The liposome-incorporating cell wall skeleton of Mycobacterium bovis bacillus Calmette-Guein can directly enhance the susceptibility of cancer cells to lymphokine-activated killer cells through up-regulation of natural-killer group 2, member D ligands

ArticleinBJU International 108(9):1520-6 · February 2011with11 Reads
DOI: 10.1111/j.1464-410X.2010.10056.x · Source: PubMed
Abstract
• To conduct a preclinical evaluation of the ability of natural killer cells to cytolyze bladder cancer cells that were modified to show enhanced expression of natural-killer group 2, member D (NKG2D) ligands by R8-liposome-bacillus Calmette-Guéin (BCG)-cell wall skeleton (CWS) treatment. • The T24 cells and RT-112 cells were co-cultured with R8-liposome-BCG-CWS and BCG for 2, 4, or 6 h, and then the surface expression of NKG2D ligands was analyzed using TaqMan real-time quantitative RT-PCR. • Peripheral blood mononuclear cells were obtained with a conventional preparation kit, and then lymphokine-activated killer (LAK) cells were generated from these purified peripheral blood mononuclear cells via interleukin-2 stimulation. • The anti-tumour effect of LAK cells against untreated and R8-liposome-BCG-CWS co-cultured with cells of the human bladder cancer cell lines T24 and RT-112 was analyzed using the cytotoxic WST-8 assay method at 4 h of culture at various effector/target (E : T) ratios. • Major histocompatibility complex class I-related chain B (MICB) expression was increased ≈1.5-fold on T24 cells and RT-112 cells with BCG. • UL-16-binding protein (ULBP) 1 expression was also increased ≈1.5-fold on T24 cells and RT-112 cells with BCG. R8-liposome-BCG-CWS increased the surface expression of MICB 2.2-fold on T24 cells but did not increase it significantly on RT-112 cells. • ULBP1 expression was increased ≈2.2-fold on RT-112 cells, although no differences were observed between the expression of ULBP2 and 3 with R8-liposome-BCG-CWS. • T24 cells that were co-cultured with R8-liposome-BCG-CWS showed an ≈1.3-fold increase in sensitivity to cytolysis by LAK cells at an E : T ratio of 4 and RT-112 cells showed an ≈1.4-fold increase at an E : T ratio of 2. • In the present study, the induction of surface NKG2D ligands by R8-liposome-BCG-CWS rendered cancer cells more susceptible to cytolysis by LAK cells. • T24 cells and RT-112 cells, even when cultured singly in the absence of immune cells, can directly respond to R8-liposome-BCG-CWS. • The results obtained in the present study may therefore indicate a novel adoptive immunotherapy against bladder cancers.
    • "Principle lipid PAMP PRR Immune response Reference CAF01 (DDA) TDB Mincle IgG2 production, Th1/Th17 activation [42] CAF04 (DDA) MMG Non-TLR Moderate levels of IFN-γ-or TNF-α-producing CD4+ T cells [43] CAF05 (DDA) TDB/Poly I:C Mincle/TLR3 CD4 and CD8 cross-primed T cell response [44] CAF06 (DDA) TDB/MPL Mincle/TLR4 CD8 T cell and humoral Ab response [44] CAF09 (DDA) MMG Non-TLR Strong cross-primed Th1, Th17, IgG1 and IgG2 response [45] AS01 (proprietary) MPL TLR4 Th1 and anti-malarial Ab response [46] AS01b (proprietary) MPL/QS-21 TLR4/non-TLR? Specific and persistent humoral, CD4 and CD8 response [47] Tecemotide (Stimuvax) [L-BLP25] MPL TLR4 Th1 and CD8 response to antigen [48] Lipovaxin-MM (proprietary) Fg115/Lipokel (proprietary) TLR5 / TLR2 – [49] ONT-10 PET lipid A TLR4 Th1 and CD8 response to antigen, IgG2b [50] pegPC/cholestrol/Octa-arginine SA Alpha-galactosyl ceramide NK associated TLRs Natural killer cell activation, tumour targeting [51] DOTAP-PIC Poly I:C TLR3 Tumour specific CTL, IFN gamma production [52] DOPE/DOTAP MP lipid A TLR4 bFGF-associated metastasis inhibition [53] PC/cholesterol BCG-CWS TLR2/TLR4 Effective carcinoma targeting [54] DPPC/cholesterol Poly I:C/oligomannose TLR3/MBL High specific IgG and IgA response [55] RAFTsomes MHCII complex microdomain CD4 High specific IgG1 production [56] PC/cholesterol Galactosyl-DLPE MGL Specific Ab and IL 4/5/6 response [57] DDA trehalose 6,6′-diester analogues Mincle TDB-Comparable T-Cell responses [58] Mannosylated ErbB2/HER2 peptide liposomes Pam3CAG/PAM2CGD TLR 1/2/6 Effective CTL anti-tumour response [59] PC/DC-chol/pegDSPE/Protamine Oblimersen G3139 oligonucleotide TLR9 NK and DC driven anti-tumour response [60] DDAB/Cholesterol Mannosylated cholesterol MBL DC CD40 and CD80 upregulation [61] approaches to be considered in this field; among them antibody derived vectors [79], viral envelope derived vectors [80] and carbon nanotube carrier vectors [81]. Each approach will, with further investigation, present its own array of advantages and disadvantages surrounding their compositions, delivery mechanics and vector-genetic material interactions. "
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  • [Show abstract] [Hide abstract] ABSTRACT: Since the first report in 1976, accumulated clinical evidence has supported intravesical BCG therapy as one of the standard methods of management of intermediate- and high-risk non-muscle invasive bladder cancer. Despite its efficacy, intravesical BCG therapy is associated with a variety of adverse events (AEs), most of which are tolerable or controllable with supportive care. However, some patients receiving intravesical BCG therapy may be experience uncommon but severe AEs, leading to cessation of BCG therapy. Not all, but most severe AEs result from either local or systemic infection with live BCG. Intravesical instillation of BCG elicits multiple immune reactions, although the precise immunological mechanism of BCG therapy is not clear. It is convenient to separate the complex reactions to the following three categories: infection of urothelial cells or bladder cancer cells, induction of immune reactions, and induction of antitumor effects. Recently, our knowledge about each category has increased. Based on this understanding, predictors of the efficacy of intravesical BCG therapy, such as urinary cytokine measurement and cytokine gene polymorphism, have been investigated. Recently, preclinical studies using a novel engineered mycobacterium vaccine have been conducted to overcome the limitations of BCG therapy. One approach is Th1 cytokine-expressing recombinant forms of BCG; another approach is development of non-live bacterial agents to avoid AEs due to live BCG infection. We also briefly describe our approach using an octaarginine-modified liposome-incorporating BCG cell wall component to develop future substitutes for live BCG.
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  • [Show abstract] [Hide abstract] ABSTRACT: The Mycobacterium bovis Bacille Calmett-Guerin cell wall skeleton (BCG-CWS) could be used to replace live BCG as a bladder cancer drug. However, because BCG-CWS is poorly soluble, has a strong-negative charge, very high molecular weight and heterogeneity in size of tens of μm, it cannot be used in such an application. We report herein on the development of a novel packaging method that permits BCG-CWS to be encapsulated into 166nm-sized lipid particles. The BCG-CWS encapsulated nano particle (CWS-NP) has a high uniformity and can be easily dispersed. Thus, it has the potential for use as a packaging method that would advance the scope of applications of BCG-CWS as a bladder cancer drug. In a functional evaluation, CWS-NP was efficiently taken up by mouse bladder tumor cells (MBT-2) in vitro and inhibited tumor growth in mice bearing MBT-2 tumors. Moreover, intravesically administered CWS-NP showed significant antitumor effects in a rat model with in naturally developed bladder cancer. An enhancement in Th1 differentiation by CWS-NP was also confirmed in human T cells. In conclusion, CWS-NP represents a promising delivery system for BCG-CWS for clinical development as a potent bladder cancer drug.
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