5-Aminolevulinic acid의 국소적 투여에 의한 자궁경부암의 광역학치료에 대한 연구

안진철 ( Jin Chul Ahn ),조현철 ( Hyun Chul Cho ),박진완 ( Jin Wan Park ),박충학 ( Choong Hak Park ),윤문희 ( Moon Hee Youn ) 대한산부인과학회 2010 Obstetrics & Gynecology Science Vol.53 No.12

목적: 자궁경부암에서 5-aminolevulinic acid (5-ALA)의 국소적 투여에 의한 광역학치료의 항암치료효과를 알아보고자 하였다. 연구 방법: Nude mouse에 인체의 자궁경부암세포인 hella cell을 이종이식 후에 일정크기의 종양이 형성되면 20% 5-ALA cream을 만들어 종양에 국소도포하고 밀봉하여 3시간마다 532 nm diode laser를 종양에 조사하여 발광되는 정도를 관찰하여 5-ALA의 생체 내 분포와 최대농도로 종양에 분포되는 시간을 알아보았다. 이렇게 알아낸 시간을 치료를 위한 최적의 투여시 간으로 간주하고 같은 방법으로 이번에는 632 nm diode laser를 이용하여 480 J/cm2의 에너지로 광역학치료를 시행하고 30일간 3일마다 그 크기를 측정하여 치료효과를 판별하였다. 이종이식된 25마리의 nude mouse를 대조군, 5-ALA만 투여한 군, laser만 조사한 군, 광역학치료 군으로 나누어 서로 비교분석하여 종양의 치료효과를 알아보았다. 결과: 5-ALA 국소투여 후 9시간 후에 가장 많이 발광되었다. 광역학치료 군에서 40% (4/10)는 종양의 크기가 감소하였지만, 이를 제외한 나머지 실험군에서는 지속적인 종양 크기 증가가 관찰되었다. 결론: 본 연구에서는 5-ALA의 국소투여 후 9시간이 치료에 가장 적절한 시간임을 알 수 있었다. 5-ALA 국소투여에 의한 광역 학치료는 대조군에 비해 유의한 종양크기의 감소가 없었으나, 반응군(4/10)의 경우는 대조군에 비해 유의한 종양의 크기 감소가 관찰되었다(P<0.05). Objective: The purpose of this study was to investigate the treatment effects of a topical application of 5-aminolevulinic acid (5-ALA) for photodynamic therapy (PDT) to treat cervical cancer. Methods: We first investigated the effects of 5-ALA cream according to application time. And to find the effective 5-ALA concentration and the distribution times in vivo, 20% 5-ALA cream was topically applied to the tumor of the nude mouse. We then observed the distribution of 5-ALA via fluorescence measurement with using a 532 nm diode laser. 25 nude mice were divided into Control, ALA, Laser, and PDT group. To evaluate the PDT effect at cancer lesion, we applied 20% 5-ALA cream to the tumor by the same method, and the PDT was done by using a 632 nm diode laser at the time of the peak level of fluorescence. We checked the changes of the volume of cancer for 30 days, and then biopsy was done. Results: The effective post-irradiation time after topical ALA application was 9 hours. In the PDT group, 40% (4/10) of the mice showed decreased tumor size. Conclusion: The maximum PpIX fluorescence at 9 hours after local applicationof 5-ALA cream was checked. And PDT group did not show any statistical difference than control group in the growth of tumor size than control group. However responding cases (4/10) of PDT group showed the meaningful decrease of tumor size than control group (P<0.05).

9-hydroxypheophorbide-a와 660 ㎚ 다이오드 레이저를 이용한 광역학치료의 항암효과와 치료기전에 대한 연구

안진철(Jin-Chul Ahn) 한국생명과학회 2009 생명과학회지 Vol.19 No.6

녹조류인 Spirulina platensis에서 추출하여 만든 새로운 광감작제와 660 ㎚의 다이오드 레이저를 이용한 광역학치료의 항암효과와 치료기전을 알아보았다. 세포 독성능은 MTT assay를 이용하였고, 세포사멸기전은 propidium iodide과 Hoechst 33342 염색법과 투과전자현미경으로 확인하였다. 또한 암세포가 이종 이식된 누드마우스 모델에서 광역학치료를 시행하여 항암효과를 확인하였다. 3종류의 클로로필 유도체중 9-hydroxypheophorbide-a (9-HpbD-a)의 세포 독성능이 가장 우수하였고, 9-HpbD-a의 적정 레이저조사 시간은 30분(3.2 J/㎠), 광감작제를 투여하고 레이저조사시간까지의 배양시간은 최소 6시간 이상임을 확인하였다. 광역학치료의 세포사멸기전은 낮은 9-HpbD-a 농도에서 세포고사가 주된 세포사멸기전이었고, 높은 농도의 9-HpbD-a에서는 세포괴사에 의한 세포사멸이 주된 기전임을 확인하였다. 투과전자현미경 하에서도 같은 양상을 관찰하였다. 그리고 암세포가 이종 이식된 누드마우스 모델에서의 광역학치료는 제1군 정상대조군과 제2군 9-HpbD-a만을 투여한 종양조직모두 지속적인 종양의 성장(100%)을 보였고, 제3군인 레이저만을 종양조직에 조사한 실험군에서는 3 마리는 치료가 되지 않았고(75.0%), 1 마리는 재발(25.0%)하였다. 제4군 광역학치료군에서 총16 마리의 종양에서 10 마리는 완치(62.5%), 4 마리는 재발(25.0%), 2 마리는 치유되지 않았음(12.5%)을 확인하였다. 9-HpbD-a와 660 ㎚ 다이오드 레이저를 이용한 광역학치료는 유의한 항암효과를 나타내었고 9-HpbD-a를 이용한 광역학치료는 새로운 치료방법으로서 향후 암치료의 유용한 치료방법으로 기대된다. A new photosensitizer, 9-Hydroxypheophorbide-a (9-HpbD-a), was derived from Spirulina platensis. We conducted a series of experiments, in vitro and in vivo, to evaluate the anticancer effect and mechanism of photodynamic therapy using 9-HpbD-a and 660 ㎚ diode lasers on a squamous carcinoma cell line. We studied the cytotoxic effects of pheophytin-a, 9-HpbD-a, 9-HpbD-a red and 660 ㎚ diode lasers in a human head and neck cancer cell line (SNU-1041). Cell growth inhibition was determined by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay. The effects of 9-HpbD was higher than those of 9-HpbD-a red or pheophytin-a in PDT. We then tested the cytotoxic effects of 9-hydroxypheophorbide-a (9-HpbD-a) in vitro. The cultured SNU-1041 cells were treated with serial concentrations of 9-HpbD-a followed by various energy doses (0, 0.1, 0.5, 3.2 J/㎠) and by various interval times (0, 3, 6, 9, 12 hr) until laser irradiation, then MTT assay was applied to measure the relative inhibitory effects of photodynamic therapy (PDT). Optimal laser irradiation time was 30 minutes and the cytotoxic effects according to incubation time after 9-HpbD-a treatment increased until 6 hours, after which it then showed no increase. To observe the cell death mechanism after PDT, SUN-1041 cells were stained by Hoechst 33342 and propidium iodide after PDT, and observed under transmission electron microscopy (TEM). The principal mechanism of PDT at a low dose of 9-HpbD-a was apoptosis, and at a high dose of 9-HpbD-a it was necrosis. PDT effects were also observed in a xenografted nude mouse model. Group Ⅰ (no 9-HpbD-a, no laser irradiation) and Group Ⅱ (9-HpbD-a injection only) showed no response (4/4, 100%), and Group Ⅲ (laser irradiation only) showed recurrence (1/4, 25%) or no response (3/4, 75%). Group Ⅳ (9-HpbD-a + laser irradiation) showed complete response (10/16, 62.5%), recurrence (4/16, 25%) or no response (2/16, 12.5%). Group Ⅳ showed a significant remission rate compared to other groups (p<0.05). These results suggest that 9-HpbD-a is a promising photosensitizer for the future and that further studies on biodistribution, toxicity and mechanism of action would be needed to use 9-HpbD-a as a photosensitizer in the clinical setting.

한국 주식가격변동의 안정 파레티안 분포가설의 검증

성삼경(Sam - Kyung Sung),안진철(Jin - Chul Ahn) 한국경영학회 1986 經營學硏究 Vol.15 No.2

Sphingomonas sp. HS362에 의한 Phenanthrene 분해특성

김수화,홍승복,강희정,안진철,정재훈,손승렬,Kim Su Hwa,Hong Seung-Bok,Kang Hee Jeong,Ahn Jin-Chul,Jeong Jae Hoon,Son Seung-Yeol 한국미생물학회 2005 미생물학회지 Vol.41 No.3

유류에 의해 오염된 토양으로부터 난분해성 물질인 phenanthrene을 유일한 탄소원과 에너지원으로 이용하며 성장하는 균주들을 분리한 후, 그중에서 분해능이 가장 우수한 균주를 선별하여 HS362라고 명명하였다. HS362는 생화학적 검사로는 Sphingomonas paucimobilis와, 16S rDNA 염기서열 분석으로는 Sphingomonas CF06과 가장 유사한 것으로 나타났고, 지방산분석 결과도 그람음성 간균인 Sphingomonas 속으로 판명되었으므로 Sphingomonas sp. HS362라고 명명하였다. 이 균은 500 ppm의 phenanthrene을 단일 탄소원으로 첨가한 경우, 10일 만에 $98{\%}$ 이상을 분해하였고,3000 ppm의 phenanthrene이 첨가된 경우에도 10일 만에 약$30{\%}$ 이상을 분해하는우수한 균임이 확인되었다. 또한 이 균은PAH들(Polycyclic aromatic hydrocarbons) 중에서 phenanthrene 이외에도 분자량이 적은 indole, naphthalene은 분해할 수 있는 반면에, 분자량이 큰 pyrene, fluoranthene은 분해하지 못하였다. Spitingomonas sp. HS362에 의한 phenanthrene 분해는$30^{\circ}C$, pH $4{\~}8$, NaCl $1{\%}$ 이하의 농도인 조건하에서 배양했을 때 가장 우수했으며, 특히 전분과 SDS, Tween 85, Triton X-100와 같은 계면활성제를 첨가해 주었을 때 분해가 증진되었다. 또한, 전배양을 통해서 phenanthrene의 분해가 증진되는 것을 볼 때에 분해효소가 유도되는 것으로 추측할 수 있었다. Spltingomonas sp. HS362는 5개의 plasmid를 가지고 있는데, 그중에서 plasmid p4를 잃었을 때에는phenanthrene을 분해하지 못하는 것으로 보아plasmid p4가 phenanthrene분해와 밀접한 관련이 있는 것으로 보인다. A phenanthrene-degrading bacterium HS362, which is capable of using phenanthrene as a sole carbon and energy source, was isolated from oil contaminated soil. This strain is a gram negative, rod shaped organism that is most closely related to Sphingomonas paucimobilis based on biochemical tests, and belongs to the genus Sphingomonas based on fatty acids analysis. It exhibited more than $99.2{\%}$ nucleotide sequence similarity of 16S rDNA to that of Sphingomonas CF06. Thus, we named this strain as Sphingomonas sp. HS362. It degraded $98{\%}$ of phenanthrene after 10 days of incubation when phenanthrene was added at 500 ppm and $30{\%}$ even when phenanthrene was added at 3000 ppm. Sphingomonas sp. HS362 could also degrade low molecular weight PAHs(Polycyclic aromatic hydrocarbons) such as indole and naphthalene, but was unable to degrade high molecular weight PAHs such as pyrene and fluoranthene. The optimum temperature and pH for phenanthrene degradation were $30^{\circ}C$ and $4{\~}8$, respectively. Sphingomonas sp. HS362 could degrade phenanthrene effectively in the concentration range of NaCl of up to $1{\%}$. Its phenanhrene degrading ability was enhanced by preculture, suggesting the possibility of induction of phenanthrene degrading enzymes. Starch and surfactants such as SDS, Tween 85, and Triton X-100 were also able to enhance phenanthrene degradation by Sphingomonas sp. HS362. It carries five plasmids and one of them, plasmid p4, is considered to be involved in the degradation of phenanthrene according to the plasmid curing experiment by growing at $42^{\circ}C$.

레이저수술기 성능평가 연구

Jaeyoung Lee(이재영),Sanggeon Park(박상건),Jin-Chul Ahn(안진철) 대한전기학회 2020 대한전기학회 워크샵 Vol.2020 No.7

ALA 및 ALA-Methylester의 투여에 따른 종양조직 및 정상조직에의 분포양상에 대한 연구

정필상(Phil-Sang),정상운(Sang Oun Jung),안진철(Jin Chul Ahn),이상준(Sang Joon Lee),윤준식(Jun Sik Yoon) 대한두경부종양학회 2003 대한두경부 종양학회지 Vol.19 No.1

Objectives: In this report, we confirmed the distributed pattern of ALA and ALA-methylester in normal and tumor-bearing region. Materials and Methods: ALA and ALA-methylester were administered to nude mouse by intratumoral, subcutaneous and intravenous injection. After injection, the fluorescence in normal and tumor region was measured by LESA (laser electronic spectrum analyzer). Results: The tumor-specificity of ALA and ALA-methylester was shown in the case of intratumoral injection. In all case, the fluorescence caused by ALA and ALA-methylester was maximally increased in 2 hours after injection. Then while the fluorescence level was rapidly decreased to control level in normal region, it was still remained in tumor region. Conclusion: According to this result, The intratumoral injection was more efficient administration method for PDT/PDD than subcutaneous and intravenous injection.

Effects of Propolis on 808 ㎚ Laser Induced Wounds

Jang-In Shin,Chung-Ku Rhee,Seung-Wan Lee,Jin-Chul Ahn(안진철) 대한의생명과학회 2009 Biomedical Science Letters Vol.15 No.3

Wound healing is a complex and highly organized biological response to injury that results in the loss of tissue integrity. Our particular interest was propolis, traditional used as an antimicrobial and an anti-inflammatory agent. The purpose of this study was to see the effects of propolis on healing of the laser induced wounds and the level of collagen formation. 808 ㎚ laser (20 J) was irradiated on the back of rats. Irradiated wounds were divided into gel control and 6 experimental groups (3 and 5% of different three kinds of propolis). Gel type of propolis (supplied by Seoul Propolis Co., Daejeon, Korea) in water based were applied on the laser wound once daily for 14 days. One fourth of rats were sacrificed on 3<SUP>rd</SUP>, 7<SUP>th</SUP>, 10<SUP>th</SUP>, and 14<SUP>th</SUP> day and each wound was evaluated for degree of wound healing and the level of collagen formation. Healing of wound was evaluated by measuring and comparing the width and depth of the wounds. The levels of collagen were increased in 3 and 5% gel groups comparing to control on 3<SUP>rd</SUP> day and the collagen level were increased more on 7<SUP>th</SUP> day. Wounds of the experimental groups showed better healing in width comparing to the healing of the control. The results of this study demonstrated that propolis accelerated healing of wounds induced by laser irradiation and good collagen formation.

AMC-HN3 인체 두경부 암세포에서 genistein과 photofrin PDT의 병행처리에 의한 세포 독성능의 증의

강정욱(Jung-Wook Kang),정필상(Phil-Sang Chung),신장인(Jang-In Shin),손승렬(Seung-Yeol Son),안진철(Jin-Chul Ahn) 한국생명과학회 2008 생명과학회지 Vol.18 No.9

본 연구에서는 두경부 암세포 주에서 genistein의 증식억제 효과를 알아보고, genistein을 광역학 치료와 병행 치료시 나타나는 세포 독성능 및 세포고사 유발에 대해 연구 하였다. 두경부 암세포 주에서 genistein의 세포독성능을 확인한 결과, 농도 및 시간 의존적으로 세포 독성능이 나타났다. 이에 genistein을 photofrin을 이용한 광역학 치료와 병행 치료 시 더 많은 세포 독성능이 나타났다. Genistein과 광역학 치료의 병행 치료 시 나타나는 형태적 변화는 광역학 치료시에는 주로 세포괴사의 형태가 관찰 되었으나, genistein과 병행 치료 시에는 세포고사가 관찰 되었다. 시간별로 관찰하였을 때, 광역학 치료를 IC??으로 처리하였을 때 3시간 때에 apoptotic body가 약 3배정도 증가하였다. 세포 내 ROS를 확인한 결과 genistein과 광역학 치료를 단독으로 처리하였을 때 세포 내 존재하는 ROS가 각각 증가 하였으며, 병행 치료하였을 때는 더 많은 ROS가 형성되는 것으로 나타났다. 즉, genistein이 세포 내 ROS를 유발하여 mitochondria가 손상되고, 또한 photofrin에 의한 광역학 치료도 mitochondria를 손상시켜 세포고사를 유도하는 것으로 추측되어 진다. Photodynamic therapy (PDT) is a treatment utilizing the generation of singlet oxygen and other reactive oxygen species (ROS), which selectively accumulated in target cells. Genistein, soy-derived phytoestrogen, is one of the anticancer agents found in soybean. In the current study, we investigated the effect of photofrin-induced PDT and genistein on apoptotic cell death in head and neck cell line (AMC-HN3) to confirm the photodynamic therapy of genistein. It was determined by MTT assay that the combination group had more cytotoxicity effect than PDT group alone. Combination of photofrin PDT and genistein induced apoptosis more when comparing with PDT alone. Our data also showed that ROS was increased in combination therapy, indicating apoptosis by mitochondrial damage. These results indicated that the combination of photofrin PDT and genistein showed more cytotoxic effect and induced apoptosis in head and neck cancer cell line.

Wistar 흰쥐에서 안드로겐 수용체 증가를 통한 비수리 추출물의 남성 갱년기 개선 효과

이성(Seong Lee),정세영(Se-Young Choung),김창희(Changhee Kim),김형지(Hyung Jee Kim),모상준(SangJoon Mo),최순향(Sun-Hyang Choi),김은영(Eun Young Kim),강다혜(Dahye Kang),김은진(Eun Jin Kim),안진철(Jin Chul Ahn) 대한약학회 2022 약학회지 Vol.66 No.6

Male menopause, also referred to as late-onset hypogonadism or andropause, is a clinical syndrome characterized by sexual dysfunction, depressive disorder, and insomnia secondary to a testosterone deficiency in older men. Lespedeza cuneata, which belongs to the Fabaceae family, possesses several biological properties, including antiinflammatory, anti-oxidant, and antidiabetic effects. We investigated the inhibitory effect of standardized L. cuneata extract (LCE) on andropause symptoms in vivo. LCE administration significantly increased serum levels of total testosterone without any effects on serum estrogen levels. Additionally, LCE significantly increased serum 17-beta hydroxysteroid dehydrogenase 13 protein levels and reduced levels of sex hormone-binding globulin (SHBG), a protein that blocks free testosterone movement into the cytosol. LCE significantly upregulated the expression of cAMP response element binding protein and androgen receptor (AR), which functions as a transcription factor to stimulate spermatogenesis-associated genes and results in increased numbers of AR-positive cells, such as Leydig and Sertoli cells in the testis. To summarize, LCE administration increases serum testosterone, reduces SHBG levels and upregulates AR expression. Therefore, LCE may be useful as alternative therapy for andropause in older men.