CD47, a "don't eat me" signal for phagocytic cells, is expressed on the surface of all human solid tumor cells. Analysis of patient tumor and matched adjacent normal (nontumor) tissue revealed that CD47 is overexpressed on cancer cells. CD47 mRNA expression levels correlated with a decreased probability of survival for multiple types of cancer. CD47 is a ligand for SIRPα, a protein expressed on macrophages and dendritic cells. In vitro, blockade of CD47 signaling using targeted monoclonal antibodies enabled macrophage phagocytosis of tumor cells that were otherwise protected. Administration of anti-CD47 antibodies inhibited tumor growth in orthotopic immunodeficient mouse xenotransplantation models established with patient tumor cells and increased the survival of the mice over time. Anti-CD47 antibody therapy initiated on larger tumors inhibited tumor growth and prevented or treated metastasis, but initiation of the therapy on smaller tumors was potentially curative. The safety and efficacy of targeting CD47 was further tested and validated in immune competent hosts using an orthotopic mouse breast cancer model. These results suggest all human solid tumor cells require CD47 expression to suppress phagocytic innate immune surveillance and elimination. These data, taken together with similar findings with other human neoplasms, show that CD47 is a commonly expressed molecule on all cancers, its function to block phagocytosis is known, and blockade of its function leads to tumor cell phagocytosis and elimination. CD47 is therefore a validated target for cancer therapies.

Elucidating the mechanism of liver tumor growth and metastasis after hepatic ischemia-reperfusion (I/R) injury of a small liver remnant will lay the foundation for the development of therapeutic strategies to target small liver remnant injury, and will reduce the likelihood of tumor recurrence after major hepatectomy or liver transplantation for liver cancer patients. In the current study, we aimed to investigate the effect of hepatic I/R injury of a small liver remnant on liver tumor development and metastases, and to explore the precise molecular mechanisms. A rat liver tumor model that underwent partial hepatic I/R injury with or without major hepatectomy was investigated. Liver tumor growth and metastases were compared among the groups with different surgical stress. An orthotopic liver tumor nude mice model was used to further confirm the invasiveness of the tumor cells from the above rat liver tumor model. Significant tumor growth and intrahepatic metastasis (5 of 6 vs. 0 of 6, P ϭ 0.015), and lung metastasis (5 of 6 vs. 0 of 6, P ϭ 0.015) were found in rats undergoing I/R and major hepatectomy compared with the control group, and was accompanied by upregulation of mRNA levels for Cdc42, ROCK (Rho kinase), and vascular endothelial growth factor, as well as activation of hepatic stellate cells. Most of the nude mice implanted with liver tumor from rats under I/R injury and major hepatectomy developed intrahepatic and lung metastases. In conclusion, hepatic I/R injury of a small liver remnant exacerbated liver tumor growth and metastasis by marked activation of cell adhesion, invasion, and angiogenesis pathways.

This study evaluated the significance of circulating bone marrow-derived endothelial progenitor cells (EPCs) in patients with hepatocellular carcinoma (HCC), a solid tumor with rich neovasculature. Eighty patients with HCC were recruited for the study, and 16 patients with liver cirrhosis and 14 healthy subjects were also included for comparison. Blood samples were taken before treatment. Total mononuclear cells were isolated from peripheral blood, preplated to eliminate mature circulating endothelial cells, and colony-forming units (CFUs) formed by circulating EPCs were counted. To validate the CFU scores, FACS quantification of EPCs using CD133, VEGFR2, and CD34 as markers was performed in 30 cases. Our study showed significantly higher mean CFU scores in patients with HCC compared to patients with cirrhosis and healthy controls (P ‫؍‬ .001 and .009, respectively). Furthermore, the CFU scores of patients with HCC positively correlated with levels of serum ␣-fetoprotein (r ‫؍‬ .303, P ‫؍‬ .017), plasma VEGF (r ‫؍‬ .242, P ‫؍‬ .035), and plasma interleukin-8 (IL-8) (r ‫؍‬ .258, P ‫؍‬ .025). Patients with unresectable HCC had higher CFU scores than patients with resectable tumors (P ‫؍‬ .027). Furthermore, for those who underwent curative surgery, higher preoperative CFU scores were observed in patients with recurrence within 1 year compared with those who were disease-free after 1 year (P ‫؍‬ .013). In conclusion, higher circulating levels of EPCs are seen in patients with advanced unresectable HCC as compared to patients with resectable HCC or those with liver cirrhosis. Our evidence supports the potential use of circulating level of EPCs as a prognostic marker in patients with HCC. (HEPATOLOGY 2006;44:836-843.)

The aim of the current study is to elucidate the mechanism of proline-rich tyrosine kinase 2 (Pyk2)-mediated cell proliferation and invasiveness in hepatocellular carcinoma (HCC) cells. Human HCC cell lines PLC and MHCC97L were stably transfected with either full-length Pyk2 or C-terminal non-kinase region of Pyk2 (PRNK). Functional studies on cell proliferation and invasion were conducted in vitro by colony formation assay, adhesion assay, migration assay and wound-healing assay. For the in vivo study, an orthotopic nude mice liver tumor model was applied to investigate the effects of Pyk2 overexpression on tumor growth and metastasis. Overexpression of Pyk2 in PLC cells resulted in an upregulation of colony formation (P = 0.021) and adhesion toward laminin (P = 0.018). Pyk2 promoted wound recovery by stimulation of actin stress fiber polymerization. In the in vivo study, transfection of PRNK in MHCC97L cells significantly decreased tumor volume (P = 0.001) and the incidence of lung metastasis (P = 0.014). Overexpression of Pyk2 promoted the activation of c-Src, formation of Pyk2/c-Src complex and activated the extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK)-signaling pathway. Pyk2 upregulated the activation of ERK1/2 that is insensitive to MAPK/ERK kinase (MEK)1/2 inhibition. On the contrary, PRNK overexpression downregulated the activation of c-Src and ERK/MAPK-signaling pathways. Immunofluorescence staining showed that the focal adhesion localization of Pyk2 is a major determinant for c-Src and ERK/MAPK activation. In conclusion, our results showed that Pyk2 promoted cell proliferation and invasiveness by upregulation of the c-Src and ERK/MAPK-signaling pathways.