆率反而因脫去鹽酸鹽的量增加而下降。在掃描式顯微鏡的觀察下，三種高分子製備出來的微球呈現良好的球形。此外，紅外線分光光譜儀、示差掃描量熱儀的結果顯示，製備劑型後藥物已脫去鹽酸鹽，粉末X光繞射呈現藥物不僅脫去鹽類且結晶度亦下降。體內藥物釋放實驗的結果發現，整體而言，以LSD-PCL-MPs釋放速度最快，LSD-ERS-MPs次之，而LSD-PLGA-MPs則最慢且慢於LSD-HCl，三種微球的藥物釋放機制皆為Higuchi動力模式，藥物釋放速度提升原因來自於藥物結晶度的下降，而不同聚合物的釋放差異來自於微球表面的型態，LSD-PLGA-MPs的緩慢釋放來自於其表面上的無孔洞產生。在4小時的漂浮試驗中，三種聚合物所製備的微球其浮力皆隨著聚合物在有機相濃度越高而降低，且在相同濃度中，漂浮力排序為LSD-ERS-MPs > LSD-PCL-MPs > LSD-PLGA-MPs，與聚合物的密度排序呈現負相關，而LSD-PLGA-MPs、LSD-ERS-MPs及LSD-PCL-MPs漂浮力最高組別，其漂浮力依序為56.9±3.3%、73.6±5.9%、67.3±1.9%，顯示其劑型有幫助藥物漂浮的效果。在Wistar雄性大鼠的體內藥物動力學試驗，發現與LSD-HCl相比，三種微球的Cmax皆上升，Tmax皆下降，推測為藥物結晶度下降導致藥物溶解速度上升釋放速度變快且較完全；且因微球具有胃滯留效果使藥物釋放延長，因此三種微球的藥物t1/2皆變長，這兩種效果使LSD-PLGA-MPs、LSD-ERS-MP及LSD-PCL-MPs相較於LSD-HCl，顯著提升2.25±0.30、2.40±0.16及2.18±0.40倍的生體可用率。 而第二部分，同樣使用三種高分子為載體，用探針式超音波震盪器油/水溶媒揮發法製備包覆魯拉西酮的LSD-PLGA-NPs、LSD-ERS-NPs及LSD-PCL-NPs，粒徑大小分別為143.8±5.6、139.4±14.4、201.6±12.4 nm，表面電位為-24.5±2.2、33.7±1.3、-30.3±5.7 mV，三者的PDI皆在0.3以下。於4℃水中及冷凍乾燥後回溶於水的安定性試驗顯示，在28天內三種奈米劑型皆有良好的安定性。而經過紅外線分光光譜儀、示差掃描量熱儀、粉末X光繞射儀的測試後發現，與微球相同，製備奈米劑型後的藥物已脫去了鹽類且結晶度下降。體外釋放顯示，三種奈米劑型在48小時的釋放比例(LSD-PLGA-NPs: 87.7±9.7%, LSD-ERS-NPs: 87.4±2.8%, LSD-PCL-NPs: 84.2±4.3%)皆比單純藥物(LSD-HCl) (59.0±8.6%)高，此結果來自於藥物結晶度的下降及奈米尺寸的優勢。以Caco-2細胞作為腸壁細胞模型，進行細胞攝取及細胞轉運試驗，以流式細胞儀分析細胞攝取的結果，奈米顆粒的細胞攝取量皆顯著大於未包覆的Coumarin 6，說明奈米顆粒有幫助細胞攝取的能力。而細胞轉運試驗顯示奈米載體有幫助細胞攝取及轉運的效果，其表觀穿透係數依序為LSD-ERS-NPs (6.76±0.66×10-6 cm/s) > LSD-PLGA-NPs (4.85±0.14×10-6 cm/s) > LSD-PCL-NPs (4.56±0.43×10-6 cm/s cm/s) > LSD-HCl (1.35±0.26×10-6 cm/s)。在Wistar雄性大鼠的體內藥物動力學試驗中，奈米劑型比LSD-HCl，皆可顯著提高Cmax及縮短Tmax，推測為藥物結晶度下降導致藥物溶解速度上升、釋放速度變快且較完全，且奈米顆粒可以促進細胞攝取及細胞轉運的效果所致，而三種奈米顆粒其Cmax依序為LSD-ERS-NPs>LSD-PLGA-NPs>LSD-PCL-NPs，與表觀穿透係數排序相同，顯示細胞轉運能力是影響體內吸收的重要因素，以上因素使LSD-PLGA-NPs、LSD-ERS-NPs及LSD-PCL-NPs相較於LSD-HCl，顯著提升藥物生體可用率達2.91±0.32、3.30±0.48及2.37±0.24倍。

Lurasidone·HCl (LSD-HCl), the hydrochloride form of lurasidone, was approved by FDA as a second generation antipsychotic drug (SGA) in 2010 and usually used in treatment for schizophrenia and bipolar disorder. Because of low solubility (0.224 mg/mL at 20℃), the bioavailability of LSD-HCl is only about 9-19%. Therefore, how to improve its bioavailability is urgently required. In this study, three potential polymers, including poly(lactic-co-glycolic acid) (PLGA), Eudragit RS100 (ERS) and poly(caprolactone) (PCL), were selected to develop microparticle and nanoparticle delivery systems for LSD-HCl. It was expected the gastroretentive properties of microparticles (MPs) and the enhanced permeability and retention effect of nanoparticles (NPs) in gastrointestinal tract could improve the bioavailability. In the first part of study, LSD loaded PLGA, ERS, and PCL MPs were prepared by o/w solvent emulsion-evaporation method. The results showed that polymer concentration affected the particle size and encapsulation efficiency (EE) of MPs. Increase of the polymer concentration increased the particle size of MPs due to raising the viscosity of organic phase. However, the EE was decreased which resulted from the presence of free-base LSD. FT-IR and DSC analysis illustrated the drug present as a free-base form in the MPs. Powder X-ray further indicated the reductive drug crystallinity after MPs preparation. Findings from in-vitro release study demonstrated that the release rate of drug was in order of LSD-PCL-MPs > LSD-ERS-MPs > LSD-HCL ~ LSD-PLGA-MPs. In 4-hour buoyancy study, the buoyancy of MPs was in order of LSD-ERS-MPs (73.6±5.9%) > LSD-PCL-MPs (67.3±1.9%) > LSD-PLGA-MPs (56.9±3.3%). In vivo pharmacokinetic study in male Wistar rats showed that MPs had higher Cmax and shorter t1/2 than LSD-HCl. It was presumed that the reductive crystallinity of drug enhanced the rate of solubility of dug, led to higher Cmax. Longer t1/2 resulted from sustain-released and gastroretentive properties. The bioavailability of LSD-PLGA-MPs, LSD-ERS-MPs, and LSD-PCL-MPs was significantly increased by 2.25±0.30, 2.40±0.16, 2.18±0.40 folds relative to LSD-HCl. In the second part of study, LSD load PLGA, ERS, PCL NPs were formulated by o/w solvent evaporation method, too. The size and zeta potential of LSD-PLGA-NPs, LSD-ERS-NPs and LSD-PCL-NPs were 143.8±5.6, 139.4±14.4, 201.6±12.4 nm and -24.5±2.2, 33.7±1.3, -30.3±5.7 mV, respectively, and all of NPs had PDI < 0.3. These NPs exhibited good stability at 4℃ in ddH2O and after lyophilization for 28 days. The same as MPs, the results of FT-IR, DSC, and powder X-ray analysis illustrated the drug as free-base form in NPs and the crystallinity was reduced. The release of drug from NPs was higher than LSD-HCl because of the advantages of nano-size and, less crystallinity of drug. The cellular uptake of NPs in Caco-2 cell was higher than free coumarin-6. In cellular transport study, the apparent permeability coefficient (Papp) of drug was in order of LSD-ERS-NPs (6.76±0.66×10-6 cm/s) > LSD-PLGA-NPs (4.85±0.14×10-6 cm/s) > LSD-PCL-NPs (4.56±0.43×10-6 cm/s cm/s) > LSD-HCl (1.35±0.26×10-6 cm/s). It indicated that NPs can improve cellular transport of LSD. In vivo pharmacokinetic study in male Wistar rats demonstrated that NPs significantly increased Cmax and shortened t1/2 as compared to LSD-HCl. The reductive crystallinity of drug and enhanced cellular uptake as well as transport abilities of NPs resulted in this result. The Cmax of NPs was in order of LSD-ERS-NPs > LSD-PCL-NPs > LSD-PLGA-NPs which showed the same trend as Papp. It suggested that the cellular transport ability of NPs plays the important role in affecting drug absorption. The bioavailability of LSD-PLGA-NPs, LSD-ERS-MPs and LSD-PCL-NPs was 2.91±0.32, 3.30±0.48, 2.37±0.24 folds higher than LSD-HCl.