Comparative RNA sequencing identifies superior cytocidal mechanisms of imatinib over rapamycin in Pulmonary Lymphangioleiomyomatosis (LAM) Tumor Cells
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ABSTRACT:
PROVIDER: PRJNA796029 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR17535800_1.fastq.gz | Fastqsanger.gz | |||
| SRR17535800_2.fastq.gz | Fastqsanger.gz | |||
| SRR17535801_1.fastq.gz | Fastqsanger.gz | |||
| SRR17535801_2.fastq.gz | Fastqsanger.gz | |||
| SRR17535802_1.fastq.gz | Fastqsanger.gz |
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Project description:We report results of RNA sequencing analysis of serum starved pulmonary LAM tumor cells comparatively treated for 24hrs at 37C and 5% CO2 with rapamycin (0.05uM and 1uM) and tyrosine kinase inhibitor (TKI) imatinib (1uM, 5uM, and 10uM). Experiments were done in duplicates per drug concentration. This study was performed to investigate the biological mechanisms underlying superior cytocidal capabilities of the TKI over FDA-approved rapamycin by inhibiting receptor tyrosine kinases on mesenchymal tumorigenic cells in Tuberous Sclerosis and Lymphangioleiomyomatosis (LAM) diseases. Total RNA isolates (RIN>8.0) were sequenced on an illumina Novaseq6000 platform at 100 base pairs to a depth of 20 milllion paired end reads. Real-Time Analysis was used for base callling converted to fastq format with bcl2fastq2. Pseudoalignment of RNA-seq reads to a kallisto index was created from human reference genome NCBI/GRCh38.p13. Differential expression was resolved using Sleuth in R and signaling pathway analysis was performed using Ingenuity Pathway Analysis (IPA). Results reveal differential inactivation of the Cell Cycle: G2/M DNA Damage Checkpoint Regulation SIgnaling Pathway only in pulmonary LAM tumor cells treated with 10uM imatinib concentration.We further determined that increasing imatinib conentrations from 1uM to 10uM induced endoplasmic reticulum inositol triphosphate receptor (IP3R) subtype switching from type I to type III typically associated with differential ER calcium efflux and apoptosis.
2022-01-15 | GSE193398 | GEO
Project description:We report results of RNA sequencing analysis of serum starved UMB1949 renal angiomyolipoma (AML) cells comparatively treated for 24hrs at 37C and 5% CO2 with rapamycin (0.05uM and 1uM) and tyrosine kinase inhibitor (TKI) imatinib (1uM and 10uM). Experiments were done in triplicates per drug concentration. This study was performed to investigate the biological mechanisms underlying superior cytocidal capabilities of the TKI over FDA-approved rapamycin by inhibiting receptor tyrosine kinases on mesenchymal tumorigenic cells in Tuberous Sclerosis and Lymphangioleiomyomatosis (LAM) diseases. Total RNA isolates (RIN>8.0) were sequenced on an illumina Novaseq6000 platform at 100 base pairs to a depth of 30 milllion single end reads. Real-Time Analysis was used for base callling converted to fastq format with bcl2fastq2. Pseudoalignment of RNA-seq reads to a kallisto index was created from human reference genome NCBI/GRCh38.p13. Reads were also mapped to this reference genome using STAR (2.5.2b) and featureCounts (v1.5.0-p3) for results comparison. Differential expression was resolved using Sleuth and DESeq2 in R and signaling pathway analysis was performed using Ingenuity Pathway Analysis (IPA). Results reveal differential inactivation of the GPVI pathway and key cell survival and viability genes only in renal AML cells treated with imatinib conentrations.We further determined that this TKI differentially induced ER calcium efflux that disrupted mitochondrial permeability leading to increased cytosolic cytochrome C, caspase 3 activation and apoptosis.
2022-01-15 | GSE193402 | GEO
Project description:Inhibition of mechanistic target of rapamycin (mTOR) constitutes the standard of care of lymphangioleiomyomatosis (LAM). However, this treatment does not eradicate diseased cells and some patients show progressive decline of lung function. This study provides preclinical evidence for beneficial therapeutic approaches in combination with rapamycin. LAM tumorigenesis is reduced in vivo (ELT3-V3 tumor xenografts) using approved drugs for other human diseases and targeting monoamine oxidase A (MAO-A; clorgyline) or MAO-B (rasagiline).
2019-10-01 | GSE129399 | GEO
Project description:Rationale: Increased matrix metalloproteinase (MMP) activity has been implicated in the pathogenesis of lymphangioleiomyomatosis (LAM). Objectives: To investigate how TSC1 or TSC2 deficiency alters MMP expression and regulation. Methods: We studied immortalized cells that lack TSC2 derived from an angiomyolipoma (AML) of a LAM patient, and a TSC2 add back derivative; and murine embryonic fibroblast cells that lack Tsc1 or Tsc2 and respective controls. Global gene expression analysis was carried out in the AML and derivative cell lines. MMP levels in the conditioned media from these cells were analyzed by zymography and ELISA. Measurements and Main Results: We found increased MMP-2 expression in cells lacking TSC1/TSC2 compared to their respective controls by zymography. MMP-2 overproduction by these cells was not affected by rapamycin treatment. Gene expression analysis confirmed increased MMP-2 gene expression that was not affected by rapamycin. Furthermore, multiple other genes were found to be over-expressed in rapamycin-treated TSC2-deficient cells compared to TSC2+ cells. Conclusions: We conclude that TSC1/TSC2 deficiency leads to MMP-2 overproduction that is rapamycin insensitive, and that several genes exhibit similar patterns suggesting TSC1/TSC2 dependent but mTOR independent pathways may be involved in the pathogenesis of LAM.
2009-10-21 | GSE16944 | GEO
Project description:Measurement of mRNA abundance from the following cells lines (red) versus universal mouse reference RNA (green). Wildtype v-Abl transformed pre-B cells were treated for 12 hours with 2.5 uM imatinib mesylate, 10ng/mL rapamycin or nothing. Compound Based Treatment: wildtype v-Abl transformed pre-B cells were treated with imatinib mesylate (IMA), rapamycin (RAP) or nothing (NONE)
2013-01-10 | GSE35040 | GEO
Project description:Measurement of mRNA abundance from the following cells lines (red) versus universal mouse reference RNA (green). Wildtype v-Abl transformed pre-B cells were treated for 12 hours with 2.5 uM imatinib mesylate, 10ng/mL rapamycin or nothing. Compound Based Treatment: wildtype v-Abl transformed pre-B cells were treated with imatinib mesylate (IMA), rapamycin (RAP) or nothing (NONE) compound_treatment_design
2013-01-10 | E-GEOD-35040 | biostudies-arrayexpress
Project description:Comparative RNA sequencing identifies superior cytocidal mechanisms of imatinib over rapamycin in UMB1949 renal angiomyolipoma cells
| PRJNA796033 | ENA
Project description:The purpose of this study was to compare changes in translation (using Gradient Encoding, described below) to changes in mRNA abundance. Lysates of wildtype v-Abl transformed pre-B cells harvested before and after 12 hours of treatment either with 2.5 uM imatinib, a v-Abl kinase inhibitor, or 10ng/mL (10.9 nM) rapamycin, an mTOR inhibitor, were fractionated by sedimentation through linear sucrose gradients. Gradient fractions were encoded such that the mRNA from successive fractions was labeled with increasing ratios of Cy5 to Cy3. mRNAs derived from fractions in the lighter portion of the gradient therefore have a lower Cy5 to Cy3 ratio, whereas those deeper in the gradient have a higher Cy5 to Cy3 ratio. The ratio of Cy5 to Cy3 for each mRNA therefore reflects its average position within the gradient. We thus encoded the sedimentation rate of each mRNA across the entire gradient. The resulting ratios were quantitatively measured for each mRNA species by hybridization to DNA microarrays, and related to the 260 nm absorbance peaks representing different numbers of ribosomes bound per mRNA Compound Based Treatment: wildtype v-Abl transformed pre-B cells were treated with imatinib mesylate (IMA), rapamycin (RAP) or nothing (NONE)
2013-01-10 | GSE35039 | GEO
Project description:The purpose of this study was to compare changes in translation (using Gradient Encoding, described below) to changes in mRNA abundance. Lysates of wildtype v-Abl transformed pre-B cells harvested before and after 12 hours of treatment either with 2.5 uM imatinib, a v-Abl kinase inhibitor, or 10ng/mL (10.9 nM) rapamycin, an mTOR inhibitor, were fractionated by sedimentation through linear sucrose gradients. Gradient fractions were encoded such that the mRNA from successive fractions was labeled with increasing ratios of Cy5 to Cy3. mRNAs derived from fractions in the lighter portion of the gradient therefore have a lower Cy5 to Cy3 ratio, whereas those deeper in the gradient have a higher Cy5 to Cy3 ratio. The ratio of Cy5 to Cy3 for each mRNA therefore reflects its average position within the gradient. We thus encoded the sedimentation rate of each mRNA across the entire gradient. The resulting ratios were quantitatively measured for each mRNA species by hybridization to DNA microarrays, and related to the 260 nm absorbance peaks representing different numbers of ribosomes bound per mRNA Compound Based Treatment: wildtype v-Abl transformed pre-B cells were treated with imatinib mesylate (IMA), rapamycin (RAP) or nothing (NONE) compound_treatment_design
2013-01-10 | E-GEOD-35039 | biostudies-arrayexpress
Project description:Lymphangioleiomyomatosis (LAM) is characterized by cystic lung destruction caused by smooth, muscle-like LAM cells which have mutations in the tumor suppressor genes Tuberous Sclerosis Complex (TSC) 1 or 2, and the capacity to metastasize. Since chemokines and their receptors function in chemotaxis of metastatic cells, we hypothesized that LAM cells may be recruited by chemokine(s) in the lung. Quantification of 25 chemokines in bronchoalveolar lavage fluid from LAM patients and healthy volunteers revealed that concentrations of MCP-1/CCL2, GROa/CXCL1 and ENA-78/CXCL5 were significantly higher in samples from LAM patients than healthy volunteers. In this transcript analysis, expression of chemokine and chemokine receptor mRNA in LAM cells differed from those in melanoma and smooth muscle cells. Subsequent immunohistochemistry of lung sections from 30 LAM patients confirmed protein expression of chemokines and these receptors varied among LAM patient and differed from that seen in breast cancer and melanoma cells. . In vitro, MCP-1/CCL2 induced selective migration of cells showing loss of heterozygosity of TSC2 from a heterogeneous populations of cells grown from explanted LAM lungs. In addition, the frequencies of single-nucleotide polymorphisms in the MCP-1 gene promoter region differed significantly in LAM patients and healthy volunteers (p=0.018), and one polymorphism was associated significantly more frequently with the decline of lung function. These observations are consistent with the notion that chemokines such as MCP-1 may serve to specify site of LAM cell metastasis. Keywords: Human patient sample comparison with cell lines The study is of case/control design with biological replication. Biopsies of nodules from 14 LAM patients (cases) are compared with cultured cell lines (controls) with similar properties.
2008-12-30 | E-GEOD-12027 | biostudies-arrayexpress