Palenzuela, D. | bioRxiv Science

bioRxiv2023

Gene Expression Profiling Unveils the Temporal Dynamics of CIGB-300-Regulated Transcriptome in AML Cells

Protein kinase CK2 activity is implicated in the pathogenesis of various hematological malignancies like Acute Myeloid Leukemia (AML) that remains challenging concerning treatment. Consequently, here we used Illumina HT-12 microarray gene RNA expression profiling to study the molecular events that might support the anti-leukemic effect of CIGB-300 peptide which targets both CK2 substrates and the CK2 catalytic subunits on HL-60 and OCI-AML3 cell lines. As a result, 185 and 812 genes appeared significantly modulated in HL-60 cells at 30 min and 3 h of incubation with CIGB-300 for p< 0.01 and FC>[boxv]1.5[boxv], respectively; while 222 and 332 genes appeared modulated in OCI-AML3 cells. Importantly, functional enrichment analysis evidenced that genes and transcription factors related to apoptosis, cell cycle, leukocyte differentiation, signaling by cytokines/interleukins, and NF-kB, TNF signaling pathways were significantly represented in AML cells transcriptomic profiles. The influence of CIGB-300 on these biological processes and pathways is dependent on the cellular background, in first place, and treatment duration. Of note, the impact of the peptide on NF-kB signaling was corroborated by the quantification of selected NF-kB target genes, as well as the measurement of p50 binding activity and soluble TNF- induction. Quantification of CSF1/M-CSF and CDKN1A/P21 by PCR supports peptide effects on differentiation and cell cycle. Overall, here we explore for the first time the temporal dynamics of the gene expression profile regulated by CIGB-300 and provide fresh molecular clues concerning the antineoplastic effect of CIGB-300 in two relevant AML backgrounds.

cancer biology↗

bioRxiv2022

HeberFERON distinctively targets Cell Cycle in the glioblastoma-derived cell line U-87MG.

BackgroundHeberFERON is a co-formulation of 2b and {gamma} interferons, based on their synergism, that have shown its clinical superiority over individual interferons in basal cell carcinomas. In Glioblastoma (GBM), HeberFERON has shown promising preclinical and clinical results. This motivated us to design a microarray experiment aimed to identify the molecular mechanisms involved into the distinctive effect of HeberFERON compared with individual interferons. MethodsTranscriptional expression profiling including a control (untreated) and three groups receiving 2b-interferon, {gamma}-interferon and HeberFERON was performed using an Illumina HT-12 microarray platform. Unsupervised methods for gene and sample grouping, identification of differentially expressed genes, functional enrichment and network analysis computational biology methods were applied to identify distinctive patterns of HeberFERON action. Validation of most distinctive genes was performed by qPCR. Cell Cycle analysis of cell treated by HeberFERON for 24h, 48h and 72h was carried out by flow cytometry. ResultsThe three treatments show different behavior based on the gene expression profiles. Enrichment analysis identified several Mitotic Cell Cycle related events, in particular from Prometaphase to Anaphase, that are exclusively targeted by HeberFERON. FOXM1 transcription factor network which is involved in several Cell Cycle phases and is highly expressed in GBMs is significantly down regulated by HeberFERON. Flow cytometry experiments corroborated the action of HeberFERON over Cell Cycle in a dose and time dependent manner with a clear cellular arrest since 24h post-treatment. Despite the fact that p53 was not down-regulated by HeberFERON several genes involved in its regulatory activity were functionally enriched. Network analysis also revealed a strong relation of p53 with genes targeted by HeberFERON. We propose a mechanistic model to explain HeberFERON distinctive action, based on the simultaneous activation of PKR and ATF3, p53 phosphorylation changes as well as its reduced MDM2 mediated ubiquitination and export from nucleus to cytoplasm. PLK1, AURKB, BIRC5 and CCNB1 genes, all regulated by FOXM1, also play central roles in this model. These and other interactions could explain a G2/M arrest and the effect of HeberFERON over the proliferation of U-87MG. ConclusionsWe proposed molecular mechanisms underlying the distinctive behavior of HeberFERON compared to individual interferon treatments, where Cell Cycle related events showed the highest relevance.

cancer biology↗