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. 2014 Oct 6;11(10):3443-51.
doi: 10.1021/mp500174p. Epub 2014 Sep 22.

Role of the phospholipase A2 receptor in liposome drug delivery in prostate cancer cells

N D Quach  1 , J N Mock, N E Scholpa, M W Eggert, C Payré, G Lambeau, R D Arnold, B S Cummings
Affiliations

Role of the phospholipase A2 receptor in liposome drug delivery in prostate cancer cells

N D Quach et al. Mol Pharm. .

Abstract

The M-type phospholipase A2 receptor (PLA2R1) is a member of the C-type lectin superfamily and can internalize secreted phospholipase A2 (sPLA2) via endocytosis in non-cancer cells. sPLA2 itself was recently shown to be overexpressed in prostate tumors and to be a possible mediator of metastasis; however, little is known about the expression of PLA2R1 or its function in prostate cancers. Thus, we examined PLA2R1 expression in primary prostate cells (PCS-440-010) and human prostate cancer cells (LNCaP, DU-145, and PC-3), and we determined the effect of PLA2R1 knockdown on cytotoxicity induced by free or liposome-encapsulated chemotherapeutics. Immunoblot analysis demonstrated that the expression of PLA2R1 was higher in prostate cancer cells compared to that in primary prostate cells. Knockdown of PLA2R1 expression in PC-3 cells using shRNA increased cell proliferation and did not affect the toxicity of cisplatin, doxorubicin (Dox), and docetaxel. In contrast, PLA2R1 knockdown increased the in vitro toxicity of Dox encapsulated in sPLA2 responsive liposomes (SPRL) and correlated with increased Dox and SPRL uptake. Knockdown of PLA2R1 also increased the expression of Group IIA and X sPLA2. These data show the novel findings that PLA2R1 is expressed in prostate cancer cells, that PLA2R1 expression alters cell proliferation, and that PLA2R1 modulates the behavior of liposome-based nanoparticles. Furthermore, these studies suggest that PLA2R1 may represent a novel molecular target for controlling tumor growth or modulating delivery of lipid-based nanomedicines.

Keywords: drug delivery; liposomes; nanoparticles; phospholipase A2 receptor; prostate cancer; secreted phospholipase A2; targeting.

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Figures

Figure 1
Figure 1
Expression of PLA2R1 in prostate cells. PLA2R1 protein expression was assessed using immunoblot analysis. The data are indicative of at least three separate experiments.
Figure 2
Figure 2
Effect of PLA2R1 inhibition on PC-3 cell growth. PLA2R1 expression was inhibited using shRNA. Scrambled shRNA was used as a control. Knockdown of PLA2R1 was verified by qRT-PCR (A) and immunoblot analysis (B). GAPDH was used as a house-keeping gene. Assessment of cell growth was performed using MTT assays (C) and crystal violet staining (D). Data in panels A–C are represented as the mean ± SEM of at least three separate experiments (n = 3/study). *Indicates a significant difference (p < 0.05) as compared to control cells. Data in panel D are indicative of at least three separate experiments.
Figure 3
Figure 3
Effect of PLA2R1 knockdown on chemotherapeutic-induced toxicity in PC-3 cells. Cells were treated with docetaxel (A), cisplatin (B), or Dox (C) for 72 h. MTT assays were used to determine the effect of knocking down PLA2R1 on chemotherapeutic-induced cytotoxicity. The effect of PLA2R1 on doxorubicin-induced toxicity was further assessed at 72 h using phase contrast microscopy at 10× magnification (D). Data in panels A–C are represented as the mean ± SD of at least three separate experiments (n = 3/study). Data in panel D are indicative of at least three separate experiments.
Figure 4
Figure 4
Effect of PLA2R1 knockdown on the toxicity of doxorubicin encapsulated in SSL and SPRL in PC-3 cells. PLA2R1 knockdown cells and those expressing scrambled shRNA were treated with 2.5 μM concentrations of Dox encapsulated in SSL or SPRL. MTT assays (A) and phase contrast microscopy at 40× magnification (B) were used to determine the effect of PLA2R1 knockdown on cytotoxicity. Data in panel A are represented as the mean ± SEM of at least three separate experiments (n = 3/study). *Indicates a significant difference (p < 0.05) as compared to cells transfected with scrambled shRNA. Data in panel E are indicative of at least three separate experiments.
Figure 5
Figure 5
Effect of PLA2R1 knockdown on DiO and doxorubicin uptake from SSL and SPRL in PC-3 cells. PC-3 cells were treated with liposomes containing Dox and DiO for 24 to 72 h. The efficiency of DiO and drug uptake via SSL (A, B) and SPRL (C, D) was quantified using flow cytometry. Data are represented as the mean ± SEM of at least three separate experiments (n = 3/study). *Indicates a significant difference (p < 0.05) as compared to cells expressing scrambled shRNA.
Figure 6
Figure 6
Effect of PLA2R1 knockdown on sPLA2 expression in PC-3 cells. PLA2R1 expression was inhibited in PC-3 cells using shRNA, and changes in the expression of various sPLA2 isoforms was determined by immunoblot analysis (A) and qRT-PCR (B). Data in panel A are indicative of at least three separate experiments. Data in panel B are represented as the mean ± SEM of at least three separate experiments (n = 3/study). *Indicates a significant difference (p < 0.05) as compared to cells expressing scrambled shRNA.
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