Toxicity Studies of Cremophor-free Paclitaxel Solid Dispersion Formulated by a Supercritical Antisolvent Process

박재현,지상철,용철순,최한곤,우종수,이원석,이원모,구윤본 대한약학회 2009 Archives of Pharmacal Research Vol.32 No.1

To evaluate the acute toxicity of a paclitaxel solid dispersion formulation, single dose studies in ICR mice were carried out for injectable excipients, paclitaxel solid dispersion powder, and Taxol®. In the dose range of excipients used for preparing paclitaxel solid dispersion, each excipient was clinically safe, and the LD50 for exicipients was higher than 2,000 mg/kg for both males and females. In this study, there were no remarkable clinical signs or deaths related to paclitaxel solid dispersion even at doses up to 160 mg/kg of paclitaxel. But Taxol® resulted in clinical signs when it contained more than 30 mg/mL paclitaxel. The LD50 for paclitaxel solid dispersion was above 160 mg/kg and the LD50 for Taxol® was 31.3 mg/kg, more than 5 times lower than that of paclitaxel solid dispersion. However, paclitaxel solid dispersion could not be administered i.v. at a dose exceeding 160 mg/kg, because of high viscosity. To evaluate the nephrotoxicity of paclitaxel solid dispersion, plasma level of creatinine and kidney weight were measured and compared to Taxol®. At the doses administered, paclitaxel solid dispersion did not change creatinine clearance, while Taxol® killed all animals at doses >15 mg/kg. To investigate membrane damage when paclitaxel formulations were injected, hemolytic activity was determined for different concentrations. Paclitaxel solid dispersion showed about 10% hemolytic activity, whereas Taxol® showed about 40% hemolytic activity when it contained 2 mg of paclitaxel. Comparisons with the LD50 value, nephrotoxicity, and hemolytic activity of Taxol® suggested that Cremophor-free paclitaxel solid dispersion as an injectable formulation is a promising approach to increasing the safety and clinical efficacy of paclitaxel for treatment of cancer.

Paclitaxel에 의한 관절연골 세포의 capase-비의존적 mitotic catastrophe 유도

Jeong Hee Im(임정희),Song Ja Kim(김송자) 한국생명과학회 2010 생명과학회지 Vol.20 No.4

Paclitaxel은 미세소관의 탈중합을 억제하는 시약으로 알려져 있다. Paclitaxel은 다양한 세포에서 세포 내 방추체를 안정화시킴으로써 유사분열 억제 및 세포사멸을 유도한다. 본 실험에서는 토끼 관절 연골세포에서 paclitaxel이 연골세포의 증식과 사멸에 미치는 효과에 대한 연구를 수행하였다. MTT assay를 수행한 결과 paclitaxel은 연골세포에서 농도 의존적으로 세포 증식을 억제한다는 것을 확인 할 수 있었으며, FACS analysis와 Western blot analysis를 수행한 결과, paclitaxel이 G2/M 정지를 유도하는 것을 확인하였다. 또한, paclitaxel이 비정상적인 세포 분열유도와 핵 단편분절 유도없이 일어나는 mitotic catastrophe 즉, caspase-3 비의존적인 세포사멸을 유도하였다. Paclitaxel을 처리한 세포에서 일어나는 이러한 mitotic catastrophe에 의한 세포 죽음은 Gl/S기의 진행을 억제하는 시약인 thymidine을 처리하는 것에 의해 억제되는 것을 확인할 수 있었다. 이러한 결과를 종합해 볼 때, paclitaxel에 의한 토끼 관절 연골 세포에서의 세포죽음은 caspase-3 비의존적인 mitotic catastrophe에 의해 일어나는 것으로 사료되어진다. Paclitaxel is known as a potent inhibitor of microtubule depolymerization. It leads to mitotic arrest and cell death by stabilizing the spindle in various cell types. Here, we investigated the effects of paclitaxel on the proliferation and cell death of rabbit articular chondroeytes. Paclitaxel inhibited proliferation in a dose- and time- dependent manner, determined by MTT assay in rabbit articular chondrocytes. We also established paclitaxel-induced G2/M arrest by fluorescent activated cell sorter (FACS) analysis. Paclitaxel increased expression of cyclin B, p53 and p2l, while reducing expression of cdc2 and cdc25C in chondrocytes, as detected by Western blot analysis. Interestingly, paclitaxel showed the mitotic catastrophe that leads to abnormal nucleus division and cell death without DNA fragmentation through activation of caspase. Cell death by mitotic catastrophe in cells treated with paclitaxel was suppressed by inhibiting G1/S arrest with 2 mM thymidine. These results demonstrate that paclitaxel induces cell death via mitotic catastrophe without activation of casepase in rabbit articular chondrocytes.

흰쥐의 소장점막에 Paclitaxel(Taxol)과 방사선조사의 효과

이경자(Kyung Ja Lee) 대한방사선종양학회 1996 Radiation Oncology Journal Vol.14 No.4

목적 : Paclitaxel(Taxol)은 미소관의 집합을 촉진시키고 분해를 방지하여 세포주기 중 유사분열을 정지시킴으로써 방사선조사와 병용할 경우 방사선감작제로서의 가능성이 있다. 정상세포 중 횐쥐의 소장에서 paclitaxel이 방사선의 독성을 증가시키는지 알기 위하여 실험을 시도하였다. 대상 및 방법 : Paclitaxel군은 paclitaxel 10mg/kg을 복강내 1회 주입하였다. 방사선조사군은 8 Gy를 전복부에 단일조사하였다. Paclitaxel과 방사선병용군은 paclitaxel(10mg/kg)을 복강내 주입 후 24시간에 방사선조사군과 동일한 방법으로 조사하였다. 실험완료 후 소장점막에서 유사분열수, apoptosis와 기타 점막의 변화를 시간별로(6시간 -5일) 관찰하였다. 결과 : Paclitaxel은 소장점막의 소낭선세포에서 유사분열을 정지시키며 약간의 apoptosis을 유발하였으며 유사분열정지는 6시간에서 최대치를 보였고 24시간에 정상으로 회복되었다. 방사선조사는 apoptosis를 유발하였으며 6시간에 최대치를 보이고 24시간에 정상으로 회복되었다. Paclitaxel과 방사선병용군에서 유사분열정지는 6시간에서 3일까지 나타났으며 apoptosis는 6시간과 24시간에 약간 보였으며 3일에 정상으로 회복되었다. 병용군에서 apoptosis의 빈도는 정상군보다 높았으나 방사선단독군에 비하여 현저하게 감소되어 방사선보호작용이 있었다. 결론 : Paclitaxel에 의한 유사분열정지는 6시간에 최대치를 보이고 24시간에 정상으로 회복 되었으며 약간의 apoptosis을 유발하였다. 방사선조사는 apoptosis를 유발하였으며 6시간에 최대치를 보이고 24시간에 정상으로 회복되었다. 정상 소장에 방사선조사를 paclitaxel주입 후 24 시간에 시행하여 apoptosis가 방사선조사군에 비하여 현저히 감소되어 paclitaxel이 방사선보호작용이 있었다 Purpose : Paclitaxel is a chemotherapeutic agent with potent microtubule stabilizing activity that arrests cell cycle in G2-M Because G2-M is the most radiosensitive Phase of the cell cycle, paclitaxel has potential as a cell cycle- specific radiosensitizer. This study was designed to investigate the ability of paclitaxel to increase the radiotoxicity in normal small bowel mucosa of the rat. Materials and Methods : A sigle intraperitoneal infusion of paclitaxel (10mg/kg), and a single irradiation(8 Gy, x-ray) to the whole abdomen and combination of radiation(8 Gr, x-ray) 24 hours after paclitaxel infusion in the rats were done. The changes of jejunal mucosa, and kinetics of mitotic arrest and apoptosis in the jejunal crypt were defined at 6 hours - 5 days after each treatment histologically. Results : Paclitaxel blocked jejunal crypt cell in mitosis and induced minmal apoptosis. Mitotic arrest by paclitaxel was peaked at 6 hours after infusion and returned to normal by 24 hours. Radiation induced apoptosis and peaked at 6 hours and returned to normal by 24 hours. Combination of paclitaxel and radiation blocked crypt cell in mitosis at 3 days and induced apoptosis slightly at 6 hours and 24 hours and returned to normal by 3 days. The incidence of apoptosis in combined group at 6 hours was slightly higher than normal control but significantly lower than radiation alone group. The major changes of jejunal mucosa were nuclear vesicle and atypia which were appeared at 6 hours - 3 days and returned to normal by 5 days The degree of the mucosal changes are not different in 3 groups except for absence of inflmmatory reaction in radiation group. Conclusion : Mitotic arrest by paclitaxel was peaked at 6 hours and returned to normal by 24 hours and paclitaxel induced minimal apoptosis. Radiation induced apoptosis, peaked at 6 hours and returned to normal by 24 hours. Radiation-induced apoptosis was less in combined group which suggested that paclitaxel have a radioprotective effect when radiation was given 24 hours after paclitaxel infusion.

초임계유체를 이용한 파클리탁셀고체분산체의 제조 및 평가

우종수,박재현,지상철 한국약제학회 2008 Journal of Pharmaceutical Investigation Vol.38 No.4

Paclitaxel is a taxane diterpene amide, which was first extracted from the stem bark of the western yew, Taxus brevifolia. This natural product has proven to be useful in the treatment of a variety of human neoplastic disorders, including ovarian cancer, breast and lung cancer. Paclitaxel is a highly hydrophobic drug that is poorly soluble in water. It is mainly given by intravenous administration. Therefore, The pharmaceutical formulation of paclitaxel (TaxolR; Bristol-Myers Squibb) contains 50% CremophorR EL and 50% dehydrated ethanol. However the ethanol/Cremophor EL vehicle required to solubilize paclitaxel in TaxolR has a pharmacological and pharmaceutical problems. To overcome these problems, new formulations for paclitaxel that do not require solubilization by CremophorR EL are currently being developed. Therefore this study utilized a supercritical fluid antisolvent (SAS) process for cremophor-free formulation. To select hydrophilic polymers that require solubilization for paclitaxel, we evaluated polymers and the ratio of paclitaxel/polymers. HP-β-CD was used as a hydrophilic polymer in the preparation of the paclitaxel solid dispersion. Although solubility of paclitaxel by polymers was increased, physical stability of solution after paclitaxel/polymer powder soluble in saline was unstable. To overcome this problem, we investigated the use of surfactants. At 1/20/40 of paclitaxel/hydrophilic polymer/surfactant weight ratio, about 10 mg/mL of paclitaxel can be solubilized in this system. Compared with the solubility of paclitaxel in water (1 μg/mL), the paclitaxel solid dispersion prepared by SAS process increased the solubility of paclitaxel by near 10,000 folds. The physicochemical properties was also evaluated. The particle size distribution, melting point and amophorization and shape of the powder particles were fully characterized by particle size distribution analyzer, DSC, SEM and XRD. In summary, through the SAS process, uniform nano-scale paclitaxel solid dispersion powders were obtained with excellent results compared with TaxolR for the physicochemical properties, solubility and pharmacokinetic behavior.

흰쥐의 간에서 Paclitaxel(Taxol)이 방사선에 미치는 효과

이경자,구혜수 梨花女子大學校 醫科大學 醫科學硏究所 1997 EMJ (Ewha medical journal) Vol.20 No.1

연구목적: Paclitaxel은 소관종합체의 형성을 안정화시키는 미소관 dr제제로서 이 작용이 주로 세포주기의 G2-M시기에 일어나기 때문에 방선 감작제의 가능성이 있는 항암제이다. 흰쥐의 간에서 paclitaxel과 방선조사를 병용하여 paclitaxel이 방사선의 효과에 미치는 여향을 파악하기 위한 실험연구이다. 방 법: 흰쥐 52마리를 대상으로 정상대조군, paclitaxel군, 방사선조사군과 paclitaxel과 방사선병용군으로 분류하였다. Paclitaxel군은 paclitaxel(10mg/kg)을 복강내에 1회 주입하였으며, 방사선조사군은 전복부에 8Gy x-ray를 단일 조사하였으며, 병용군은 paclitaxel(10mg/kg)을 주입 후 24시간에 방사선조사(8Gy)를 시행하였다. 실험완료 후 6시간, 24시간, 3일 및 5일에 간의 병리 조직학적 변화를 관찰하였다. 결 과: Paclitaxel군은 유사분열수가 6시간에 증가되어 24시간에 정상으로 회복되었다. 방사선조사군은 조사 후 6시간과 24시간에 apoptosis가 발현하였으며 3일 후 정상으로 회복되었다. 간세포의 괴사는 paclitaxel군, 방사선조사군 및 병용군에서 모두 나타났으며 괴사의 정도는 3군에서 차이가 없었다. 결 론: Paclitaxel주입 후 6시간에 유사분열중지가 유발되었으며 24시간에 정상으로 회복되었고 aoptosis는 발현되지 않았다. 방선조사 후 6시간과 24시간에 apoptosis가 유발되었으며 3일에 정상으로 회복되었다. 간세포의 괴사는 paclitaxel군, 방사선조사군 및 병용군에서 모두 나타났으며 3군에서 정도의 차이ㅏ 없었다. 따라서 paclitaxel을 방사선조사 24시간 전에 주입하여 간세포에서 방사선감작제의 효과는 없었다. Objectives : Paclitaxel(Taxol) si a chemotherapeutic agent with potent microtubule stabilizing activities that arrests cell cycle in G2-M. Since D2-m is the most radiosensitive phase of the cell cycle, paclitaxel has potential as a cell cycle-specific radiosensitizer. This study was designed to investigate the effects of paclitaxel to radiotoxicity in normal rat liver. Materials & Methods : A single intraperitoneal infusion of paclitaxel(10mg/kg), and a single irradiation(8Gy, x-ray) to the whole abdomen, and combination of irradiation(8Gy,x-ray)24 hours after paclitaxel infusion were done in Sprague-Dawley rats. The incidence of mitosis, apoptosis and parenchymal changes of the liver were evaluated at 6 hours, 24 hours, 3 and 5 days, respectively. Results : Paclitaxel and irradiation significantly increased mitosis at 6 hours and apoptosis was increased by irradiation at 6 and 24 hours. Increased numbers of apoptosis at 3 days by paclitaxel alone was not significantly different from control. Combination of paclitaxel and irradiation showed significantly increased numbers of mitosis and apoptosis at 6 hours. The degree of necrosis of hepatocyte was not significantly different between 3 groups. Conclusion : Since the incidence of mitosis, apoptosis and hepatocyte necrosis were not increased by paclitaxel infusion 24 hours before irradiation, paclitaxel did not show radiosensitizing effect in this experimental condition. Studies with conditions similar to clinical situation will be the next stop to define the radiosensitizing effects of paclitaxel.

Rapamycin/Paclitaxel이 사람과 흰쥐 평활근세포에서 Heme oxygenase-1의 발현에 미치는 영향

박철보,정헌택,류현열 고신대학교(의대) 고신대학교 의과대학 학술지 2005 고신대학교 의과대학 학술지 Vol.20 No.1

Background : Both rapamycin and paclitaxel are currently being used in drug-eluting stent to block the proliferation of vascular endothelial and smooth muscle cells. Heme oxygenase-1 (HO-1) has also known to be anti-proliferative against vascular cells. In this study, it focused on the role of HO-1 in cytoprotection and antiproliferation of paclitaxel. Methods : Proliferation of cells and metabolic activity of HO-1 were evaluated with the tetrazolium-based assay. HO-1 protein expression was shown by western blot analysis of vascular smooth muscle cells (VSMCs), or endothelial cells (ECs), using anti-HO-1 polyclonal antibody from untreated cells and cells exposed to rapamycin and paclitaxel. To determine whether the inhibition of paclitaxel on platelet-derived growth factor (PDGF)-dependent proliferation may be mediated by its induction of HO-1 we exposed cells to paclitaxel, PDGF and ZnPP, an inhibitor of HO activity. Survival rate was checked to find the anti-apoptotic effect of paclitaxel against tumor necrosis factor (TNF)-a-mediated apoptosis, in the presence or abscence of ZnPP. Result : Rapamycin induces HO-1 protein in rat VSMCs and human ECs. Exposure of VSMCs to paclitaxel for 12h resulted in a dose-dependent increase in HO-1 ativity. Ten nM of paclitaxel led to a maimal expression of HO-1 protein at 12h after exposure. The addition of ZnPP abrogated the suppression effect of paclitaxel on PDGF-stimulated cell proliferation, and blocked paclitaxel-mediated suppression of TNF-a-mediated apoptosis. Hemoglobin, a scavenger of CO, abrogated the paclitaxel induced cytoprotection. Conclusion : Paclitaxel induced the expression of HO-1 gene in rat VSMCs and human ECs, in dose-dependent and time-dependent manner, like rapamycin.

흰쥐의 위점막에 Paclitaxel (Taxol)과 방사선조사의 효과

이경자(Kyung-Ja Lee),구혜수(Heasoo Koo) 대한방사선종양학회 1999 Radiation Oncology Journal Vol.17 No.4

목 적 : Paclitaxel (Taxol)은 미소관의 집합을 촉진시키고 분해를 방지하는 미소관 억제제로서 이 작용은 세포주기 중 방사선에 예민한 G2/M 시기에 일어나기 때문에 방사선조사와 병용할 경우 방사선감작제 가능성이 있다. Paclitaxel과 방사선조사를 병용하여 흰쥐의 위점막에서 paclitaxel이 방사선의 효과에 미치는 영향을 파악하기 위하여 본 연구를 시행하였다. 대상 및 방법 : 실험군은 paclitaxel 단독군, 방사선조사 단독군 및 paclitaxel과 방사선조사 병용군의 세 군으로 분류하여 paclitaxel 단독군은 paclitaxel (10 mg/kg)을 복강내 1회 주입하였고, 방사조사 단독군은 8 Gy를 전복부에 단일조사하였으며, paclitaxel과 방사선조사 병용군은 paclitaxel (10 mg/kg)을 복강내 주입 후 24시간에 방사선조사 단독군과 동일한 방법으로 방사선조사하였다. 실험 완료 후 위점막 내 유사분열수, apoptosis 와 기타 점막의 변화를 시간별로(6, 24시간, 3일 및 5일) 비교관찰하였다. 결 과 : Paclitaxel 주입 후 위점막 내 유사분열은 증가하지 않았고 apoptosis는 6시간에 5.75%였고 3일까지 비슷하게 지속되었다. Paclitaxel 주입 후 경미한 위선의 확장이 24시간에 관찰되었고, 세포의 비정형이 24시간과 3일에 보였으나 5일에는 정상으로 회복되었다. 방사선조사 단독군에서 apoptosis 는 6시간에 6.0%로 가장 많았으며 24시간에 1.25%로 감소되었고 5일까지 지속되었다. 위선의 확장과 세포의 비정형은 방사선조사 후 6시간부터 5일까지 계속해서 경미하게 보였다. Paclitaxel과 방사선조사 병용군은 apoptosis가 6, 24시간, 3일 및 5일에 각각 5.5, 4.5, 4.0, 4.0%로 방사선조사 단독군에 비하여 24시간부터 많아졌고 3일에는 통계학적으로 유의한 차이가 있었다. 위선의 확장과 세포의 비정형은 palcitaxel과 방사선조사 병용군에서 방사선조사 단독군에 비하여 증가되지 않았다. 결 론 : 흰쥐의 위점막은 paclitaxel 주입 후 24시간에 방사선조사를 시행한 결과 apoptosis를 종말점으로 볼 때 paclitaxel의 첨가효과만을 보여 주었다. Purpose : Paclitaxel is a chemotherapeutic agent with potent microtubule stabilizing activity that arrests cells in G2- M phase. Because G2 and M are the most radiosensitive phase of the cel l cycle, paclitaxel has potential role as a cell-cycle specific radiosensitizer. This study was performed to see the effects of paclitaxel on the radiation-induced damage of gastric mucosa of the rat. Materials and Methods : The rats were divided into the three groups i.e., paclitaxel alone group, radiation alone group and, a combination of paclitaxel and radiation in combined group. A single intraperitoneal infusion of paclitaxel (10 mg/kg) was done in paclitaxel alone group. In radiation alone group, a si ngle fraction of irradiation (8 Gy, x-ray) to the whole abdomen and, a combination of a single fraction of irradiation (8 Gy, x -ray) to the whole abdomen was given 24 hrs after paclitaxel infusion in combined group of paclitaxel and radiation. The inciden ce of mitosis and apoptosis as well as histologic changes of the gastric mucosa were evaluated at 6 hrs, 24 hrs, 3 days and 5 days after treatment. Results : The number of the mitosis was not increased by paclitaxel infusion. The incidence of the apoptosis was similar from 6 hrs to 3 days after paclitaxel infusion and was decreased at 5 days. Paclitaxel induced minimal glandular dilatation and cellular atypia of gastric mucosa at 24 hrs and 3 days. In irradiation group, the incidence of apoptosis was 6.0% in 6 hrs and 1.25% in 24 hrs after irradiation and minimal glandular dilatation and cellular atypia were noted throughout the experimental period. The incidence of apoptosis in the combined group of paclitaxel and irradiation (4.5%) was significantly higher than irradiation alone group (1.25%) at 3 days ( p<0.05). Conclusion : Paclitaxel had no effect on mitotic arrest in gastric mucosa of the rat. Increased number of apoptosis in combined paclitaxel and irradiation group suggested the additive effects of paclitaxel on irradiation.

α-Tocopheryl succinate potentiates the paclitaxel-induced apoptosis through enforced caspase 8 activation in human H460 lung cancer cells

Soo-Jeong Lim,Moon Kyung Choi,Min Jung Kim,Joo Kyoung Kim 생화학분자생물학회 2009 Experimental and molecular medicine Vol.41 No.10

Paclitaxel is one of the chemotheraputic drugs widely used for the treatment of nonsmall cell lung cancer (NSCLC) patients. Here, we tested the ability of α-tocopheryl succinate (TOS), another promising anticancer agent, to enhance the paclitaxel response in NSCLC cells. We found that sub-apoptotic doses of TOS greatly enhanced paclitaxel-induced growth suppression and apoptosis in the human H460 NSCLC cell lines. Our data revealed that this was accounted for primarily by an augmented cleavage of poly(ADP-ribose) polymerase (PARP) and enhanced activation of caspase- 8. Pretreatment with z-VAD-FMK (a pan-caspase inhibitor) or z-IETD-FMK (a caspase-8 inhibitor) blocked TOS/paclitaxel cotreatment-induced PARP cleavage and apoptosis, suggesting that TOS potentiates the paclitaxel-induced apoptosis through enforced caspase 8 activation in H460 cells. Furthermore, the growth suppression effect of TOS/paclitaxel combination on human H460, A549 and H358 NSCLC cell lines were synergistic. Our observations indicate that combination of paclitaxel and TOS may offer a novel therapeutic strategy for improving paclitaxel drug efficacy in NSCLC patient therapy as well as for potentially lowering the toxic side effects of paclitaxel through reduced drug dosage. Paclitaxel is one of the chemotheraputic drugs widely used for the treatment of nonsmall cell lung cancer (NSCLC) patients. Here, we tested the ability of α-tocopheryl succinate (TOS), another promising anticancer agent, to enhance the paclitaxel response in NSCLC cells. We found that sub-apoptotic doses of TOS greatly enhanced paclitaxel-induced growth suppression and apoptosis in the human H460 NSCLC cell lines. Our data revealed that this was accounted for primarily by an augmented cleavage of poly(ADP-ribose) polymerase (PARP) and enhanced activation of caspase- 8. Pretreatment with z-VAD-FMK (a pan-caspase inhibitor) or z-IETD-FMK (a caspase-8 inhibitor) blocked TOS/paclitaxel cotreatment-induced PARP cleavage and apoptosis, suggesting that TOS potentiates the paclitaxel-induced apoptosis through enforced caspase 8 activation in H460 cells. Furthermore, the growth suppression effect of TOS/paclitaxel combination on human H460, A549 and H358 NSCLC cell lines were synergistic. Our observations indicate that combination of paclitaxel and TOS may offer a novel therapeutic strategy for improving paclitaxel drug efficacy in NSCLC patient therapy as well as for potentially lowering the toxic side effects of paclitaxel through reduced drug dosage.

Relation of Poly(ADP-ribose) Polymerase Cleavage and Apoptosis Induced by Paclitaxel in HeLa S₃ Uterine Cancer Cells

장정현,김광연,안순철,권헌영,Chang, Jeong-Hyun,Kim, Kwang-Youn,Ahn, Soon-Cheol,Kwon, Heun-Young Korean Society of Life Science 2007 생명과학회지 Vol.17 No.8

Paclitaxel이 암세포에서 세포예정사를 유발할지라도, 아직 정확한 기전은 잘 알려져 있지 않다. 이에 본 연구에서는 HeLa $S_{3}$ 자궁암세포에서의 paclitaxel이 어떠한 영향을 미치는지 알아보고자 한다. 그리하여 방법으로는 세포독성검사, apoptotic cells의 형태학적 변화(DAPI 염색 ), western blot 분석법을 사용하여 수행하였다. 본 연구의 결과로 paclitaxel은 HeLa $S_{3}$ 세포에서 세포독성을 보이며 특히 paclitaxel의 $IC_{50}$ 값은 약 1 ${\mu}M$이며, paclitaxel 처리한 HeLa $S_{3}$ 세포에서 형태학적 변화(분절화)를 관찰하였고, flow cytometric 분석에서는 G2/M기가 차단되어 paclitaxel은 세포주기 특히 Sub-$G_{1}$기를 조절함을 알 수 있다. 그리고 Paclitaxel을 처리한 HeLa $S_{3}$ 세포에서는 PARP cleavage를 유발하였고 Bc1-2의 감소와도 관련되었다. Although paclitaxel induces apoptosis of cancer cells, its exact mechanism of action is not yet known. The present study has been performed to determine whether influence of paclitaxel in HeLa $S_{3}$ uterine cancer cells. Three assays were employed in this study: cell cytotoxicity, morphological assessments of apoptotic cells (DAPI staining assay), and western blot analysis. The results indicated that paclitaxel has cytotoxic effects in HeLa $S_{3}$ cells. Especially, the $IC_{50}$ value of paclitaxel was about 1 ${\mu}M$. And morphological changes (fragmentation) of cells were observed by paclitaxel in HeLa $S_{3}$ cells. The flow cytometric analysis of paclitaxel-treated cells indicated a block of G2/M phase. The results that pacli-taxel regulates the cell cycle, especially Sub-$G_{1}$ phase. Paclitaxel induces apoptosis of HeLa $S_{3}$ cells via PARP-dependent fashion, and this apoptosis is related to disappearance of Bcl-2 proteins.

Effect of Ethyl Pyruvate on Paclitaxel-Induced Neuropathic Pain in Rats

최성수,고원욱,남재식,신진우,임정길,서정훈 대한통증학회 2013 The Korean Journal of Pain Vol.26 No.2

Background:Although paclitaxel is a widely used chemotherapeutic agent for the treatment of solid cancers, side effects such as neuropathic pain lead to poor compliance and discontinuation of the therapy. Ethyl pyruvate (EP) is known to have analgesic effects in several pain models and may inhibit apoptosis. The present study was designed to investigate the analgesic effects of EP on mechanical allodynia and apoptosis in dorsal root ganglion (DRG) cells after paclitaxel administration. Methods:Rats were randomly divided into 3 groups: 1) a control group, which received only vehicle; 2) a paclitaxel group, which received paclitaxel; and 3) an EP group, which received EP after paclitaxel administration. Mechanical allodynia was tested before and at 7 and 14 days after final paclitaxel administration. Fourteen days after paclitaxel treatment, DRG apoptosis was determined by activated caspase-3 immunoreactivity (IR). Results:Post-treatment with EP did not significantly affect paclitaxel-induced allodynia, although it tended to slightly reduce sensitivities to mechanical stimuli after paclitaxel administration. After paclitaxel administration, an increase in caspase-3 IR in DRG cells was observed, which was co-localized with NF200-positive myelinated neurons. Post-treatment with EP decreased the paclitaxel-induced caspase-3 IR. Paclitaxel administration or post-treatment with EP did not alter the glial fibrillary acidic protein IRs in DRG cells. Conclusions:Inhibition of apoptosis in DRG neurons by EP may not be critical in paclitaxel-induced mechanical allodynia.