Max Planck Institute of Biochemistry, Department of Molecular Biology, Am Klopferspitz 18, 82152-, Martinsried, Germany.
University of Lorraine, NGERE Unit, Faculté de Médecine, 9 avenue de la Forêt de Haye, 54505, Vandoeuvre-lès-Nancy, France.
Chembiochem. 2022 Jun 3;23(11):e202100637. doi: 10.1002/cbic.202100637. Epub 2022 Apr 20.
The promise of personalized medicine for monogenic and complex polygenic diseases depends on the availability of strategies for targeted inhibition of disease-associated polymorphic protein variants. Loss of function variants, including non-synonymous single nucleotide variants (nsSNVs) and insertion/deletion producing a frameshift, account for the vast majority of disease-related genetic changes. Because it is challenging to interpret the functional consequences of nsSNVs, they are considered a big barrier for personalized medicine. A method for inhibiting the specific expression of nsSNVs without editing the human genome will facilitate the elucidation of the biology of nsSNVs, but such a method is currently lacking. Here, I describe the phenomenon of membrane anchorage-induced (MAGIC) knockdown of allele-specific inhibition of protein and mRNA expression upon inner membrane tethering of point mutation-specific monoclonal antibodies (mAb). This phenomenon is likely mediated by a mechanism distinct from the protein degradation pathways, as the epitope-specific knockdown is replicated upon intracellular expression of a membrane-anchored single domain intrabody that lacks the Fc domain of the mAb. By harnessing the MAGIC knockdown of epitope-containing protein targets, I report a novel approach for inhibiting the expression of amino-acid-altering germline and somatic nsSNVs. As a proof-of-concept, I show the inhibition of human disease-associated variants namely, FGFR4 p.G388R, KRAS p.G12D and BRAF p.V600E protein variants. This method opens up a new avenue for not just therapeutic suppression of undruggable protein variants, but also for functional interrogation of the nsSNVs of unknown significance.
个性化医学对单基因和复杂多基因疾病的承诺取决于针对疾病相关多态性蛋白变异体的靶向抑制策略的可用性。功能丧失变异体，包括非同义单核苷酸变异体（nsSNV）和导致移码的插入/缺失，占与疾病相关遗传变化的绝大多数。由于难以解释 nsSNV 的功能后果，因此它们被认为是个性化医学的一大障碍。一种抑制特定 nsSNV 表达而不编辑人类基因组的方法将有助于阐明 nsSNV 的生物学特性，但目前缺乏这种方法。在这里，我描述了膜锚定诱导（MAGIC）现象，即在点突变特异性单克隆抗体（mAb）将内膜锚定后，等位基因特异性抑制蛋白和 mRNA 表达。这种现象可能是由一种不同于蛋白降解途径的机制介导的，因为在缺乏 mAb 的 Fc 结构域的膜锚定单域内体的细胞内表达中，重复了表位特异性敲低。通过利用 MAGIC 对含表位蛋白靶标的敲低，我报告了一种抑制氨基酸改变的种系和体细胞 nsSNV 表达的新方法。作为概念验证，我展示了对人类疾病相关变体的抑制，即 FGFR4 p.G388R、KRAS p.G12D 和 BRAF p.V600E 蛋白变体。该方法为抑制不可成药的蛋白变体的治疗性抑制以及对未知意义的 nsSNV 的功能研究开辟了新途径。
