P190B RhoGAP has pro-tumorigenic functions during MMTV-Neu mammary tumorigenesis and metastasis.

INTRODUCTION: Rho GTPases are overexpressed and hyperactivated in human breast cancers. Deficiency of p190B RhoGAP, a major inhibitor of the Rho GTPases, inhibits mouse mammary tumor virus long terminal repeat (MMTV)-Neu/ErbB2 mammary tumor formation and progression in part through effects within the stromal environment, suggesting that p190B function is pro-tumorigenic. To further investigate the potential pro-tumorigenic actions of p190B, we examined the effects of exogenous p190B expression within the mammary epithelium on MMTV-Neu tumor formation and progression. METHODS: Tetracycline-regulatable p190B transgenic mice were bred to MMTV-Neu mice, and the effects of exogenous p190B expression on tumor latency, multiplicity, growth rates, angiogenesis, and metastasis were examined. The effects of exogenous p190B expression on cell-matrix adhesion and invasion were tested using non-transformed primary mammary epithelial cells (MECs). Rho GTPase activity, oxidative stress as an indicator of reactive oxygen species (ROS) production, and downstream signaling pathways were analyzed. RESULTS: Altered p190B expression resulted in a 2-fold increase in tumor multiplicity and a 3-fold increase in metastases compared to control mice indicating that exogenous p190B expression in the mammary epithelium promotes MMTV-Neu mammary tumor formation and progression. Interestingly, non-transformed primary MECs expressing exogenous p190B displayed increased adhesion to laminin and type IV collagen and formed invasive structures in a three-dimensional culture assay. Ras related C3 botulinum toxin 1 (Rac1)-GTP levels were elevated in p190B transgenic tumors whereas Ras homologous A (RhoA) and cell division cycle 42 (Cdc42)-GTP levels were not significantly altered. Rac1 activity affects production of ROS, which regulate transformation, metastasis, and oxidative stress. Protein carbonylation, which is indicative of oxidative stress, was elevated 1.75-fold in p190B transgenic tumors as compared to control tumors suggesting that exogenous p190B expression may affect Rac1-dependent ROS production. CONCLUSIONS: These studies indicate that paradoxically, p190B RhoGAP, a major inhibitor of the Rho GTPases in vitro, has pro-tumorigenic functions that enhance MMTV-Neu induced mammary tumor formation and metastasis. Furthermore, exogenous p190B expression enhances cell adhesion and invasion, which may facilitate metastasis. Rac1 activity and oxidative stress are elevated in tumors expressing exogenous p190B suggesting that p190B may promote tumorigenesis through a Rac1/ROS dependent mechanism.

Haploinsufficiency for p190B RhoGAP inhibits MMTV-Neu tumor progression.

INTRODUCTION: Rho signaling regulates key cellular processes including proliferation, survival, and migration, and it has been implicated in the development of many types of cancer including breast cancer. P190B Rho GTPase activating protein (RhoGAP) functions as a major inhibitor of the Rho GTPases. P190B is required for mammary gland morphogenesis, and overexpression of p190B in the mammary gland induces hyperplastic lesions. Hence, we hypothesized that p190B may play a pivotal role in mammary tumorigenesis. METHODS: To investigate the effects of loss of p190B function on mammary tumor progression, p190B heterozygous mice were crossed with an MMTV-Neu breast cancer model. Effects of p190B deficiency on tumor latency, multiplicity, growth, preneoplastic progression and metastasis were evaluated. To investigate potential differences in tumor angiogenesis between the two groups, immunohistochemistry to detect von Willebrand factor was performed and quantified. To examine gene expression of potential mediators of the angiogenic switch, an angiogenesis PCR array was utilized and results were confirmed using immunohistochemistry. Finally, reciprocal transplantation of tumor fragments was performed to determine the impact of stromal deficiency of p190B on tumor angiogenesis. RESULTS: P190B deficiency reduced tumor penetrance (53% of p190B+/-Neu mice vs. 100% of p190B+/+Neu mice formed tumors) and markedly delayed tumor onset by an average of 46 weeks. Tumor multiplicity was also decreased, but an increase in the number of preneoplastic lesions was detected indicating that p190B deficiency inhibited preneoplastic progression. Angiogenesis was decreased in the p190B heterozygous tumors, and expression of a potent angiogenic inhibitor, thrombospondin-1, was elevated in p190B+/-Neu mammary glands. Transplantation of p190B+/-Neu tumor fragments into wild-type recipients restored tumor angiogenesis. Strikingly, p190B+/+Neu tumor fragments were unable to grow when transplanted into p190B+/-Neu recipients. CONCLUSIONS: These data suggest that p190B haploinsufficiency in the epithelium inhibits MMTV-Neu tumor initiation. Furthermore, p190B deficiency in the vasculature is responsible, in part, for the inhibition of MMTV-Neu tumor progression.

Rearrangement of 3-membered 1,1,2-trifluorobromonium and iodonium ions and comparison of trifluorochloronium to fluorocarbenium ions.

Reactions of chlorine (Cl(2)) with 4-halo-1,1,2-trifluorobut-1-enes (1, 2, or 3) give open-ion intermediates A and E that are in equilibrium. The open-chloronium ions (E) rearrange to a five-membered-ring halonium ion during ionic chlorination of 3 when the number-4 halo-substituent is iodine. Three-membered-ring bromonium and iodonium ions from alkenes 1, 2, or 3 are rather symmetrical and similar in structure. Quantum chemical calculations show that five-membered-ring halonium ion intermediates are 11 to 27 kcal/mol more stable than the three-membered-ring halonium ions or the open-ions A and E. The five-membered-ring intermediates lead to rearranged products. Rearranged products increase as the number-4 halogen (Z) becomes more nucleophilic (Z: Cl < Br < I). Open chloronium ions from ionic chlorination of terminal fluorovinyl alkenes are compared to the open ions generated by protons to similar alkenes.