A role for phospholipase C-gamma-mediated signaling in tumor cell invasion.

The invasive and metastatic transformation of cancers often results in death. However, the mechanisms that promote this transformation remain unclear. Two closely related receptors, the epidermal growth factor receptor (EGFR) and ErbB2, are overexpressed in a significant percentage of breast and prostate carcinomas, among others, with this up-regulated signaling correlating with tumor progression. Previous studies in our laboratory have demonstrated that an EGFR-phospholipase C (PLC)gamma-mediated motility-associated signaling pathway is rate-limiting for tumor cell invasion in vitro and in vivo in one model of prostate carcinoma. Therefore, we investigated whether this PLCgamma signaling pathway also was rate-limiting for invasion in other tumor cell lines and types and whether this EGFR activity is subsumed by the closely related ErbB2. We determined the effects of PLCgamma signal abrogation by pharmacological (U73122) and molecular (expression of the dominant-negative PLCz) means on the in vitro invasiveness of tumor cells. Inhibition of PLCgamma signaling concomitantly decreased invasiveness of de novo-occurring transgenic adenocarcinoma mouse prostate (TRAMP) lines and the human breast cancer cell lines MDA-468 and MDA-231; these lines present up-regulated EGFR signaling. Because the prostate and breast cancer lines usually present autocrine stimulatory loops involving EGFR, we also examined transgenic adenocarcinoma mouse prostate C1 and MDA-468 treated with the EGFR-specific kinase inhibitor PD153035 to determine whether invasiveness is dependent on EGFR signaling. PD153035 reduced invasiveness to levels similar to those seen with U73122, suggesting that the autocrine EGFR stimulatory loop is functioning to promote invasiveness. To determine whether this signaling pathway also promotes invasiveness of ErbB2-overexpressing tumors, we examined the human breast carcinoma line MDA-361; again, U73122 inhibition of PLCgamma decreased invasiveness. In all situations, the inhibition of PLCgamma signaling did not decrease mitogenic signaling. Thus, the motility-associated PLCgamma signaling pathway is a generalizable rate-limiting step for tumor cell progression.

Transcriptional suppression of matrix metalloproteinase-2 gene expression in human astroglioma cells by TNF-alpha and IFN-gamma.

Matrix metalloproteinases (MMPs) are a family of zinc-dependent endopeptidases that function in the turnover of extracellular matrix components during development. In addition, MMPs also contribute to pathological conditions associated with inflammation, angiogenesis, and tumor invasion. A 72-kDa type IV collagenase, also referred to as gelatinase A or MMP-2, has been proposed to potentiate the invasion and metastasis of malignant tumors. In particular, MMP-2 activity has been shown to constitute an important component of human astroglioma invasion. We investigated the influence of various cytokines, both proinflammatory and immunosuppressive, on MMP-2 gene expression in two human astroglioma cell lines (U251-MG and CRT). Our results indicate that the cell lines constitutively express high levels of MMP-2 mRNA, protein, and bioactivity as assessed by ribonuclease protection assay, immunoblotting, and zymography assays, respectively. The proinflammatory cytokines TNF-alpha and IFN-gamma individually can inhibit constitutive MMP-2 expression, and function in an additive manner for near-complete inhibition of MMP-2 expression. Inhibition of MMP-2 mRNA levels by TNF-alpha and IFN-gamma is not due to destabilization of the MMP-2 message; rather, inhibition is mediated at the transcriptional level. Furthermore, TNF-alpha/IFN-gamma inhibition of MMP-2 expression results in decreased invasiveness of the human astroglioma cells through an extracellular matrix. These results raise the possibility that TNF-alpha and IFN-gamma may have beneficial effects in attenuating astroglioma invasive properties.