IL-6RA

前言近年来，干细胞在临床应用方面愈加广泛，但人体内的干细胞比例和数量极低，无法满足临床需求，所以体外扩增培养干细胞变得越来越重要，由于伦理和工艺的原因，干细胞的治疗还面临很多的问题，造血干/祖细胞及各系血细胞的表面标志较为清晰，细胞表型特征可进行定量选择，可分选，且发挥功能可相对游离，不需“生物支架”神经、血管、外科移植等复杂的“下游”工艺。因此是干细胞增殖分化的最佳模型，也便于直接用于临床。造血干细胞有高度的自我更新和多种分化潜能，可以产生所有成熟的血细胞，如红细胞、白细胞、血小板和淋巴细胞等，并且可以重建整个造血系统，造血干细胞体外扩增既要保持自我更新能力的同时还要阻止其分化，所以这是一项极具挑战性的技术。近年来大量实验表明，造血干细胞的分化依赖细胞因子，下面就为大家简单整理了一下造血干细胞培养用到的因子及其作用。● 干细胞因子（SCF）干细胞因子（stemcellfactor，SCF）是一种通过锚定并表达于所有HSC表面的酪氨酸受体c-Kit而发挥作用的因子，c-Kit表达缺陷将导致HSC扩增数量减少。目前几乎用于培养HSC实验中的所有细胞因子组合都包含SCF。另外，SCF与FL3同属于酪氨酸激酶受体TKR家族，具有扩增原始造血细胞的协同作用，通过与特异性TKR结合，向细胞内传递信号，启动早期祖细胞的分裂和增殖，使细胞走出G0期开始扩增，抑制凋亡。● 血小板生成素（TPO）TPO最初被认为是巨核细胞系特异性生长因子，属于特异性作用细胞因子,可维持巨核细胞增殖、分化、成熟、分裂及形成有功能的血小板，是扩增巨核细胞系的首选因子。近年来实验证实，在体外研究中TPO在促进HSC增殖中也起到重要的作用，并且与其他细胞因子组合使用可提高集落形成单位总数和CD34+干细胞的扩增倍数。特别是在有FL3组合中，可维持脐带血CD34+干细胞长期生长和扩增。● 白细胞介素-3白细胞介素-3（IL-3）称为肥大细胞生长因子，是一个具有多效性的细胞因子，主要由激活的T淋巴细胞产生，能够刺激多能HSC和不同谱系定向祖细胞的增殖与分化。IL-3诱导的干细胞表面受体异二聚体化后，可结合许多信号转导蛋白，如Janus激酶信号转导子与转录激活子（JAK/STAT）通路等，从而激发了一股下传的信号流，参与了干细胞的增殖调控。IL-3还可激活细胞外信号调节激酶（ERK）途径和c-jun氨基末端酶（JNK）途径，诱导干细胞的生长、增殖与存活。● 白细胞介素-6IL-6 是一种多向性细胞因子，通过调节免疫和炎症反应，在宿主防御中发挥重要作用。IL-6 由 T细胞、单核细胞、成纤维细胞、内皮细胞和角质形成细胞产生，具有多种生物学功能。它可以促进 B 细胞分化和抗体产生，协同 IL-3 在巨核细胞发育和血小板产生中发挥作用，诱导肝急性期蛋白的表达，调节骨代谢。IL-6 通过 IL-6 受体系统传递信号，IL-6 受体系统由 IL-6Rα和 gp 130 两条链组成。在胚胎干细胞中STAT3是维持胚胎干细胞不分化状态的决定性分子，而IL-6是JAK/STAT3信号通路最初的启动因子。● lFLT3配体（FL）FLT3配体是调节早期造血细胞增殖的生长因子。FLT3-配体与表达酪氨酸激酶受体FLT3的细胞结合。FLT3-配体本身并不刺激早期造血细胞的增殖，而是与其他CSFs和白细胞介素协同诱导生长和分化。与SCF不同，FLT3-配体对肥大细胞没有作用。已鉴定出FLT3-配体的多种亚型。主要的生物活性形式作为跨膜蛋白(209a.)的胞外区域锚定在细胞表面。所述膜结合异构体可被蛋白质裂解生成具有生物活性的可溶性异构体。● Fms样酪氨酸激酶3CD34+CD38dim细胞高度表达的Fms样酪氨酸激酶3（FL3），通过与其特异性酪氨酸激酶活性的受体（tyrosinekinase-linkedreceptors，TKR）结合，向细胞内传递信号。FL3作用于HSC/HPC，通过与在细胞表面的有TKR结合而发挥造血调控作用。FL3也是十分重要的早期祖细胞刺激因子，对HSC/HPC体外扩增有明显的促进作用，能阻止CD34+干细胞在体外扩增时逐渐定向分化，耗净HPC的倾向。● 转化生长因子-β转化生长因子-β、转化生长因子-β1、β2 和 β3 三种哺乳动物亚型通过同一受体发出信号，引起相似的生物学反应。它们是多功能细胞因子，调节细胞增殖、生长、分化和运动，以及细胞外基质的合成和沉积。转化生长因子-β（transforminggrowthfactor-β，TGF-β）由骨髓基质细胞产生，抑制早期HSC/HPC进入S期，使大多数HSC/HPC处于G0期。● 巨噬细胞炎性蛋白-1α巨噬细胞炎性蛋白-1β（MIP-1β）与巨噬细胞炎性蛋白-1α（macrophageinflammatoryprotein1α，MIP-1α）相伴而生，是MIP-1α的天然拮抗剂，可以解除MIP-1α对早期HSC/HPC的抑制作用，阻止HSC重新返回静止状态。● p38p38作为一个信号传递分子属于丝裂原激活蛋白激酶（MAPK）家族的一员，在常氧条件下抑制HSC的体外扩增。有实验表明，当HSC在加有TPO、SCF、FL3的无血清培养液中，由于氧化应激而激活p38和p16，导致鼠HSC数量明显的减少。● 粒-巨噬细胞-集落刺激因子（GM-CSF）粒细胞-巨噬细胞集落刺激因子是临床上用于各种原因引起的白细胞或粒细胞减少症的药物。目前使用的细胞动员剂是“粒一巨噬细胞集落刺激因子（ GM--CSF ）”，GM--CSF除能增加外周血造血干细胞的数量外，还有辅助心脏功能等作用。● 粒细胞-集落刺激因子（G-CSF）粒细胞集落刺激因子的作用一般有抗原呈递、增强巨噬细胞作用、促进造血干细胞增殖等。粒细胞集落刺激因子是骨髓干细胞的强力动员剂，能刺激自体骨髓干细胞增殖，动员骨髓干细胞从骨髓到外周血。● 促红细胞生成素（EPO）红细胞生成素（EPO）是造血分化中的主要刺激因子，能促进造血干细胞向原始红细胞分化、加速幼红细胞的分裂增殖、促进血红蛋白的合成，同时在红细胞诱导分化研究中也发挥重要的作用。造血干细胞具有自我更新和多向分化潜能，不同因子的搭配对造血干细胞的增殖效果不同，利用造血干细胞可定向诱导产生多种细胞，这无疑也给NK细胞的增殖提供了新的思路。目前NK细胞可以由胚胎干细胞、iPSC诱导而来，但是无论是胚胎干细胞还是诱导多能干细胞，他们都要转化为造血干细胞后才能进而分化为NK细胞，所以造血干细胞在这其中扮演了不可或缺的桥梁。由干细胞衍生的NK细胞优点是可以随需随用，同质性强，低细胞因子释放，杀伤活性强，所以市面上至今对于干细胞来源的NK的探索仍然保持着极高的热情，下面是干细胞→NK细胞的有关文献，我们主要关注了文献中提到的因子。一、人胚胎干细胞来源的NK细胞将hESCs与小鼠骨髓基质细胞系M210-B4 在含RPMI 1640的培养基中共培养， 15%胎牛血清， 2 mM l -谷氨酰胺， 1%非必需氨基酸， 1%青霉素/链霉素和0.1 mM β-巯基乙醇，培养基每2 ~ 3天更换1次。17 ~ 20 d后，制备单细胞悬液及CD34+CD45+细胞如前所述分离的细胞转移到与小鼠胎肝来源的基质细胞系共培养AFT024在培养基中含有1:2的杜尔贝可混合物改良Eagle培养基/Ham F12， 20%热灭活人血清AB， 2mM l -谷氨酰胺，1%青霉素/链霉素，5 ng/mL亚硒酸钠，50μM乙醇胺，25μM-巯基乙醇，20 mg/mL抗坏血酸，IL-3，干细胞因子, IL-15, fms样酪氨酸激酶3配体和IL-7。细胞用新鲜培养液培养，每隔5 ~ 6天更换一半培养液。培养30 ~ 35 d，收集细胞，过滤70μm滤波器，并用于进一步分析。二、人多能干细胞来源的NK细胞收集18~21d细胞进行CD34+ CD45+祖细胞富集，10万个CD34+CD45+细胞、1 ml NK细胞、起始细胞因子（白细胞介素IL-3, IL- 7, IL-15，干细胞因子[SCF]和fms样酪氨酸激酶受体-3配体[FLT3L]置于EL08-1D2基质上。每4-5天用0.5 ml含细胞因子的培养基更新NK细胞培养物。在EL08-1D2细胞培养28~35d时检测成熟NK细胞。自从基因治疗的概念提出以来, 造血干细胞一直被认为是理想的靶细胞，造血干细胞移植治疗可以治疗很多血液系统疾病，并且多种恶性血液系统疾病只有造血干细胞移植治疗才有可能彻底治愈，此外，地中海贫血等造血系统的基因遗传疾病，也可通过对造血干细胞基因编辑进行治疗。所以造血干细胞的体外培养工艺变得尤为重要。以下是同立海源关于造血干细胞培养所用到的因子列表，北京同立海源拥有十余年重组蛋白研发生产经验，可提供满足您需求的细胞因子产品，欢迎大家垂询。同立海源干细胞相关因子部分数据展示图1：Recombinant Human GM-CSF stimulates proliferation of  TF-1(浓度：ng/ml）图2：Recombinant Human FLT3 Ligand stimulates proliferation of OCI AML-5(浓度：ng/ml）图3：Recombinant Human IL-3 stimulates proliferation of TF-1(浓度：ng/ml)图4：Recombinant Human SCF stimulates proliferation of Mo7e(浓度：ng/ml)扫码申请试用试用说明：1.活动时间：即日起-2023年7月31日 ；2.活动对象：未订购申请样品的新老终端客户；3.试用规格：50μg/支（GM-CSF、Vitronectin蛋白为100μg/支）；4.免费试用名额有限，每个客户限申请一次，（注：同一导师或同一公司为同一客户，不累计赠送）；5.为了让更多用户享有试用机会，拒绝浪费，每个客户申请试用样品种类不超过5个，每个产品种类限制1支；6.申请要求：申请信息完整，能够近期开展实验；7.其他说明：试用结束后，需反馈试用结果；8.本次活动最终解释权归北京同立海源生物科技有限公司所有。END关于同立海源北京同立海源生物科技有限公司，成立于2011年，专注细胞和基因治疗（CGT）上游GMP级原料试剂研发，致力于为生命科学提供可靠的产品与服务。产品涉及细胞分选磁珠试剂，真核/原核重组蛋白、无血清培养基、细胞培养试剂盒、转染试剂、基因编辑工具酶等。为细胞和基因治疗药物、抗体药物开发、细胞储存等生物制药和IVD领域提供核心原料试剂与服务。公司建有2000m2的研发实验室及GMP级洁净车间，包括真核与原核蛋白表达工程平台、细胞培养技术开发平台、体外诊断试剂生产平台，通过ISO13485和ISO9001双认证，部分产品已获美国FDA DMF备案。

Tocilizumab is an important anti-rheumatic agent used in the treatment of adult patients with moderate to severe active rheumatoid arthritis who have had an inadequate response to one or more Disease-Modifying Anti-Rheumatic Drugs (DMARDs) Phase I study conducted by Dr. Reddy’s demonstrated pharmacokinetic equivalence and similarity in pharmacodynamic parameters, safety and immunogenicity between Dr. Reddy’s proposed biosimilar candidate (DRL_TC) and European Union (EU) reference medicinal product* and United States (U.S.) reference product** by the intravenous route (IV) In December 2022, the company had announced the successful completion of the Phase I study of DRL_TC via the subcutaneous route Global Phase III study is being initiated to compare efficacy, safety, tolerability and immunogenicity of DRL_TC with the reference product HYDERABAD, India--(BUSINESS WIRE)-- Dr. Reddy's Laboratories Ltd. (BSE: 500124, NSE: DRREDDY, NYSE: RDY, NSEIFSC: DRREDDY; hereafter referred to as “Dr. Reddy’s”), a global pharmaceutical company, announced that its tocilizumab biosimilar candidate, DRL_TC, successfully met its primary and secondary endpoints in a Phase I study. This Phase I study used an intravenous (IV) formulation to evaluate the pharmacokinetic equivalence, safety and immunogenicity of Dr. Reddy’s tocilizumab biosimilar candidate in comparison to reference products. The Phase I study entitled ‘A Phase I, Double Blind, Randomized, Parallel-group, Single dose, Three arm, Comparative Pharmacokinetic and Pharmacodynamic Study of Dr. Reddy’s Tocilizumab (DRL_TC), USA sourced Reference Tocilizumab (Actemra®) and EU sourced Reference Tocilizumab (RoActemra®) Administered by the Intravenous Route to Normal Healthy Male Volunteers’ met all primary and secondary endpoints. Pharmacokinetic equivalence of DRL_TC to the EU reference medicinal product* and the U.S. reference product** was successfully demonstrated. The clinical trial also confirmed the similarity between DRL_TC and the EU* and U.S.** reference products in terms of pharmacodynamic parameters and found no noteworthy differences in safety and immunogenicity across these three treatment groups. The successful outcome of this study represents an important milestone in Dr. Reddy’s commitment to make high-quality biosimilar products more accessible and affordable to healthcare providers and patients around the world. Dr. Reddy’s is developing the proposed tocilizumab biosimilar as both intravenous and subcutaneous formulations. Dr. Jayanth Sridhar, Global Head of Biologics at Dr. Reddy’s, said: “Tocilizumab is an important anti-rheumatic agent that has a unique place in treating patients with rheumatoid arthritis and other diseases. By developing the formulation in both subcutaneous and intravenous formulations, we aim to reach more patients around the world. With our recent milestones in our proposed biosimilars of tocilizumab and rituximab, our partner’s launch of pegfilgrastim in the U.S and Europe, we look forward to maintaining our momentum as part of our goal to serve over 1.5 billion patients by 2030.” Dr Reddy’s has already demonstrated pharmacokinetic equivalence and similarity in pharmacodynamic parameters, safety and immunogenicity by subcutaneous route. The company is now initiating a global Phase III study with the aim of comparing the efficacy, safety, tolerability and immunogenicity of DRL_TC with the reference product in patients with moderate to severe active rheumatoid arthritis. *EU reference medicinal product is RoActemra® **U.S. reference product is Actemra® Actemra® and RoActemra® are registered trademarks of Chugai Seiyaku Kabushiki Kaisha Corp., a member of the Roche Group. About Dr. Reddy’s biosimilars programme: Dr. Reddy’s biosimilars business is part of our key strategic initiatives expected to drive both near-term and long-term growth. Over the last 20 years, our Biologics team has developed into a fully integrated organisation with robust capabilities in the development, manufacture and commercialisation of a range of biosimilar products in oncology and immunology. We have a current portfolio of six commercial products marketed in India and over 27 Emerging Markets. In addition, we have several products in the pipeline in oncology and auto-immune diseases in various stages of development for global launches across regulated as well as emerging markets. In December 2022, we announced the successful completion of Phase I study and initiation of Phase III study of DRL_TC, our proposed biosimilar of tocilizumab via the subcutaneous route, for global markets. In January 2023, we announced the successful completion of the full set of clinical studies of our rituximab biosimilar for filing in the U.S. and Europe. We are also ramping up manufacturing capacity to support our global expansion plans. About Dr. Reddy’s: Dr. Reddy’s Laboratories Ltd. (BSE: 500124, NSE: DRREDDY, NYSE: RDY, NSEIFSC: DRREDDY) is a global pharmaceutical company headquartered in Hyderabad, India. Established in 1984, we are committed to providing access to affordable and innovative medicines. Driven by our purpose of ‘Good Health Can’t Wait’, we offer a portfolio of products and services including APIs, generics, branded generics, biosimilars and OTC. Our major therapeutic areas of focus are gastrointestinal, cardiovascular, diabetology, oncology, pain management and dermatology. Our major markets include – USA, India, Russia & CIS countries, China, Brazil and Europe. As a company with a history of deep science that has led to several industry firsts, we continue to plan ahead and invest in businesses of the future. As an early adopter of sustainability and ESG actions, we released our first Sustainability Report in 2004. Our current ESG goals aim to set the bar high in environmental stewardship; access and affordability for patients; diversity; and governance. For more information, log on to: . Disclaimer: This press release may include statements of future expectations and other forward-looking statements that are based on the management’s current views and assumptions and involve known or unknown risks and uncertainties that could cause actual results, performance or events to differ materially from those expressed or implied in such statements. In addition to statements which are forward-looking by reason of context, the words "may", "will", "should", "expects", "plans", "intends", "anticipates", "believes", "estimates", "predicts", "potential", or "continue" and similar expressions identify forward-looking statements. Actual results, performance or events may differ materially from those in such statements due to without limitation, (i) general economic conditions such as performance of financial markets, credit defaults, currency exchange rates, interest rates, persistency levels and frequency / severity of insured loss events, (ii) mortality and morbidity levels and trends, (iii) changing levels of competition and general competitive factors, (iv) changes in laws and regulations and in the policies of central banks and/or governments, (v) the impact of acquisitions or reorganization, including related integration issues, and (vi) the susceptibility of our industry and the markets addressed by our, and our customers’, products and services to economic downturns as a result of natural disasters, epidemics, pandemics or other widespread illness, including coronavirus (or COVID-19), and (vii) other risks and uncertainties identified in our public filings with the Securities and Exchange Commission, including those listed under the "Risk Factors" and "Forward-Looking Statements" sections of our Annual Report on Form 20-F for the year ended March 31, 2022. The company assumes no obligation to update any information contained herein.

▎Armstrong国家药监局药品审评中心官网2023年1月7日的公开信息显示，金宇博沃润泽生物技术有限公司的治疗用生物制品托珠单抗注射液申报上市，成为继百奥泰、珠海丽珠之后第三个在国内申报上市的托珠单抗生物类似药。托珠单抗（Tocilizumab）最早由日本中外制药株式会社联合开发，该抗体靶向IL-6受体（IL-6R），商品名雅美罗（Actemra）。托珠单抗采用哺乳动物细胞表达，特异性阻断人白介素6（IL-6）受体从而阻断促炎症反应的细胞因子风暴。在体内，IL-6结合IL-6R和gp130蛋白从而形成三聚体复合物。其中，IL-6R分为膜结合型和可溶型两种形式，但两种形式均引发经典的JAK-STAT信号通路。IL-6引起的下游信号通路，可以产生局部和全身性的炎症反应。因此，对IL-6信号通路的阻断，可以产生“广谱”的临床获益。图片来源-2020 nature review托珠单抗先后于2005、2009、2010、2013年在日本、欧洲、美国和中国获批上市。适应症包括成人类风湿关节炎、多关节幼年特发性关节炎、巨细胞关节炎、全身型幼年特发性关节炎、细胞因子释放综合征。在国内获批的适应症为成人类风湿关节炎、全身型幼年特发性关节炎和细胞因子释放综合征。鉴于托珠单抗在细胞因子风暴控制中的关键作用，并且COVID-19的重症患者的细胞因子风暴造成的炎症反应是造成死亡率的重要因素，在新冠爆发后，国内外启动了多个托珠单抗治疗新冠的临床研究。2022年12月22日，FDA批准了罗氏的托珠单抗原研药用于治疗住院成年患者的新型冠状病毒肺炎。2023年1月3日在顶级医学期刊《美国医学会杂志》（JAMA）上发表的研究数据进一步证明了托珠单抗在危重型新型冠状病毒肺炎治疗中的关键作用。该研究基于4869名重症患者的180天随访结果证明IL-6受体拮抗剂（tocilizumab, sarilumab）可提高重症新型冠状病毒感染者6个月生存率的概率大于99.9%，抗血小板药物提高6个月生存率的概率为95%，研究涉及的其他疗法要么无效，要么可能对患者有害。过去几年，托珠单抗在全球的的销售额均接近或者超过40亿美金。原研药没有放弃创新的脚步，2022年11月21日，中国国家药监局药品审评中心（CDE）官网公示显示，罗氏（Roche）递交了托珠单抗皮下注射液的上市申请，并获得受理。国内研发方面，百奥泰、丽珠集团、金宇博沃润泽、海正药业、均有布局托珠单抗生物仿制药的研发。其中，百奥泰进展最快，于2021年11月8日获NDA受理。丽珠集团紧随其后，于2021年12月22日获NDA受理。未来，随着其适应症的逐渐拓展，托珠单抗的竞争将变得更加激烈。2021年4月，百奥泰将托珠单抗类似药国际化权益授权给Biogen，后者支付3000万美元预付款、9000万美元里程碑金额，以及两位数比例的销售分成。可见托珠单抗类似药的商业价值受到业界认可。公开数据显示，金宇博沃润泽生物技术有限公司是上市公司-金宇生物（生物股份）的全资子公司，目前的产业化基地落地江苏省丹阳市。此次申报上市的托珠单抗生物仿制药，临床批件转让自迈博太科药业。预计在产业化基地建成前，双方将基于MAH指导原则进行生产和商业化销售。