Generation and characterization of EphA1 receptor tyrosine kinase reporter knockout mice.

Eph receptor tyrosine kinases (RTKs) are a highly conserved family of signaling proteins with functions in cellular migration, adhesion, apoptosis, and proliferation during both adult and embryonic life. Here, we describe a knock-in mouse in which EphA1 expression is disrupted via the insertion of an internal ribosome entry site (IRES)-human placental alkaline phosphatase (ALPP) reporter cassette into exon II of the EphA1 gene. This was shown to successfully knockout expression of endogenous EphA1 and enforce expression of the ALPP reporter by the EphA1 promoter. Staining for the ALPP reporter protein demonstrated an epithelially restricted expression pattern in mouse tissues. In EphA1 null mice, two separate phenotypes were identified: abnormal tail development manifesting as a kinky tail was found in approximately 80% of homozygous adults. A second, distinct abnormality present in approximately 18% of females was characterized by imperforate uterovaginal development with hydrometrocolpos and caused by a resistance of cells to apoptosis during reproductive tract canalization. These results indicate a possible role for EphA1 in tissue patterning and hormone-induced apoptotic processes.

A Phase 1 study of the safety, pharmacokinetics and anti-leukemic activity of the anti-CD123 monoclonal antibody CSL360 in relapsed, refractory or high-risk acute myeloid leukemia.

Acute myeloid leukemia (AML) blasts express high levels of interlekin-3 (IL-3) receptor-α (CD123). CSL360 is a recombinant, chimeric immunoglobulin G1 (IgG1), anti-CD123 monoclonal antibody (MoAb) that neutralizes IL-3 and demonstrates anti-leukemic activity in vitro. This phase 1 study assessed safety, pharmacokinetics and bioactivity of weekly intravenous CSL360 for 12 weeks in 40 patients with advanced AML across five dose levels (0.1-10.0 mg/kg). Other than mild infusion reactions, CSL360 was well tolerated. The maximal tolerated dose was not reached. The half-life was 4.9 days, and the area under the curve (AUC) and maximum concentration (Cmax) increased proportionally with dose. Doses ≥ 3.0 mg/kg resulted in complete saturation and down-regulation of CD123 and abolition of ex vivo proliferative responsiveness to IL-3, indicating adequate blockade of IL-3 signaling. Two patients responded, with one remaining in complete remission after 17 doses. CSL360 bound CD123 specifically, but did not induce anti-leukemic activity in most patients. While safe, MoAb blockade of CD123 function is insufficient as a therapeutic strategy.

Tumor progression in hepatocellular carcinoma: relationship with tumor stroma and parenchymal disease.

BACKGROUND: Encapsulation in hepatocellular carcinoma is associated with decreased invasiveness and improved survival in several series. Although active fibrogenesis by myofibroblasts has been demonstrated in the capsule, it is unclear if the capsule results from a general increase in peritumoral fibrosis, or an inherently less invasive tumor phenotype. The relationship between collagen deposition within tumor stroma, presence of cirrhosis and invasiveness also needs clarification. METHODS: We performed immunohistochemistry for collagens I, III, IV and VI on sections of encapsulated and non-encapsulated hepatocellular carcinoma, arising in cirrhotic and non-cirrhotic livers. Staining was graded semi-quantitatively in tumor stromal elements and adjacent parenchymal sinusoids. The relationship of this staining with encapsulation, cirrhosis, and vascular invasion was analyzed. RESULTS: Formation of a discrete capsular layer was associated with reduced vascular invasion, but not with a pervasive increase in peritumoral fibrosis. Increased collagen I content of tumor stroma and adjacent parenchymal sinusoids was associated with non-encapsulated tumors and vascular invasion. The presence of cirrhosis had little effect on capsule composition. CONCLUSIONS: Encapsulation of hepatocellular carcinoma reflects reduced invasiveness, rather than increased peritumoral collagen synthesis, which may instead enhance invasion. Increased intratumoral collagen I protein is also associated with increased tumor invasiveness. Pre-existing cirrhosis has little effect on tumor progression, possibly because the characteristics of cirrhosis are overwhelmed by tumor-induced changes in the adjacent parenchyma.