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High-Throughput Examination of Therapy-Induced Alterations in Redox Metabolism in Spheroid and Microtumor Models

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Photodynamic Therapy

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2451))

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Abstract

The capacity of cancer cells to adjust their metabolism to thrive in new environments and in response to treatments has been implicated in the acquisition of treatment resistance. To optimize therapeutic strategies such as photodynamic therapy (PDT)-based combination treatments, methods to characterize the plasticity of cancer metabolism in response to treatments are required. This protocol provides a method for high-throughput and label-free tracking of metabolic redox states in cancer tissues, leveraging the autofluorescent properties of nicotinamide dinucleotide (NAD(P)H) and oxidized flavoprotein adenine dinucleotide (FAD). The methodology is optimized to be applied to 3D spheroid/microtumor/organoid cultures, regardless of the culture type (e.g., adherent or suspension cultures) and morphology. The exploitation of these methods may elucidate mechanisms of metabolic adaptation and perturbations in redox homeostasis, and chart the overall tumor health in both 3D culture models and ex vivo tissues following cancer therapies, such as PDT.

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References

  1. Lehuédé C, Dupuy F, Rabinovitch R, Jones RG, Siegel PM (2016) Metabolic plasticity as a determinant of tumor growth and metastasis. Cancer Res 76:5201–5208

    Article  PubMed  Google Scholar 

  2. Vander Heiden MG, DeBerardinis RJ (2017) Understanding the intersections between metabolism and cancer biology. Cell 168:657–669

    Article  CAS  PubMed  Google Scholar 

  3. Vander Heiden MG (2011) Targeting cancer metabolism: a therapeutic window opens. Nat Rev Drug Discov 10:671–684

    Article  CAS  PubMed  Google Scholar 

  4. Castano AP, Mroz P, Hamblin MR (2006) Photodynamic therapy and anti-tumour immunity. Nat Rev Cancer 6:535–545

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Dolmans DEJGJ, Fukumura D, Jain RK (2003) Photodynamic therapy for cancer. Nat Rev Cancer 3:380–387

    Article  CAS  PubMed  Google Scholar 

  6. Hilf R (2007) Mitochondria are targets of photodynamic therapy. J Bioenerg Biomembr 39:85–89

    Article  CAS  PubMed  Google Scholar 

  7. Kessel D (2014) Reversible effects of photodamage directed toward mitochondria. Photochem Photobiol 90:1211–1213

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Pogue BW, O’Hara JA, Demidenko E, Wilmot CM, Goodwin IA, Chen B, Swartz HM, Hasan T (2003) Photodynamic therapy with verteporfin in the radiation-induced fibrosarcoma-1 tumor causes enhanced radiation sensitivity. Cancer Res 63:1025–1033

    CAS  PubMed  Google Scholar 

  9. Kessel D, Oleinick NL (2009) Initiation of autophagy by photodynamic therapy. Methods Enzymol 453:1–16

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Kessel D, Vicente MGH, Reiners JJ (2006) Initiation of apoptosis and autophagy by photodynamic therapy. Autophagy 2:289–290

    Article  CAS  PubMed  Google Scholar 

  11. Broekgaarden M, Weijer R, van Gulik TM, Hamblin MR, Heger M (2015) Tumor cell survival pathways activated by photodynamic therapy: a molecular basis for pharmacological inhibition strategies. Cancer Metastasis Rev 34:643–690

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Fingar VH (1996) Vascular effects of photodynamic therapy. J Clin Laser Med Surg 14:323–328

    Article  CAS  PubMed  Google Scholar 

  13. Ferrario A, von Tiehl KF, Rucker N, Schwarz MA, Gill PS, Gomer CJ (2000) Antiangiogenic treatment enhances photodynamic therapy responsiveness in a mouse mammary carcinoma. Cancer Res 60:4066–4069

    CAS  PubMed  Google Scholar 

  14. Semenza GL (2012) Hypoxia-inducible factors in physiology and medicine. Cell 148:399–408

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Broekgaarden M, Weijer R, Krekorian M, Ijssel B, Kos M, Alles LK, Wijk AC, Bikadi Z, Hazai E, Gulik TM et al (2016) Inhibition of hypoxia-inducible factor 1 with acriflavine sensitizes hypoxic tumor cells to photodynamic therapy with zinc phthalocyanine-encapsulating cationic liposomes. Nano Res 6:1639–1662

    Article  Google Scholar 

  16. Weijer R, Broekgaarden M, Krekorian M, Alles LK, van Wijk AC, Mackaaij C, Verheij J, van der Wal AC, van Gulik TM, Storm G et al (2016) Inhibition of hypoxia inducible factor 1 and topoisomerase with acriflavine sensitizes perihilar cholangiocarcinomas to photodynamic therapy. Oncotarget 7:3341–3356

    Article  PubMed  Google Scholar 

  17. Lamberti MJ, Pansa MF, Vera RE, Fernández-Zapico ME, Rumie Vittar NB, Rivarola VA (2017) Transcriptional activation of HIF-1 by a ROS-ERK axis underlies the resistance to photodynamic therapy. PLoS One 12:e0177801

    Article  PubMed  PubMed Central  Google Scholar 

  18. Georgakoudi I, Quinn KP (2012) Optical imaging using endogenous contrast to assess metabolic state. Annu Rev Biomed Eng 14:351–367

    Article  CAS  PubMed  Google Scholar 

  19. Heikal AA (2010) Intracellular coenzymes as natural biomarkers for metabolic activities and mitochondrial anomalies. Biomark Med 4:241–263

    Article  CAS  PubMed  Google Scholar 

  20. Pogue BW, Pitts JD, Mycek MA, Sloboda RD, Wilmot CM, Brandsema JF, O’Hara JA (2001) In vivo NADH fluorescence monitoring as an assay for cellular damage in photodynamic therapy. Photochem Photobiol 74:817–824

    Article  CAS  PubMed  Google Scholar 

  21. Zhang Z, Blessington D, Li H, Busch TM, Glickson J, Luo Q, Chance B, Zheng G (2004) Redox ratio of mitochondria as an indicator for the response of photodynamic therapy. J Biomed Opt 9:772–778

    Article  CAS  PubMed  Google Scholar 

  22. Walsh AJ, Castellanos JA, Nagathihalli NS, Merchant NB, Skala MC (2016) Optical imaging of drug-induced metabolism changes in murine and human pancreatic cancer organoids reveals heterogeneous drug response. Pancreas 45:863–869

    Article  CAS  PubMed  Google Scholar 

  23. Daemen A, Peterson D, Sahu N, McCord R, Du X, Liu B, Kowanetz K, Hong R, Moffat J, Gao M et al (2015) Metabolite profiling stratifies pancreatic ductal adenocarcinomas into subtypes with distinct sensitivities to metabolic inhibitors. Proc Natl Acad Sci U S A 112:E4410–E4417

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Pampaloni F, Reynaud EG, Stelzer EHK (2007) The third dimension bridges the gap between cell culture and live tissue. Nat Rev Mol Cell Biol 8:839–845

    Article  CAS  PubMed  Google Scholar 

  25. Tanner K, Gottesman MM (2015) Beyond 3D culture models of cancer. Sci Transl Med 7:283ps9

    Article  PubMed  PubMed Central  Google Scholar 

  26. Cannon TM, Shah AT, Skala MC (2017) Autofluorescence imaging captures heterogeneous drug response differences between 2D and 3D breast cancer cultures. Biomed Opt Express 8:1911–1925

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Shah AT, Heaster TM, Skala MC (2017) Metabolic imaging of head and neck cancer organoids. PLoS One 12:e0170415

    Article  PubMed  PubMed Central  Google Scholar 

  28. Broekgaarden M, Bulin A-L, Frederick J, Mai Z, Hasan T (2019) Tracking photodynamic- and chemotherapy-induced redox state perturbations in 3D culture models of pancreatic cancer: a tool for identifying therapy-induced metabolic changes. J Clin Med 8:1399

    Article  CAS  PubMed Central  Google Scholar 

  29. Bulin A-L, Broekgaarden M, Simeone D, Hasan T (2019) Low dose photodynamic therapy harmonizes with radiation therapy to induce beneficial effects on pancreatic heterocellular spheroids. Oncotarget 10:2625–2643

    Article  PubMed  PubMed Central  Google Scholar 

  30. Broekgaarden M, Anbil S, Bulin A-L, Obaid G, Mai Z, Baglo Y, Rizvi I, Hasan T (2019) Modulation of redox metabolism negates cancer-associated fibroblasts-induced treatment resistance in a heterotypic 3D culture platform of pancreatic cancer. Biomaterials 222:119421

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Skala MC, Riching KM, Gendron-Fitzpatrick A, Eickhoff J, Eliceiri KW, White JG, Ramanujam N (2007) In vivo multiphoton microscopy of NADH and FAD redox states, fluorescence lifetimes, and cellular morphology in precancerous epithelia. Proc Natl Acad Sci U S A 104:19494–19499

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Celli JP, Rizvi I, Evans CL, Abu-Yousif AO, Hasan T (2010) Quantitative imaging reveals heterogeneous growth dynamics and treatment-dependent residual tumor distributions in a three-dimensional ovarian cancer model. J Biomed Opt 15:051603

    Article  PubMed  PubMed Central  Google Scholar 

  33. Rizvi I, Celli JP, Evans CL, Abu-Yousif AO, Muzikansky A, Pogue BW, Finkelstein D, Hasan T (2010) Synergistic enhancement of carboplatin efficacy with photodynamic therapy in a three-dimensional model for micrometastatic ovarian cancer. Cancer Res 70:9319–9328

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Bulin A-L, Broekgaarden M, Hasan T (2017) Comprehensive high-throughput image analysis for therapeutic efficacy of architecturally complex heterotypic organoids. Sci Rep 7:16445

    Article  Google Scholar 

  35. Broekgaarden M, Rizvi I, Bulin A-L, Petrovic L, Goldschmidt R, Celli JP, Hasan T (2018) Neoadjuvant photodynamic therapy augments immediate and prolonged oxaliplatin efficacy in metastatic pancreatic cancer organoids. Oncotarget 9:13009–13022

    Article  PubMed  PubMed Central  Google Scholar 

  36. Zanoni M, Piccinini F, Arienti C, Zamagni A, Santi S, Polico R, Bevilacqua A, Tesei A (2016) 3D tumor spheroid models for in vitro therapeutic screening: a systematic approach to enhance the biological relevance of data obtained. Sci Rep 6:19103

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Molla A, Couvet M, Coll J-L (2017) Unsuccessful mitosis in multicellular tumour spheroids. Oncotarget 8:28769–28784

    Article  PubMed  PubMed Central  Google Scholar 

  38. Lee GY, Kenny PA, Lee EH, Bissell MJ (2007) Three-dimensional culture models of normal and malignant breast epithelial cells. Nat Methods 4:359–365

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Glidden MD, Celli JP, Massodi I, Rizvi I, Pogue BW, Hasan T (2012) Image-based quantification of benzoporphyrin derivative uptake, localization, and Photobleaching in 3D tumor models, for optimization of PDT parameters. Theranostics 2:827–839

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Author information

Authors and Affiliations

  1. Wellman Center for Photomedicine, Department of Dermatology, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA

    Mans Broekgaarden, Anne-Laure Bulin & Tayyaba Hasan

  2. Institute for Advanced Biosciences, INSERM U1209, CNRS UMR 5309, Université de Grenoble Alpes, Grenoble, France

    Mans Broekgaarden

  3. Synchrotron Radiation for Biomedicine, INSERM UA07, Université de Grenoble Alpes, Grenoble, France

    Anne-Laure Bulin

Authors
  1. Mans Broekgaarden
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  2. Anne-Laure Bulin
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  3. Tayyaba Hasan
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Corresponding author

Correspondence to Tayyaba Hasan .

Editor information

Editors and Affiliations

  1. Institute for Advanced Biosciences, Université de Grenoble Alpes, Grenoble, France

    Mans Broekgaarden

  2. van ‘t Hoff Institute for Molecular Sciences, University of Amsterdam, Amsterdam, The Netherlands

    Hong Zhang

  3. British Columbia Cancer Research Centre, Vancouver, BC, Canada

    Mladen Korbelik

  4. Laser Research Centre, University of Johannesburg, Doornfontein, South Africa

    Michael R. Hamblin

  5. Jiaxing Key Laboratory for Photonanomedicine and Experimental Therapeutics, Department of Pharmaceutics, Jiaxing University College of Medicine, Jiaxing, Zhejiang, China

    Michal Heger

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Broekgaarden, M., Bulin, AL., Hasan, T. (2022). High-Throughput Examination of Therapy-Induced Alterations in Redox Metabolism in Spheroid and Microtumor Models. In: Broekgaarden, M., Zhang, H., Korbelik, M., Hamblin, M.R., Heger, M. (eds) Photodynamic Therapy. Methods in Molecular Biology, vol 2451. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2099-1_6

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  • DOI: https://doi.org/10.1007/978-1-0716-2099-1_6

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2098-4

  • Online ISBN: 978-1-0716-2099-1

  • eBook Packages: Springer Protocols

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