Keratinocytes coordinate inflammatory responses and regulate development of secondary lymphedema.

Epidermal changes are histological hallmarks of secondary lymphedema, but it is unknown if keratinocytes contribute to its pathophysiology. Using clinical lymphedema specimens and mouse models, we show that keratinocytes play a primary role in lymphedema development by producing T-helper 2 (Th2) -inducing cytokines. Specifically, we find that keratinocyte proliferation and expression of protease-activated receptor 2 (PAR2) are early responses following lymphatic injury and regulate the expression of Th2-inducing cytokines, migration of Langerhans cells, and skin infiltration of Th2-differentiated T cells. Furthermore, inhibition of PAR2 activation with a small molecule inhibitor or the proliferation inhibitor teriflunomide (TF) prevents activation of keratinocytes stimulated with lymphedema fluid. Finally, topical TF is highly effective for decreasing swelling, fibrosis, and inflammation in a preclinical mouse model. Our findings suggest that lymphedema is a chronic inflammatory skin disease, and topically targeting keratinocyte activation may be a clinically effective therapy for this condition.

TGF-ß1 mediates pathologic changes of secondary lymphedema by promoting fibrosis and inflammation.

BACKGROUND: Secondary lymphedema is a common complication of cancer treatment, and previous studies have shown that the expression of transforming growth factor-beta 1 (TGF-ß1), a pro-fibrotic and anti-lymphangiogenic growth factor, is increased in this disease. Inhibition of TGF-ß1 decreases the severity of the disease in mouse models; however, the mechanisms that regulate this improvement remain unknown. METHODS: Expression of TGF-ß1 and extracellular matrix molecules (ECM) was assessed in biopsy specimens from patients with unilateral breast cancer-related lymphedema (BCRL). The effects of TGF-ß1 inhibition using neutralizing antibodies or a topical formulation of pirfenidone (PFD) were analyzed in mouse models of lymphedema. We also assessed the direct effects of TGF-ß1 on lymphatic endothelial cells (LECs) using transgenic mice that expressed a dominant-negative TGF-ß receptor selectively on LECs (LECDN-RII ). RESULTS: The expression of TGF-ß1 and ECM molecules is significantly increased in BCRL skin biopsies. Inhibition of TGF-ß1 in mouse models of lymphedema using neutralizing antibodies or with topical PFD decreased ECM deposition, increased the formation of collateral lymphatics, and inhibited infiltration of T cells. In vitro studies showed that TGF-ß1 in lymphedematous tissues increases fibroblast, lymphatic endothelial cell (LEC), and lymphatic smooth muscle cell stiffness. Knockdown of TGF-ß1 responsiveness in LECDN-RII resulted in increased lymphangiogenesis and collateral lymphatic formation; however, ECM deposition and fibrosis persisted, and the severity of lymphedema was indistinguishable from controls. CONCLUSIONS: Our results show that TGF-ß1 is an essential regulator of ECM deposition in secondary lymphedema and that inhibition of this response is a promising means of treating lymphedema.

Pilot Study of Anti-Th2 Immunotherapy for the Treatment of Breast Cancer-Related Upper Extremity Lymphedema.

Recent studies suggest that Th2 cells play a key role in the pathology of secondary lymphedema by elaborating cytokines such as IL4 and IL13. The aim of this study was to test the efficacy of QBX258, a monoclonal IL4/IL13 neutralizing antibody, in women with breast cancer-related lymphedema (BCRL). We enrolled nine women with unilateral stage I/II BCRL and treated them once monthly with intravenous infusions of QBX258 for 4 months. We measured limb volumes, bioimpedance, and skin tonometry, and analyzed the quality of life (QOL) using a validated lymphedema questionnaire (Upper Limb Lymphedema 27, ULL-27) before treatment, immediately after treatment, and 4 months following treatment withdrawal. We also obtained 5 mm skin biopsies from the normal and lymphedematous limbs before and after treatment. Treatment was well-tolerated; however, one patient with a history of cellulitis developed cellulitis during the trial and was excluded from further analysis. We found no differences in limb volumes or bioimpedance measurements after drug treatment. However, QBX258 treatment improved skin stiffness (p < 0.001) and improved QOL measurements (Physical p < 0.05, Social p = 0.01). These improvements returned to baseline after treatment withdrawal. Histologically, treatment decreased epidermal thickness, the number of proliferating keratinocytes, type III collagen deposition, infiltration of mast cells, and the expression of Th2-inducing cytokines in the lymphedematous skin. Our limited study suggests that immunotherapy against Th2 cytokines may improve skin changes and QOL of women with BCRL. This treatment appears to be less effective for decreasing limb volumes; however, additional studies are needed.