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Whole Transcriptome Sequencing Analysis of Cancer Stem/Progenitor Cells Obtained from Mouse Lung Adenocarcinomas

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Lung Cancer

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2279))

Abstract

Interrogation and characterization of lung cancer stem cells (CSCs) that are implicated in lung oncogenesis is crucial for our understanding of inter- and intra-tumor heterogeneity and the aggressive nature of the disease, including tumor resistance and relapse in response to conventional therapy. Here, we describe an in vitro surrogate model, namely the “sphere-forming assay,” for the derivation, enrichment, and propagation of lung stem/progenitor cells with CSC properties, including self-renewal, tumor initiation capacity and propagation, and differentiation into cells of the tumor bulk, from a murine Kras-mutant lung adenocarcinoma cell line. Self-renewing cancer stem/progenitor cells, in the form of 3D in vitro spheres, can be phenotypically interrogated using downstream techniques such as gene expression analysis (e.g., whole transcriptome sequencing and quantitative real time PCR), which is the focus of this chapter.

Ansam Sinjab and Reem Daouk Co-contributing first authors.

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References

  1. Reynolds BA, Rietze RL (2005) Neural stem cells and neurospheres--re-evaluating the relationship. Nat Methods 2(5):333–336

    Article  CAS  Google Scholar 

  2. Daouk R, Hassane M, Bahmad HF et al (2019) Genome-wide and phenotypic evaluation of stem cell progenitors derived from Gprc5a-deficient murine lung adenocarcinoma with somatic Kras mutations. Front Oncol 9:207

    Article  Google Scholar 

  3. Bolger AM, Lohse M, Usadel B (2014) Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30(15):2114–2120

    Article  CAS  Google Scholar 

  4. Kim D, Langmead B, Salzberg SL (2015) HISAT: a fast spliced aligner with low memory requirements. Nat Methods 12(4):357–360

    Article  CAS  Google Scholar 

  5. Pertea M, Pertea GM, Antonescu CM et al (2015) StringTie enables improved reconstruction of a transcriptome from RNA-seq reads. Nat Biotechnol 33(3):290–295

    Article  CAS  Google Scholar 

  6. Love MI, Huber W, Anders S (2014) Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol 15(12):550

    Article  Google Scholar 

  7. Bahmad HF, Cheaito K, Chalhoub RM et al (2018) Sphere-formation assay: three-dimensional in vitro culturing of prostate cancer stem/progenitor sphere-forming cells. Front Oncol 8:347–347

    Article  Google Scholar 

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Correspondence to Humam Kadara .

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Sinjab, A., Daouk, R., Abou-Kheir, W., Kadara, H. (2021). Whole Transcriptome Sequencing Analysis of Cancer Stem/Progenitor Cells Obtained from Mouse Lung Adenocarcinomas. In: Santiago-Cardona, P.G. (eds) Lung Cancer. Methods in Molecular Biology, vol 2279. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1278-1_15

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  • DOI: https://doi.org/10.1007/978-1-0716-1278-1_15

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1277-4

  • Online ISBN: 978-1-0716-1278-1

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