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Homo sapiens strain:human embryonic kidney cells

ABSTRACT: HEK293 Transcriptome during rAAV production (HEK293 cells from Mass Biologics in BalanCD HEK293 Medium)

PROVIDER: PRJNA1254762 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR33327456_1.fastq.gz	Fastqsanger.gz
	SRR33327456_2.fastq.gz	Fastqsanger.gz
	SRR33327457_1.fastq.gz	Fastqsanger.gz
	SRR33327457_2.fastq.gz	Fastqsanger.gz
	SRR33327458_1.fastq.gz	Fastqsanger.gz

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AAV production system proteomics

Project description:We performed absolute quantification (AQUA) of viral proteins by targeted quantitative proteomics and conducted tandem mass tag (TMT)-based proteomics on the three rAAV and wtAAV production systems to identify potential factors limiting rAAV productivity in HEK293 cells

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Project description:HEK293 Transcriptome during rAAV production (ATCC HEK293 cells adapted in AMBIC media)

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Expression data from human embryonic kidney cells (HEK293) cultivated in high and low glucose

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Project description:Gene expression profiling of human embryonic kidney (HEK293) cells was performed to determine the effect of high and low glucose on gene expression. Microarrays were used to identify distinct classes of genes up-regulated in HEK293 cells during cultivation for 7 days in medium with physiological (low) glucose compared to high glucose. Human embryonic kidney cells (HEK293) were cultivated for 7 days in commercially available DMEM (supplemented with 10 % FCS) containing high glucose (450 mg/dl) or low glucose (100 mg/dl). Cells were harvested, total RNA was extracted and microarray gene expression profiling was performed to compare differential gene expression between low and high glucose conditions. Two biological replicates for each condition were made (high glucose: HEK-high-gluc-1 and HEK-high-gluc-2; and low glucose: HEK-low-gluc-1 and HEK-low-gluc-2).

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Transcriptomics-informed pharmacology identifies epigenetic and cell cycle regulators as enhancers of AAV production

Project description:Recombinant adeno-associated virus (rAAV) is a widely used viral vector for gene therapy. Despite its clinical efficacy, the manufacturing of rAAV faces challenges in productivity and quality, leading to limited availability. To address the growing demand, next-generation process development should be informed by a mechanistic understanding of the cellular response to rAAV. In this study, we performed transcriptomic analysis of 5 cell lines with variable capacities for rAAV production. Using an intersectional approach, we assessed the transcriptional response to rAAV production and compared transcriptional profiles between high and baseline producers to identify possible targets for enhancing production. Modulation of cell cycle and nucleosome components suggested a reduction of proliferative capacity and a shift toward DNA replication to support rAAV production. During rAAV production, we observed upregulation of several core functions including transcription, stress response, and Golgi and endoplasmic reticulum organization. Conversely, inhibitors of DNA-binding proteins and mitochondrial components were consistently downregulated during rAAV production. We next performed a drug connectivity analysis of these results and identified 5 classes of drugs predicted to enhance rAAV production. Validation studies confirmed the efficacy of HDAC and microtubule inhibitors. Our data uncover novel and previously identified pathways that may enhance rAAV productivity, potentially enabling a path to engineer improved processes and cell lines for higher yields and better quality rAAV production.

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Project description:Single-cell transcriptomics of transfection-based rAAV production in HEK293 cells

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Unveiling small non-coding RNA dynamics during recombinant Adeno-associated virus production.

Project description:Recombinant adeno-associated viruses (rAAVs) are a cornerstone of modern gene therapy, offering precise delivery and stable transgene expression with minimal immunogenicity. Despite clinical advancements, large-scale rAAV production faces significant challenges such as low yield, high costs, and variability, hindering its scalability. Recent research suggests that miRNAs and snoRNAs can modulate viral replication, transcriptional regulation, and host defense mechanisms, making them attractive targets for improving rAAV production. This study examines the expression profiles of microRNAs (miRNAs) and small nucleolar RNAs (snoRNAs) during rAAV plasmid transfection and production in HEK293F cells. We used microarrays to detail differential expression during rAAV production versus mock transfection and basal expression and identified significantly up- and down-regulated small non-coding RNAs (ncRNAs).

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