CAJ | 학술논문

本文报道了有关人表皮生长因子(hEGF)及其受体(EGFR)的基础研究结果，以及在此基础上进行新药开发的有关工作：hEGF的基因合成、表达载体的构建和转化宿主细胞、表达产物的纯化和中试规模三批样品的生产和检验等的有关研究结果、hEGF滴眼液的动物实验和临床实验等新药研制过程。有关基础研究表明，编码51个氨基酸的hEGF可以在α因子前导肽的引导下在酵母体系中分泌性表达，表达产物可促进角膜缘上皮细胞的增殖，可促进角膜碱烧伤的愈合，可用于口腔溃疡和皮肤烧伤的治疗，可对大鼠十二指肠溃疡有预防作用等，并用之制备了抗血清以测定血尿中EGF的浓度。研究表明EGF在大于l0ng/ml浓度下对神经胶质瘤细胞株BT325和人阴道上皮癌细胞株A431的生长具有抑制作用，并用差异显示方法对此现象进行了分子机制的探讨。研究肾癌EGFR表达和DNA含量检测具有临床意义。对EGF的晶体进行了初步的结构分析。中试产品检测表明hEGF相对分子质量为6000、等电点为4.6、氨基端15个氨基酸顺序正确、具有EGF的免疫原性和生物活性。产品中无酵母细胞的残余DNA。动物实验表明EFG在体内无蓄积，有明确的促进角膜上皮细胞增殖的作用，无急毒和长毒副作用，对眼无刺激和局部毒性作用。在四家医院进行的多中心双盲的临床实验以证实该滴眼液安全性、耐受量和有效性。在200例角膜移植和247例翼状胬肉切除术中有明显促进角膜损伤恢复的作用，其效果强于国外产阳性对照药素高捷疗(Solcoseryl Eye Gel)。所有这些研制工作促成了利用酵母体系表达的人表皮生长因子制成的滴眼液获得了国家I类新药证书，这是我国首次批准的酵母体系表达的基因工程药物。
본문보도료유관인표피생장인자(hEGF)급기수체(EGFR)적기출연구결과，이급재차기출상진행신약개발적유관공작：hEGF적기인합성、표체재체적구건화전화숙주세포、표체산물적순화화중시규모삼비양품적생산화검험등적유관연구결과、hEGF적안액적동물실험화림상실험등신약연제과정。유관기출연구표명，편마51개안기산적hEGF가이재α인자전도태적인도하재효모체계중분비성표체，표체산물가촉진각막연상피세포적증식，가촉진각막감소상적유합，가용우구강궤양화피부소상적치료，가대대서십이지장궤양유예방작용등，병용지제비료항혈청이측정혈뇨중EGF적농도。연구표명EGF재대우l0ng/ml농도하대신경효질류세포주BT325화인음도상피암세포주A431적생장구유억제작용，병용차이현시방법대차현상진행료분자궤제적탐토。연구신암EGFR표체화DNA함량검측구유림상의의。대EGF적정체진행료초보적결구분석。중시산품검측표명hEGF상대분자질량위6000、등전점위4.6、안기단15개안기산순서정학、구유EGF적면역원성화생물활성。산품중무효모세포적잔여DNA。동물실험표명EFG재체내무축적，유명학적촉진각막상피세포증식적작용，무급독화장독부작용，대안무자격화국부독성작용。재사가의원진행적다중심쌍맹적림상실험이증실해적안액안전성、내수량화유효성。재200례각막이식화247례익상노육절제술중유명현촉진각막손상회복적작용，기효과강우국외산양성대조약소고첩료(Solcoseryl Eye Gel)。소유저사연제공작촉성료이용효모체계표체적인표피생장인자제성적적안액획득료국가I류신약증서，저시아국수차비준적효모체계표체적기인공정약물。
This article reviews the resultsof the basic research about epidermal growth factor and its receptor,and the development of the novel drug, EGF eyedrop, that containing chemically synthesised EGF gene,the construction of EGF expression vector, the transformation of the host cells , the purification of the recombinant protein EGF, the preparation of three batches of the EGF product and identification, the preclinical and clinical trials. Relevant studies show that recombinant EGF consisting of 51 amino acids can be secreted into the medium under the control of the α factor leading sequence in the yeast cells.The EGF can accelerate the growth of corneal-limbal epithelial cells and the healing of an alkali burned corneal. The EGF can be used in curing oral cavity ulcer and skin burned wound. And it has the preventive effects on experimental duodenal ulcer of rat. The antiserum was made for test of the con- centration of blood EGF and urine EGF by RIA. Data from studies demonstrate the inhibition effect of EGF on the growth of tumor cells, such as A431 and BT325 cells in the presence of high EGF concentration (＞10 ng/ml). The expression of EGFR and DNA ploidy in renal carcinoma has clinical significance. Crystallization and preliminary x-ray diffraction studies of the EGF has been made. The MW of the EGF product is 6000, and the pI is about 4. 6 and it has correct N-terminal amino acids sequences , immunogenicity and biological activity. There is no vestige of the DNA of the yeast cells. Animal experiments reveal that there is no cumulation of the EGF in the body, and EGF can promote corneal epithelial healing. There is no toxicological effect during cornea wound healing of rabbit. A randomized, double-blind, placebo-controlled, multi-center clinical trial was conducted in four hospitals to assess safety,ocular tolerance and efficacy of an ophthalmic solution of EGF for 200 cases of cornea transplantation and 247 cases of nebulae. Unequivocal results were obtained as the eyedrop really accelerate the wounded cornea healing. So, the EGF eyedrop as a novel drug of class I is approved by the National Drug Administration, and this is the first gene engineering drug that come from yeast expression system in China.