A 3D Cell Death Assay to Quantitatively Determine Ferroptosis in Spheroids

Demuynck, Robin; Efimova, Iuliia; Lin, Abraham; Declercq, Heidi; Krysko, Dmitri V.

doi:10.3390/cells9030703

Cited by 23 publications

(32 citation statements)

References 50 publications

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“…ICD refers to an immunological feature of cell death and is observed in immunogenic apoptosis and immunogenic necroptosis, as well as in mixed cell death types (2). The role of PDT in the induction of ferroptosis 132 133 in cancer cells needs to be further clarified 134 . Importantly, only a fraction of cancer cells can be reached by light during PDT because light can penetrate only to a limited depth.…”

Section: Main Principles Of Pdtmentioning

confidence: 99%

Targeting immunogenic cancer cell death by photodynamic therapy: past, present and future

Alzeibak

Mishchenko

Shilyagina

et al. 2021

J Immunother Cancer

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310

213

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The past decade has witnessed major breakthroughs in cancer immunotherapy. This development has been largely motivated by cancer cell evasion of immunological control and consequent tumor resistance to conventional therapies. Immunogenic cell death (ICD) is considered one of the most promising ways to achieve total tumor cell elimination. It activates the T-cell adaptive immune response and results in the formation of long-term immunological memory. ICD can be triggered by many anticancer treatment modalities, including photodynamic therapy (PDT). In this review, we first discuss the role of PDT based on several classes of photosensitizers, including porphyrins and non-porphyrins, and critically evaluate their potential role in ICD induction. We emphasize the emerging trend of ICD induction by PDT in combination with nanotechnology, which represents third-generation photosensitizers and involves targeted induction of ICD by PDT. However, PDT also has some limitations, including the reduced efficiency of ICD induction in the hypoxic tumor microenvironment. Therefore, we critically evaluate strategies for overcoming this limitation, which is essential for increasing PDT efficiency. In the final part, we suggest several areas for future research for personalized cancer immunotherapy, including strategies based on oxygen-boosted PDT and nanoparticles. In conclusion, the insights from the last several years increasingly support the idea that PDT is a powerful strategy for inducing ICD in experimental cancer therapy. However, most studies have focused on mouse models, but it is necessary to validate this strategy in clinical settings, which will be a challenging research area in the future.

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Section: Main Principles Of Pdtmentioning

confidence: 99%

Targeting immunogenic cancer cell death by photodynamic therapy: past, present and future

Alzeibak

Mishchenko

Shilyagina

et al. 2021

J Immunother Cancer

Self Cite

310

213

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“…After incubation overnight, the cells were stained with 3.3 µM Sytox Green nucleic acid stain (molecular probes) and stimulated with 2.5 µM RASselective lethal 3 (RSL3) (Sigma Aldrich) for different durations (1, 3, 6 and 24 hours). Cell death was analyzed as described in Demuynck et al 36 Fluorescence was measured on a Tecan Spark 20M multimode microplate reader.…”

Section: Materials and Methods Cell Lines And Cell Culturementioning

confidence: 99%

Vaccination with early ferroptotic cancer cells induces efficient antitumor immunity

Efimova

Catanzaro

Meeren

et al. 2020

J Immunother Cancer

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286

262

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BackgroundImmunotherapy represents the future of clinical cancer treatment. The type of cancer cell death determines the antitumor immune response and thereby contributes to the efficacy of anticancer therapy and long-term survival of patients. Induction of immunogenic apoptosis or necroptosis in cancer cells does activate antitumor immunity, but resistance to these cell death modalities is common. Therefore, it is of great importance to find other ways to kill tumor cells. Recently, ferroptosis has been identified as a novel, iron-dependent form of regulated cell death but whether ferroptotic cancer cells are immunogenic is unknown.MethodsFerroptotic cell death in murine fibrosarcoma MCA205 or glioma GL261 cells was induced by RAS-selective lethal 3 and ferroptosis was analyzed by flow cytometry, atomic force and confocal microscopy. ATP and high-mobility group box 1 (HMGB1) release were detected by luminescence and ELISA assays, respectively. Immunogenicity in vitro was analyzed by coculturing of ferroptotic cancer cells with bone-marrow derived dendritic cells (BMDCs) and rate of phagocytosis and activation/maturation of BMDCs (CD11c+CD86+, CD11c+CD40+, CD11c+MHCII+, IL-6, RNAseq analysis). The tumor prophylactic vaccination model in immune-competent and immune compromised (Rag-2−/−) mice was used to analyze ferroptosis immunogenicity.ResultsFerroptosis can be induced in cancer cells by inhibition of glutathione peroxidase 4, as evidenced by confocal and atomic force microscopy and inhibitors’ analysis. We demonstrate for the first time that ferroptosis is immunogenic in vitro and in vivo. Early, but not late, ferroptotic cells promote the phenotypic maturation of BMDCs and elicit a vaccination-like effect in immune-competent mice but not in Rag-2−/− mice, suggesting that the mechanism of immunogenicity is very tightly regulated by the adaptive immune system and is time dependent. Also, ATP and HMGB1, the best-characterized damage-associated molecular patterns involved in immunogenic cell death, have proven to be passively released along the timeline of ferroptosis and act as immunogenic signal associated with the immunogenicity of early ferroptotic cancer cells.ConclusionsThese results pave the way for the development of new therapeutic strategies for cancers based on induction of ferroptosis, and thus broadens the current concept of immunogenic cell death and opens the door for the development of new strategies in cancer immunotherapy.

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“…The cytotoxic and proapoptotic potential of compound 1 was also tested and confirmed on a different tumor cell line, the acute T cell leukemia Jurkat ( Figure S6, Supplementary Materials). Since PS exposure is an event that characterizes both caspase-dependent and -independent types of cell death [35], to exclude that beside apoptosis different forms of regulated cell death concur to compound 1 antitumor potential, we used specific cell death inhibitors: (zVAD-fmk, a pan-caspase blocker), necroptosis [necrostatin-1 s (Nec1s), a RIPK1 inhibitor] or ferroptosis [ferrostatin-1 (fer-1), an inhibitor of reactive oxygen species and lipid peroxidation and deferoxamine (DFO), an iron chelator to characterize apoptosis, necroptosis or ferroptosis, respectively [36,37]. Only zVAD-fmk was able to rescue cells after exposure to compound 1.…”

Section: Phosphatidylserine Exposure and Cell Death Analysismentioning

confidence: 99%

Curcumin-1,2,3-Triazole Conjugation for Targeting the Cancer Apoptosis Machinery

et al. 2020

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The burden of neoplastic diseases is widely recognized as a severe cause of mortality. The clinical inadequacy of most anticancer therapeutics urgently prompted intense drug discovery efforts toward the identification of new chemical entities endowed with a potent and safe antitumor profile. In this scenario, targeting cancer cells apoptosis machinery has emerged as a relevant strategy, useful for tackling the emergence of drug resistance. On this basis, a small library of naturally inspired hybrid molecules was obtained by combining, through a click chemistry approach, “privileged” synthons such as curcumin scaffold and 1,2,3-triazole building block. Compound 1, bearing a para-fluoro phenyl moiety, showed low-micromolar potency against T acute lymphoblastic leukemia cell growth. More in-depth biologic studies demonstrated, for this analog, cell death-inducing properties associated with its capability to simultaneously activate both the receptor and the mitochondrial apoptosis cascades. This peculiar behavior offers promises for achieving an expanded anticancer effect, namely intense cytotoxic response coupled with reduced predisposition of chemoresistance insurgence. Altogether, this study allowed the identification of compound 1 as a lead compound worth to be progressed as an anticancer drug candidate.

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A 3D Cell Death Assay to Quantitatively Determine Ferroptosis in Spheroids

Cited by 23 publications

References 50 publications

Targeting immunogenic cancer cell death by photodynamic therapy: past, present and future

Targeting immunogenic cancer cell death by photodynamic therapy: past, present and future

Vaccination with early ferroptotic cancer cells induces efficient antitumor immunity

Curcumin-1,2,3-Triazole Conjugation for Targeting the Cancer Apoptosis Machinery

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