Effect of doxycycline on proliferation, MMP production, and adhesion in LAM-related cells

Abstract

Matrix metalloproteinases (MMPs) have been implicated in lung cyst formation in lymphangioleiomyomatosis (LAM). As doxycycline inhibits MMP activity in vivo, some patients take doxycycline, as one report has suggested a possible benefit in LAM. However, there have been no randomized controlled clinical trials of doxycycline for LAM, and any mechanism of action is unclear. Here, we examine previously proposed mechanisms of actions. Cell proliferation and adhesion were examined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction and Cytomatrix cell adhesion kits. Apoptosis was examined by TdT-mediated dUTP nick end labeling (TUNEL) assay. MMP-2 expression was examined by quantitative real-time PCR and zymography in doxycycline-treated ELT3 cells and tumor growth using angiomyolipoma-derived tumor xenografts in nude mice. In ELT3 cells, ≥25 μg/ml doxycycline decreased proliferation, increased apoptosis, and caused a change in cell morphology associated with redistribution of actin stress filaments. Reduction in proliferation was also seen in human angiomyolipoma-derived cells. Cell adhesion to ECM proteins was decreased by doxycycline at 50 μg/ml and prevented detachment of already adherent cells. There was no effect of doxycycline on MMP-2 expression or activity in vitro. In the xenograft model, doxycycline (30 mg·kg−1·day−1) had no effect on tumor growth, final tumor weight, or tumor lysate MMP levels. Doxycycline at doses ≥ 25 μg/ml inhibited cell proliferation and adhesion, possibly by a toxic effect. Doxycycline had no effect on MMP-2 expression or activity or tumor growth in the xenograft model. Any possible in vivo effect is unlikely to be mediated by MMP-2 or reduced cell proliferation.

Faslodex Inhibits Estradiol-Induced Extracellular Matrix Dynamics and Lung Metastasis in a Model of Lymphangioleiomyomatosis

“…Therefore, despite tremendous progress in LAM, the urgent need remains for improved therapeutic strategies. Our promising preclinical results with Faslodex add to a growing list of agents approved by the United States Food and Drug Administration for their potential efficacy in LAM, including simvastatin (37), chloroquine (38), and doxycycline (39). Faslodex may be of particular benefit in premenopausal women, in whom higher estradiol concentrations may be associated with higher rates of lung function decline.…”

Doxiciclina em pacientes com linfangioleiomiomatose: segurança e eficácia no bloqueio de metaloproteinases

“…(23,24) Chang et al demonstrated the in vitro effect of doxycycline on LAM cell adhesion, as well as on MMP-2 and MMP-9, revealing a reduction in cell proliferation, although very high doses of doxycycline were required and there was no significant blockade of MMP expression. (25) These findings, however, are questionable, because, to date, there have been no pathophysiological studies involving animal models of the LAM lung. Although the mechanism of action of doxycycline remains unclear, it is speculated that it inhibits MMPs by inducing TIMP activity.…”

LAM cells biology and lymphangioleiomyomatosis

“…Clinical trials have shown that administration of doxycycline to LAM patients resulted in lower levels of MMP-2, MMP-9 in serum and urine, decreased symptoms and improved life quality [36,37]. However, in vitro studies showed that there was no significant reduction in the level and activity of MMPs in the ELT3 -/-cells, derived from an animal model of LAM, the Eker rat, after doxycycline cells treatment [38]. Thus, the efficacy of doxycycline to inhibit MMPs activity in LAM patients needs to be further investigated.…”

“…mRNA of gp100, and MART-1 (spindle-shaped cells) and estrogen receptors was detected in these cells by RT-PCR. Cells were cultured in the presence of epidermal growth factor [52] and used to study the role of chemokines in LAM and angiomyolipoma [29] as well as the effects of doxycycline on the proliferation, production of MMPs and adhesion of LAM-related cells [38]. Yu et al [53] described a line derived from human angiomyolipoma.…”