摻鐵硫化鈣磁性奈米粒子應用於癌症熱治療之研究

Abstract

本研究為利用摻鐵硫化鈣磁性奈米粒子作為癌症熱治療之研究。此奈米粒子具有良好之磁性，生物可分解性，以及生物相容性。實驗利用共沉法以及在氮氣中經過熱處理來和成摻鐵硫化鈣奈米粒子。此磁性粒子晶體結構經由X光繞射分析與硫化鈣類似。經由Debye-Scherrer方程式計算以及由穿透式電子顯微鏡可得知奈米粒子大小約40奈米。 由SQUID測得CaS為反磁性，一旦鐵離子摻入其磁性將轉成順磁性。當加熱至600oC 將轉換為鐵磁性。在磁場作用下具有足夠的能力使溫度在6分鐘內上升至42.5oC。尤其是在800oC加熱1小時，因為晶格中有更多的鐵離子取代鈣離子，其磁滯曲線面積將隨著加熱溫度的上升而增加。產生的熱能也隨著熱處理溫度的增加而上升。在37oC以及磁場條件 (f = 750 KHz, H = 10 Oe) 時，SAR值為45.47 W/g，經由LDH分析體外生物相容性證明對於3T3纖維母細胞並沒有細胞毒性。而在體外細胞熱療的結果也顯示摻鐵硫化鈣納米粒子能夠產生足夠的熱量，進而消滅CT - 26腫瘤細胞。 此外將進一步利用PVP高分子接合劑以及二氧化矽高分子對摻鐵硫化鈣磁性奈米粒子進行表面改質。改質後的磁性粒子鐵濃度11.6 wt%，其晶體結構也與CaS類似，平均粒徑約47.5nm並可以均勻得分散在水溶液中。經由紅外光譜分析可測得二氧化矽主要吸收峰。在37 °C時，磁性粒子濃度 10 mg/ml的條件下，SAR值為37.92 W/g。在磁場作用下具有足夠的能力使溫度在15分鐘內上升至45oC。細胞毒性測試也顯示了良好的生物相容性結果，表示此奈米粒子不影響細胞活力。 表面改質之摻鐵硫化鈣奈米粒子在體外細胞熱治療結果顯示在外加磁場下， 能夠使腫瘤細胞之細胞毒性上升60%， 明顯比未加磁性粒子或是未加磁場之控制組高。另外在小鼠動物實驗中，在Balb/c小鼠的皮下注入奈米粒子以及暴露在外加磁場下，經由15天的治療期後，腫瘤體積明顯出現減少趨勢。因此本研究新開發的摻鐵硫化鈣以及改質後的磁性奈米粒子是一種相當具有開發潛力的材料，期望在未來可以更深入的應用在癌症熱治療之研究。

In this study, a magnetic iron-doped calcium sulfide (Fe-CaS) nanoparticle was newly developed and studied for the purpose of hyperthermia due to its promising magnetic property, adequate biodegradation rate and relatively good biocompatibility. Fe-CaS nanoparticles were synthesized by a wet chemical co-precipitation process with heat treatment in an N2 atmosphere, and were subsequently cooled in N2 and exposured to air at a low temperature. The crystal structure of the Fe-CaS nanoparticles was similar to that of the CaS, which was identified by an X-ray diffractometer (XRD). The particle size was less than 40 nm based on a Debye-Scherrer equation and transmission electron microscope (TEM) examination. Magnetic properties obtained from the SQUID magnetometer demonstrated that the synthesized CaS was a diamagnetic property. Once the Fe ions were doped, the synthesized Fe-CaS converted into paramagnetism which showed no hysteresis loop. Having been heated above 600oC in N2, the Fe-CaS showed a promising magnetic property to produce enough energy to increase the temperature for hyperthermia. 10 mg/ml of the Fe-CaS was able to generate heat to elevate the media temperature over 42.5oC within 6 minutes. The area of the hysteresis loop increased with the increasing of the treated temperature, especially at 800oC for 1 hour. This is because more Fe ions replaced Ca ions in the lattice at the higher heat treatment temperature. The heat production was also increasing with the increasing of heat treatment temperature, which resulted in an adequate specific absorption ratio (SAR) value, which was found to be 45.47 W/g at 37oC under an alternative magnetic field of f = 750 KHz, H = 10 Oe.

The in vitro biocompatibility test of the synthesized Fe-CaS nanoparticles examined by the LDH assay showed no cytotoxicity to 3T3 fibroblast. The result of in vitro cell hyperthermia shows that under magnetic field the Fe-CaS nanoparticles were able to generate heat and kill the CT-26 cancer cells significantly. Furthermore, the sulfide-based magnetic Fe-doped CaS nanoparticles modified with a silica layer were then investigated. A polyvinyl pyrrolidone polymer was used as the coupling agent. The developed nanoparticles contained 11.6 wt% iron concentration, and their x-ray diffraction pattern was similar to those of CaS and Fe-CaS nanoparticles. The average particle size was approximately 47.5 nm and homogeneously dispersed in aqueous solutions. The major absorption bands of silica were observed from the FTIR spectrum. The magnetic properties and heating efficiency were also examined. The specific absorption ratio of nanoparticles at a concentration of 10 mg/ml at 37°C in an ethanol carrier fluid was 37.92 W/g and the nanoparticles would raise the temperature to over 45°C within 15 min. A cytotoxicity analysis revealed that the nanoparticles had good biocompatibility, which indicated that the nanoparticles did not affect cell viability. The therapeutic effects of the nanoparticles were investigated using in-vitro and animal studies. Cells seeded with nanoparticles and treated under an AC magnetic field revealed a percentage of cytotoxicity (60%) that was significantly higher from that in other groups. In the animal study, during a hyperthermia period of 15 days, tumor-bearing Balb/c mice that were subcutaneously injected with nanoparticles and exposed to an AC magnetic field manifested a reduction in tumor volume. The newly developed Fe-CaS nanoparticles and silica-modified Fe-CaS nanoparticles can thus be considered a promising and attractive hyperthermia thermoseed.