Screening for Compounds That Induce Apoptosis of Cancer Cells Grown as Multicellular Spheroids

Herrmann, Richard; Fayad, Walid; Schwarz, Stephan; Berndtsson, Maria; Linder, Stig

doi:10.1177/1087057107310442

Cited by 75 publications

(68 citation statements)

References 25 publications

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“…HCT116 spheroids of homogeneous size (500 lm at 5 days) were generated by the hanging drop technique. 22,25 Spheroids contained a core portion with a lower cell packaging density compared to the peripheral areas (Fig. 1a).…”

Section: Tumor Cell Proliferation In Multicellular Spheroidsmentioning

confidence: 99%

“…Spheroids were prepared using a modification of our previously described method. 22 A cell suspension containing 10,000 cells was added to each well of poly-HEMA-coated 96-well plates, and the wells were overfilled by adding an additional 170 ll media to acquire a convex surface curvature. Plasticine spacers (3 mm) were placed in the corners of each plate to prevent the lids from touching the media.…”

Section: Cell Culturementioning

confidence: 99%

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Restriction of cisplatin induction of acute apoptosis to a subpopulation of cells in a three‐dimensional carcinoma culture model

Fayad

Brnjic

Berglind

et al. 2009

Intl Journal of Cancer

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Cisplatin is a clinically important chemotherapeutical agent used to treat epithelial malignancies. High concentrations (20-100 lM) of cisplatin have been used in numerous studies to induce apoptosis of carcinoma cells grown in monolayer culture over 24-48 hr. These conditions may not be relevant to 3-D tumor tissue in vivo and the importance of apoptosis for tumor response is controversial. We here studied the effects of cisplatin on a 3-D colon carcinoma in vitro model (multicellular spheroids). Cisplatin at a dose of 40 lM induced active caspase-3 preferentially in the peripheral 30 lm cell layer of spheroids, mainly during late stages (72-96 hr). The p53 response to cisplatin was also largely confined to peripheral cell layers. Despite the use of a high cisplatin concentration, a significant fraction of the cells in the spheroids survived treatment. A high proportion of surviving cells stained positive for b-galactosidase, a marker of premature senescence. Cells growtharrested by cisplatin treatment showed a higher spontaneous cell death rate than untreated proliferating cells. We propose that acute apoptosis is of minor significance for the overall response of carcinoma cells to cisplatin treatment. ' UICCKey words: cisplatin; apoptosis; senescence; p21Cip1; p27Kip1 Platinum derivatives are important chemotherapeutical agents for many epithelial malignancies, including testicular, ovarian, squamous cell and colon cancers.1,2 In the low intracellular chloride concentrations, cisplatin undergoes equation to form molecules that are reactive with cellular macromolecules. Cisplatin forms covalent bonds to the N7 positions of DNA purines leading to formation intra-or interstrand crosslinks, but also reacts with RNA and protein. 3,4 Treatment of tumor cells in vitro with cisplatin results in apoptosis within 24-48 hr.4-6 Dysregulation of apoptosis pathways is generally assumed to be important for resistance to cisplatin.6 Cisplatin-induced calcium signaling, leading to activation of calpain and cleavage of Bid, 7 and sustained activation of N-terminal-c-Jun kinase, 8 is important for apoptosis signaling. Recent work has shown that cisplatin is able to induce caspase activation in enucleated cells, demonstrating that cisplatin can trigger apoptosis pathways that are independent of its effects on nuclear DNA.9,10 This cytoplasmic pathway involves reactive oxygen species (ROS) and calcium signaling.10 According to one model, ROS induces activation of Bak, leading to VDAC1 and Bax activation. 11Despite the very large number of studies in this area (2,970 articles on MedLine on cisplatin/apoptosis March 2009), it is controversial whether apoptosis is the primary mode of cell death of human carcinoma cells in response to DNA damaging therapeutic drugs, and also whether apoptosis is necessary for the therapeutic effect of such drugs. 10,[12][13][14][15][16] The importance of apoptosis appears to be most pronounced in short-term experiments (1-2 days) and is of lower significance when cell viability is examined after lo...

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Section: Tumor Cell Proliferation In Multicellular Spheroidsmentioning

confidence: 99%

Section: Cell Culturementioning

confidence: 99%

Restriction of cisplatin induction of acute apoptosis to a subpopulation of cells in a three‐dimensional carcinoma culture model

Fayad

Brnjic

Berglind

et al. 2009

Intl Journal of Cancer

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“…This library has been preselected for compounds with antiproliferative activity in the NCI60 cell line screen. Apoptosis was assessed as generation of an endogenous apoptosis product, caspase-cleaved keratin 18 (ccK18) [18]. In a secondary screen, 29 compounds were tested both for their ability to induce apoptosis and to induce loss of cell viability ( Figure 1C).…”

Section: Screening Multicellular Tumor Spheroids For Apoptosisinducinmentioning

confidence: 99%

“…Therefore, MCS are better suited than monolayer cultures for drugs screening projects aimed to identify compounds active on solid tumors. We have established a screening procedure with good assay performance for the identification of drugs that induce apoptosis of tumor cells grown in MCS [18]. We here describe a novel small molecule (NSC647889) doi: 10.7243/2049-7962-2-19 capable of inducing an unprecedented extent of apoptosis in MCS.…”

Section: Introductionmentioning

confidence: 99%

Massive induction of apoptosis of multicellular tumor spheroids by a novel compound with a calmodulin inhibitor-like mechanism

Mohanty¹,

Fayad²,

Olofsson³

et al. 2013

J Cancer Ther Res

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“…An interesting approach consists to use multicellular tumor spheroid models which are of intermediate complexity between in vivo tumors and in vitro monolayer cultures. Multicellular spheroids can be generated in 96-well plates format for HTS, from a variety of tumor cell lineages [52,53]. Similar approaches can be established to study heterologous cell interactions in solid tumors and the cellular tumor environment [54].…”

Section: Natural Product-biological Target: a Pas De Deux In Drug Dismentioning

confidence: 99%

Ready for a comeback of natural products in oncology

Bailly

2009

Biochemical Pharmacology

100

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(186 words)Since the late 1990s and the rapid expansion of monoclonal antibodies and synthetic protein kinase inhibitors in oncology, anticancer natural products fell out of fashion with the pharmaceutical industry. But in 2007 with the approval of three new drugs derived from natural products, the emergence of promising antitumor compounds from microorganisms (e.g. alvespimycin, salinosporamide) and the growing importance of new formulations of known natural product-derived drugs (nanoparticle formulations, oral forms), we are witnessing a new wave for natural products in oncology. The recent approval of the microtubule-targeted epothilone derivative ixabepilone (Ixempra ® ), the DNA-alkylating marine alkaloid trabectedin (Yondelis ® ) and the inhibitor of mTOR protein kinase temsirolimus (Torisel ® ) is emblematic of the evolution of the field which combines the long established finding of conventional cytotoxic agents and the emergence of molecularly targeted therapeutics. These three examples also illustrate the increasing importance of microbial sources for the discovery of medically useful natural products. The contribution of innovative biological targets is also highlighted here, with references to proteasome inhibitors and novel approaches such as manipulation of mRNA splicing. Altogether, these observations plead for the return of natural products in oncology.

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Screening for Compounds That Induce Apoptosis of Cancer Cells Grown as Multicellular Spheroids

Cited by 75 publications

References 25 publications

Restriction of cisplatin induction of acute apoptosis to a subpopulation of cells in a three‐dimensional carcinoma culture model

Restriction of cisplatin induction of acute apoptosis to a subpopulation of cells in a three‐dimensional carcinoma culture model

Massive induction of apoptosis of multicellular tumor spheroids by a novel compound with a calmodulin inhibitor-like mechanism

Ready for a comeback of natural products in oncology

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