Ofloxacin as a Disruptor of Actin Aggresome "Hirano Bodies": A Potential Repurposed Drug for the Treatment of Neurodegenerative Diseases

Abstract

There is a growing number of aging populations that are more prone to the prevalence of neuropathological disorders. Two major diseases that show a late onset of the symptoms include Alzheimer's disorder (AD) and Parkinson's disorder (PD), which are causing an unexpected social and economic impact on the families. A large number of researches in the last decade have focused upon the role of amyloid precursor protein, Aβ-plaque, and intraneuronal neurofibrillary tangles (tau-proteins). However, there is very few understanding of actin-associated paracrystalline structures formed in the hippocampus region of the brain and are called Hirano bodies. These actin-rich inclusion bodies are known to modulate the synaptic plasticity and employ conspicuous effects on long-term potentiation and paired-pulse paradigms. Since the currently known drugs have very little effect in controlling the progression of these diseases, there is a need to develop therapeutic agents, which can have improved efficacy and bioavailability, and can transport across the blood-brain barrier. Moreover, finding novel targets involving compound screening is both laborious and is an expensive process in itself followed by equally tedious Food and Drug Administration (FDA) approval exercise. Finding alternative functions to the already existing FDA-approved molecules for reversing the progression of age-related proteinopathies is of utmost importance. In the current study, we decipher the role of a broad-spectrum general antibiotic (Ofloxacin) on actin polymerization dynamics using various biophysical techniques like right-angle light scattering, dynamic light scattering, circular dichroism spectrometry, isothermal titration calorimetry, scanning electron microscopy, etc. We have also performed in silico docking studies to deduce a plausible mechanism of the drug binding to the actin. We report that actin gets disrupted upon binding to Ofloxacin in a concentration-dependent manner. We have inferred that Ofloxacin, when attached to a drug delivery system, can act as a good candidate for the treatment of neuropathological diseases.

Keywords: SEM; actin; biophysical studies; in silico; neurodegenerative diseases; ofloxacin; repurposable drugs.

FIGURE 1

Constant wavelength synchronous analysis (CWSF) profile for actin control in (A) polymerization buffer (PB), (B) G-actin buffer (GB), (C) water, and (D) Ofloxacin control in PB. Black color represents CWSF at 0 h, red color represents CWSF at 3 h, blue color represents CWSF at 6 h, green color represents CWSF at 24 h, pink color represents CWSF at 30 h, and the cyan color represents CWSF at T48 h.

FIGURE 2

Right-angle light scattering (RLS) profile for actin control and actin treated with Ofloxacin in (A) PB, (B) GB, and (C) water. The orange bar stands for actin aggregates at 0 h, green for 6 h, purple for 12 h, yellow for 18 h, blue for 24 h, and pink for 48 h.

FIGURE 3

Profile for dynamic light scattering (DLS) scattering of (A) untreated actin in PB; (B) actin treated with Ofloxacin in PB; (C) untreated actin control in GB; (D) actin treated with Ofloxacin in GB; (E) untreated actin control in water; and (F) treated actin with Ofloxacin at 300 μM.

FIGURE 4

CD Spectroscopic data analyzed for actin control and actin treated with Ofloxacin using CAPITO software. (A) PB; (B) GB; and (C) water. The blue curve is representative of actin control, whereas the green curve is representative of actin treated with Ofloxacin at 50 μM, and the red curve is representative of actin treated with Ofloxacin at 100 μM in the respective buffer.

FIGURE 5

Pie chart representation of the structural distribution of treated and untreated actin. (A) Actin control in PB. (B) Actin treated with Ofloxacin (50 μM) in PB. (C) Actin control in GB. (D) Actin treated with Ofloxacin (50 μM) in GB. (E) Actin control in water. (F) actin treated with Ofloxacin (50 μM) in water.

FIGURE 6

SEM images for actin (a) dialyzed in G-actin buffer 20 μm; (b) treated with Ofloxacin in G-actin buffer 20 μm; (c) dialyzed in water 2 μM; and (d) treated with Ofloxacin in water 2 μM.

FIGURE 7

Graphical representation of actin polymerization dynamics in the presence and absence of Ofloxacin at different concentrations observed in PB. (A) Actin control; (B) treated actin at 3 μM; (C) treated actin at 6 μM; (D) treated actin at 9 μM; (E) treated actin at 15 μM; (F) treated actin at 30 μM; (G) treated actin at 45 μM; (H) treated actin at 60 μM; (I) treated actin at 90 μM. The red straight line indicates the single exponential curve fit analysis while the black dots represent the RLS intensity recorded every 5 s of actin–Ofloxacin interaction.

FIGURE 8

Graphical representation of representative kinetic profiles in PB for actin treated with Ofloxacin: (A) Actin control; (B) extent of disintegration (y0); (C) amplitude (A2); and (D) time constant (t1).

FIGURE 9

Isothermal calorimetric profile for actin aggregates treated with Ofloxacin in PB. (A) Model: one-site binding; (B) model: sequential two-site binding; (C) model: sequential three-site binding; and (D) model: sequential four-site binding.

FIGURE 10

Actin polymer interaction with Ofloxacin. Hexamer actin. ChainA is shown salmon, ChainB in purple, ChainC in cyan, ChainD in lemon, ChainE in pink, and ChainF in orange.

FIGURE 11

Cartoon representation of the mechanism of action of Ofloxacin on actin aggregates. Actin aggregates upon treatment with Ofloxacin result in binding at two sites on actin. Large oligomers are formed in the intermediate phase followed by the formation of smaller-sized oligomers of monomeric actin.