CVnCoV Vaccine (CureVac)

一场旷日持久的mRNA疫苗专利纷争，最终以和解的方式落幕。  日前，德国生物科技公司CureVac宣布与辉瑞和BioNTech达成协议，解决并终止关于mRNA相关疫苗的所有未决专利纠纷。根据协议条款，CureVac和葛兰素史克（GSK）将获得总计7.4亿美元的付款，以及该疫苗在美国未来销售收益的个位数百分比特许权使用费。此外，CureVac还将从GSK获得5000万美元的付款，用于货币化部分美国产品许可费。 同日，GSK也宣布从这项和解中获得3.7亿美元的预付款，其中3.2亿美元以现金支付，其余部分正在通过修改其与CureVac的协议来实现这一价值。此外，GSK还将获得BioNTech和辉瑞在美国销售的mRNA疫苗产品的1%特许权使用费。 值得一提的是，此次和解正值BioNTech寻求以约12.5亿美元收购竞争对手CureVac之际。如果BioNTech成功完成对CureVac的收购，二者在美国以外的mRNA专利诉讼也将得到解决，同时GSK还将有权获得额外的1.3亿美元现金和在美国以外销售上述mRNA疫苗产品的1%特许权使用费。 强强联手却失市场先机 彼时，Moderna和辉瑞/BioNTech都在快速推进mRNA疫苗研发，GSK作为全球疫苗市场的重要玩家也出手跟进，在2020年7月与德国mRNA先驱企业CureVac达成战略合作。 根据协议，双方将制造和商业化多达五种基于mRNA的疫苗和针对传染病病原体的单克隆抗体（mAbs）。与此同时，GSK还对CureVac进行1.3亿英镑的股权投资，并预付1.04亿英镑。 GSK认为，CureVac在mRNA技术方面的领导地位及其mRNA制造能力，补充了GSK在疫苗方面现有的科学领导地位，包括GSK的自我扩增mRNA（SAM）疫苗技术平台；进一步巩固了GSK在mAbs创新方面不断增长的能力，与其对免疫学科学的研发重点保持一致。GSK判断，基于mRNA的疫苗和治疗技术有望在未来发挥重要作用。 随着Moderna和辉瑞/BioNTech的mRNA疫苗在2020年底获得紧急使用许可，这对本是强强联手的组合失去了市场先机，但二者并未放弃。 2021年2月，GSK和CureVac宣布新增一项1.5亿欧元的合作，共同开发下一代mRNA疫苗。该疫苗将具有多价方法的潜力，以解决一种疫苗中的多种新兴变体。彼时，GSK还计划在2021年支持生产多达1亿剂CureVac的CVnCoV，但最终也因有效率问题而搁浅。 2024年7月，CureVac做出一项重大战略调整，将其剩余的项目出售给合作伙伴GSK，全面转向肿瘤领域。CureVac获得4亿欧元的预付款，以及高达10.5亿欧元的里程碑付款。 尽管在研产品折戟，但CureVac却牢牢掌握着关于非修饰mRNA序列设计和脂质纳米颗粒（LNP）递送的基础专利。从2022年起，CureVac在美国、德国、荷兰等多地对辉瑞/BioNTech提起专利侵权诉讼，指控其mRNA疫苗使用了CureVac的专有技术。 今年3月，欧洲专利局（EPO）作出一项关键裁决，维持了CureVac一项核心专利的有效性。这项名为EP 3 708 668 B1的专利涵盖了一项基础发明，对提高mRNA药物编码蛋白质的表达效果至关重要。CureVac彼时的股价还因此飙升11%，而BioNTech则面临专利侵权风险。 或是为了解决后顾之忧，今年6月，BioNTech宣布以12.5亿美元全股票交易收购CureVac，结束这两家mRNA先驱长达20年的竞争关系。BioNTech认为，此次收购CureVac补充了其在mRNA设计、管理配方和mRNA制造方面的能力和已拥有的技术，有助于加强基于mRNA的癌症免疫疗法候选药物的研发，是其实施肿瘤学战略的另一个重要里程碑。 这次收购要约显然触及到GSK的利益，其强硬介入也是此次和解能够达成的主要原因。面对昔日合作伙伴CureVac的倒戈相向，GSK的无奈显露无遗。但作为这次纠纷和解的关键受益者，GSK收获了实打实的现金收入和未来相关产品1%的销售分成，也并非是这场博弈棋局的完全输家。 专利纠纷直指创新博弈 在相关mRNA疫苗问世后，全球掀起了一场“专利混战”，谁是这类产品专利权的真正拥有者，这不仅是一个科学问题，更是一个价值百亿美元的商业问题。 值得注意的是，尽管GSK与CureVac的利益瓜葛暂时告一段落，但其同辉瑞和BioNTech的专利纠纷仍在继续。GSK表示，该和解协议不会影响其在美国和欧洲对辉瑞和BioNTech的专利执行，针对二者侵犯相关专利的诉讼仍将继续。 据报道，GSK于2024年4月对辉瑞与BioNTech提起的另一起诉讼仍在进行中。该案涉及5项mRNA疫苗技术专利，GSK称这些专利源于前十年的研究，2015年随诺华全球疫苗业务大部分被其收购而归为己有。GSK称辉瑞与BioNTech从这些侵权中获得了数十亿美元的收入并继续收益，而从未获得许可。GSK彼时表态，愿在商业合理条件下授权专利，以保障疫苗可及性。 GSK的专利战矛头并非只对准辉瑞与BioNTech。2024年10月，GSK也对Moderna提起了两起专利诉讼，指控Moderna侵犯了其批准的mRNA疫苗的多项专利。诉讼特别涉及Moderna的Spikevax，以及彼时刚获批准的呼吸道合胞病毒(RSV) mRNA注射产品。GSK声称Moderna未经许可就违反了其拥有知识产权的产品，同样表态授权意愿。 有机构预测，mRNA疫苗和治疗药物年销售额预计到2029年将高达1189亿美元，预测期内（2024-2029年）复合年增长率为16.80%。在业内看来，GSK的一系列专利诉讼旨在巩固其在mRNA领域的话语权，弥补传统疫苗技术劣势。 今年上半年，疫苗业务为GSK贡献了41.86亿英镑（54.43亿美元）的营收，同比仅增长1%。具体来看，GSK相关产品的销售业绩依旧疲软：带状疱疹疫苗Shingrix销售额为17.20亿英镑（约22.36亿美元），同比下降了1%；仅脑膜炎疫苗Meningitis（包括Bexsero和Menveo）实现了两位数增长，报告期内斩获7.29亿英镑（约9.47亿美元）的销售额，同比增长21%。 此外，GSK的RSV疫苗Arexvy的销售额为1.44亿英镑（约1.87亿美元），同比下降了39%。据悉，这一产品在美国遭遇了不利的市场环境影响，导致销售表现不如预期。 业界普遍认为，mRNA疫苗领域的专利纠纷反映了这一颠覆性技术商业化后的深层矛盾，其核心是数百亿美元市场利益与创新归属权的博弈。当前，mRNA专利混战是技术爆发期的必然阵痛，长远来看，资本整合和司法裁决将重塑“专利地图”，头部企业控制力增强，下半年需关注GSK独立诉讼进展及Moderna在德国的上诉结果。 编辑：范晓艳 版式编辑：于成林 审校：马飞、张松 赛诺菲回应波立达停供！恒瑞、康方、君实等围战PCSK9 区别化学药，中成药、生物制品首发价怎么定？ 上半年医疗设备集采：从数据洞察市场“变”与“稳” www.yyjjb.com.cn 洞悉行业趋势 长按关注医药经济报 《中国处方药》 学术公众号 聚焦药学学术和循证研究 长按关注中国处方药 《医药经济报》 终端公众号 记录药品终端产经大事件 长按关注21世纪药店

作者：梁嘉维美工：何国红 罗真真排版：马超前期系列报道系统梳理了三优生物"九大超亿万级分子发现平台"中的五大核心系统：全人源单抗库、共同轻链抗体库、人源化单域抗体库、ADC分子库及环状多肽分子库，这些技术体系持续为全球生物医药研发提供多元化分子解决方案。2025年，三优生物战略级创新平台——mRNA mAb抗体开发平台即将正式发布。本文将从八大技术维度深度解构这一革命性平台：1）演进脉络：追踪mRNA技术从基础研究到治疗转化的里程碑突破；2）作用机理：解析mRNA疗法的分子作用路径与脂质纳米颗粒（LNP）递送系统；3）应用图谱：综述当前mRNA技术在治疗领域的核心应用场景；4）平台架构：首度披露三优mRNA创新平台的技术蓝图；5）建设进程：复盘平台从概念验证到系统搭建的进阶之路；6）全流程服务：从靶点序列解析到高效抗体生成的标准化路径；7）技术优势：融合mRNA快速表达与传统抗体精准筛选的双重优势；8）实证案例：mRNA mAb技术在难成药靶点领域的突破性实践。一、mRNA技术的发展历程mRNA技术的发展历程堪称现代医学史上的重要篇章（Fig 1）。1961年，科学家们首次揭示了mRNA的分子结构，这一发现为后续研究奠定了理论基础。1990年，Wolff团队通过小鼠模型成功实现mRNA编码蛋白的体内表达，标志着mRNA治疗应用的开端。1999年标志着mRNA技术迈入临床转化阶段，全球首个基于mRNA工程化树突细胞的抗肿瘤疫苗进入临床试验（NCT00004211）。2005年，Karikó团队对mRNA的核苷酸进行了修饰，有效减弱了mRNA的免疫原性问题，这项重大贡献最终荣获2023年诺贝尔生理学或医学奖。2013年CureVac公司完成首个人用mRNA疫苗（CV7201，NCT02241135）临床评估后，该技术在2020年COVID-19大流行中迎来爆发式发展——Moderna与辉瑞/BioNTech研发的mRNA疫苗以创纪录速度获紧急授权，重新定义了传染病防控范式。▲ Fig 1 mRNA与LNP历史发展的里程碑Hou, X., Zaks, T., Langer, R. et al. Lipid nanoparticles for mRNA delivery.二、mRNA分子机制与递送系统从分子机制层面，mRNA疗法通过工程化设计的合成mRNA指导宿主细胞合成靶蛋白。其技术框架包含三大核心要素：1) 序列优化：采用宿主偏好密码子提升翻译效率，引入修饰核苷（如Ψ）降低免疫原性，5'UTR/3'UTR结构增强mRNA稳定性；2) 体外转录(IVT)：完成设计的序列通过质粒DNA模板进行精密体外转录；3) 递送系统：为确保mRNA体内的有效递送，技术采用LNP作为载体系统。LNP由可电离脂质（Ionizable Lipid）、DSPC、胆固醇及PEG-lipid组成（Fig 2），通过微流控技术实现mRNA的高效包封，其表面经PEG修饰可延长体内循环时间。经注射给药后，LNP通过胞吞作用进入靶细胞，经内涵体逃逸机制将mRNA释放至细胞质。释放的mRNA被核糖体识别翻译，可产生各类治疗性蛋白（如抗体、酶替代蛋白）、疫苗抗原或基因编辑元件（Fig 3）。▲ Fig 2 脂质纳米微粒（LNP）结构与组成Albertsen,X., Kulkarni, J. et al. The role of lipid components in lipid nanoparticles for vaccines and gene therapy.▲ Fig 3 mRNA-LNP货载内吞与靶点蛋白形成Vishweshwaraiah, Y.& Dokholyan, N. mRNA vaccines for cancer immunotherapy.三、mRNA应用领域在应用拓展方面，mRNA技术已形成多维度治疗矩阵：1) 传染病疫苗：覆盖SARS-CoV-2、流感、HIV等病毒防控；2) 肿瘤疫苗：癌症疫苗训练免疫系统攻击肿瘤特定抗原，具体应用如靶向黑色素瘤、胶质母细胞瘤等肿瘤特异性抗原；3) 蛋白替代疗法：通过传递mRNA产生疾病缺陷蛋白，如纠正囊性纤维化等遗传性蛋白缺陷；4) 癌症免疫治疗：通过mRNA表达免疫检查点抑制剂或细胞因子；5) 基因编辑载体：mRNA传递表达CRISPR-Cas9或其他基因编辑工具进行精准DNA修改，如T淋巴细胞治疗剂，自体T细胞治疗药物，CD4+T细胞治疗药物，干细胞治疗等；6) 再生医学：通过mRNA表达生长因子刺激组织修复。四、创新平台：三优mRNA mAb分子产生系统三优mRNA mAb平台通过LNP递送系统实现突破性创新，基于mRNA精准编码特性，在宿主细胞内高效合成抗原蛋白，激活机体产生高亲和力保护性抗体，并依托三优平台整合的标准化筛选体系，完成从抗体发现到成药性优化的全流程开发，最终产出可大规模应用的抗体药物。该平台构建四大创新支柱（Fig 4）——第一，基于mRNA-LNP的抗原保真表达系统，确保靶蛋白完整三维结构呈现；第二，融合传统免疫策略与磁列阵技术，建立跨物种（小鼠/羊驼/兔）多维免疫文库，候选药物兼容性大大提升；第三，mRNA免疫路径兼具自我佐剂效应，将抗原制备周期从常规重组蛋白的4-6个月压缩至6周；第四，体内原位表达机制规避体外折叠偏差，解决了因体外表达错误折叠导致的抗体非特异性识别问题。基于此技术矩阵，平台突破多跨膜蛋白、高度糖基化修饰靶点、复杂结构抗原及神经毒性蛋白等传统"不可成药"靶点的开发壁垒，开辟了抗体药物研发的新范式。▲ Fig 4 三优生物mRNA mAb平台四大核心优势五、三优mRNA mAb抗体开发平台的发展历史▲ Fig 5 三优生物mRNA mAb平台发展历程三优生物在抗体技术领域的发展历程展现了持续创新的技术实力（Fig 5）。自2015年成功建立自主知识产权的噬菌体展示技术平台以来，公司不断突破技术边界，在2016至2023年间先后开发并商业化磁列阵小鼠、羊驼和兔免疫库系统，构建了完善的抗体发现技术体系，为后续mRNA mAb平台的研发奠定了坚实基础。2024年成为三优mRNA技术创新的重要里程碑。6月，三优生物前瞻性地提出mRNA mAb抗体开发平台概念；8月即快速启动体外平台建设，重点开发高纯度mRNA IVT试剂盒，并开展LNP递送系统的优化筛选工作。2024年的12月，三优开展mRNA mAb体内平台的建设。这些关键技术的突破为平台建设提供了重要支撑。进入2025年，平台建设取得突破性进展。5月，三优生物成功完成技术验证，使用40余个不同类别靶点验证了平台的可靠性，特别是针对跨膜蛋白等传统难成药靶点取得了显著成果。6月，这一革命性的mRNA mAb抗体开发平台将正式投入商业化运营，其核心优势包括：开发周期较传统方法缩短，抗体亲和力提升，难成药靶点项目成功率大幅提高。该平台的推出标志着抗体药物开发进入全新时代，预计到2025年底将完成100+靶点的技术验证，持续引领行业创新发展。六、三优mRNA制备平台服务流程三优生物为客户提供从基因序列到先导抗体的12周全流程定制开发服务（Fig 6）。▲ Fig 6 三优生物mRNA mAb平台服务流程阶段①-②：基于客户提供的靶点抗原或基因序列，三优首先采用AI驱动的密码子优化算法进行序列优化，确保获得高表达效率的mRNA序列。阶段③：优化后的序列将作为模板进行质粒构建和线性化处理，通过严格的内毒素检测和质量控制后，进入IVT环节。阶段④：IVT系统利用噬菌体的T7 RNA聚合酶，以DNA作为模板、NTP作为底物、RNA Cap1作为共转录帽子，实现mRNA的体外转录。其中，T7 RNA聚合酶能够实现高产量mRNA合成并支持超长转录本生产，其性能显著优于其他类型的RNA聚合酶。而经优化的5'端帽子结构可显著增强核糖体结合能力，使mRNA翻译效率提升达5-8倍。与未加帽RNA（15分钟内即被核酸酶降解）相比，合成加帽mRNA在胞内稳定性可维持48小时以上，其半衰期延长超180倍。生成mRNA转录子的稳定性与高产量为后续的LNP包装与体内免疫奠定坚实基础。阶段⑤：在LNP包装阶段，三优提供多样的LNP递送系统进行mRNA包装，如SM102、MC3、LP01，并通过多项关键质控指标确保产品质量，包括mRNA完整性分析、LNP粒径检测、包封率测定以及体外表达验证等。只有完全符合质量标准的mRNA制剂才会进入动物免疫环节。阶段⑥-⑦：抗体开发阶段整合了多物种免疫策略和高通量筛选技术平台。三优采用小鼠、羊驼、兔进行免疫，使用噬菌体展示技术进行抗体筛选，并通过真核表达系统完成抗体鉴定。获得先导抗体后，三优将根据客户需求开展系统的成药性评估，包括体外亲和力测定、特异性检测、生物学活性评价，以及体内药效学研究和药代动力学分析。此外，为确保抗体药物的可开发性，三优还提供完整的工艺开发服务，涵盖细胞株构建、上下游工艺开发、制剂工艺优化和分析方法建立等关键环节。整个流程采用严格的质量控制体系，结合AI算法优化和多技术平台协同，为客户提供高效、可靠的一站式抗体开发解决方案。七、平台核心优势01快速设计与产生mRNA序列可根据靶点抗原基因快速合成。此路径无需复杂蛋白表达和纯化流程，显著缩短抗体发现周期，详情见Fig 6。02多物种协同免疫增强免疫原性mRNA递送至细胞后直接表达抗原蛋白，模拟自然感染过程，更易激发强烈的体液免疫（B细胞抗体）和细胞免疫（T细胞应答）。而mRNA凭借其自身佐剂效应可在体内显著增强免疫原性，无需额外添加佐剂成分。除具备成本优势外，mRNA免疫周期仅需6周即可达到高效保护效果，较传统免疫开发路径缩短4周时间（表 1）。此外，mRNA免疫可与传统免疫路径结合，除了进行单独免疫外，亦可与蛋白、细胞、VLP等其他抗原交叉免疫，实现更好的免疫效果。这种并行免疫模式不仅缩短了开发周期，与三优现有的多物种免疫策略（小鼠、羊驼和兔）融合后，更使抗体多样性大幅提升，能够获得传统方法难以产生的特殊表位抗体。▼ 表1：mRNA免疫与其他传统免疫路径对比03颠覆性开发效率三优mRNA mAb平台将传统数月甚至数年的抗体开发周期缩短至12周（Fig 6）。1周便可以实现从靶点到mRNA分子的体外转录，10周从mRNA分子到单克隆抗体产生。一体化解决方案覆盖了mRNA设计→抗原表达→抗体发现→生产优化全流程，并支持个性化定制，快速响应肿瘤新抗原等特殊需求（具体流程请回顾上一节）。04全面质量控制评价体系三优mRNA mAb抗体开发平台建立了严格的全流程质量控制体系，确保各环节产出质量符合最高标准。在mRNA制备环节，我们采用琼脂糖凝胶电泳（表2），严格控制如mRNA完整性（目标条带清晰无杂带且大小正确）、纯度（OD260/280比值2.0-2.3）的基本关键指标。同时亦可检测mRNA加帽效率、dsRNA与T7 RNA聚合酶残留量、内毒素等进阶项目。▼ 表2：mRNA分子体外转录质量标准LNP递送系统需通过多项核心检测方可放行（表3），包括粒径（POI ±20 nm，65 - 125 nm）、PDI值（<0.2）、包封率（>85%）、内毒素水平（<4EU/mg）等。这些严格的标准确保递送系统的安全性和有效性。▼ 表3：mRNA-LNP递送系统质量标准在抗体筛选阶段，我们实施多级质量评估体系：首先进行理化特性分析，包括SEC-HPLC纯度（>95%）、SDS-PAGE和质谱分子量确认；其次开展功能活性检测，涵盖亲和力（SPR/BLI分析）、特异性（交叉反应性实验）和生物学活性（细胞水平效价测定）；最后进行稳定性评估，考察不同条件下的质量变化趋势。这套完善的质量控制体系确保最终获得的候选分子具有优异的成药性潜力。八、CCR8靶点案例解析01抗CCR8抗体的研发背景CCR8作为A类G蛋白偶联受体，具有七次跨膜结构域，在肿瘤浸润性调节性T细胞（Tregs）中呈现高表达特征，同时也可在Th2细胞、固有淋巴细胞、皮肤驻留记忆T细胞、树突状细胞及嗜酸性粒细胞中检测到。在健康成人中，CCR8+ T细胞主要定位于皮肤组织，而在血液或肠道组织中极少存在。其内源性配体CCL1通过介导T细胞皮肤归巢、促进Treg存活及肿瘤趋化作用参与免疫调控。由于CCR8在肿瘤微环境Tregs中的选择性过表达，该靶点成为通过清除免疫抑制性细胞重塑抗肿瘤免疫的热门研究方向。02临床开发进展与作用机制全球目前有5款抗CCR8抗体处于临床开发阶段，其中包括针对非小细胞肺癌等实体瘤的II期候选药物LM-108——该抗体通过Fc段工程化改造增强抗体依赖性细胞毒性（ADCC效应）。现有抗CCR8疗法主要通过阻断CCL1与受体的结合，抑制Treg向肿瘤组织的募集及其免疫抑制功能，从而恢复机体抗肿瘤免疫应答。然而，传统抗体开发面临重大挑战：CCR8复杂的天然构象难以通过体外重组表达系统准确复现，导致抗原制备效率低下且易引发非特异性抗体应答。03mRNA技术的突破性应用基于mRNA的抗原生产技术为上述难题提供了创新解决方案：通过LNP递送编码CCR8的mRNA，可在体内直接表达具有完整跨膜结构及正确翻译后修饰的天然构象蛋白。这种"原位抗原生成"策略不仅能确保免疫系统识别真实表位，还可通过动态折叠过程呈现膜蛋白的天然构象多样性，从而筛选出针对"不可成药"GPCR靶点的高质量抗体。04抗CCR8抗体的开发流程本研究采用整合mRNA技术与多物种免疫策略开发抗CCR8抗体：1) 抗原制备：通过体外转录系统构建线性化CCR8 mRNA质粒模板，经LNP包封后实现体内靶向递送，确保跨膜蛋白的天然构象表达；2) 抗体库构建：采用CCR8 mRNA-LNP复合物免疫小鼠并建立免疫库；3) 噬菌体筛选：经过2-3轮严格噬菌体展示生物淘选，筛选获得CCR8特异性结合克隆；4) 功能验证：在真核表达系统中重组表达候选抗体并进行验证。05关键结果1) mRNA制备及质检利用三优自主开发的体外转录方法，成功转录出mRNA样品，并经过凝胶及CE检测显示纯度与分子大小均达到合格标准，检测dsRNA及T7 RNA聚合酶残留物含量均符合标准（表4 & Fig 7-9）。▼ 表4：CCR8-mRNA质检结果▲ Fig 7 凝胶电泳检测mRNA大小▲ Fig 8 CE检测mRNA纯度▲ Fig 9 ELISA检测dsRNA及T7 RNA聚合酶残留物含量2) LNP包装及质检采用行业顶尖的LNP包装方案，对mRNA进行LNP包装，并且有多种LNP可供选择。案例中的SM102-LNP包装后包装效率达到96.83%，粒径大小为78.56nM，符合交付标准（表5 & Fig 10）。▼ 表5：CCR8-mRNA-LNP质检结果▲ Fig 10 LNP粒径大小及Zeta电位检测结果3) huCCR8-mRNA-LNP小鼠免疫后抗CCR8抗体血清滴度分析采用三剂次小鼠免疫方案，通过huCCR8-mRNA-LNP免疫成功诱导高滴度抗CCR8抗体生成。血清学检测显示，免疫后血清可特异性结合CCR8过表达HEK293细胞（Fig 11），而野生型HEK293对照组未检测到显著结合信号（Fig 12），证实了免疫应答的靶点特异性。该结果从两方面验证了平台优势：① mRNA-LNP递送系统能够高效诱导功能性抗体产生；② 基于天然构象抗原的免疫策略可精准维持抗体-抗原表位识别的特异性，为靶向GPCR的抗体开发提供了关键实验证据。▲ Fig 11 huCCR8-mRNA-LNP不同给药途径诱导的小鼠血清抗体滴度分析通过静脉注射（IV）、肌肉注射（IM）及联合注射（IV+IM）三种途径对小鼠进行三剂次huCCR8-mRNA-LNP免疫后，采用流式细胞术定量分析血清抗体与huCCR8-FL-His-EGFP-HEK293-A2细胞的结合能力。▲ Fig 12 由huCCR8-mRNA-LNP或CCR8过表达细胞系小鼠免疫产生的抗CCR8抗体的非特异性结合分析通过静脉注射（IV）、肌肉注射（IM）、联合注射（IV+IM）及腹腔注射（IP）四种途径对小鼠进行三剂次huCCR8-mRNA-LNP或huCCR8-HEK293细胞系免疫后，采用流式细胞术定量分析血清抗体与HEK292细胞的结合能力。九、总结展望作为分子医学领域的革命性技术，mRNA正重塑治疗格局。而三优生物的mRNA mAb平台为抗体发现提供与传统免疫路径兼容的全新解决方案，并通过结合智能化技术，不断拓展mRNA的应用，开创人类抗体产生的新高度。SANYOU Bio Solution for mRNA-based Molecular Generation SystemPrevious series reports have systematically outlined five core systems among Sanyou Bio's "Nine artificial intelligence hundred-trillion-level molecule library": the fully human antibody library, common light chain antibody library, humanized single-domain antibody library, ADC molecule library, and cyclic peptide molecule library. These technological systems continue to provide diversified molecular solutions for global biopharmaceutical R&D. In 2025, a new Sanyou Bio's strategic innovation platform—the mRNA mAb Antibody discovery Platform—will be officially launched. This article will deconstruct this revolutionary platform from eight technical dimensions:1. Evolutionary Trajectory: Tracing milestone breakthroughs in mRNA technology from basic research to therapeutic translation.2. Mechanism of Action: Decoding the molecular pathways of mRNA therapeutics and the lipid nanoparticle (LNP) delivery system.3. Application Landscape: Summarizing core application scenarios of mRNA technology in therapeutics.4. Platform Architecture: First disclosure of Sanyou's mRNA innovation platform blueprint.5. Development Process: Reviewing the platform's journey from concept validation to systematic construction.6. End-to-End Services: Standardized workflow from target sequence analysis to high-efficiency antibody generation.7. Technical Advantages: Combining the dual benefits of rapid mRNA expression and traditional antibody precision screening.8. Case Studies: Breakthrough practices of mRNA mAb technology in undruggable targets.1. The Evolution of mRNA TechnologyThe development of mRNA technology is a pivotal chapter in modern medical history (Fig 1). In 1961, scientists first revealed the molecular structure of mRNA, laying the theoretical foundation for subsequent research. In 1990, Wolff's team achieved in vivo expression of mRNA-encoded proteins in mouse models, marking the dawn of mRNA therapeutic applications.In 1999, mRNA technology entered clinical translation, with the world's first mRNA-engineered dendritic cell-based anti-tumor vaccine entering clinical trials (NCT00004211). In 2005, Karikó's team modified mRNA nucleotides, effectively mitigating mRNA immunogenicity—a breakthrough that earned the 2023 Nobel Prize in Physiology or Medicine.After CureVac completed the first human mRNA vaccine (CV7201, NCT02241135) clinical evaluation in 2013, the technology experienced explosive growth during the 2020 COVID-19 pandemic. Moderna and Pfizer/BioNTech's mRNA vaccines received emergency authorization at record speed, redefining infectious disease prevention paradigms.▲ Fig 1 Milestones in mRNA and LNP Development.Hou, X., Zaks, T., Langer, R. et al. Lipid nanoparticles for mRNA delivery.2. Molecular Mechanism and Delivery System of mRNAAt the molecular level, mRNA therapeutics utilize engineered synthetic mRNA to instruct host cells to produce target proteins. The technical framework comprises three core elements:1. Sequence Optimization: Host-preferred codons enhance translation efficiency; modified nucleotides (e.g., Ψ) reduce immunogenicity; 5'UTR/3'UTR structures improve mRNA stability.2. In Vitro Transcription (IVT): Designed sequences undergo precise IVT using plasmid DNA templates.3. Delivery System: LNPs, composed of ionizable lipids, DSPC, cholesterol, and PEG-lipid (Fig 2), ensure efficient mRNA delivery. LNPs encapsulate mRNA via microfluidics, with PEGylation extending circulation time. Post-injection, LNPs enter target cells via endocytosis, releasing mRNA into the cytoplasm through endosomal escape. The mRNA is then translated by ribosomes to produce therapeutic proteins (e.g., antibodies, enzymes), vaccine antigens, or gene-editing components (Fig 3).▲ Fig 2 Structure and Composition of Lipid Nanoparticles (LNPs).Albertsen,X., Kulkarni, J. et al. The role of lipid components in lipid nanoparticles for vaccines and gene therapy.▲ Fig 3 mRNA-LNP Endocytosis and Target Protein Formation.Vishweshwaraiah, Y.& Dokholyan, N. mRNA vaccines for cancer immunotherapy.3. Applications of mRNA TechnologymRNA technology has established a multi-dimensional therapeutic matrix:1) Infectious Disease Vaccines: Targeting SARS-CoV-2, influenza, HIV, etc.2) Cancer Vaccines: Training the immune system to attack tumor-specific antigens (e.g., melanoma, glioblastoma).3) Protein Replacement Therapy: Correcting genetic protein deficiencies (e.g., cystic fibrosis).4) Cancer Immunotherapy: Expressing immune checkpoint inhibitors or cytokines via mRNA.5) Gene Editing Vectors: Delivering CRISPR-Cas9 or other tools for precise DNA modification (e.g., T-cell therapies, stem cell treatments).6) Regenerative Medicine: Stimulating tissue repair via mRNA-expressed growth factors.4. Innovation Platform: Sanyou’s mRNA mAb Molecular Generation SystemSanyou’s mRNA mAb platform leverages LNP delivery for breakthrough innovation. By precisely encoding antigens in host cells, it efficiently synthesizes target proteins, elicits high-affinity antibody production, and finally integrates standardized screening to streamline antibody discovery and optimization.The platform features four pillars (Fig 4):1. mRNA-LNP Antigen Fidelity System: Ensures native 3D protein structures.2. Multi-Species Immunization: Combines traditional strategies with magnetic array technology for diverse antibody libraries (mouse/alpaca/rabbit).3. Rapid Antigen Production: mRNA’s self-adjuvant effect shortens the production cycle from 4-6 months to 6 weeks.4. In Situ Expression: Avoids misfolding issues seen in vitro, enabling targeting of "undruggable" proteins (e.g., multi-transmembrane, glycosylated, or neurotoxic targets).▲ Fig 4 Four Core Advantages of Sanyou’s mRNA mAb Platform.5. Development History of Sanyou’s mRNA mAb Platform▲ Fig 5 Development Timeline of Sanyou’s mRNA mAb Platform.Sanyou’s antibody technology milestones (Fig 5) began in 2015 with its proprietary phage display platform. From 2016–2023, magnetic array libraries (mouse/alpaca/rabbit) were commercialized, laying the groundwork for mRNA mAb development.2024 marked key advancements: June: Conceptualization of mRNA mAb platform. August: Launch of IVT system development and LNP optimization. December: In vivo platform construction. By 2025, the platform achieved validation across 40+ targets (including challenging transmembrane proteins) and will commence operations in June, reducing development cycles and boosting success rates for undruggable targets.6. Sanyou’s mRNA Preparation Service WorkflowA 12-week, end-to-end customized service (Fig 6) includes: ▲ Fig 6 Service Workflow of Sanyou’s mRNA mAb Platform.Stage ①-②：Based on the target sequence provided by the customer, Sanyou employs AI-driven codon optimization to enhances mRNA expression efficiency.Stage ③：Optimized sequence undergoes plasmid construction and linearization. Linearized templates will pass a series of stringent QC before IVT step.Stage ④：The IVT system utilizes T7 RNA polymerases of bacteriophage, DNA templates, NTP, RNA Cap 1 as co-translational capping to achieve in vitro transcription of mRNA. Among these, T7 RNA polymerases enables high-yield mRNA synthesis and supports ultra-long transcript production—outperforming other polymerases. In addition, an optimized 5′ cap enhances ribosome binding, significantly boosting mRNA translation efficiency. Unlike uncapped RNA (degraded in 15 minutes), synthetic capped mRNA remains stable for over 48 hours. The stability and high-quality of mRNA transcript products guarantee the efficiency of LNP packaging and animal immunization.Stage ⑤：Sanyou provides a variety of LNP delivery systems to encapsulate the mRNA transcripts, such as SM102, MC3, LP01. Only those reach the releasing criteria (size, integrity, encapsulation rate, expression validation) will continue as immunogens for the animal immunization.Stage ⑥-⑦：Sanyou integrates multi-species immunization (mouse/alpaca/rabbit), with advanced high-throughput screening technology platforms. The screening process utilizes phage display technology for efficient antibody selection, followed by thorough characterization through eukaryotic expression systems to verify functionality. Once lead antibody candidates are identified, Sanyou conducts systematic developability assessments tailored to specific client requirements. These evaluations encompass comprehensive in vitro testing including affinity measurements (using SPR/BLI techniques), specificity profiling, and biological activity assays, as well as in vivo studies involving pharmacodynamic efficacy testing and pharmacokinetic analysis.7. Platform Advantages01Rapid Design and ProductionmRNA synthesis bypasses protein expression/purification, shortening discovery cycles (Fig 6).02Enhanced ImmunogenicityThe mRNA delivery system enables direct antigen protein expression within host cells, effectively mimicking natural infection pathways to robustly activate both humoral immunity (B-cell antibody production) and cellular immunity (T-cell responses). Notably, mRNA's intrinsic adjuvant properties significantly enhance immunogenicity in vivo without requiring supplemental adjuvant components. Beyond its cost-efficiency advantages, this mRNA immunization approach achieves high protective efficacy within just 6 weeks - a 4-week reduction compared to conventional immunization protocols (Table 1).Furthermore, mRNA immunization demonstrates exceptional compatibility with traditional immunization methods. In addition to standalone administration, it can be strategically combined with protein-based, cellular, or VLP antigens in prime-boost regimens to achieve superior immune outcomes. This hybrid immunization paradigm not only accelerates development timelines but, when integrated with Sanyou's established multi-species immunization platform (mice/alpacas/rabbits), dramatically expands antibody diversity. The combined approach enables generation of antibodies targeting unique epitopes that are typically inaccessible through conventional methods, particularly for complex antigens like membrane proteins or heavily glycosylated targets.▼ Table 1：mRNA vs. Traditional Immunization.03Ultra-EfficiencySanyou's revolutionary mRNA mAb Platform has successfully compressed the traditional antibody development timeline from several months or even years down to just 12 weeks (Fig 6). Target-to-mRNA synthesis can be completed within merely 1 week via advanced in vitro transcription technology, followed by efficient monoclonal antibody generation in the subsequent 10 weeks. The platform delivers a fully integrated solution that seamlessly encompasses the entire antibody development continuum: from AI-powered mRNA design and in vivo antigen expression to comprehensive antibody discovery and production optimization. This end-to-end system not only maintains rigorous scientific standards but also offers exceptional flexibility for personalized customization, enabling rapid response to specialized requirements such as tumor neoantigen targeting and other urgent therapeutic needs (for detailed workflow, please refer to the previous section).04Comprehensive Quality Control SystemSanyou Bio's mRNA mAb platform implements a comprehensive quality control system throughout the entire antibody development pipeline to ensure consistently high-quality outputs. For mRNA preparation, we employ multiple analytical techniques including agarose gel electrophoresis to verify mRNA integrity (clear target bands of correct size without impurities) and purity (optimal OD260/280 ratios of 2.0-2.3), while also monitoring critical parameters such as capping efficiency, dsRNA contamination, residual T7 RNA polymerase, and endotoxin levels (Table 2). ▼ Table 2：mRNA IVT Quality Standards.Our LNP delivery systems must meet stringent release criteria including controlled particle size (65-125 nm ±20 nm), low polydispersity (PDI <0.2), high encapsulation efficiency (>85%), and minimal endotoxin content (<4 EU/mg) to ensure both safety and functionality (Table 3). ▼ Table 3: mRNA-LNP Quality Standards.During antibody screening, we conduct rigorous multi-dimensional evaluations: physicochemical characterization through SEC-HPLC (>95% purity), SDS-PAGE, and mass spectrometry; functional assessment of affinity (SPR/BLI), specificity (cross-reactivity), and biological activity (cell-based assays); plus comprehensive stability testing under various conditions. This meticulous quality framework at every stage - from mRNA synthesis to final antibody characterization - guarantees the development of therapeutic candidates with optimal properties and clinical potential, demonstrating our commitment to delivering reliable, high-performance antibody products.8. CCR8 mRNA mAb Case Study01BackgroundCCR8, a class A GPCR with seven transmembrane domains, is highly expressed on tumor-infiltrating Tregs but also present on Th2 cells, innate lymphoid cells, skin-resident memory T cells, dendritic cells, and eosinophils. In healthy adults, CCR8+ T cells are predominantly localized to the skin and are rarely detected in blood or intestinal tissues. Its primary ligand, CCL1, facilitates T-cell skin homing, Treg survival, and tumor chemotaxis. Due to its selective overexpression in tumor Tregs, CCR8 is an attractive therapeutic target for their depletion.02Clinical Development and Mechanism of ActionCurrently, five anti-CCR8 antibodies are in clinical development globally, including the Phase II candidate LM-108—an ADCC-enhanced monoclonal antibody being evaluated for solid tumors such as non-small cell lung cancer. These antibodies exert their therapeutic effect by blocking CCL1 binding, thereby disrupting Treg recruitment and immunosuppressive activity to restore antitumor immunity. However, conventional antibody development has been hampered by challenges in producing CCR8’s complex native conformation through in vitro methods.03Groundbreaking Application of mRNA TechnologymRNA-based antigen production offers a promising solution to this hurdle, enabling proper protein folding and presentation to generate high-quality, druggable antibodies for clinical translation.04Technical Route for CCR8 mRNA Preparation and Antibody DiscoveryThe study combines mRNA technology with multi-species immunization pathways for anti-CCR8 antibody development:a) Antigen preparation: a linearized plasmid for CCR8 mRNA was designed and synthesized, which subsequently underwent in vitro transcription, followed by LNP packaging for in vivo delivery.b) Antibody library construction: mouse immune antibody libraries was constructed by injecting the above antigen.c) Phage display biopanning: 2–3 rounds of panning to enrich CCR8-specific binders.d) Function verification: candidate antibodies were then validated in a eukaryotic expression system, yielding multiple lead molecules with confirmed in vitro activity against CCR8.05Key Results1) mRNA synthesis and quality analysisThrough our IVT technology developed in-house, we successfully generated mRNA samples that passed all quality tests. Gel electrophoresis and capillary electrophoresis (CE) confirmed the mRNA's purity and correct molecular size. The samples also met specifications for dsRNA contamination and T7 RNA polymerase residues (Table 4 & Fig 7-9).▼ Table 4: CCR8-mRNA QC Results.▲ Fig 7 The size of mRNA verified by agarose gel electrophoresis▲ Fig 8 mRNA purity tested by CE▲ Fig 9 dsRNA & T7 RNA polymerase residues tested by ELISA2) LNP packaging and quality analysisWe employed industry-leading LNP formulations for mRNA packaging, with multiple LNP options available. In this case, SM102-LNP demonstrated 96.83% encapsulation efficiency and 78.56nm particle size, meeting all release specifications (Table 5 & Fig 10).▼ Table 5：CCR8-mRNA-LNP QC Results▲ Fig 10 LNP size and zeta potential3) Anti-CCR8 antibody serum titer after huCCR8-mRNA-LNP immunizationmRNA immunization protocol employed a three-dose administration regimen to maximize immune response (Fig. 11). Serum analysis revealed robust anti-CCR8 antibody titers following huCCR8-mRNA-LNP immunization, demonstrating specific binding to CCR8-overexpressing HEK293 cells. No detectable binding was observed in wild-type HEK293 controls, confirming the immune response's specificity for CCR8 (Fig. 12). These results validate the huCCR8-mRNA-LNP platform's ability to generate target-specific antibodies while maintaining excellent antigen specificity. ▲ Fig 11 Mouse serum titer after 3 doses of huCCR8-mRNA-LNP immunization via three different routes: IV, IM, and IV+IM; the binding of mouse serum with huCCR8-FL-His-EGFP-HEK293-A2 cells was measured by FACS. IV, intravenous; IM, intramuscular; BMS-986340-CHO-K1S (P243254): reference antibody for huCCR8. ▲ Fig 12 Non-specific binding of anti-CCR8 antibody extracted from mice immunized with huCCR8-mRNA-LNP or CCR8 over-expressed cell line via four different routes: IV, IM, IV+IM, and IP; the binding of mouse serum with wild-type HEK293 cells was measured by FACS. IV, intravenous; IM, intramuscular; IP, intraperitoneal; BMS-986340-CHO-K1S (P243254): reference antibody for huCCR8.9. ConclusionAs a revolutionary technology, mRNA is reshaping therapeutics. 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