Cancer-associated fibroblasts enhance cancer progression when activated by tumor cells through mechanisms not yet fully understood. Blocking mammary tumor cell-derived lysyl oxidase-like 2 (LOXL2) significantly inhibited mammary tumor cell invasion and metastasis in transgenic and orthotopic mouse models. Here, we discovered that tumor-derived LOXL2 directly activated stromal fibroblasts in the tumor microenvironment. Genetic manipulation or antibody inhibition of LOXL2 in orthotopically grown mammary tumors reduced the expression of ?-smooth muscle actin (?-SMA). Using a marker for reticular fibroblasts, it was determined that expression of ?-SMA was localized to fibroblasts recruited from the host tissue. This marker also revealed that the matrix present in tumors with reduced levels of LOXL2 was more scattered compared with control tumors which exhibited matrices with dense, parallel alignments. Importantly, in vitro assays revealed that tumor-derived LOXL2 and a recombinant LOXL2 protein induced fibroblast branching on collagen matrices, as well as increased fibroblast-mediated collagen contraction and invasion of fibroblasts through extracellular matrix. Moreover, LOXL2 induced the expression of ?-SMA in fibroblasts grown on collagen matrices. Mechanistically, it was determined that LOXL2 activated fibroblasts through integrin-mediated focal adhesion kinase activation. These results indicate that inhibition of LOXL2 in tumors not only reduces tumor cell invasion but also attenuates the activation of host cells in the tumor microenvironment. Implications: These findings reveal new insight into the mechanisms of fibroblast activation, a novel function of LOXL2, and further highlight the importance of generating LOXL2-targeted therapies for the prevention of tumor progression and metastasis.
Tumor metastasis is a highly complex, dynamic, and inefficient process involving multiple steps, yet it accounts for more than 90% of cancer-related deaths. Although it has long been known that fibrotic signals enhance tumor progression and metastasis, the underlying molecular mechanisms are still unclear. Identifying events involved in creating environments that promote metastatic colonization and growth are critical for the development of effective cancer therapies. Here, we show a critical role for lysyl oxidase (LOX) in establishing a milieu within fibrosing tissues that is favorable to growth of metastastic tumor cells. We show that LOX-dependent collagen crosslinking is involved in creating a growth-permissive fibrotic microenvironment capable of supporting metastatic growth by enhancing tumor cell persistence and survival. We show that therapeutic targeting of LOX abrogates not only the extent to which fibrosis manifests, but also prevents fibrosis-enhanced metastatic colonization. Finally, we show that the LOX-mediated collagen crosslinking directly increases tumor cell proliferation, enhancing metastatic colonization and growth manifesting in vivo as increased metastasis. This is the first time that crosslinking of collagen I has been shown to enhance metastatic growth. These findings provide an important link between ECM homeostasis, fibrosis, and cancer with important clinical implications for both the treatment of fibrotic disease and cancer.
Emerging evidence implicates lysyl oxidase (LOX), an extracellular matrix-modifying enzyme, in promoting metastasis of solid tumors. We investigated whether LOX plays an important role in the metastasis of colorectal cancer (CRC).
More than 90% of cancer patient mortality is attributed to metastasis. In this study, we investigated a role for the lysyl oxidase-related enzyme lysyl oxidase-like 2 (LOXL2) in breast cancer metastasis, in both patient samples and in vivo models. Analysis of a published microarray data set revealed that LOXL2 expression is correlated with metastasis and decreased survival in patients with aggressive breast cancer. In immunocompetent or immunocompromised orthotopic and transgenic breast cancer models we showed that genetic, chemical or antibody-mediated inhibition of LOXL2 resulted in decreased metastasis. Mechanistic investigations revealed that LOXL2 promotes invasion by regulating the expression and activity of the extracellular proteins tissue inhibitor of metalloproteinase-1 (TIMP1) and matrix metalloproteinase-9 (MMP9). We found that LOXL2, TIMP1, and MMP9 are coexpressed during mammary gland involution, suggesting they function together in glandular remodeling after weaning. Finally, we found that LOXL2 is highly expressed in the basal/myoepithelial mammary cell lineage, like many other genes that are upregulated in basal-like breast cancers. Our findings highlight the importance of LOXL2 in breast cancer progression and support the development of anti-LOXL2 therapeutics for the treatment of metastatic breast cancer.
Tumor cell metastasis is facilitated by "premetastatic niches" formed in destination organs by invading bone marrow-derived cells (BMDCs). Lysyl oxidase (LOX) is critical for premetastatic niche formation. LOX secreted by hypoxic breast tumor cells accumulates at premetastatic sites, crosslinks collagen IV in the basement membrane, and is essential for CD11b+ myeloid cell recruitment. CD11b+ cells adhere to crosslinked collagen IV and produce matrix metalloproteinase-2, which cleaves collagen, enhancing the invasion and recruitment of BMDCs and metastasizing tumor cells. LOX inhibition prevents CD11b+ cell recruitment and metastatic growth. CD11b+ cells and LOX also colocalize in biopsies of human metastases. Our findings demonstrate a critical role for LOX in premetastatic niche formation and support targeting LOX for the treatment and prevention of metastatic disease.
Identification of key molecules that drive angiogenesis is critical for the development of new modalities for the prevention of solid tumor progression. Using multiple models of colorectal cancer, we show that activity of the extracellular matrix-modifying enzyme lysyl oxidase (LOX) is essential for stimulating endothelial cells in vitro and angiogenesis in vivo. We show that LOX activates Akt through platelet-derived growth factor receptor ? (PDGFR?) stimulation, resulting in increased VEGF expression. LOX-driven angiogenesis can be abrogated through targeting LOX directly or using inhibitors of PDGFR?, Akt, and VEGF signaling. Furthermore, we show that LOX is clinically correlated with VEGF expression and blood vessel formation in 515 colorectal cancer patient samples. Finally, we validate our findings in a breast cancer model, showing the universality of these observations. Taken together, our findings have broad clinical and therapeutic implications for a wide variety of solid tumor types.
Sunitinib is a potent and clinically approved tyrosine kinase inhibitor that can suppress tumour growth by inhibiting angiogenesis. However, conflicting data exist regarding the effects of this drug on the growth of metastases in preclinical models. Here we use 4T1 and RENCA tumour cells, which both form lung metastases in Balb/c mice, to re-address the effects of sunitinib on the progression of metastatic disease in mice. We show that treatment of mice with sunitinib prior to intravenous injection of tumour cells can promote the seeding and growth of 4T1 lung metastases, but not RENCA lung metastases, showing that this effect is cell line dependent. However, increased metastasis occurred only upon administration of a very high sunitinib dose, but not when lower, clinically relevant doses were used. Mechanistically, high dose sunitinib led to a pericyte depletion effect in the lung vasculature that correlated with increased seeding of metastasis. By administering sunitinib to mice after intravenous injection of tumour cells, we demonstrate that while sunitinib does not inhibit the growth of 4T1 lung tumour nodules, it does block the growth of RENCA lung tumour nodules. This contrasting response was correlated with increased myeloid cell recruitment and persistent vascularisation in 4T1 tumours, whereas RENCA tumours recruited less myeloid cells and were more profoundly devascularised upon sunitinib treatment. Finally, we show that progression of 4T1 tumours in sunitinib treated mice results in increased hypoxia and increased glucose metabolism in these tumours and that this is associated with a poor outcome. Taken together, these data suggest that the effects of sunitinib on tumour progression are dose-dependent and tumour model-dependent. These findings have relevance for understanding how anti-angiogenic agents may influence disease progression when used in the adjuvant or metastatic setting in cancer patients.
The tripartite motif family protein 27 (TRIM27) is a transcriptional repressor that interacts with, and attenuates senescence induction by, the retinoblastoma-associated protein (RB1). High expression of TRIM27 was noted in several human cancer types including breast and endometrial cancer, where elevated TRIM27 expression predicts poor prognosis. Here, we investigated the role of TRIM27 expression in cancer development.
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