Kortt A A, Dolezal O, Power B E, Hudson P J
CSIRO Health Science and Nutrition, 343 Royal Parade, Vic. 3052, Parkville, Australia.
Biomol Eng. 2001 Oct 15;18(3):95-108. doi: 10.1016/s1389-0344(01)00090-9.
Recombinant antibody fragments can be engineered to assemble into stable multimeric oligomers of high binding avidity and specificity to a wide range of target antigens and haptens. This review describes the design and expression of diabodies (dimers), triabodies (trimers) and tetrabodies (tetramers). In particular we discuss the role of linker length between V-domains and the orientation of the V-domains to direct the formation of either diabodies (60 kDa), triabodies (90 kDa) or tetrabodies (120 kDa), and how the size, flexibility and valency of each molecules is suited to different applications for in vivo imaging and therapy. Single chain Fv antibody fragments joined by polypeptide linkers of at least 12 residues irrespective of V-domains orientation predominantly form monomers with varying amounts of dimer and higher molecular mass oligomers in equilibrium. A scFv molecule with a linker of 3-12 residues cannot fold into a functional Fv domain and instead associates with a second scFv molecule to form a bivalent dimer (diabody, approximately 60 kDa). Reducing the linker length below three residues can force scFv association into trimers (triabodies, approximately 90 kDa) or tetramers ( approximately 120 kDa) depending on linker length, composition and V-domain orientation. A particular advantage for tumour targeting is that molecules of 60-100 kDa have increased tumour penetration and fast clearance rates compared with the parent Ig (150 kDa). We highlight a number of cancer-targeting scFv diabodies that have undergone successful pre-clinical trials for in vivo stability and efficacy. We also briefly review the design of multi-specific Fv modules suited to cross-link two or more different target antigens. Bi-specific diabodies formed by association of different scFv molecules have been designed as cross-linking reagents for T-cell recruitment into tumours (immunotherapy), viral retargeting (gene therapy) and as red blood cell agglutination reagents (immunodiagnostics). The more challenging trispecific multimers (triabodies) remain to be described.
重组抗体片段可被设计组装成具有高结合亲和力和特异性的稳定多聚体寡聚体，以针对多种靶抗原和半抗原。本综述描述了双抗体（二聚体）、三抗体（三聚体）和四抗体（四聚体）的设计与表达。特别地，我们讨论了V结构域之间连接子长度以及V结构域的方向在引导形成双抗体（60 kDa）、三抗体（90 kDa）或四抗体（120 kDa）中的作用，以及每个分子的大小、灵活性和价态如何适合体内成像和治疗的不同应用。由至少12个残基的多肽连接子连接的单链Fv抗体片段，无论V结构域的方向如何，主要形成具有不同量二聚体和更高分子量寡聚体处于平衡状态的单体。具有3 - 12个残基连接子的scFv分子不能折叠成功能性Fv结构域，而是与第二个scFv分子缔合形成二价二聚体（双抗体，约60 kDa）。将连接子长度减少到三个残基以下可迫使scFv缔合成三聚体（三抗体，约90 kDa）或四聚体（约120 kDa），这取决于连接子长度、组成和V结构域方向。肿瘤靶向的一个特别优势是，与亲本Ig（150 kDa）相比，60 - 100 kDa的分子具有更高的肿瘤穿透性和更快的清除率。我们重点介绍了一些针对癌症的scFv双抗体，它们已成功进行了体内稳定性和疗效的临床前试验。我们还简要回顾了适用于交联两种或更多种不同靶抗原的多特异性Fv模块的设计。由不同scFv分子缔合形成的双特异性双抗体已被设计为用于将T细胞募集到肿瘤中的交联试剂（免疫疗法）、病毒重靶向试剂（基因疗法）以及作为红细胞凝集试剂（免疫诊断）。更具挑战性的三特异性多聚体（三抗体）仍有待描述。
