Ricerc@Sapienza

Chronic lymphocytic leukemia (CLL) is the most common human leukemia and the majority of patients show a deletion in the region 13q that encodes for mir15-a and mir16-1 that are normally responsible for the targeting of the antiapoptotic B cell lymphoma 2 protein (BCL2) which is consequently overexpressed in CLL and determines this pathology. In the proposed project, we will evaluate the delivery and the uptake efficiency of mir15-a and miR-16-1 through new modified ferritin-based nanoparticles. Compared with traditional systems, the humanized Archaeal ferritin (HumAfFt) offers several advantages and may be an ideal tool for an efficient and targeted delivery of nucleic acids. This chimeric ferritin is endowed with the recognition motifs typical of Human H homopolymer and the unique oligomerization properties of Archaeoglobus fulgidus ferritin. Based on an friendly protocol of purification, it is produced in large amounts, is thermostable and can be readily modified to efficiently encapsulate small nucleic acids into its cavity. Two different approaches will be used for mir-15a and 16-1 loading: i) physical entrapment by Mg2+ induced protein polymerization around nucleic acids and ii) chemical coupling of the thiol terminated miRNA with a cysteine residue located inside the ferritin cavity. Mir15-a and mir16-1 loaded HumAFt will be then tested for uptake efficiency on MEG01 cells, a very simple and powerful model displaying the classic CLL deletion 13q to study the expression of the target genes and the consequent apoptotic induction. We observed that HumAFt deliver efficiently and under physiological conditions fluorescent probes to HeLa cells through the CD71 receptor, overexpressed in many tumoral cell lines. The key deliverable of our project is a new ferritin-based RNAs nanocarrier, as a good alternative in the delivery of nucleic acids into cancer cell of human leukemia overcoming the significant experimental deficiency of traditional transfection agents.