Micalet Auxtine, Tappouni Luke J, Peszko Katarzyna, Karagianni Despoina, Lam Ashley, Counsell John R, Quezada Sergio A, Moeendarbary Emad, Cheema Umber
UCL Centre for 3D Models of Health and Disease, Department of Targeted Intervention, Division of Surgery and Interventional Science, University College London, Charles Bell House, 43-45 Foley Street, London W1W 7TS, United Kingdom.
Department of Mechanical Engineering, University College London, Gower Street, London WC1E 6BT, United Kingdom.
Matrix Biol Plus. 2023 Nov 15;19-20:100137. doi: 10.1016/j.mbplus.2023.100137. eCollection 2023 Dec.
Cancer cells remodel their local physical environment through processes of matrix reorganisation, deposition, stiffening and degradation. Urokinase-type plasminogen activator (uPA), which is encoded by the gene, is an extracellular proteolytic enzyme known to be involved in cancer progression and tumour microenvironment (TME) remodelling. Perturbing uPA therefore has a strong potential as a mechano-based cancer therapy. This work is a bioengineering investigation to validate whether 1) uPA is involved in matrix degradation and 2) preventing matrix degradation by targeting uPA can reduce cancer cell invasion and metastasis.
To this aim, we used an engineered 3D model, termed the tumouroid, that appropriately mimics the tumour's native biophysical environment (3 kPa). A CRISPR-Cas9 mediated uPA knockout was performed to introduce a loss of function mutation in the gene coding sequence. Subsequently, to validate the translational potential of blocking uPA action, we tested a pharmacological inhibitor, UK-371,801. The changes in matrix stiffness were measured by atomic force microscopy (AFM). Invasion was quantified using images of the tumouroid, obtained after 21 days of culture.
We showed that uPA is highly expressed in invasive breast and colorectal cancers, and these invasive cancer cells locally degrade their TME. (uPA) gene knock-out (KO) completely stopped matrix remodelling and significantly reduced cancer invasion. Many invasive cancer gene markers were also downregulated in the KO tumouroids. Pharmacological inhibition of uPA showed similarly promising results, where matrix degradation was reduced and so was the cancer invasion.
This work supports the role of uPA in matrix degradation. It demonstrates that the invasion of cancer cells was significantly reduced when enzymatic breakdown of the TME matrix was prevented. Collectively, this provides strong evidence of the effectiveness of targeting uPA as a mechano-based cancer therapy.
癌细胞通过基质重组、沉积、硬化和降解过程重塑其局部物理环境。由该基因编码的尿激酶型纤溶酶原激活剂（uPA）是一种细胞外蛋白水解酶，已知其参与癌症进展和肿瘤微环境（TME）重塑。因此，干扰uPA作为一种基于机械作用的癌症治疗方法具有很大潜力。这项工作是一项生物工程研究，旨在验证：1）uPA是否参与基质降解；2）通过靶向uPA防止基质降解是否能减少癌细胞的侵袭和转移。
为实现这一目标，我们使用了一种名为肿瘤类器官的工程化三维模型，该模型能恰当模拟肿瘤的天然生物物理环境（3千帕）。进行了CRISPR-Cas9介导的uPA基因敲除，以在编码序列中引入功能缺失突变。随后，为验证阻断uPA作用的转化潜力，我们测试了一种药理抑制剂UK-371,801。通过原子力显微镜（AFM）测量基质硬度的变化。使用培养21天后获得的肿瘤类器官图像对侵袭进行定量分析。
我们发现uPA在侵袭性乳腺癌和结直肠癌中高表达，并且这些侵袭性癌细胞会局部降解其TME。（uPA）基因敲除（KO）完全停止了基质重塑，并显著降低了癌症侵袭。许多侵袭性癌症基因标志物在KO肿瘤类器官中也下调。对uPA的药理抑制显示出类似的有前景的结果，其中基质降解减少，癌症侵袭也减少。
这项工作支持了uPA在基质降解中的作用。它表明，当TME基质的酶促分解被阻止时，癌细胞的侵袭显著减少。总体而言，这为靶向uPA作为一种基于机械作用的癌症治疗方法的有效性提供了有力证据。
