랫드에 대한 DWP-311의 급성피하독성시험

곽승준,김형식,천선아,임소영,박현선,한하수,홍채영,안미영,이병무,Kwack, Seung-Jun,Kim, Hyung-Sik,Chun, Sun-Ah,Lim, So-Young,Park, Hyun-Sun,Han, Ha-Su,Hong, Chae-Young,Ahn, Mi-Young,Lee, Byung-Mu 한국독성학회 1998 Toxicological Research Vol.14 No.3

The acute toxicity of DWP-311 was investigated in Sprague-Dawley rats. DWP-311 was subcutaneously administratered at dose levels of 595, 1,070, 1,930, 3,470, and 6,250mg/kg. In this study, we daily examined numbers of deaths, clinical signs, body weights, and pathological examinations for 7 days after administration of DWP-311. The results indicate that DWP-311 did not show any toxic effect in rats and the oral $LD_{50}$ value was over 6,250mg/kg in Sprague-Dawley rats.

랫드에 대한 KDRD-002 의 아급성경구독성시험

곽승준(Seung Jun Kwack),김형식(Hyung Sik Kim),이소영(So Young Lee),천선아(Sun Ah Chun),홍채영(Che Young Hong),한하수(Ha Su Han),최병천(Byung Chun Choi),이병무(Byung Mu Lee) 한국응용약물학회 1996 Biomolecules & Therapeutics(구 응용약물학회지) Vol.4 No.4

The subacute toxicity was investigated in Sprague-Dawley rats orally treated with KDRD-010 at the doses of 0.056, 0.28, and 1.4 g/㎏ for one month. There were no clinical signs and pathological changes compared with control group. Body weights were not significantly changed between control and treatment groups. In hematological and biochemical serum parameters, all mean values appear to be within the normal range. In pathological examinations, hemorrhages of lung was observed in one male rat at low dose group and one female rat at high dose group of KDRD-010, but it was not considered to be caused by KDRD-010. These results suggest that KDRD-010 dose not induce any significant subacute oral toxicities in Sprague-Dawley rats.

랫드에서 인체 재조합 적혈구 조혈인자 , rHu-EPO 의 급성정맥독성시험

곽승준(Seung Jun Kwack),김형식(Hyung Sik Kim),임소영(So Young Lim),천선아(Sun Ah Chun),홍채영(Chae Young Hong),박현선(Hyun Sun Park),김원배(Won Bae Kim),김병문(Byoung Moon Kim),안병옥(Byoung Ok Ahn),이병무(Byung Mu Lee) 한국응용약물학회 1996 Biomolecules & Therapeutics(구 응용약물학회지) Vol.4 No.4

Acute intravenous toxicities of rHu-EPO (recombinant human erythropoietin) were investigated in Sprague-Dawley rats. Seven days after administration of rHu-EPO, we examined the clinical signs, mortalities, body weight and etc. No clinical signs and mortalities of toxicity were observed in animals. Also, a significant change of body weights was not observed. These results suggest that LD_(50) value was >25,000 unit/kg in Sprague-Dawley rats and the acute intravenous toxicities of rHu-EPO were not significant.

3-MCPD의 생식ㆍ발생독성에 관한 연구

곽승준(Seung Jun Kwack),김순선(Soon Sun Kim),최요우(Yo Woo Choi),이규식(Gyu Seek Rhee),손경희(Kyung Hee Sohn),이이다(Rhee Da Lee),채수영(Soo Young Chae),정용현(Yong-Hyun Chung),유일재(Il Je Yu),박귀례(Kui Lea Park) 한국독성학회 2004 Toxicological Research Vol.20 No.1

3-Monochloro-1,2-propanediol(3-MCPD) is a toxic compound, often present in different foods containing acid hydrolyzed(AH) protein, like seasonings and savory food products. The purpose of the present study was to investigate the effects of 3-MCPD on male fertility, sperm and testosterone secretion. In vivo male fertility test was performed for observing the adverse effects of 3-MCPD on the function of male reproductive system and pregnancy outcome. 0.01, 0.05, 0.25, 1 and 5 mg/kg b.w. of 3-MCPD was given daily by gavage to groups of 15 adult male SD rats for 4 weeks. At the end of pre-treatment period, males were mated overnight with normal females. Following morning, males demonstrating successful induction of pregnancy were sacrificed on that day to assess<br/> sperm parameters and histopathology of reproductive organs. The resulting pregnant females were sacrificed on day 20 of gestation to evaluate pregnancy outcome. As a result, four-week paternal<br/> administration with 3-MCPD resulted in adverse effects on male fertility and pregnancy outcome without remarkable histopathological changes in testes and epididymides; sperm motility, copulation index and fertility index were markedly decreased in the treated group and numbers of live fetuses showed<br/> steep dose-response curves. Also, spermatogenesis was investigated in this experiment. However, no effect was observed on production of sperm in testes treated with 3-MCPD for 4 weeks. Hormone assay was performed for observing the effects of 3-MCPD on testosterone and luteinizing hormone (LH) in blood and testes of male SD rats and cultured primary Leydig cell. In result, significant changes of related hormones did not observed by treatment of 3-MCPD. These results indicated that paternal treatment with 3-MCPD induced spermatotoxic effect, which caused an antifertility on male.

Thimerosal의 발생독성에 관한 연구

곽승준(Seung Jun Kwack),이규식(Gyu Seek Rhee),김순선(Soon Sun Kim),손경희(Kyung Hee Sohn),김소희(So Hee Kim),채수영(Soo Young Chae),최요우(Yo Woo Choi),원용혁(Yong Hyuck Won),박귀례(Kui Lea Park) 한국독성학회 2003 Toxicological Research Vol.19 No.4

The purpose of our study was to evaluate the toxicity of the thimerosal in embryos and neonates. Thimerosal (also known as mercurothiolate) is a mercury-containing compound used in<br/> trace amounts to prevent bacteria and other organisms from contaminating vaccines, especially in opened multi-dose vials. The toxicity of mercury is well known and those most at risk occurrs in<br/> unborn babies and newborn babies. Test methods included in vitro whole embryo culture (WEC) system and in vivo test of neonatal toxicity in Wistar rats. Ethylmercury and methylmercury were used as positive controls for the evaluating of toxic effects of mercury. In WEC assay, treated concentrations of thimerosal, ethylmercury and methylmercury were up to 0.01, 0.025, 0.05, 0.1, 0.25, 0.5, 1, 2.5 and 5 mg/ml, respectively. All compounds didn’t show any morphological abnormalities, but showed retardation<br/> of growth and development in dose dependent manner (>0.5㎍/ml). These data indicated that thimerosal showed developmental toxicity in vitro. In vivo neonatal toxicity, Wistar rats were administered<br/> subcutaneously with thimerosal, ethylmercury, or methylmercury (5, 25, 50, 250, and 500 mg/kg) during from postnatal day (PND) 4 to 25. Significant effects of these compounds on relative organ<br/> weights and organ morphology were not observed in this experiment. However, accumulation of mercury was detected in the kidney and testis when treated with thimerosal, ethylmercury, or methylmercury. These results suggest that thimerosal may be a harmful compound to embryo and neonate, but used concentration of thimerosal in these experiments is much higher than that of clinical application. Further investigation is needed on the safety of vaccine components, i.e. a thimerosal using in vitro and in vivo tests in the future.

생식ㆍ발생독성시험의 방법적 고찰과 최신 연구 동향

곽승준(Seung Jun Kwack),조대현(Dae Hyun Cho) 한국독성학회 2005 Toxicological Research Vol.21 No.4

Reproductive and developmental toxicology is concerned with various physical or chemical agents interfering with fertility in both gender or normal growth of offsprings. Reproductive and developmental toxicology is rather a complex science, with many fields, i.e., various endpoints are involved and many different mechanisms of action. For that reason, diverse aspects must be considered when attempting to assess possible adverse health effects in the area of reproductive and developmental toxicology. The thalidomide tragedy made it clear to regulatory authorities around the world that systematic, comprehensive evaluation of the reproductive cycle was needed to adequately evaluate the potential of medicinal drugs to impair the process of reproduction or the development of embryos, fetuses, and children. International Conference on Harmonization of Technical Requirements for the Registration of Pharmaceuticals for Human Use (ICH) and U.S. Food and Drug Administration (FDA) developed a guideline to assess the reproductive and developmental toxicity. Also these guidelines have since been applied to the detection and regulation of environmental toxicants, food additives, and so on. Although it was hoped that testing procedures of guideline would be updated constantly to reflect the current state of the science in reproductive and developmental toxicology, it was not until this decade that regulatory guidelines and testing methods have been altered in a significant way. In this paper, we would like to present the recommended approaches and recent trends for improvement of testing guidelines or experimental methods in reproductive and developmental toxicology.

3-MCPD의 생식˙발생독성에 관한 연구

곽승준(Seung Jun Kwack),김순선(Soon Sun Kim),최요우(Yo Woo Choi),이규식(Gyu Seek Rhee),손경희(Kyung Hee Sohn),이이다(Rhee Da Lee),채수영(Soo Young Chae),정용현(Yong-Hyun Chung1),유일재(Il Je Yu1),박귀례(Kui Lea Park) 한국독성학회 2004 Toxicological Research Vol.20 No.2

3-Monochloro-1,2-propanediol(3-MCPD) is a toxic compound, often present in different foods containing acid hydrolyzed(AH) protein, like seasonings and savory food products. The purpose of the present study was to investigate the effects of 3-MCPD on male fertility, sperm and testosterone secretion. In vivo male fertility test was performed for observing the adverse effects of 3-MCPD on the function of male reproductive system and pregnancy outcome. 0.01, 0.05, 0.25, 1 and 5 mg/kg b.w. of 3-MCPD was given daily by gavage to groups of 15 adult male SD rats for 4 weeks. At the end of pre-treatment period, males were mated overnight with normal females. Following morning, males demonstrating successful induction of pregnancy were sacrificed on that day to assess sperm parameters and histopathology of reproductive organs. The resulting pregnant females were sacrificed on day 20 of gestation to evaluate pregnancy outcome. As a result, four-week paternal administration with 3-MCPD resulted in adverse effects on male fertility and pregnancy outcome without remarkable histopathological changes in testes and epididymides; sperm motility, copulation index and fertility index were markedly decreased in the treated group and numbers of live fetuses showed steep dose-response curves. Also, spermatogenesis was investigated in this experiment. However, no<br/> effect was observed on production of sperm in testes treated with 3-MCPD for 4 weeks. Hormone assay was performed for observing the effects of 3-MCPD on testosterone and luteinizing hormone (LH) in blood and testes of male SD rats and cultured primary Leydig cell. In result, significant changes of related hormones did not observed by treatment of 3-MCPD. These results indicated that paternal treatment with 3-MCPD induced spermatotoxic effect, which caused an antifertility on male.

아데노바이러스 유전자치료벡터의 생식독성 연구

이규식,곽승준,김순선,이이다,석지현,채수영,정수연,김승희,이승훈,박귀례,Rhee, Gyu-Seek,Kwack, Seung-Jun,Kim, Soon-Sun,Lee, Rhee-Da,Seok, Ji-Hyun,Chae, Soo-Young,Chung, Soo-Youn,Kim, Seung-Hee,Lee, Seung-Hoon,Park, Kui-Lea 한국미생물학회 2007 미생물학회지 Vol.43 No.3

유전자치료W터의 주입시 생식세포를 통한 다음 세대로의 전달 가능성은 안전성 측면에서 관심을 중대시키고 있다. 특히 전립선암이나 난소암의 치료시 바이러스를 생식기관에 인접한 부위에 주입하여야 하므로 그 가능성이 높다. 따라서 본 연구에서는 유전자치료에 많이 이용되는 아데노바이러스를 매개로하여 tumor suppressor 유전자인 p53을 발현하는 아데노바이러스 벡터를 제조하여 이를 투여시 생식장기를 포함한 주요장기조직에의 분포와 germ cell을 통한 차세대로의 전달 가능성 등의 생식독성을 조사하였다. In vivo biodistribution study를 위하여 $Ad-CMV-{\beta}-gal$흑은 Ad-CMV-p53를 마우스 암 수의 복강에 주사한 후 생식장기를 포함한 주요 장기에서 아데노바이러스 유래 DNA검출 및 RNA발현 여부를PCR과 RT-PCR로 각각 확인하였다. 그 결과 간 및 비장과 같은 일반 장기에서도 주입한 외부유전자의 DNA가 검출되거나RNA가 발현되었을 뿐만 아니라, 정낭, 전립선, 부고환, 난소 및 자궁 등의 생식장기에서도 주입한 외부유전자가 검출되거나 발현되는 것으로 나타났다. Real-time PCR을 이용하여 각 장기에서의 투여된 아데노바이러스 벡터는 시간 의존적으로 감소되는 것을 정량하였다. Ad-CMV-p53를 암 수 마우스의 난소와 고환에 각각 직접 주사하여 교배시킨 후 그 후세대의 DNA를 분리하여 주입한 아데노바이러스 유래의 DNA를 검색한 결과, 어떠한 차세대에서도 주입한 아데노바이러스 유래의 DNA가 검출되지 않았다. 한편 생식장기에서의 PCR및 RT-PCR signal유래 vector의 위치를 확인하기 위해 매우 감도가 높은 in-situ PCR로 조사한 결과 고환의 경우 간질조직으로의 전달은 일어나나 정세관 내에는 아데노바이러스 벡터가 전달되지 않으며, 난소에서도 아데노바이러스벡터는 난포내의 난자에 전달되지 않고 기질조직에 존재하는 것으로 확인되었다. 결론적으로 복제 능력 이 결여된 아데노바이러스를 매개로 한 유전자치료제는 생식 장기에서 검출되더라도 다음 세대로 전달될 가능성은 대단히 낮음을 제시한다. The possibility of inadvertent introduction of therapeutic gene expressing viral vectors has raised safety concerns about germ-line infection. Particularly, for indications such as prostate cancer and ovarian cancer, the proximity of the point of viral administration to organs of the reproductive system raises concerns regarding inadvertent germ-line transmission of genes carried by the virus vector. To evaluate the safety of in vivo adenovirus mediated gene transfer, we explored the biodistribution, persistance and potential germ-line transmission of p53-expressing adenovirus (Ad-CMV-p53). Both male and female Balb/c mice were injected with $1{\times}10^9$ PFU of Ad-CMV-p53. The PCR analysis showed that there were detectable vector sequences in liver, kidney, spleen, seminal vesicle, epididymis, prostate, ovary, and uterus. The RT-PCR analysis for detecting inserted gene, p53 showed that Ad-CMV-p53 viral RNA were present in spleen, prostate and ovary. Direct injected male and female mice of adenovirus vector into testis and ovary were mated and their of offspring were evaluated for germ-line transmission of the adenoviral vector. The PCR and RT-PCR analysis showed no evidence of germline transmission, although vector sequences were detected in DNA extracted from gonadal tissues. Real-time PCR result confirmed a significant decrease of adenovirus in gonad tissues 1 week after injection. We have also analysed the cell specific localization of viral DNA in gonad tissues by using in-situ PCR. Positive signals were detected in interstitial tissue but not in seminiferous tubule in sperm. In the case of ovary, adenovirus signal were localized to the stromal tissue, but no follicular signals were observed. Together, these data provide strong evidence that the risk of the Inadvertent germ-line transmission of vector sequences following intraperitoneal or direct injection into genito-urinary system of adenovirus is extremely low.

랫드에서 인체 재조합 적혈구 조혈인자, rHuEPO의 13주 정맥투여 아만성독성에 관한 연구

김형식,곽승준,천선아,박현선,한하수,임소영,안미영,김원배,안병옥,홍성렬,이병무,Kim, Hyung-Sik,Kwack, Seung-Jun,Chun, Sun-Ah,Park, Hyun-Sun,Han, 한하수,Lim, So-Young,Ahn, Mi-Young,Kim, Won-Bae,Ahn, Byoung-Ok,Hong, Sung-Youl,Lee, Byung-Mu 한국독성학회 1998 Toxicological Research Vol.14 No.3

A recombinant human erythropoietin (rHuEPO) was administered intravenously at dosage levels of 0, 100, 500, and 2500IU/kg/day for a period of 3 weeks. There were no observed clinical signs and deaths related to treatment in all groups tested. Decreases in body weight gain and food consumption were observed only in males of 2,5000IU/kg group after 2 weeks. In hematological parameters, erythrocyte content, hematocrit values and hemoglobin concentration were dose- dependently increased in rHuEPO treated groups. The ratio between kidney weight and whole body weight was significantly increased in females of 500 and 2,500IU/kg groups. The spleen weight was also increased in both sexes of 500 and 2,500IU/kg groups. However, the absolute weight change of other organs was not observed. In histopathological examinations, the renal tubular basophilia was observed only in males and females of 2,500IU/kg groups. From these results, it is concluded that the no-observed adverse effect level (NOAEL) of rHuEPO is 100 IU/kg in rats in the present study.

비글개에서 인체 재조합 적혈구 조혈인자, rHu-EPO의 급성독성에 관한 연구

조명행,성하정,김형식,곽승준,천선아,임소영,김원배,김병문,안병옥,이병무,Cho, Myung-Hang,Seong, Ha-Jung,Kim, Hyung-Sik,Kwack, Seung-Jun,Chun, Sun-Ah,Lim, So-Young,Kim, Won-Bae,Kim, Byoung-Moon,Ahn, Byoung-Ok,Lee, Byung-Mu 한국독성학회 1996 Toxicological Research Vol.12 No.2

The acute toxicity of rHu-EPO, newly developed recombinant erythropoietin, was tested in beagle dogs. rHu-EPO, when administered intravenously at 25, 000 IU/kg, did not cause any death. Also, rHu-EPO did not induce any change of body weight, food intake and clinical signs compared to controls. There were no significant changes in hematological, urine analysis and pathological examination. These results showed that rHu-EPO did not induce any remarkable toxic response and the $LD_50$ was greater than 25, 000 IU/kg in beagle dogs.