title: Rationally designed aberrant kinase-targeted endogenous protein nanomedicine against oncogene mutated/amplified refractory chronic myeloid leukemia.
abstract: Deregulated protein kinases play a very critical role in tumorigenesis, metastasis, and drug resistance of cancer. Although molecularly targeted small molecule kinase inhibitors (SMI) are effective against many types of cancer, point mutations in the kinase domain impart drug resistance, a major challenge in the clinic. A classic example is chronic myeloid leukemia (CML) caused by BCR-ABL fusion protein, wherein a BCR-ABL kinase inhibitor, imatinib (IM), was highly successful in the early chronic phase of the disease, but failed in the advanced stages due to amplification of oncogene or point mutations in the drug-binding site of kinase domain. Here, by identifying critical molecular pathways responsible for the drug-resistance in refractory CML patient samples and a model cell line, we have rationally designed an endogenous protein nanomedicine targeted to both cell surface receptors and aberrantly activated secondary kinase in the oncogenic network. Molecular diagnosis revealed that, in addition to point mutations and amplification of oncogenic BCR-ABL kinase, relapsed/refractory patients exhibited significant activation of STAT5 signaling with correlative overexpression of transferrin receptors (TfR) on the cell membrane. Accordingly, we have developed a human serum albumin (HSA) based nanomedicine, loaded with STAT5 inhibitor (sorafenib), and surface conjugated the same with holo-transferrin (Tf) ligands for TfR specific delivery. This dual-targeted "transferrin conjugated albumin bound sorafenib" nanomedicine (Tf-nAlb-Soraf), prepared using aqueous nanoprecipitation method, displayed uniform spherical morphology with average size of ∼150 nm and drug encapsulation efficiency of ∼74%. TfR specific uptake and enhanced antileukemic activity of the nanomedicine was found maximum in the most drug resistant patient sample having the highest level of STAT5 and TfR expression, thereby confirming the accuracy of our rational design and potential of dual-targeting approach. The nanomedicine induced downregulation of key survival pathways such as pSTAT5 and antiapoptotic protein MCL-1 was demonstrated using immunoblotting. This study reveals that, by implementing molecular diagnosis, personalized nanomedicines can be rationally designed and nanoengineered by imparting therapeutic functionality to endogenous proteins to overcome clinically important challenges like molecular drug resistance.
authors.affiliation: Amrita Center for Nanosciences and Molecular Medicine, Amrita Vishwavidyapeetham University, Cochin, India.
authors.forename: Archana P
authors.initials: AP
authors.lastname: Retnakumari
authors.forename: Prasanna Lakshmi
authors.initials: PL
authors.lastname: Hanumanthu
authors.forename: Giridharan L
authors.initials: GL
authors.lastname: Malarvizhi
authors.forename: Raghuveer
authors.initials: R
authors.lastname: Prabhu
authors.forename: Neeraj
authors.initials: N
authors.lastname: Sidharthan
authors.forename: Madhavan V
authors.initials: MV
authors.lastname: Thampi
authors.forename: Deepthy
authors.initials: D
authors.lastname: Menon
authors.forename: Ullas
authors.initials: U
authors.lastname: Mony
authors.forename: Krishnakumar
authors.initials: K
authors.lastname: Menon
authors.forename: Pavithran
authors.initials: P
authors.lastname: Keechilat
authors.forename: Shantikumar
authors.initials: S
authors.lastname: Nair
authors.forename: Manzoor
authors.initials: M
authors.lastname: Koyakutty
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D001549:Benzamides; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects; D004337:Drug Carriers; D016503:Drug Delivery Systems; D015195:Drug Design; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D005434:Flow Cytometry; D016044:Fusion Proteins, bcr-abl / Q000235:genetics; D006801:Humans; D000068877:Imatinib Mesylate; D017404:In Situ Hybridization, Fluorescence; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D050997:Nanomedicine; D053758:Nanoparticles; D009536:Niacinamide / Q000031:analogs & derivatives* / Q000494:pharmacology; D010671:Phenylurea Compounds / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D010879:Piperazines / Q000494:pharmacology; D011487:Protein Conformation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D011990:Receptors, Transferrin / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050799:STAT5 Transcription Factor / Q000235:genetics / Q000378:metabolism; D012709:Serum Albumin / Q000378:metabolism; D000077157:Sorafenib; D014168:Transferrin / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D000970:Antineoplastic Agents; D001549:Benzamides; D004337:Drug Carriers; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D010671:Phenylurea Compounds; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D011743:Pyrimidines; D012333:RNA, Messenger; D011990:Receptors, Transferrin; D050799:STAT5 Transcription Factor; D012709:Serum Albumin; D014168:Transferrin; D009536:Niacinamide; D000068877:Imatinib Mesylate; D000077157:Sorafenib; D011505:Protein-Tyrosine Kinases; D016044:Fusion Proteins, bcr-abl