The antimicrobial peptide cathelicidin inhibits development
The antimicrobial peptide cathelicidin inhibits development of colitis-associated colon cancer. However, the role of cathelici-din in colon cancer metastasis remains unknown. We hypoth-esized that cathelicidin is effective in inhibiting colon cancer metastasis. Human colon cancer HT-29 DAP5 were injected intravenously into nude mice. Control HA-tagged adeno-asso-ciated virus (HA-AAV) or cathelicidin-overexpressing AAV (CAMP-HA-AAV) were injected intravenously into nude mice on the same day. Four weeks later, the nude mice were as-sessed for lung and liver metastases. Human colon cancer SW620 cells were used to study the effect of cathelicidin on cell migration and cytoskeleton. Incubation of SW620 cells with cathelicidin dose-dependently reduced cell migration, dis-rupted cytoskeletal structure, and reduced bIII-tubulin (TUBB3) mRNA expression. The addition of the P2RX7 antag-onist KN62, but not the FPRL1 antagonist WRW4, prevented the LL-37-mediated inhibition of cell migration and TUBB3 mRNA expression. The CAMP-HA-AAV-overexpressing group showed significantly reduced human CK20 protein (by 60%) and TUBB3 mRNA expression (by 40%) in the lungs and liver of the HT-29-loaded nude mice, compared to the HA-AAV control group. Intraperitoneal injection of KN62 reversed the CAMP-HA-AAV-mediated inhibition of human CK20 and TUBB3 expression in the lungs and liver of HT-29-loaded nude mice. In conclusion, cathelicidin inhibits colon cancer metastasis via a P2RX7-dependent pathway.
Low survival rates of advanced colon cancer (stage IIIC 53% and stage IV 11% in 5 years; American Cancer Society) and high treatment resistance have driven discovery of novel drug targets and therapeutic approaches. Cathelicidin (human gene CAMP, human protein LL-37, mouse gene Camp, and mouse protein mCRAMP) is an antimicrobial peptide that possesses therapeutic effects in inflammatory bowel dis-ease, Clostridium difficile infection, and obesity.1–4 Recent studies have also shown that cathelicidin is involved in malignancies. The tumoral expression of endogenous cathelicidin varies among different types of cancer.5,6 For example, cathelicidin expression is diminished in gastric and colon cancers,7,8 while it is increased in breast, ovarian, and lung cancers.9–11 Endogenous cathelicidin modulates azoxyme-thane (AOM)-mediated colon cancer in mice.7 Cathelicidin sup-presses gastric cancer cell proliferation via the bone morphogenetic
protein-mediated pathway.8 The biological functions of cathelicidin are largely mediated by its receptors, which include FPRL1 and P2RX7.12
Cathelicidin and its analog FK-16 induce p53-dependent apoptosis in human colon cancer HCT116 cells.7,13 Other cathelicidin analogs (FF/CAP18 and Ceragenin CSA13) inhibit HCT116 cell proliferation without relying on the p53-dependent mechanism in vitro.14,15 Inter-estingly, cathelicidin does not inhibit the viability of colon cancer HT-29 cells directly, but inhibits tumor-associated fibroblasts (TAFs) through suppression of epithelial-mesenchymal transition (EMT) and disruption of the cytoskeleton.16 The inhibition of TAFs, in turn, reduces their support of colon cancer cell proliferation. All of the evidence suggests that cathelicidin is a potential target for colon cancer. However, the role of cathelicidin and its receptor in metastatic colon cancer is unknown.
Metastatic colon cancer cells often possess mesenchymal characteris-
tics, as exemplified by bIII-tubulin expression, which is associated with increased cell migratory behavior.17,18 We hypothesize that cath- elicidin acts through receptor-dependent modulation of the cytoskel-eton in the colon cancer cells, leading to a reduction of colon cancer cell migration. To address this hypothesis, a nude mouse model and various cell-based assays were included in this study.
Systemic Overexpression of Cathelicidin Inhibited Colon Cancer Metastasis in Nude Mice
To determine whether cathelicidin inhibits colon cancer metastasis, the nude mice were injected intravenously with human colon cancer HT-29 cells (Figure 1A). Control hemagglutinin-expressing adeno-associ-ated viruses (HA-AAVs) and human cathelicidin-overexpressing
Correspondence: Mingjun Sun, MD, PhD, Department of Gastroenterology, First
Affiliated Hospital of China Medical University, North Nanjing Street 155, She-
E-mail: [email protected]
Correspondence: Hon Wai Koon, PhD, Vatche and Tamar Manoukian Division of
Digestive Diseases, David Geffen School of Medicine, University of California, Los
E-mail: [email protected]
Molecular Therapy: Oncolytics
AAVs (CAMP-HA-AAVs) were injected into nude mice intravenously on the same day as the HT-29 injection, as described in a previous study.16