Suppression of breast cancer cell growth by a monoclonal antibody targeting cleavable ErbB4 isoforms.

ErbB4 isoforms mediate different cellular activities depending on their susceptibility to proteolytic cleavage. The biological significance of ErbB4 cleavage in tumorigenesis, however, remains poorly understood. Here, we describe characterization of a monoclonal antibody (mAb 1479) that selectively recognizes the ectodomain of cleavable ErbB4 JM-a isoforms both in vitro and in vivo. mAb 1479 was used to analyse ErbB4 JM-a expression and ectodomain shedding in a series of 17 matched breast cancer/histologically normal peripheral breast tissue pairs. ErbB4 ectodomain was observed in 75% of tumors expressing ErbB4 but only in 18% of normal breast tissue samples expressing ErbB4. Difference in the relative quantity of ErbB4 ectodomain between normal and tumor tissue pairs was statistically significant (P=0.015). Treatment with mAb 1479 suppressed ErbB4 function by inhibiting ErbB4 tyrosine phosphorylation and ectodomain shedding, and by stimulating ErbB4 downregulation and ubiquitination. mAb 1479 suppressed both anchorage-dependent and -independent growth of human breast cancer cell lines that naturally express cleavable ErbB4 JM-a. These findings indicate that ErbB4 ectodomain shedding is enhanced in breast cancer tissue in vivo, and that mAb 1479 represents a potential drug candidate that suppresses breast cancer cell growth by selectively binding cleavable ErbB4 isoforms.

Vascular endothelial growth factor plays a major role in development of experimental obliterative bronchiolitis.

Obliterative bronchiolitis (OB) is the major limitation for long-term survival of lung allograft recipients. The exact molecular and cellular mechanisms contributing to obliterative lesion formation are unknown. Pathological characteristics of OB are epithelial damage, peribronchial inflammation, and increasing obliteration of bronchioli. Vascular endothelial growth factor (VEGF) is an angiogenic growth factor that exerts proinflammatory effects by increasing endothelial permeability and inducing expression of endothelial adhesion molecules. We investigated the role of VEGF in the development of OB in rat tracheal allografts and the role of VEGF receptors (VEGFR)-1 and -2 in the development of OB in mouse tracheal allografts. In nontreated allografts, with increasing loss of epithelium and airway occlusion, VEGF messenger RNA (mRNA) and protein expression vanished in the epithelium and increased in smooth muscle cells and mononuclear inflammatory cells compared with syngeneic grafts. Intragraft VEGF overexpression by adenoviral transfer of a mouse VEGF164 gene led to a decrease in epithelial necrosis but increased luminal occlusion by >50% compared with AdLacZ-treated rat tracheal allografts. When compared with the control immunoglobulin (Ig)G group, simultaneous treatment with antibodies against VEGFR-1 and -2 significantly lowered the degree of luminal occlusion of mouse tracheal allografts.

Amplification of HER-2 in gastric carcinoma: association with Topoisomerase IIalpha gene amplification, intestinal type, poor prognosis and sensitivity to trastuzumab.

BACKGROUND: HER-2/neu gene amplification has predictive value in breast cancer patients responding to trastuzumab. We wanted to investigate the frequency and clinical significance of HER-2/neu amplification in gastric carcinoma. PATIENTS AND METHODS: The frequency of HER-2/neu and Topoisomerase IIalpha gene amplification was studied in adenocarcinomas of the stomach (n=131) and the gastroesophageal junction (n=100) by chromogenic in situ hybridization (CISH). Sensitivity of a gastric cancer cell line N87 with HER-2/neu amplification to trastuzumab was studied by a cell viability assay and compared with that of a HER-2 amplified breast cancer cell line SKBR-3. Growth inhibition of N87 cells was also verified in vivo in N87 xenograft tumors. RESULTS: HER-2/neu amplification was present in 16 (12.2%) of the 131 gastric and in 24 (24.0%) of the 100 gastroesophageal adenocarcinomas. Co-amplification of Topoisomerase IIalpha was present in the majority of gastric (63%) and esophagogastric junction cancers (68%) with HER-2/neu amplification. HER-2/neu amplification was more common in the intestinal histologic type of gastric cancer (21.5%) than in the diffuse (2%) or the mixed/anaplastic type (5%, P=0.0051), but it was not associated with gender, age at diagnosis or clinical stage. Presence of HER-2/neu amplification was associated with poor carcinoma-specific survival (P=0.0089). HER-2/neu targeting antibody trastuzumab inhibited the growth of a p185(HER-2/neu) overexpressing gastric and breast carcinoma cell lines (N87 and SKBR-3) with equal efficacy. CONCLUSIONS: HER-2/neu amplification is common in the intestinal type of gastric carcinoma, and it is associated with a poor outcome. HER-2 might be a useful target in this disease, and this hypothesis deserves to be investigated in clinical trials.

Suppression of viability and acetyl-LDL metabolism in RAW 264 macrophage-like and smooth muscle cells by bisphosphonates in vitro.

Etidronate and clodronate are bisphosphonates that inhibit the development of experimental atherosclerosis. Etidronate decreases the intimamedia thickness of carotid artery even in man. Liposome-encapsulated bisphosphonates inhibit the cellular metabolism of atherogenic, modified low-density lipoprotein (acetyl-LDL) by cultured macrophages. In the present study, the effects of new bisphosphonate tiludronate and nitrogen-containing bisphosphonate alendronate on cell viability and cellular uptake and degradation of acetyl-LDL were investigated in vitro with macrophages and arterial smooth muscle cells, which have a significant role in atherogenesis. Tiludronate and alendronate decreased the viability of RAW 264 macrophages at high concentration (1,000 microM; p < 0.05), while liposome-encapsulated drugs suppressed the viability at concentrations of 30-300 microM. At concentrations greater than or equal to 10 microM, tiludronate and alendronate inhibited the uptake and degradation of acetyl-LDL by RAW 264 cells in a concentration-dependent manner (p < 0.001). None of the bisphosphonates affected the viability of smooth muscle cells, and none but alendronate at a high concentration (1,000 microM) inhibited the uptake and degradation of acetyl-LDL by smooth muscle cells. The results show that tiludronate and alendronate inhibit the atherogenic activity of macrophages in vitro, as shown previously with etidronate and clodronate, providing further evidence for the antiatherogenic effects of bisphosphonates.