公开(公告)号：US09623115B2

公开(公告)日：2017-04-18

申请号：US13295647

申请日：2011-11-14

Applicant: Chien-Hsing Chang ,  David M. Goldenberg

Inventor： Chien-Hsing Chang ,  David M. Goldenberg

IPC: A61K47/48 ,  C07K16/28 ,  A61K51/08 ,  C07K16/30 ,  C07K16/40 ,  C07K16/24 ,  B82Y5/00 ,  B82Y10/00 ,  B82Y30/00 ,  C07K16/00 ,  C07K16/44 ,  C07K16/46 ,  A61K39/00

CPC classification number: C07K16/2887 ,  A61K47/60 ,  A61K47/65 ,  A61K47/6813 ,  A61K47/6853 ,  A61K47/6891 ,  A61K2039/505 ,  B82Y5/00 ,  B82Y10/00 ,  B82Y30/00 ,  C07K16/00 ,  C07K16/2803 ,  C07K16/283 ,  C07K16/2833 ,  C07K16/2863 ,  C07K16/3007 ,  C07K16/44 ,  C07K16/468 ,  C07K2317/31 ,  C07K2317/55 ,  C07K2317/73 ,  C07K2317/732 ,  C07K2317/734 ,  C07K2319/00 ,  C07K2319/30 ,  C07K2319/70

Abstract: Disclosed herein are compositions and methods of use of dock and lock (DNL) complexes comprising a first antibody or fragment that binds to a stem cell antigen and a second antibody or fragment thereof that binds to an antigen on a diseased or damaged tissue or organ. The DNL complexes are of use for targeting stem cells to diseased or damaged organs or tissues and may be used to treat a variety of diseases or conditions that are responsive to stem cell therapy.

公开(公告)号：US08932593B2

公开(公告)日：2015-01-13

申请号：US13419614

申请日：2012-03-14

Applicant: Chien-Hsing Chang ,  David M. Goldenberg ,  William J. McBride ,  Edmund A. Rossi

Inventor： Chien-Hsing Chang ,  David M. Goldenberg ,  William J. McBride ,  Edmund A. Rossi

IPC: C07K16/28 ,  C07K14/475 ,  C07K14/52 ,  C07K14/54 ,  C07K14/505 ,  C07K16/40 ,  B82Y5/00 ,  B82Y10/00 ,  C07K16/30

CPC classification number: C12N9/12 ,  B82Y5/00 ,  B82Y10/00 ,  C07K16/18 ,  C07K16/2803 ,  C07K16/2887 ,  C07K16/3007 ,  C07K16/3092 ,  C07K2317/31 ,  C07K2317/55 ,  C07K2319/00 ,  C07K2319/70 ,  C12Y207/11011

Abstract: The present invention concerns methods and compositions for stably tethered structures of defined compositions, which may have multiple functionalities and/or binding specificities. Particular embodiments concern homodimers comprising monomers that contain a dimerization and docking domain attached to a precursor. The precursors may be virtually any molecule or structure, such as antibodies, antibody fragments, antibody analogs or mimetics, aptamers, binding peptides, fragments of binding proteins, known ligands for proteins or other molecules, enzymes, detectable labels or tags, therapeutic agents, toxins, pharmaceuticals, cytokines, interleukins, interferons, radioisotopes, proteins, peptides, peptide mimetics, polynucleotides, RNAi, oligosaccharides, natural or synthetic polymeric substances, nanoparticles, quantum dots, organic or inorganic compounds, etc. Other embodiments concern tetramers comprising a first and second homodimer, which may be identical or different. The disclosed methods and compositions provide a facile and general way to obtain homodimers, homotetramers and heterotetramers of virtually any functionality and/or binding specificity.

公开(公告)号：US08865176B2

公开(公告)日：2014-10-21

申请号：US13549906

申请日：2012-07-16

Applicant: Chien-Hsing Chang ,  David M. Goldenberg ,  William J. McBride ,  Edmund A. Rossi

Inventor： Chien-Hsing Chang ,  David M. Goldenberg ,  William J. McBride ,  Edmund A. Rossi

IPC: C07K14/71 ,  C07K16/00 ,  C07K16/26 ,  C07K16/28 ,  A61K47/48 ,  A61K31/00 ,  B82Y10/00 ,  B82Y5/00 ,  C07K16/30 ,  G01N33/58 ,  B82Y15/00 ,  C12N9/96 ,  G01N33/543 ,  A61K38/00 ,  A61K39/00

CPC classification number: A61K47/48438 ,  A61K31/00 ,  A61K38/00 ,  A61K47/48415 ,  A61K47/48484 ,  A61K47/485 ,  A61K47/48538 ,  A61K47/48546 ,  A61K47/48561 ,  A61K47/48569 ,  A61K47/6811 ,  A61K47/6817 ,  A61K47/6829 ,  A61K47/6833 ,  A61K47/6843 ,  A61K47/6845 ,  A61K47/6849 ,  A61K47/6851 ,  A61K2039/505 ,  B82Y5/00 ,  B82Y10/00 ,  B82Y15/00 ,  C07K14/475 ,  C07K16/18 ,  C07K16/22 ,  C07K16/28 ,  C07K16/2803 ,  C07K16/2833 ,  C07K16/2851 ,  C07K16/2863 ,  C07K16/2881 ,  C07K16/2887 ,  C07K16/2896 ,  C07K16/3007 ,  C07K16/3092 ,  C07K16/32 ,  C07K2317/31 ,  C07K2317/55 ,  C07K2317/76 ,  C07K2319/30 ,  C07K2319/35 ,  C07K2319/70 ,  C07K2319/74 ,  C12N9/12 ,  C12N9/22 ,  C12N9/96 ,  C12Y207/11001 ,  G01N33/54353 ,  G01N33/588 ,  Y02A50/491

Abstract: The present invention concerns methods and compositions for making and using bioactive assemblies of defined compositions, which may have multiple functionalities and/or binding specificities. In particular embodiments, the bioactive assembly is formed using dock-and-lock (DNL) methodology, which takes advantage of the specific binding interaction between dimerization and docking domains (DDD) and anchoring domains (AD) to form the assembly. In various embodiments, one or more effectors may be attached to a DDD or AD sequence. Complementary AD or DDD sequences may be attached to an adaptor module that forms the core of the bioactive assembly, allowing formation of the assembly through the specific DDD/AD binding interactions. Such assemblies may be attached to a wide variety of effector moieties for treatment, detection and/or diagnosis of a disease, pathogen infection or other medical or veterinary condition.

Abstract translation: 本发明涉及用于制备和使用具有多种功能和/或结合特异性的限定组合物的生物活性组合物的方法和组合物。 在特定的实施方案中，生物活性组件使用对接和锁定（DNL）方法形成，其利用二聚化和对接结构域（DDD）和锚定结构域（AD）之间的特异性结合相互作用形成组件。 在各种实施方案中，一个或多个效应物可以连接到DDD或AD序列。 互补AD或DDD序列可以连接到形成生物活性组件核心的适配器模块，从而允许通过特定的DDD / AD结合相互作用形成组装体。 这些组合物可以连接到各种各样的效应部分，用于治疗，检测和/或诊断疾病，病原体感染或其他医学或兽医病症。

公开(公告)号：US20130177532A1

公开(公告)日：2013-07-11

申请号：US13589575

申请日：2012-08-20

Applicant: Chien-Hsing Chang ,  David M. Goldenberg ,  William J. McBride ,  Edmund A. Rossi

Inventor： Chien-Hsing Chang ,  David M. Goldenberg ,  William J. McBride ,  Edmund A. Rossi

IPC: A61K47/48 ,  A61K38/19 ,  A61K38/18 ,  A61K38/21

CPC classification number: A61K47/6835 ,  A61K38/1816 ,  A61K38/193 ,  A61K38/212 ,  A61K47/60 ,  A61K47/65 ,  A61K47/6853 ,  A61K51/1048 ,  B82Y5/00 ,  B82Y10/00 ,  C07K2319/00 ,  C07K2319/70

Abstract: The present invention concerns methods and compositions for forming PEGylated complexes of defined stoichiometry and structure. In preferred embodiments, the PEGylated complex is formed using dock-and-lock technology, by attaching a target agent to a DDD sequence and attaching a PEG moiety to an AD sequence and allowing the DDD sequence to bind to the AD sequence in a 2:1 stoichiometry, to form PEGylated complexes with two target agents and one PEG moiety. In alternative embodiments, the target agent may be attached to the AD sequence and the PEG to the DDD sequence to form PEGylated complexes with two PEG moieties and one target agent. In more preferred embodiments, the target agent may comprise any peptide or protein of physiologic or therapeutic activity. The PEGylated complexes exhibit a significantly slower rate of clearance when injected into a subject and are of use for treatment of a wide variety of diseases.

公开(公告)号：US08435540B2

公开(公告)日：2013-05-07

申请号：US13412816

申请日：2012-03-06

Applicant: Chien-Hsing Chang ,  David M. Goldenberg ,  William J. McBride ,  Edmund A. Rossi

Inventor： Chien-Hsing Chang ,  David M. Goldenberg ,  William J. McBride ,  Edmund A. Rossi

IPC: A61K38/21 ,  C07K17/08 ,  C07K14/52 ,  C07K14/56 ,  C07K14/565

CPC classification number: C12N9/1205 ,  A61K39/395 ,  A61K47/60 ,  A61K47/64 ,  B82Y10/00 ,  B82Y30/00 ,  C07K16/00 ,  C07K16/005 ,  C07K16/283 ,  C07K16/3007 ,  C07K16/468 ,  C07K2317/31 ,  C07K2317/55 ,  C07K2317/73 ,  C07K2319/00 ,  C07K2319/70 ,  Y02A50/386

Abstract: The present invention concerns methods and compositions for PEGylated complexes of defined stoichiometry and structure. Preferably, the PEGylated complex is formed using dock-and-lock technology, by attaching a therapeutic agent to a DDD sequence and a PEG moiety to an AD sequence, allowing the DDD sequence to bind to the AD sequence in a 2:1 stoichiometry, to form PEGylated complexes with two therapeutic agents and one PEG moiety. Alternatively, the therapeutic agent may be attached to the AD sequence and the PEG to the DDD sequence to form PEGylated complexes with two PEG moieties and one therapeutic agent. In more preferred embodiments, the therapeutic agent may comprise any peptide or protein of physiologic or therapeutic activity, preferably a cytokine, more preferably interferon-α2b. The PEGylated complexes exhibit a significantly slower rate of clearance when injected into a subject and are of use for treatment of a wide variety of diseases.

Abstract translation: 本发明涉及具有限定化学计量和结构的聚乙二醇化配合物的方法和组合物。 优选地，使用对接和锁定技术形成聚乙二醇化复合物，通过将治疗剂连接到AD序列和PEG部分到AD序列，允许DDD序列以2:1化学计量比结合AD序列， 以形成具有两个治疗剂和一个PEG部分的PEG化复合物。 或者，治疗剂可以连接到AD序列，并将PEG连接到DDD序列以形成具有两个PEG部分和一种治疗剂的聚乙二醇化复合物。 在更优选的实施方案中，治疗剂可以包含生理或治疗活性的任何肽或蛋白质，优选细胞因子，更优选干扰素-α2b。 当注射到受试者中时，聚乙二醇化复合物表现出明显较慢的清除率，并且可用于治疗各种各样的疾病。

公开(公告)号：US08349332B2

公开(公告)日：2013-01-08

申请号：US13086786

申请日：2011-04-14

Applicant: Chien-Hsing Chang ,  David M. Goldenberg ,  Edmund A. Rossi

Inventor： Chien-Hsing Chang ,  David M. Goldenberg ,  Edmund A. Rossi

IPC: A61K39/395 ,  A61K39/00 ,  A61K39/44 ,  C07K16/28 ,  C07K16/46 ,  C12P21/08

CPC classification number: A61K47/6807 ,  A61K39/3955 ,  A61K47/6811 ,  A61K47/6813 ,  A61K47/6815 ,  A61K47/6849 ,  A61K47/6851 ,  A61K47/6867 ,  A61K47/6885 ,  A61K51/1027 ,  A61K51/1045 ,  C07K14/47 ,  C07K2317/31 ,  C07K2317/732 ,  C07K2317/734 ,  C07K2319/33 ,  C07K2319/70 ,  C07K2319/74

Abstract: Disclosed herein are compositions and methods of use comprising hexavalent DNL complexes. Preferably, the complexes comprise anti-CD20 and/or anti-CD22 antibodies or fragments thereof. More preferably, the anti-CD20 antibody is veltuzumab and the anti-CD22 antibody is epratuzumab. Administration of the subject hexavalent DNL complexes induces apoptosis and cell death of target cells in diseases such as B-cell lymphomas or leukemias, autoimmune disease or immune dysfunction disease. In most preferred embodiments, the DNL complexes increase levels of phosphorylated p38 and PTEN, decrease levels of phosphorylated Lyn, Akt, ERK, IKKα/β and IκBα, increase expression of RKIP and Bax and decrease expression of Mcl-1, Bcl-xL, Bcl-2, and phospho-BAD in target cells. The subject DNL complexes show EC50 values for inhibiting tumor cell growth in the low nanomolar or even sub-nanomolar concentration range.

Abstract translation: 本文公开了包含六价DNL复合物的组合物和使用方法。 优选地，复合物包含抗CD20和/或抗-CD22抗体或其片段。 更优选地，抗CD20抗体是veltuzumab，抗CD22抗体是依帕珠珠单抗。 主题六价DNL复合物的给药诱导疾病如B细胞淋巴瘤或白血病，自身免疫疾病或免疫功能障碍疾病中靶细胞的凋亡和细胞死亡。 在最优选的实施方案中，DNL复合物增加磷酸化p38和PTEN的水平，降低磷酸化Lyn，Akt，ERK，IKKα/ bgr的水平; 和Bα，增加RKIP和Bax的表达，并降低靶细胞中Mcl-1，Bcl-xL，Bcl-2和phospho-BAD的表达。 受试者DNL复合物显示在低纳摩尔或甚至亚纳摩尔浓度范围内抑制肿瘤细胞生长的EC 50值。

公开(公告)号：US08153433B1

公开(公告)日：2012-04-10

申请号：US13287395

申请日：2011-11-02

Applicant: David M. Goldenberg ,  Zhengxing Qu ,  Chien-Hsing Chang ,  Edmund A. Rossi ,  Jeng-Dar Yang ,  Diane Nordstrom

Inventor： David M. Goldenberg ,  Zhengxing Qu ,  Chien-Hsing Chang ,  Edmund A. Rossi ,  Jeng-Dar Yang ,  Diane Nordstrom

IPC: C12N5/10

CPC classification number: C07K16/2896 ,  A61K2039/505 ,  C07K14/005 ,  C07K14/4747 ,  C07K16/00 ,  C07K16/18 ,  C07K16/2803 ,  C07K16/3007 ,  C07K2317/24 ,  C07K2317/31 ,  C12N5/0694 ,  C12N15/85 ,  C12N15/86 ,  C12N2500/90 ,  C12N2501/48 ,  C12N2510/02 ,  C12N2710/20022 ,  C12N2740/10043 ,  C12P21/02

Abstract: Disclosed are compositions and methods for increasing the longevity of a cell culture and permitting the increased production of proteins, preferably recombinant proteins, such as antibodies, peptides, enzymes, growth factors, interleukins, interferons, hamiones, and vaccines. Cells transfected with an apoptosis-inhibiting gene or vector, such as a triple mutant Bcl-2 gene, can survive longer in culture, resulting in extension of the state and yield of protein biosynthesis. Such transfected cells exhibit maximal cell densities that equal or exceed the maximal density achieved by the parent cell lines. Transfected cells can also be pre-adapted for growth in serum-free medium, greatly decreasing the time required to obtain protein production in serum-free medium. In certain methods, the pre-adapted cells can be used for protein production following transformation under serum-free conditions. The method preferably involves eukaryotic cells, more preferably mammalian cells.

Abstract translation: 公开了用于增加细胞培养物的寿命并允许增加蛋白质生产的组合物和方法，优选重组蛋白质，例如抗体，肽，酶，生长因子，白细胞介素，干扰素，hamiones和疫苗。 用凋亡抑制基因或载体如三重突变体Bcl-2基因转染的细胞可以在培养物中更长时间地存活，导致蛋白质生物合成的状态和产量的延长。 这样的转染细胞表现出最大的细胞密度，其等于或超过由亲本细胞系实现的最大密度。 转染的细胞也可以预先适应于无血清培养基中生长，大大减少了在无血清培养基中获得蛋白质产生所需的时间。 在某些方法中，预先调节的细胞可以在无血清条件下转化后用于蛋白质生产。 该方法优选涉及真核细胞，更优选哺乳动物细胞。

公开(公告)号：US20120009149A1

公开(公告)日：2012-01-12

申请号：US13219940

申请日：2011-08-29

Applicant: Chien-Hsing Chang ,  David M. Goldenberg ,  Edmund A. Rossi

Inventor： Chien-Hsing Chang ,  David M. Goldenberg ,  Edmund A. Rossi

IPC: A61K38/21 ,  A61K38/20 ,  A61P37/02 ,  A61K39/395

CPC classification number: A61K47/48423 ,  A61K39/395 ,  A61K47/65 ,  A61K47/6813 ,  A61K47/6853 ,  A61K51/1048 ,  A61K2039/505 ,  B82Y5/00 ,  B82Y10/00 ,  B82Y30/00 ,  C07K16/2803 ,  C07K16/283 ,  C07K16/2833 ,  C07K16/2863 ,  C07K16/2887 ,  C07K16/3007 ,  C07K16/44 ,  C07K16/468 ,  C07K2317/31 ,  C07K2317/55 ,  C07K2317/732 ,  C07K2317/734 ,  C07K2319/00 ,  C07K2319/30 ,  C07K2319/70

Abstract: The present invention concerns methods and compositions for forming cytokine-antibody complexes using dock-and-lock technology. In preferred embodiments, the cytokine-MAb DNL complex comprises an IgG antibody attached to two AD (anchor domain) moieties and four cytokines, each attached to a DDD (docking and dimerization domain) moiety. The DDD moieties form dimers that bind to the AD moieties, resulting in a 2:1 ratio of DDD to AD. The cytokine-MAb complex exhibits improved pharmacokinetics, with a significantly longer serum half-life than either naked cytokine or PEGylated cytokine. The cytokine-MAb complex also exhibits significantly improved in vitro and in vivo efficacy compared to cytokine alone, antibody alone, unconjugated cytokine plus antibody or cytokine-MAb DNL complexes incorporating an irrelevant antibody. In more preferred embodiment the cytokine is G-CSF, erythropoietin or INF-α2b.

Abstract translation: 本发明涉及使用基座和锁定技术形成细胞因子 - 抗体复合物的方法和组合物。 在优选的实施方案中，细胞因子-MAb DNL复合物包含连接到两个AD（锚结构域）部分的IgG抗体和四个细胞因子，每个连接到DDD（对接和二聚化结构域）部分。 DDD部分形成与AD部分结合的二聚体，导致DDD与AD的比例为2:1。 细胞因子-MAb复合物表现出改善的药代动力学，其具有比裸细胞因子或聚乙二醇化细胞因子显着更长的血清半衰期。 细胞因子-MAb复合物与单独的细胞因子，单独的抗体，非结合的细胞因子+抗体或掺入不相关抗体的细胞因子-MAb DNL复合物相比，也显示出显着改善的体外和体内功效。 在更优选的实施方案中，细胞因子是G-CSF，红细胞生成素或INF-α2b。

公开(公告)号：US20110243841A1

公开(公告)日：2011-10-06

申请号：US13074351

申请日：2011-03-29

Applicant: Chien-Hsing Chang ,  David M. Goldenberg

Inventor： Chien-Hsing Chang ,  David M. Goldenberg

IPC: A61K39/395 ,  A61K51/00 ,  A61K38/19 ,  A61K38/21 ,  A61K38/20 ,  C07K16/28 ,  A61P37/06

CPC classification number: C07K16/2833 ,  A61K31/4965 ,  A61K47/484 ,  A61K47/48423 ,  A61K47/4843 ,  A61K47/6807 ,  A61K47/6813 ,  A61K47/6815 ,  A61K47/6849 ,  A61K47/6881 ,  A61K47/6885 ,  C07K16/18 ,  C07K16/2803 ,  C07K16/2851 ,  C07K16/2863 ,  C07K16/2887 ,  C07K16/30 ,  C07K16/3007 ,  C07K16/3092 ,  C07K16/44 ,  C07K2317/24 ,  C07K2317/31 ,  C07K2317/35 ,  C07K2317/51 ,  C07K2317/52 ,  C07K2317/522 ,  C07K2317/55 ,  C07K2317/73 ,  C07K2317/734 ,  C07K2317/75 ,  C07K2317/77 ,  C07K2319/70 ,  C12N15/113 ,  C12N2310/14 ,  C12N2310/3513 ,  C12N2320/32 ,  Y02A50/403

Abstract: Disclosed herein are methods and compositions comprising anti-CD74 and/or anti-HLA-DR antibodies for treatment of GVHD and other immune dysfunction diseases. In preferred embodiments, the anti-CD74 and/or anti-HLA-DR antibodies are effective to deplete antigen-presenting cells, such as dendritic cells. Most preferably, administration of the therapeutic compositions depletes all subsets of APCs, including mDCs, pDCs, B cells and monocytes, without significant depletion of T cells. In alternative embodiments, administration of the therapeutic compositions suppresses proliferation of allo-reactive T cells, while preserving cytomegalovirus (CMV)-specific, CD8+ memory T cells. The compositions and methods provide a novel conditioning regimen for preventing aGVHD and/or treating chronic GVHD, without altering preexisting anti-viral immunity.

Abstract translation: 本文公开了包含用于治疗GVHD和其他免疫功能障碍疾病的抗CD74和/或抗HLA-DR抗体的方法和组合物。 在优选的实施方案中，抗CD74和/或抗HLA-DR抗体有效地消耗抗原呈递细胞，例如树突状细胞。 最优选地，治疗组合物的给药消耗了APC的所有子集，包括mDC，pDC，B细胞和单核细胞，而不会明显消耗T细胞。 在替代实施方案中，治疗组合物的施用抑制同种异体反应性T细胞的增殖，同时保留巨细胞病毒（CMV）特异性CD8 +记忆T细胞。 组合物和方法提供了用于预防aGVHD和/或治疗慢性GVHD的新型调节方案，而不改变预先存在的抗病毒免疫。

公开(公告)号：US20110236404A1

公开(公告)日：2011-09-29

申请号：US12542792

申请日：2009-08-18

Applicant: David M. Goldenberg ,  Chien-Hsing Chang ,  Edmund A. Rossi ,  Diane Nordstrom

Inventor： David M. Goldenberg ,  Chien-Hsing Chang ,  Edmund A. Rossi ,  Diane Nordstrom

IPC: A61K39/00 ,  C12N5/10 ,  C12N9/00 ,  C12P21/00 ,  C07K2/00 ,  C07K14/54 ,  C07K14/555 ,  C07K14/575 ,  C07K14/52 ,  C07K14/475 ,  C07K14/535 ,  C07K14/505 ,  C07K14/545 ,  C07K14/55 ,  C07K14/56 ,  C07K14/565 ,  C07K14/57 ,  C07K14/485 ,  A61P37/04

CPC classification number: C07K14/4747 ,  A61K2039/505 ,  C07K16/00 ,  C07K16/18 ,  C07K16/2803 ,  C07K16/3007 ,  C07K2317/24 ,  C07K2317/31 ,  C12N5/0694 ,  C12N2500/90 ,  C12N2501/48 ,  C12N2510/02 ,  C12N2740/16043 ,  C12P21/02

Abstract: Disclosed are compositions and methods for increasing the longevity of a cell culture and permitting the increased production of proteins, preferably recombinant proteins, such as antibodies, peptides, enzymes, growth factors, interleukins, interferons, hormones, and vaccines. Cells transfected with an apoptosis-inhibiting gene or vector, such as a triple mutant Bcl-2 gene, can survive longer in culture, resulting in extension of the state and yield of protein biosynthesis. Such transfected cells exhibit maximal cell densities that equal or exceed the maximal density achieved by the parent cell lines. Transfected cells can also be pre-adapted for growth in serum-free medium, greatly decreasing the time required to obtain protein production in serum-free medium. In certain methods, the pre-adapted cells can be used for protein production following transfection under serum-free conditions. In preferred embodiments, the cells of use are SpESF or SpESF-X cells.

Abstract translation: 公开了用于增加细胞培养物的寿命并允许增加蛋白质生产的组合物和方法，优选重组蛋白，例如抗体，肽，酶，生长因子，白细胞介素，干扰素，激素和疫苗。 用凋亡抑制基因或载体如三重突变体Bcl-2基因转染的细胞可以在培养物中更长时间地存活，导致蛋白质生物合成的状态和产量的延长。 这样的转染细胞表现出最大的细胞密度，其等于或超过由亲本细胞系实现的最大密度。 转染的细胞也可以预先适应于无血清培养基中生长，大大减少了在无血清培养基中获得蛋白质产生所需的时间。 在某些方法中，预先修饰的细胞可用于在无血清条件下转染后的蛋白质生产。 在优选的实施方案中，使用的细胞是SpESF或SpESF-X细胞。