Pharmaceutical Biotechnology Growth Factor By Yuqiong Xia 5/12/2017
生物技术制药 2 技术基础 生物技术药物 生物技术制药概念药物的开发药物的生产细胞因子生长因子激素类药物血液制品和治疗性酶抗体 疫苗和佐剂核酸药物 药物的发现 专利的申请临床前试验临床试验生物药物的来源生物药物的生产和纯化生物药物的分析干扰素白介素造血生长因子
创伤和创伤愈合 胰岛素样生长因子 IGF 表皮生长因子 EGF 生长因子 GF 血小板源性生长因子 PDGF 3 促进细胞有丝分裂的因子包括白介素 干扰素 集落刺激因子 促红细胞生成素 促血小板生成素 神经营养因子等 成纤维细胞生长因子 FGF 转化生长因子 TGF 神经营养因子 NGF
Outline Wound and wound-healing Insulin-like growth factor Epidermal growth factor Platelet-derived growth factor Fibroblast growth factor Transforming growth factor Neurotrophic factor 4
Wound healing 5 http://img.tfd.com/mk/h/x2604-h-08.png
Wound healing inflammatory reactions Secrete growth factors Stimulate cell growth Promote wound healing Scar 6
Wound healing Secrete growth factors FGF, TGF, PDGF, IGF-1, EGF Stimulate cell growth Fibroblasts ( 成纤维细胞 which produce collagen and elastin precursors, and ground substance) Epithelial cells ( 上皮细胞 e.g. skin cells) Vascular endothelial cells ( 血管内皮细胞 ) 7
Wound healing Promote wound healing Granulation ( 肉芽生成 growth of connective tissue ( 结缔组织 ) and small blood vessels at the healing surface) Epithelialization ( 上皮化过程 ) 8 http://bme240.eng.uci.edu/students/07s/ngunn/images/day5_with_text.jpg
Wound acute wound healing quickly on their own chronic wound healing slowly often requiring medication Such as ulcer ( 溃疡 ) 9 https://cimg2.ibsrv.net/cimg/www.woundcarecenters.org/609x375_85-1/242/acute-wounds-13242.jpg http://www.mayoclinic.org/-/media/kcms/gbs/patient-consumer/images/2013/11/15/17/42/ds00242_-ds00958_im02752_r7_ulcersthu_jpg.jpg
chronic wound Caused by some influence which disrupts the normal healing process The influence include Diabetes ( 糖尿病 ) Malnutrition ( 营养不良 ) rheumatoid arthritis ( 风湿性关节炎 ) Ischaemia ( 局部缺血 ) 10
Types of ulcers Ulcers Decubitus ulcer ( 褥疮性溃疡 ) Diabetic ulcers ( 糖尿病性溃疡 ) 11 Varicose ulcers ( 静脉曲张性溃疡 ) Rodent ulcers ( 侵蚀性溃疡 ) Peptic ulcers ( 消化性溃疡 ) Description Due to constant pressure, often associated with bed-ridden patients Due to complications of diabetes Due to defective circulation An ulcerous cancer (basal cell carcinoma), often on face Due to digestion of the mucosa ( 黏膜 )by acid and pepsin ( 胃蛋白酶 ); may occur in e.g. the duodenum ( 十二指肠 ), or the stomach ( 胃 )
Why are acute wound easier to heal than chronic wound inflammatory reactions Secrete growth factors Stimulate cell growth Promote wound healing Acute wound secretes several folds more growth factors than chronic wound Scar 12
Outline Wound and wound-healing Insulin-like growth factor Epidermal growth factor Platelet-derived growth factor Fibroblast growth factor Transforming growth factor Neurotrophic factor 13
Insulin-like growth factor Structure similar to insulin Two types IGF1 IGF2 IGF synthesis In liver Enter blood: In blood, most IGFs are bound by IGF bound protein (IGFBP) Act on target cells through endocrine mode 14 https://en.wikipedia.org/wiki/insulin-like_growth_factor_1#dwarfism
Biological activity of IGF1 Primary mediator of the effects of growth hormone (GH) 15 http://www.nature.com/nrgastro/journal/v11/n10/images/nrgastro.2014.102-f1.jpg
Biological activity of IGF1 Insulin-like effects Bind to insulin receptors 100-fold less well than insulin Regulate cell growth and development, especially in nerve cells, as well as cellular DNA synthesis 16
Clinical significance of IGF1 Laron dwarfism ( 拉伦侏儒症 ) The patients can express normal level of GH Height below 3 standard deviations (SD) IGF-1 levels below 3 SD, but lower than 3SD Mecasermin (brand name Increlex) is a synthetic analog of IGF-1 which is approved for the treatment of growth failure. IGF-1 has been manufactured using both yeast and E. coli. 17 http://blog.csdn.net/rns521/article/details/6953591
Clinical significance of IGF1 Acromegaly 肢端肥大症 Caused by elevated GH and insulin-like growth factor 1 (IGF-1) levels Diagnostic test IGF-1 in blood, 10-1000 ng/ml screening test for growth hormone deficiency and excess in acromegaly ( 肢端肥大症 ) and gigantism ( 巨人症 ) 18
Outline Wound and wound-healing Insulin-like growth factor Epidermal growth factor Platelet-derived growth factor Fibroblast growth factor Transforming growth factor Neurotrophic factor 19
Epidermal growth factor (EGF) 53 AA, nonglycosylation 2 beta-strand, 3 S-S bond Biological activity Stimulate the growth of endothelial cells( 内皮细胞 ), epithelial cells ( 表皮细胞 ) and fibroblasts ( 成纤维细胞 ) Produced mainly by monocytes and ectodermal cells ( 外胚层细胞 ) 20 https://en.wikipedia.org/wiki/epidermal_growth_factor#medical_uses
EGFR (EGF receptor) EGFR is also receptor of TGFα EGF signaling EGF binds to EGF EGFR dimerization and activation Various cellular events are triggered Promote cell growth 21 https://en.wikipedia.org/wiki/epidermal_growth_factor_receptor
Various cellular events Activate EGFR s tyrosine kinase activity Activate Ras pathway EGF-EGFR complex translocate to nucleus 22
EGFR with cancer Several cancer cell types express a truncated EGFR squamous-cell carcinoma of the lung 肺部鳞状细胞癌 (80% of cases), anal cancers 肛门癌, glioblastoma 恶性胶质瘤 (50%) and epithelial tumors of the head and neck (80-100%) 头颈上皮细胞肿瘤 The EGFR can lead to cellular transformation Tyrosine kinase inhibitors (PD153035) may represent effective chemotherapeutic agents for such cancers 23 http://www.selleckchem.com/products/pd-153035-hydrochloride.html
Problems with EGF as drug EGF were involved in malignant ( 恶性的 )growth Disappointing results from a rigorous clinical trial in which EGF was topically ( 局部地 )administered to acute, experimentally-induced, controlled wounds in healthy volunteers 24
EGF for DFU ( 糖尿病足部溃疡 ) Diabetic patients Decreased concentrations of growth factors, notably EGF Impaired natural wound healing Chronic non-healing wounds, which in later stages can require limb amputation ( 截肢 ). DFUs are the most difficult to treat and at highest risk of amputation. 25 MEDICC Review, January 2013, Vol 15, No 1, Page 11
EGF for DFU ( 糖尿病足部溃疡 ) The recombinant EGF drug: Heberprot-P Improve healing and reduce amputations in patients with severe DFU using topical administration Patient A Patient B 1 4 1 4 2 5 Before treatment 11.9 cm 2 2 Before treatment 21.8 cm 2 5 3 6 After treatment of several weeks 1.1 cm 2 3 After treatment of several weeks 0.5 cm 2 6 26 MEDICC Review, January 2013, Vol 15, No 1, Page 11 At week 11, healed At week 11, healed
Outline Wound and wound-healing Insulin-like growth factor Epidermal growth factor Platelet-derived growth factor Fibroblast growth factor Transforming growth factor Neurotrophic factor 27
Platelet-derived growth factor (PDGF) Dimeric glycoproteins: AA, BB or AB A 110AA or 125 AA BB B 109AA In platelets: 70% AB, 20% BB, 10% AA 28 https://en.wikipedia.org/wiki/platelet-derived_growth_factor
Biological activity of PDGF Promote uncontrollable cancer growth Promote blood vessel formation (angiogenesis), the growth of blood vessels from already-existing blood vessel tissue Promote wound healing Activated platelet release PDGF, which stimulate the growth of fibroblasts 成纤维细胞, smooth muscle cells and glial cells 神经胶质细胞 29 https://en.wikipedia.org/wiki/platelet-derived_growth_factor
Cell signaling in chemotaxis ( 趋药性 ) and cell proliferation during wound healing PDGF, VEGF and TGF-β integrally recruit osteoprogenitor cells ( 骨先质细胞 ) The cells then differentiated to different cells in response to bone morphogenic protein (BMP) 30 http://www.antibodyreview.com/article_images/19707367/biologics/2-3/p345-2721393/btt-2-345f1.jpg
Clinical application of PDGF 31 To heal chronic ulcers ( 慢性溃疡 ) REGRANEX Gel is the first and only FDA-approved recombinant PDGF (BB) therapy for diabetic neuropathic ( 神经性 ) ulcers Topical administration Produced in yeast Excipients: methylparaben( 对羟基苯甲酸苯酯 ), propylparaben( 对羟基苯甲酸丙酯 ) and m-cresol ( 间甲酚 ), all three are preservatives Side effects Cancer!
Outline Wound and wound-healing Insulin-like growth factor Epidermal growth factor Platelet-derived growth factor Fibroblast growth factor Transforming growth factor Neurotrophic factor 32
Fibroblast growth factor (FGF) FGF family FGF1,, FGF20 18-28 kda 140AA core Bind strongly to heparin ( 肝素 ) 33 https://en.wikipedia.org/wiki/fibroblast_growth_factor
Biological activity of FGF Promote wound healing Stimulate growth of a range of cell types Promote repair to damaged myocardial ( 心肌 ) tissue in animal models Promote embryonic development ( 胚胎发育 ) Overexpression of FGF is a feature of many tumor types Brain, breast, prostate( 前列腺 ), thyroid( 甲状腺 ) and skin 34
Outline Wound and wound-healing Insulin-like growth factor Epidermal growth factor Platelet-derived growth factor Fibroblast growth factor Transforming growth factor Neurotrophic factor 35
Transforming growth factor (TGF) Two types: TGFα, TGF β TGF α exhibits high amino acid homology with EGF Induces effects by binding to EGFR Synthesized by various body tissues, as well as monocytes and keratinocytes ( 角化细胞 ) Also synthesized by some tumor cell types 36 https://www.researchgate.net/profile/femke_heindryckx/publication/26721989/figure/fig6/as:279211613933575@1443580644151/figure-4-transforming-growthfactor-alpha-tgf-alpha-and-epidermal-growth-factor-egf.png
TGF β Three types: TGF β1, β2, β3 Affect tissue remodeling ( 组织重塑 ) and wound healing Induce growth of connective tissue ( 结缔组织 ), cartilage ( 软骨 )and bone Affect haemopoiesis Inhibit growth of epithelial ( 上皮 ) and haemopoietic cells Clinical trials: Anti-cancer agent due to inhibition of various human leukaemia cell lines 37
TGF β in wound healing 角化细胞 4 2 1 3 2 1. Platelets release TGF-β. 2. Leukocytes, macrophages, fibroblasts, and keratinocytes ( 角化细胞 ) produce TGF 3. Stimulate infiltration of inflammatory cells, fibroplasia, matrix deposition, and angiogenesis. 4. endogenous TGF-β has been shown to inhibit reepithelialization 38 http://physrev.physiology.org/content/physrev/83/3/835/f4.large.jpg?download=true
Outline Wound and wound-healing Insulin-like growth factor Epidermal growth factor Platelet-derived growth factor Fibroblast growth factor Transforming growth factor Neurotrophic factor 39
Neurotrophic factor A family of biomolecules, mostly are peptides or small proteins Support the growth, survival, and differentiation of both developing and mature neurons 40
Structure of a neuron 树突 : 与靶细胞结合 轴突末梢 : 与靶细胞结合 细胞体 郎氏结 轴突 : 传递信息, 有时接收信息 施旺细胞 细胞核 髓鞘 41 https://owlcation.com/stem/structure-of-a-neuron
Structure of a neuron 突触 突触 42 突触 https://owlcation.com/stem/structure-of-a-neuron
How does NTF work Target cell NTF CAM 和 NTF 诱导轴突定向生长 细胞表面黏附分子 CAMS 43
Types of NTFs Neurotrophin family Nerve growth factor (NGF) Neurotrophin-3 (NT-3) Brain-derived neurotrophic factor (BDNF) CNTF family Ciliary neurotrophic factor (CNTF) 睫状神经因子 GDNF family Glial cell line-derived neurotrophic factor (GDNF) 胶质细胞系源性神经营养因子 44
NGF Use the high-affinity receptor TrkA to promote myelination ( 髓鞘形成 ) and the differentiation of neurons Stimulate growth and differentiation of B and T lymphocytes 45
BDNF Predominantly localized within the CNS Predictably, most BDNF responsive neurons are located in (or project into) the CNS Promote the survival of embryonic retina ganglion cells ( 视网膜神经节细胞 ), dopaminergic neurons ( 多巴胺能神经元 ), as well as cholinergic neurons of the basal forebrain( 基底前脑胆碱能神经元 ), embryonic spinal motor neurons ( 胚胎脊髓运动神经元 )and cortical neurons( 皮质神经元 ) 46
Clinical application of NTF Alzheimer s disease (AD) and Parkinson s disease Death of CNS neurons 黑质 47 https://ghr.nlm.nih.gov/art/large/alzheimer-disease-brain.jpeg http://www.calchemsynthesis.com/wp-content/uploads/2017/04/parkinsons-disease.jpg
Clinical application of NTF NTF may cure these diseases NTF cannot penetrate BBB (blood-brain barrier) 48 http://study.com/academy/lesson/blood-brain-barrier-definition-function.html
Clinical application of NTF To overcome BBB (blood-brain barrier) 1. Direct injection 2. Cranial infusion pumps ( 颅内输注泵 ) 3. Polymer-encapsulated cells, which secrete NTF 49 http://www.thevaccinereaction.org/wp-content/uploads/2016/02/blood-brain-barrier.jpg
Clinical application of NTF Infusion of NGF and NT-4/5 to aged rats appears to improve their spatial memory ability. Further studies in rodents have shown that physical exercise increased BDNF expression, which may explain why exercise is often a predictor of high mental function during aging. Promising treatment for Alzheimer s disease 50
Clinical application of NTF Pre-clinical studies show that direct injection of GDNF to sites adjacent to the substantia nigra ( 黑质 ) prevents degeneration of these neurons when they are experimentally damaged by cutting their axons Promising treatment for Parkinson s disease 51
Summary Wound healing IGF1 (growth hormone) EGF (cancer, DFU) PDGF (cancer, wound healing) FGF (cancer, wound healing) TGF alfa (cancer) TGF beta (tissue remodeling, haemopoiesis) NTF (neuron, Alzheimer's disease, Parkinson's disease) 52