Treatment of tlr-4 mediated diseases and conditions with aptamers targeting tlr-4
An aptamer and pathological technology, applied in the field of treating TLR-4-mediated diseases and pathological conditions with an aptamer targeting TLR-4
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example 1
[0588] Example 1. Primary pharmacodynamics in vitro
[0589] Antagonism of hTLR-4 activation by aptamers
[0590] hTLR-4 activation assay was performed on HEK-blue-hTLR4 cells. The use of ultrapure LPS (0.1 ng / ml) as an important aspect related to the mechanism of kidney stone formation can also be drawn from these studies.
[0591] A TLR-4 agonist was used to activate cells, and a natural LPS antagonist (LPS-RS, 200 ng / ml) was used as a positive control for antagonistic activity against hTLR-4. hTLR-4 activation was quantified by measuring secreted embryonic alkaline phosphatase (SEAP) 24 hours after addition of ligand to the incubation medium. Poly AG nucleotides (38x) (38x(AG)) were used as control ssDNA (as scrambler). 38x(AG), is an oligonucleotide ssDNA with a fixed sequence, 38 times that of A-G. It was designed in the laboratory so it is a control aptamer that does not have any 3D structure or has a very limited and unstable When an aptamer interacts with a protei...
example 2
[0607] Example 2. Primary pharmacodynamics in vivo. Efficacy in a Rodent Model of Stroke
[0608] The animal model used in the study consisted of the following:
[0609] a) Permanent middle cerebral artery occlusion (pMCAO) by ligation and temporary middle cerebral artery occlusion (tMCAO) by ligation in mice (Chen et al. (1986) Stroke 17(4):738-743 ),
[0610] b) Transient intraluminal middle cerebral artery occlusion (tMCAO) in rats (Justicia et al. (2001) J Cereb Blood Flow Metab 21(9):1097-1104),
[0611] c) Permanent middle cerebral artery occlusion by electrocoagulation in rats and mice (Morancho et al., Neuropathol Appl Neurobiol 2012).
[0612] In all models, unilateral focal ischemic lesions were surgically induced in the cerebral cortex by permanent or temporary middle cerebral artery occlusion (MCAO). Following the recommendations of the STAIR stroke preclinical investigation (STAIR group: Update of the Stroke Therapy Academic Industry Roundtable Preclinical Rec...
example 3
[0640] Example 3. ApTOLL Effects in Humans
[0641] 3.1 Double-blind, placebo-controlled, randomized Phase Ia clinical study of ApTOLL for the treatment of acute ischemic stroke in healthy volunteers to assess tolerability and pharmacokinetics
[0642] Maximum recommended starting dose for humans
[0643] Calculation of the Maximum Recommended Starting Dose (MRSD) to be administered in Healthy Subjects:
[0644] NOAEL (No Observed Adverse Effect Level):
[0645] Rats: No side effects were observed with a higher dose of 50mg / kg / day intravenously injected for 14 days.
[0646] Cynomolgus monkeys: No side effects were observed with a higher dose of 13.9mg / kg / day (intravenous bolus injection) for 14 days.
[0647] The HED (Human Equivalent Dose) is calculated from the NOAEL, taking into account the conversion of animal doses to human equivalent doses based on body surface area. A correction factor of 10 is taken into account:
[0648] Rat: 50mg / kg x 0.162 / 10=0.81mg / kg
[064...
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