Linking hypoxia, DNA damage and proliferation in multicellular tumor spheroids

Riffle, Stephen; Pandey, Ram Naresh; Albert, Morgan L.; Hegde, Rashmi S.

doi:10.1186/s12885-017-3319-0

Cited by 106 publications

(102 citation statements)

References 43 publications

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“…Three‐dimensional cell culture systems have become more prominent in preclinical studies due to their ability to more closely represent tissue compartments and enable longer experimental time frames . Tumor spheroids, for example, can recapitulate aspects of the tumor microenvironment that ultimately influence drug response . A673 spheroid cultures were treated with ORY‐1001 and SP2509 for 10 days with a dose range between 0.3 and 10 µM (Figure and B).…”

Section: Resultsmentioning

confidence: 99%

“…27 Tumor spheroids, for example, can recapitulate aspects of the tumor microenvironment that ultimately influence drug response. 28 A673 spheroid cultures were treated with ORY-1001 and SP2509 for 10 days with a dose range between 0.3 and 10 µM (Figure 2 and 2B). Again, no effect on growth was observed with the irreversible inhibitor of KDM1A catalytic function ORY-1001, whereas SP2509 was active, albeit at higher concentrations compared with 2D cultures (Figure 2 and 2B).…”

Section: Resultsmentioning

confidence: 99%

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Catalytic inhibition of KDM1A in Ewing sarcoma is insufficient as a therapeutic strategy

Romo-Morales

Aładowicz

Blagg

et al. 2019

Pediatric Blood & Cancer

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Background Ewing sarcoma and desmoplastic small round cell tumors (DSRCT) are rare and clinically aggressive sarcomas usually characterized by oncogenic fusion proteins involving EWS. Emerging studies of Ewing sarcoma have demonstrated EWS‐FLI1‐driven chromatin remodeling as a key aspect of tumorigenicity. In particular, the lysine‐specific demethylase KDM1A/LSD1 is linked to transcriptional regulation of target genes orchestrated by the EWS portion of the fusion protein interacting with repressive chromatin‐remodeling complexes. Consistent with this model, depletion of KDM1A supports it is a molecular therapeutic target in Ewing sarcoma cells, but effective drugs need to be identified. Procedure A comprehensive phenotypic analysis of the effects of catalytic KDM1A inhibitors ORY‐1001 and GSK2879552, including clinically relevant doses, was carried out in 2D and 3D spheroid models of Ewing sarcoma and DSRCT. Results Catalytic inhibition of KDM1A did not affect cell viability in 2D and 3D assays and had no impact on invasion in a 3D assay. Conclusions Overall, evidence presented here does not support inhibition of KDM1A catalytic demethylase activity as an effective therapeutic strategy for Ewing sarcoma or DSRCT. However, roles of KDM1A beyond its demethylase activity should be considered for these sarcomas.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Catalytic inhibition of KDM1A in Ewing sarcoma is insufficient as a therapeutic strategy

Romo-Morales

Aładowicz

Blagg

et al. 2019

Pediatric Blood & Cancer

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“…Prime examples are the investigation of effects of hypoxia or resistance to chemotherapy and radiotherapy [20,21]. Indeed, since spheroids are avascular, cells in the center of spheroids become hypoxic and have a reduced uptake of oxygen if they are larger than 100 m in size [22], and the size of spheroids has been reported to affect hepatocyte viability and function in the 3D culture [23].…”

Section: Discussionmentioning

confidence: 99%

Long-term 3D culture of the SCC4 cell line using three different culture methods and initial seeding densities

Rustamov

Rudolf

Yagublu

et al. 2017

JCB

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Abstract. INTRODUCTION:In recent years, many different methods were introduced for generation of 3D cell culture. However, many currently available three-dimensional techniques are not suitable for certain cell lines and sometimes showed a lack of reproducibility. Therefore, specific protocols for cell lines are needed. In this work, we demonstrate different methods of generating 3D cell culture for SCC4 tongue cancer cell line and discuss their applicability. MATERIALS AND METHODS:Using three different methods, tumor spheroids were generated from SCC4 cells and cultured for 20 days. To investigate the influence of initial seeding density on spheroid morphology and size during long term culture, the same set of different cell numbers was used for each method. Using phase-contrast microscopy, spheroids were monitored until day 20 and their sizes were determined. RESULTS: We observed that spheroids were formed within 24 hours regardless of the method and initial cell density. Further, in all groups the spheroid size was maximal at day 2, followed by a decline until day 20. Spheroids remained stable until day 20 independent of initial seeding concentration in all groups. DISCUSSION: We have generated long-term culture spheroids of SCC4 cells. The size of the spheroids can be influenced by varying the initial cell seeding density until day 20. This may be useful if different sizes of spheroids are required, e.g. in hypoxia research.

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“…Although immortalized MSCs, or well characterized cell lines, could bypass the lack of 71 primary cells and avoid the variability involved with use of primary human MSCs (hMSCs) 72 samples, they are rarely employed to make spheroids [27,28]. Cell lines would also allow better 73 standardization of the spheroid formation protocol.…”

mentioning

confidence: 99%

“…157 genes were used as endogenous genes for normalization. Primer sequences (S1 Table) Among different methods to form MSC-spheroids, we followed an approach based on 179 cell aggregation in methylcellulose-based medium [27] ( Fig 1A). To establish a protocol that 180 is simple, reproducible and compatible with hematopoietic cell culture, two commercial 181 methylcelluloses commonly used for hematopoietic cell assays were tested.…”

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confidence: 99%

A comparative study of the capacity of mesenchymal stromal cell lines to form spheroids

Deynoux

Sunter

Ducrocq

et al. 2019

Preprint

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FM) 2 18 Abstract 19 Mesenchymal stem cells (MSCs)-derived spheroid models favor maintenance of stemness, ex 20 vivo expansion and transplantation efficacy. Spheroids may also be considered as useful 21 surrogate models of the hematopoietic niche. However, accessibility to primary cells, from bone 22 marrow (BM) or adipose tissues, may limit their experimental use and the lack of consistency 23 in methods to form spheroids may affect data interpretation. In this study, we aimed to create a 24 simple model by examining the ability of cell lines, from human (HS-27a and HS-5) and murine 25 (MS-5) BM origins, to form spheroids, compared to primary human MSCs (hMSCs). Our 26 protocol efficiently allowed the spheroid formation from all cell types within 24 hours. Whilst 27 hMSCs-derived spheroids began to shrink after twenty-four hours, the size of spheroids derived 28 from cell lines remained constant during three weeks. The difference was partially explained 29 by the balance between proliferation and cell death, which could be triggered by hypoxia and 30 induced oxidative stress. Our results demonstrate that, unlike hMSCs, MSC cell lines make 31 reproductible spheroids that are easily handled. Thus, this model could help in understanding 32 mechanisms involved in MSC functions and may provide a simple model by which to study 33 cell interactions in the BM niche. 3 34 Introduction 35 Over the last two decades, extensive studies have attempted to characterize 36 mesenchymal stem cell (MSC). Initially described in the bone marrow (BM), MSCs were later 37 found in almost all adult and fetal tissues [1]. Their classification rapidly suffered from a lack 38 of clear phenotypical definition. Therefore, in 2006, the International Society for Cellular 39 Therapy (ISCT) defined MSCs according to three minimal criteria: adherence to plastic, 40 specific cell surface markers and multipotent potential. Indeed, MSCs are classically described 41 as stem cells that are able to differentiate into osteoblasts, adipocytes and chondroblasts [2], 42 making them an attractive source of cells in regenerative medicine. Subsequent studies have 43 also established their ability to differentiate into cardiomyocytes [3], neurons [4], epithelial 44 cells [5] and hepatocytes [6]. The discovery of the multiple functions of MSC, such as those 45 involved in the anti-inflammatory response [7] and in injury repair [8,9] confirmed them as 46 promising cellular tools in regenerative medicine. 47 Furthermore, MSCs represent a key component of the BM microenvironment 48 supporting normal hematopoiesis through the regulation of stem cell renewal and differentiation 49 processes, but also fueling malignant cells and protecting them from therapeutic agents [10]. 50 As such, primary MSCs have often been used as feeder layers in long-term co-culture of 51 hematopoietic cells in vitro in preclinical studies [11]. With the aim of standardization, the 52 murine MS-5 cell line became the gold-standard for both normal or malignant hematopoietic 53 cell culture [1...

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Linking hypoxia, DNA damage and proliferation in multicellular tumor spheroids

Cited by 106 publications

References 43 publications

Catalytic inhibition of KDM1A in Ewing sarcoma is insufficient as a therapeutic strategy

Catalytic inhibition of KDM1A in Ewing sarcoma is insufficient as a therapeutic strategy

Long-term 3D culture of the SCC4 cell line using three different culture methods and initial seeding densities

A comparative study of the capacity of mesenchymal stromal cell lines to form spheroids

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