Human prostate cancer initiating cells isolated directly from localized cancer do not form prostaspheres in primary culture
Chemical Therapeutic Program, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, Maryland 21231, USA. The Prostate
(Impact Factor: 3.57).
09/2012; 72(13):1478-89. DOI: 10.1002/pros.22503
Recent experimental studies suggest that hierarchical expansion from a minor population of cancer cells with an unlimited self-renewal capacity, termed cancer initiating cells (CICs), drives both lethality and heterogeneity of prostate cancer. Human prostate CICs have been established from only two primary prostate cancer patients, with the remaining established CIC lines being derived from metastatic sites from <10 patients. This suggests that the established CIC lines are significant "outliers" and may not be representative of the prostate CICs seen clinically. Thus, there is an urgent need to develop new approaches to achieve the "routine" establishment of CIC containing lines, particularly derived from primary prostate cancers.
In the present studies, we confirmed that in serum free, high Ca(2+) (i.e., DMEN: F12) growth factor defined (GFD) media plus androgen, a large (n = 10) series of established human prostate cancer cell lines derived from both localized and metastatic sites characteristically self-associate in suspension and grow as unattached spheroids, termed prostaspheres which contain CICs based upon their self-renewal in vitro and tumorigenicity in vivo.
Unfortunately, however, while dissociated single cells from human primary prostate cancer tissues are viable, contain CICs as documented by their ability to take and proliferate as xenografts, and produce prostaspheres when plated with serum free, high Ca(2+) /GFD-media plus androgen onto standard tissue culture flask, these prostasphere do not contain CICs.
The development of reproducibly methods to culture CICs isolated directly from localized cancers is still an urgent unmeet need of the prostate cancer research community.
Available from: W Nathaniel Brennen
- "From these explanted cells, outgrowth of fibroblast-like prostate cancer-derived stromal cells (PrCSCs) (Figure 1A, B) was observed that had a similar morphology to human bone marrow-derived MSCs (hBM-MSCs) (Figure 1C, D). If a portion of the same cellular suspension was cultured in keratinocyte serum-free media (K-SFM), basal-like prostate-derived epithelial cells (PrECs) were obtained [25-28]. Both hBM-MSCs and PrCSCs stained positive for alpha-smooth muscle actin (aSMA) (Figure 1E, G) and vimentin (Vim) (Figure 1F, H), but not cytokeratins 5 (CK5) (Figure 1I, K) or 8 (CK8) (Figure 1J, L). "
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ABSTRACT: Circulating bone marrow-derived Mesenchymal Stem Cells (BM-MSCs) have an innate tropism for tumor tissue in response to the inflammatory microenvironment present in malignant lesions. The prostate is bombarded by numerous infectious and inflammatory insults over a lifetime. Chronic inflammation is associated with CXCL12, CCL5, and CCL2, which are highly overexpressed in prostate cancer. Among other cell types, these chemoattractant stimuli recruit BM-MSCs to the tumor. MSCs are minimally defined as plastic-adhering cells characterized by the expression of CD90, CD73, and CD105 in the absence of hematopoietic markers, which can differentiate into osteoblasts, chondrocytes, and adipocytes. MSCs are immunoprivileged and have been implicated in tumorigenesis through multiple mechanisms, including promoting proliferation, angiogenesis, and metastasis, in addition to the generation of an immunosuppressive microenvironment. We have demonstrated that MSCs represent 0.01-1.1% of the total cells present in core biopsies from primary human prostatectomies. Importantly, these analyses were performed on samples prior to expansion in tissue culture. MSCs in these prostatectomy samples are FAP-, CD90-, CD73-, and CD105-positive, and CD14-, CD20-, CD34-, CD45-, and HLA-DR-negative. Additionally, like BM-MSCs, these prostate cancer-derived stromal cells (PrCSCs) were shown to differentiate into osteoblasts, adipocytes and chondrocytes. In contrast to primary prostate cancer-derived epithelial cells, fluorescently-labeled PrCSCs and BM-MSCs were both shown to home to CWR22RH prostate cancer xenografts following IV injection. These studies demonstrate that not only are MSCs present in sites of prostate cancer where they may contribute to carcinogenesis, but these cells may also potentially be used to deliver cytotoxic or imaging agents for therapeutic and/or diagnostic purposes.
Available from: Shunqi Wang
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ABSTRACT: This study was to enrich prostate cancer stem cells (PrCSC) from primary prostate cancer cultures (PPrCC). Primary prostate cancer cells were amplified in keratinocyte serum-free medium with epidermal growth factor (EGF) and bovine pituitary extract (BPE), supplemented with leukemia inhibitory factor (LIF), stem cell factor (SCF) and cholera toxin. After amplification, cells were transferred into ultra-low attachment dishes with serum-free DMEM/F12 medium, supplemented with EGF, basic fibroblast growth factor (bFGF), bovine serum albumin (BSA), insulin, and N2 nutrition. Expression of cell-type-specific markers was determined by RT-qPCR and immunostaining. Tumorigenicity of enriched PrCSC was determined by soft agar assay and xenograft assay in NOD/SCID mice. Biopsy samples from 19 confirmed prostate cancer patients were used for establishing PPrCC, and 18 cases (95%) succeeded. Both basal marker (CK5) and luminal markers (androgen receptor and CK8) strongly co-expressed in most of PPrCC, indicating their basal epithelial origin. After amplification under adherent culture condition in vitro, transient amplifying cells were the dominant cells. Sphere formation efficiency (SFE) of passaged PPrCC was about 0.5%, which was 27 times lower than SFE of LNCaP (13.67%) in the same condition. Compared with adherent cells from PPrCC, prostasphere from PPrCC showed up regulated stem cell markers and increased tumorigenic potential in soft-agar assay. However, spheroid cells from PPrCC prostasphere failed to initiate tumor in xenograft assay in 6 months. Thus, PPrCC can be established and amplified from prostate cancer biopsy samples. Our modified sphere culture system can enrich PrCSC from PPrCC.
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ABSTRACT: Tumors constitute complex ecosystems with multiple interactions among neoplastic cells displaying various phenotypes and functions and where the tumoral niche is built with an active participation of the host environment that also impacts the malignant progression of the tumor cells. Irrespective of the cell of origin of prostate adenocarcinoma, mounting evidences support the existence of a hierarchy within neoplastic prostate cells that contributes to the heterogeneity of these tumors. At the origin of this hierarchy are small populations of tumor cells with high self-renewal potential and also capable of generating progeny tumor cells that lose self-renewal properties as they acquire more differentiated phenotypes. These cancer stem cells (CSC) depend on active gene networks that confer them with their self-renewal capacity through symmetrical divisions whereas they can also undergo asymmetrical division and differentiation either as stochastic events or in response to environmental cues. Although new experimental evidences indicate that this is can be a reversible process, thus blurring the distinction between CSCs and non-CSCs, the former are considered as the drivers of tumor growth and evolution, and thus a prime target for therapeutic intervention. Of particular importance in prostate cancer, CSCs may constitute the repository population of androgen-insensitive and chemotherapy-resistant tumor cells responsible for castration-resistant and chemotherapy-insensitive tumors, thus their identification and quantification in primary and metastatic neoplasms could play important roles in the management of this disease.
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