Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190401, Israel.
Department of Cancer Biology, Mayo Clinic Comprehensive Cancer Center, Jacksonville, FL 32224, USA.
Biomolecules. 2024 Sep 20;14(9):1187. doi: 10.3390/biom14091187.
Matrix metalloproteinases (MMPs) are significant drivers of many diseases, including cancer, and are established targets for drug development. Tissue inhibitors of metalloproteinases (TIMPs) are endogenous MMP inhibitors and are being pursued for the development of anti-MMP therapeutics. TIMPs possess many attractive properties for drug candidates, such as complete MMP inhibition, low toxicity, low immunogenicity, and high tissue permeability. However, a major challenge with TIMPs is their rapid clearance from the bloodstream due to their small size. This study explores a method for extending the plasma half-life of the N-terminal domain of TIMP2 (N-TIMP2) by appending it with a long, intrinsically unfolded tail containing Pro, Ala, and Thr (PATylation). We designed and produced two PATylated N-TIMP2 constructs with tail lengths of 100 and 200 amino acids (N-TIMP2-PAT and N-TIMP2-PAT). Both constructs demonstrated higher apparent molecular weights and retained high inhibitory activity against MMP-9. N-TIMP2-PAT significantly increased plasma half-life in mice compared to the non-PATylated variant, enhancing its therapeutic potential. PATylation offers distinct advantages for half-life extension, such as fully genetic encoding, monodispersion, and biodegradability. It can be easily applied to N-TIMP2 variants engineered for high affinity and selectivity toward individual MMPs, creating promising candidates for drug development against MMP-related diseases.
基质金属蛋白酶（MMPs）是许多疾病（包括癌症）的重要驱动因素，也是药物开发的既定靶点。金属蛋白酶组织抑制剂（TIMPs）是内源性 MMP 抑制剂，正在被开发用于抗 MMP 治疗药物。TIMP 具有许多作为候选药物的诱人特性，如完全抑制 MMP、低毒性、低免疫原性和高组织通透性。然而，TIMP 面临的一个主要挑战是由于其体积小，它们在血液中迅速清除。本研究通过将 TIMP2 的 N 端结构域（N-TIMP2）与含有 Pro、Ala 和 Thr（PATylation）的长、固有无规尾缀合，探索了延长其血浆半衰期的方法。我们设计并生产了两种带有 100 和 200 个氨基酸（N-TIMP2-PAT 和 N-TIMP2-PAT）的 PATylated N-TIMP2 构建体。这两种构建体都显示出更高的表观分子量，并保持对 MMP-9 的高抑制活性。与非 PATylated 变体相比，N-TIMP2-PAT 显著延长了血浆半衰期，增强了其治疗潜力。PATylation 为半衰期延长提供了明显的优势，如完全遗传编码、单分散性和生物降解性。它可以很容易地应用于针对单个 MMP 具有高亲和力和选择性的 N-TIMP2 变体，为开发针对与 MMP 相关的疾病的药物提供了有前途的候选药物。
