CD147-Cyclophilin a Interactions Promote Proliferation and Survival of Cutaneous T-Cell Lymphoma

Abstract

CD147, a transmembrane glycoprotein that belongs to the immunoglobulin superfamily, and cyclophilin A (CypA), one of the binding partners of CD147, are overexpressed in tumor cells and associated with the progression of several malignancies, including both solid and hematological malignancies. However, CD147 and CypA involvement in cutaneous T-cell lymphoma (CTCL) has not been reported. In this study, we examined CD147 and CypA expression and function using clinical samples of mycosis fungoides (MF) and Sézary syndrome (SS) and CTCL cell lines. CD147 and CypA were overexpressed by tumor cells of MF/SS, and CypA was also expressed by epidermal keratinocytes in MF/SS lesional skin. Serum CypA levels were increased and correlated with disease severity markers in MF/SS patients. Anti-CD147 antibody and/or anti-CypA antibody suppressed the proliferation of CTCL cell lines, both in vitro and in vivo, via downregulation of phosphorylated extracellular-regulated kinase 1/2 and Akt. These results suggest that CD147-CypA interactions can contribute to the proliferation of MF/SS tumor cells in both a autocrine and paracrine manner, and that the disruption of CD147-CypA interactions could be a new therapeutic strategy for the treatment of MF/SS.

Keywords: CD147; Sézary syndrome; cutaneous T-cell lymphoma; cyclophilin A; mycosis fungoides.

Figure 1

CD147 is overexpressed on tumor cells of mycosis fungoides (MF) and Sézary syndrome (SS). (A) Quantitative RT-PCR was performed to measure expression levels of CD147 using mRNA extracted from lesional skin of early MF (n = 10), advanced MF/SS (n = 13), and healthy skin (n = 12). (B) CD147 expression was analyzed by flow cytometry in CD4 + CD7- T cells from 4 SS patients, and CD4+ T cells from 4 healthy controls. Representative plots (left) and mean fluorescence intensity (MFI) (right) are shown. The measured values from individual patients are plotted with dots. Bars represent mean ± standard deviation. * p < 0.05, ** p < 0.01. (C) CD147 staining in MF/SS lesional skin (n = 12). Representative results are shown (original magnification ×400; scale bar = 100 μm). (D) CD147 expression was analyzed by flow cytometry in human cutaneous T-cell lymphoma cell lines (HH, Hut78, and MJ cells).

Figure 2

Cyclophilin A (CypA) is expressed on tumor cells and epidermal keratinocytes in mycosis fungoides (MF) and Sézary syndrome (SS). (A) Quantitative RT-PCR was performed to measure expression levels of CypA using mRNA extracted from lesional skin of early MF (n = 10), advanced MF/SS (n = 13), and healthy skin (n = 12). Bars represent mean ± 2 standard deviation. (B) CypA staining in MF/SS lesional skin (n = 7). Representative results are shown (original magnification ×200 in left three panels, and ×400 in the most right panel; scale bar = 50 μm in left three panels, and 100 μm in the right most panel. (C) Human cutaneous T-cell lymphoma cell lines (HH, Hut78, and MJ cells) were cultured for 24, 48, and 72 h. Supernatant CypA levels were evaluated by enzyme-linked immunosorbent assay. One representative result from two independent experiments. Data are presented as mean ± standard deviation (n = 4). * p < 0.05, ** p < 0.01, *** p < 0.005.

Figure 3

Serum cyclophilin A (CypA) levels are increased and correlated with disease severity markers in patients with mycosis fungoides (MF) and Sézary syndrome (SS). (A) Serum CypA levels in early MF patients (n = 21), advanced MF/SS patients (n = 23), and healthy control (n = 19). (B) Serum CypA levels in MF/SS patients (n = 7) before and after treatment. (C,D) Correlations between serum CypA levels and serum lactate dehydrogenase (LDH) levels (n = 44) (C) and serum soluble IL-2 receptor (sIL-2R) levels (n = 29) (D) in MF/SS patients. The measured values from individual patients are plotted with dots. Bars represent mean ± standard deviation. * p < 0.05.

Figure 4

CD147-CypA interactions promoted the proliferation of cutaneous T-cell lymphoma (CTCL) cell lines. (A,B) CTCL cell lines (HH, Hut78, and MJ cells) were cultured with anti-CD147 antibody (0.8, 8 μg/mL) or anti-CypA antibody (0.07, 0.7 μg/mL) for 48 h. Viable cells were counted. Data are presented as mean ± standard deviation. (C,D) Western blotting analysis was conducted on the lysates of HH cells treated with anti-CD147 antibody (1 μg/mL) or anti-CypA antibody (0.7 μg/mL) for 0, 1, 3, or 6 h. Phosphorylation of AKT, ERK1/2, p38 MAPK, and JNK was measured. A representative picture of three independent experiments is shown. Band intensities of phosphorylated AKT, ERK1/2, p38 MAPK, and JNK at each time were calculated by Image J software. n = 3. Data are presented as mean ± standard deviation. (E) HH cells (1.0 × 10⁷) were injected into Scid-beige mice with PBS or anti-CD147 antibody (8 μg/mL). Each reagent was injected on days 0, 4, 7, 11, 14, and 18. Tumor size was calculated on days 4, 7, 11, 14, 18, and 21. Data are presented as means ± standard error of the mean (n = 28). * p < 0.05, ** p < 0.01, *** p < 0.005, **** p < 0.0001.

Figure 5

Schematic model of the main findings of this study.