nanomedicine Ruth Prassl Gottfried Schatz Research Center Cell Signaling, Metabolism and Aging Drug Delivery & Imaging Contact: Dr. Ruth Prassl Medical University of Graz GSFZ - Biophysics Nanomedicine Group Neue Stiftingtalstraße 6 A-8010 Graz Phone: +43 (0)316 385-71695 E-Mail: [email protected]Liposomes for micro-RNA delivery 1
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Liposomes for micro-RNA delivery - Nano2Clinic · Liposomes for micro-RNA delivery 1. ... Confer high stability to miRNA molecules ... Translational Aspects ..... 1st Food and Drug
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nanomedicine
Ruth Prassl
Gottfried Schatz Research CenterCell Signaling, Metabolism and Aging
Drug Delivery & Imaging
Contact:
Dr. Ruth Prassl
Medical University of GrazGSFZ - BiophysicsNanomedicine GroupNeue Stiftingtalstraße 6A-8010 Graz
energy uptake = energy expenditureBC + EXbasal exercise
consumption
+
expenditure
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Energy balance
weight maintenance
+thermogenesis
heat
+ Tthermo-genesis
Active brown adipose tissue exists in adult humans!J. Nedergaard, T. Bengtsson, B. Cannon, Am. J. Physiol. Endocrinol. Metab 293, E444-452 (2007) K. A. Virtanen et al., N. Engl. J. Med 360, 1518-1525 (2009); W. D. van Marken Lichtenbelt et al., N. Engl. J. Med 360, 1500-1508 (2009) ;A. M. Cypess et al., N. Engl. J. Med 360, 1509-1517 (2009)
brite/brownadipocyte
(BAT)
whiteadipocyte
Energy storage
Excess Energy
(WAT)
whiteadipocyte
briteadipocyte
MicroRNA
to convertadipocytes
adipocyteprogenitor
MicroRNA-26 Family
shifts human adipogenesisto brown characteristics
Karbiener et al., Stem Cells (2013), Karbiener et al. Meth.Mol.Bio. (2015)8
Turning fat-storing white adipocytes into fat-burning adipocytes (brown-like, brite or beige)
Strategy to fight overweight and obesity
• small multilocular lipid droplets
• high density of mitochondria
• positive for UCP1 protein in mitochondria
• massive energy dissipation
via thermogenesis
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• Size: 100 - 130 nm
• Zeta potential: liposome 44 ± 10 mV
liposome/miRNA complexes 42 ± 10 mV
Functional Unit
Neutral Lipid DOPE
Cationic LipidDOTAP
(DC-Cholesterol)
StabilizingPolymer
DSPE-PEG2000
% molar ratio 45/49/6
Lipid concentration(mg/ml)
3.0-7.0
miRNA (nM) 11-22
+
+
++
+
+
Toxicity : MTX and LDH
Screening of Lipid Formulation and miRNA/liposome complexes
Nat
ive
PA
GE
miRNA + + + + +liposome - + + + +
triton - - + + -nuclease - - - + +
miRNA-26a (22nt)
successful liposomeloading
effective microRNAprotection
Transfection of adipocytes – “Browning“
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human Multipotent Adipocyte Derived stem (hMADs) cells
Readouts: Elevated levels of UCP1
• Enhanced levels of miR-26a• Repressed target level of ADAM17• Long-term activation of PPARγ• Increased energy expenditure in terms of oxygen consumption
rela
tive
UC
P1
leve
ls
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white adipocyte
brite / beigeadipocyte
UCP1+ adipocyteUCP1- adipocyte
HPF: HiPerfect Transfection ReagentmiR-NTC : non targeting control
White adipocytes
Brown/brite adipocytes
UCP1 Immunostaining
Best Formulation …..
Transfection of adipocytes – “Browning“
RT-qPCR results
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Next Steps .....
Route of Application
subcutaneous implantation
microperfusion pumpAnimal models
Peptides
Aptamers
Active targeting of adipocytes
Alzet® Minipump
• Evaluation of in vivo activity of miR-26a• Evaluation of the composition of adipose tissues• Evaluation of the metabolic parameter
1st Food and Drug Administration (FDA) approved RNAi therapeutic in 2018Liposomal Patisiran (marketed as Onpattro® by Alnylam Pharmaceuticals) for the treatment of hereditary transthyretin-mediated (ATTR) amyloidosis.
MRX34 (MiRNA Therapeutics) a liposomal miR-34 mimic was the first miRNA therapeutic that has entered a phase 1 clinical trial in 2013 in patients with advanced solid tumors. Halted in 2016 following immune-related adverse effects.
Miravirsen (Roche/Santaris) a liposomal antagomir targeting miR-122 to treat hepatitis C is in phase 2
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Health Improvement
Treating cancer, obesity, related metabolic disorders…
Conclusion
Liposomes are a versatile and flexible nanoparticle platform
Used as universal delivery system for nucleic acids