During the progression of epithelial tumors, the extracellular matrix (ECM) accumulates collagen stabilized by covalent cross-links that enhance ECM stiffness and trigger tumor cell invasion and dissemination. However, the mechanisms by which tumor cells regulate collagen cross-link formation have not been fully defined. Here, we found that lung adenocarcinoma metastasis is driven by high expression of lysyl hydroxylase 2 (LH2), an enzyme that hydroxylates telopeptidyl lysine residues on collagen, and that LH2 expression in tumor cells increases hydroxylysine aldehyde-derived collagen cross-links (HLCCs) and decreases lysine aldehyde-derived collagen cross-links (LCCs) without changing the total amount of cross-links in the tumor stroma, indicating that malignant progression is associated with a change in the type of collagen cross-link. Furthermore, we showed that LH2 is a secreted protein, which challenges the current belief that LH2 localizes strictly on the endoplasmic reticulum. On the basis of these preliminary results, we postulate that LH2 expression in tumor cells functions as a regulatory switch, controlling the relative abundance of distinct types of collagen cross-links that influence the metastatic fate of tumor cells, and that,in addition to hydroxylating collagen ? chains on the endoplasmic reticulum, LH2 is secreted and promotes metastasis by hydroxylating collagen molecules in the extracellular space. To test these hypotheses, we will pursue 2 specific aims.
Aim 1 : Determine whether ectopic LH2 expression promotes metastasis in mice that develop lung adenocarcinoma from the expression of mutant K-ras. We will intratracheally deliver lentiviruses that co-express Cre recombinase and LH2 to K-rasLSL-G12D mice to induce ectopic LH2 expression and the recombination of a conditional K-rasG12D allele in the same lung epithelial cells. As a comparison, we will also administer lentiviruses co-expressing Cre and lysyl oxidase (LOX) or LOX-like-2 (LOXL2), which are collagen-modifying enzymes that have known pro-metastatic roles.
Aim 2 : Determine whether LH2 promotes metastasis by regulating the type of collagen cross-link in the tumor stroma. We will ascertain whether LH2-induced metastasis and stromal mechanical properties are attenuated by mutation of the LH2 hydroxylase domain or by deletion of the N-terminal signal peptide and are different from those induced by LOX, which regulates the total amount, but not the type, of collagen cross-link. In summary, the novelty of our proposal rests in the implementation of unique mouse modeling approaches to test an original hypothesis that links a particular type of collagen cross-link to metastasis regulation. If proven correct, these findings will challenge the current paradigm of the way in which tumor cells regulate collagen cross-linking to increase stromal stiffness and promote metastasis, and they will provide a rationale for future efforts to develop LH2 antagonists to test whether LH2 inhibition prevents metastasis in lung cancer patients.

Public Health Relevance

Our long term goal is to better understand the biological basis for lung cancer metastasis, the most common cause of death from this disease, and to develop therapeutic strategies to prevent metastasis. During malignant disease progression, the extracellular matrix (ECM) of epithelial tumors accumulates inter-molecular cross-links between collagen strands; these cross-links enhance ECM stiffness and trigger tumor cell invasion and dissemination. In the proposed study, we will bring together investigators with expertise in functional genomic screens, collagen biochemistry, in vivo microscopy, and mouse modeling of human lung cancer to determine the mechanisms by which tumor cells regulate the type of collagen cross-links in tumors; findings from these studies may have broad clinical implications because the enzyme under investigation that regulates the type of collagen cross-links has a potentially druggable catalytic domain and is expressed at high levels in multiple tumor types. .

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
Project #
Application #
Study Section
Tumor Microenvironment Study Section (TME)
Program Officer
Ault, Grace S
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
University of Texas MD Anderson Cancer Center
Internal Medicine/Medicine
United States
Zip Code
Guo, Hou-Fu; Cho, Eun Jeong; Devkota, Ashwini K et al. (2017) A scalable lysyl hydroxylase 2 expression system and luciferase-based enzymatic activity assay. Arch Biochem Biophys 618:45-51
Chen, Yulong; Terajima, Masahiko; Banerjee, Priyam et al. (2017) FKBP65-dependent peptidyl-prolyl isomerase activity potentiates the lysyl hydroxylase 2-driven collagen cross-link switch. Sci Rep 7:46021
Pankova, Daniela; Chen, Yulong; Terajima, Masahiko et al. (2016) Cancer-Associated Fibroblasts Induce a Collagen Cross-link Switch in Tumor Stroma. Mol Cancer Res 14:287-95
Terajima, Masahiko; Taga, Yuki; Chen, Yulong et al. (2016) Cyclophilin-B Modulates Collagen Cross-linking by Differentially Affecting Lysine Hydroxylation in the Helical and Telopeptidyl Domains of Tendon Type I Collagen. J Biol Chem 291:9501-12
Chen, Yulong; Terajima, Masahiko; Yang, Yanan et al. (2015) Lysyl hydroxylase 2 induces a collagen cross-link switch in tumor stroma. J Clin Invest 125:1147-62
Ahn, Young-Ho; Yang, Yanan; Gibbons, Don L et al. (2011) Map2k4 functions as a tumor suppressor in lung adenocarcinoma and inhibits tumor cell invasion by decreasing peroxisome proliferator-activated receptor ?2 expression. Mol Cell Biol 31:4270-85
Ahn, Young-Ho; Kurie, Jonathan M (2009) MKK4/SEK1 is negatively regulated through a feedback loop involving the E3 ubiquitin ligase itch. J Biol Chem 284:29399-404
Yang, Yanan; Iwanaga, Kentaro; Raso, Maria Gabriela et al. (2008) Phosphatidylinositol 3-kinase mediates bronchioalveolar stem cell expansion in mouse models of oncogenic K-ras-induced lung cancer. PLoS One 3:e2220
Iwanaga, Kentaro; Yang, Yanan; Raso, Maria Gabriela et al. (2008) Pten inactivation accelerates oncogenic K-ras-initiated tumorigenesis in a mouse model of lung cancer. Cancer Res 68:1119-27
Khatlani, T S; Wislez, M; Sun, M et al. (2007) c-Jun N-terminal kinase is activated in non-small-cell lung cancer and promotes neoplastic transformation in human bronchial epithelial cells. Oncogene 26:2658-66

Showing the most recent 10 out of 14 publications