B4GALT1 Is a New Candidate to Maintain the Stemness of Lung Cancer Stem Cells

Abstract

Background: According to the cancer stem cells (CSCs) hypothesis, a population of cancer cells with stem cell properties is responsible for tumor propagation, drug resistance, and disease recurrence. Study of the mechanisms responsible for lung CSCs propagation is expected to provide better understanding of cancer biology and new opportunities for therapy. Methods: The Lung Adenocarcinoma (LUAD) NCI-H460 cell line was grown either as 2D or as 3D cultures. Transcriptomic and genome-wide chromatin accessibility studies of 2D vs. 3D cultures were carried out using RNA-sequencing and Assay for Transposase Accessible Chromatin with high-throughput sequencing (ATAC-seq), respectively. Reverse transcription polymerase chain reaction (RT-PCR) was also carried out on RNA extracted from primary cultures derived from malignant pleural effusions to validate RNA-seq results. Results: RNA-seq and ATAC-seq data disentangled transcriptional and genome accessibility variability of 3D vs. 2D cultures in NCI-H460 cells. The examination of genomic landscape of genes upregulated in 3D vs. 2D cultures led to the identification of 2D cultures led to the identification of Beta-1,4-galactosyltranferase 1 (B4GALT1) as the top candidate. B4GALT1 as the top candidate. B4GALT1 was validated as a stemness factor, since its silencing caused strong inhibition of 3D spheroid formation. Conclusion: Combined transcriptomic and chromatin accessibility study of 3D vs. 2D LUAD cultures led to the identification of B4GALT1 as a new factor involved in the propagation and maintenance of LUAD CSCs.

Reverse transcriptase inhibition potentiates target therapy in BRAF-mutant melanomas: effects on cell proliferation, apoptosis, DNA-damage, ROS induction and mitochondrial membrane depolarization

“…Anti-rabbit and anti-mouse were purchased from AbCam (Cambridge, UK). Melanoma cells were lysed with RIPA buffer purchased by Sigma-Aldrich (St. Louis, MO, USA) and total proteins were run using Invitrogen Bolt Bis-Tris 4-12% Plus gels precast polyacrylamide gels as previously described (12,26,27).…”

In Vitro Biophysical and Biological Characterization of Lipid Nanoparticles Co-Encapsulating Oncosuppressors miR-199b-5p and miR-204-5p as Potentiators of Target Therapy in Metastatic Melanoma

“…Total RNA was extracted using TRIzol method according to manufacturer's instruction [44] and quantitated by Qubit Fluorometer [45] (ThermoFisher Scientific, Foster City, CA, USA). Quantitative Real Time-PCR (qRT-PCR) was performed by TaqMan Gene Expression Assays (Catalog number 4444556) (Applied Biosystems, Foster City, CA, USA).…”

Reverse transcriptase inhibition potentiates target therapy in BRAF-mutant melanomas: effects on cell proliferation, apoptosis, DNA-damage, ROS induction and mitochondrial membrane depolarization

“…All results shown are presented as mean values from three independent experiments. Quantitative analysis for curve fitting and p-value estimation (significance p < 0.05) were performed by GraphPad Prism 7 (San Diego, CA, USA) [33].…”