Background: MicroRNAs (miRs) are small non coding RNAs involved in cell homeostasis. miRs are deregulated in colorectal cancer (CRC). Our study aimed at identifying miRs with a driver role in carcinogenesis altered by similar mechanisms in both human and mouse CRC. Goal of the study was to use CRC mouse models for the pre-clinical development of anti-miRs as therapeutic drugs. Methods: Azoximetane (AOM)/Dextran-Sulfate (DSS) treated mice or CDX2-CRE/APC-/- mice were used to study inflammation-associated and sporadic APC-related CRC. Human Inflammatory Bowel Disease associated (n=30), and sporadic (n=90) CRC with their matched normal tissues were collected according to Good Clinical Practice recommendation and subjected to RNA extraction using Trizol. miR and gene expression profiling was assessed by nCounter technology (Nanostring Seattle). Anti-miR-135b and scrambled probes for in vivo studies were synthesized by Girindus. Results: miRs profiling from AOM/DSS and CDX2-CRE/APC-/- CRC. revealed that miR-135b is one of the most up-regulated miRs in both models. In humans miR-135b over-expression was found in both IBD and sporadic CRC and was associated with reduced Progression Free Survival and Overall Survival in CRC patients. Molecular studies in Mouse Embryo Fibroblast and human CRC cell lines highlighted the role of two major pathways in the upstream activation of miR-135b: APC-β-Catenin and SRC-PI3K. MiR-135b up-regulation resulted in reduced apoptosis and increased invasion and metastasis due to the down-regulation of TGFRB2, DAPK1, APC and HIF1AN. Silencing of miR-135b in vivo reduced tumor multiplicity and tumor load in the AOM/DSS CRC model. Mice treated with anti-miR-135b showed well differentiated tumors and microacinar pattern while tumors in the control groups showed low differentiation and adenomatous pattern. Conclusions: Our data suggest that miR-135b is a key molecule whose activation is downstream of oncogenes and oncosuppressor genes frequently altered in CRC. Our study defines specific pathways that converge on the activation of the same microrna. The "in vivo" silencing of miR-135 shows preclinical efficacy with low toxicity and represents the first in vivo study for the use of antimiRs in CRC treatment.

Anti-miR-135b in colon cancer treatment: Results from a preclinical study.

GAFA', Roberta;LANZA, Giovanni;Croce CM
2012

Abstract

Background: MicroRNAs (miRs) are small non coding RNAs involved in cell homeostasis. miRs are deregulated in colorectal cancer (CRC). Our study aimed at identifying miRs with a driver role in carcinogenesis altered by similar mechanisms in both human and mouse CRC. Goal of the study was to use CRC mouse models for the pre-clinical development of anti-miRs as therapeutic drugs. Methods: Azoximetane (AOM)/Dextran-Sulfate (DSS) treated mice or CDX2-CRE/APC-/- mice were used to study inflammation-associated and sporadic APC-related CRC. Human Inflammatory Bowel Disease associated (n=30), and sporadic (n=90) CRC with their matched normal tissues were collected according to Good Clinical Practice recommendation and subjected to RNA extraction using Trizol. miR and gene expression profiling was assessed by nCounter technology (Nanostring Seattle). Anti-miR-135b and scrambled probes for in vivo studies were synthesized by Girindus. Results: miRs profiling from AOM/DSS and CDX2-CRE/APC-/- CRC. revealed that miR-135b is one of the most up-regulated miRs in both models. In humans miR-135b over-expression was found in both IBD and sporadic CRC and was associated with reduced Progression Free Survival and Overall Survival in CRC patients. Molecular studies in Mouse Embryo Fibroblast and human CRC cell lines highlighted the role of two major pathways in the upstream activation of miR-135b: APC-β-Catenin and SRC-PI3K. MiR-135b up-regulation resulted in reduced apoptosis and increased invasion and metastasis due to the down-regulation of TGFRB2, DAPK1, APC and HIF1AN. Silencing of miR-135b in vivo reduced tumor multiplicity and tumor load in the AOM/DSS CRC model. Mice treated with anti-miR-135b showed well differentiated tumors and microacinar pattern while tumors in the control groups showed low differentiation and adenomatous pattern. Conclusions: Our data suggest that miR-135b is a key molecule whose activation is downstream of oncogenes and oncosuppressor genes frequently altered in CRC. Our study defines specific pathways that converge on the activation of the same microrna. The "in vivo" silencing of miR-135 shows preclinical efficacy with low toxicity and represents the first in vivo study for the use of antimiRs in CRC treatment.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11392/1732584
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