Transferrin receptor-targeted liposomes encapsulating anti-BCR-ABL siRNA or asODN for chronic myeloid leukemia treatment.
The present work aimed at the development and application of transferrin receptor (TrfR)-targeted sterically stabilized liposomes encapsulating anti-BCR-ABL siRNA or asODN. Transferrin was coupled to the surface of liposomes encapsulating siRNA or asODN through the postinsertion method. Cell association and internalization were assessed by flow cytometry and confocal microscopy, respectively. BCR-ABL mRNA and Bcr-Abl protein levels were evaluated by qRT-PCR and Western blot, respectively. Cell viability was assessed using the resazurin reduction method. The amount of coupled transferrin and the size and stability over time of the liposomes were very satisfactory and reproducible. The siRNA encapsulation yield was dependent on the concentration of the encapsulation buffer used (20 or 300 mM), as opposed to asODN encapsulation yield which was high for both concentrations tested. Cell association and internalization studies were performed in leukemia cell lines treated with liposomes coupled to Trf (Trf-liposomes) or albumin (BSA-liposomes) or with nontargeted liposomes (NT-liposomes) encapsulating fluorescently labeled siRNA (Cy3-siRNA). These experiments clearly indicated that BSA- and NT-liposomes have no ability to promote the delivery of the encapsulated nucleic acids and that the Trf-liposomes deliver the nucleic acids by a Trf receptor-dependent mechanism. The Trf-liposomes encapsulating siRNA or asODN promote sequence-specific down-regulation of the BCR-ABL mRNA, although a certain extent of nonspecific sequence effects at the protein and cell viability level were observed. Overall, our results indicate that Trf-liposomes encapsulating gene silencing tools allow combining molecular and cellular targeting, which is a valuable approach for cancer treatment.
Mendonça
Liliana S
LS
Department of Biochemistry, University of Coimbra, Coimbra, Portugal.
Firmino
Filipe
F
Moreira
João N
JN
Pedroso de Lima
Maria C
MC
Simões
Sérgio
S
D015153:Blotting, Western; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D015536:Down-Regulation / Q000187:drug effects / Q000235:genetics; D016503:Drug Delivery Systems / Q000379:methods*; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000737:chemistry; D016044:Fusion Proteins, bcr-abl / Q000037:antagonists & inhibitors* / Q000235:genetics; D006801:Humans; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000188:drug therapy* / Q000235:genetics*; D008081:Liposomes; D018613:Microscopy, Confocal; D020319:Oligodeoxyribonucleotides, Antisense / Q000008:administration & dosage / Q000235:genetics / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics* / Q000494:pharmacology; D011990:Receptors, Transferrin / Q000378:metabolism*
D005456:Fluorescent Dyes; D008081:Liposomes; D020319:Oligodeoxyribonucleotides, Antisense; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011990:Receptors, Transferrin; D016044:Fusion Proteins, bcr-abl