Lysyl oxidase-like 2 promotes hepatocellular carcinoma metastasis by remodelling the tumor and metastatic microenvironments

Abstract

Hepatocellular carcinoma is a common and lethal malignancy worldwide. High death rate in HCC is mainly due to metastasis. This study revealed the detailed molecular mechanisms by which primary HCC promoted metastasis through formation of tumor and metastatic niches via lysyl oxidase-like 2 (LOXL2). LOXL2 is a secretory protein that belongs to the LOX family which is consisted of 5 members. LOX family members are amine oxidases that cross-link components in extracellular matrix (ECM) such as collagen and elastin. Among the five LOX family members, LOXL2 is the sole member over-expressed in HCC tissues. Remarkably, we detected a significant up-regulation of LOXL2 protein in the sera of 80 HCC patients as compared to normal individuals. Clinicopathological analysis showed that up-regulation of LOXL2 in human HCC was significantly associated with aggressive HCC feature, direct liver invasion. Interestingly, we found that LOXL2 expression in HCC cells was positively regulated by the hypoxia/HIF-1α and TFGβ/SMAD4-mediated pathways and negatively regulated by miR-26 and miR-29, two microRNAs families that are under-expressed in HCC. Abrogation of the LOXL2 pathway suppressed collagen cross-linking, bone marrow cell invasion, and HCC cell invasion in vitro. Stable knockdown of LOXL2 repressed tumor growth and intrahepatic metastasis in orthotopic tumor implantation model. Knockdown of LOXL2 also reduced collagen cross-linking and tumor stiffness in vivo. LOXL2-mediated change of tension within the tumor microenvironment activated a key cell motility regulator, rho-kinase (ROCK), and ROCK-mediated events such as actin polymerization and focal adhesion formation. In addition, we found that primary HCC tumor facilitated metastasis by directing bone marrow-derived cells (BMDCs) to colonize the distal metastatic sites prior to the arrival of HCC cells. We observed a profound reduction of collagen cross-linking, CD11b+ BMDC recruitment, and subsequent metastasis in the lungs of the mice that were orthotopically implanted with the LOXL2 knockdown HCC tumors. Conclusion: Our study highlighted three important advances in understanding the cancer biology of hepatocellular carcinoma. First, LOXL2 is overexpressed in HCC clinical samples and could be detected in sera of patients, indicating that LOXL2 may be a good diagnostic marker for HCC patients. Second, we delineated a complex interlinked signaling network that involves multiple regulators including HIF-1α/TGFβ/ microRNAs which converge to control the expression of LOXL2 leading to HCC progression. Third, we showed that LOXL2 played an imperative role in the tumor microenvironment and metastatic niches formation leading to HCC progression. We comprehensively unveiled the molecular mechanisms which drive LOXL2 dysregulation and the functional implications of LOXL2 in human HCC.