Vincent J. Huber

Inhibition of Aquaporin 4 by antiepileptic drugs

The potential of antiepileptic drugs (AEDs) to inhibit the water transport properties of aquaporin 4 (AQP4) was investigated using a combination of in silico and in vitro screening methods. Virtual docking studies on 14 AEDs indicated a range of docking energies that spanned approximately 40 kcal/mol, where the most stabilized energies were consistent with that of the previously identified AQP4 inhibitor acetazolamide. Nine AEDs and one bio-active metabolite were further investigated in a functional assay using AQP4 expressing Xenopus oocytes. Seven of the assayed compounds were found to inhibit AQP4 function, while three did not. A linear correlation was indicated between the in silico docking energies and the in vitro AQP4 inhibitory activity at 20 microM.

Aquaporins in drug discovery and pharmacotherapy.

Identification of the aquaporin (AQP) protein family more than twenty years ago has ushered in an era where water and neutral solute trafficking is considered a prime target for pharmacological intervention. Using AQP modulation as a basis for the treatment of human disorders has been suggested by phenotype analysis involving specific AQP-null animals, as well as by pathohistological studies. Based on those reports, a wide variety of disorders, such as cerebral edema, cancer and malaria, are considered indications for AQP modulators. Recent studies have also identified several small molecule AQP modulators that can be used to test those hypotheses in disease models. We believe these studies and compounds form the basis from which future treatments and diagnostic protocols of aquaporin-based disorders will be developed.

Identification of aquaporin 4 inhibitors using in vitro and in silico methods.

The in vitro inhibitory effects and in silico docking energies of 18 compounds with respect to aquaporin 4 (AQP4) were investigated. More than half of the compounds tested showed inhibitory activity in the in vitro functional assay and included the 5-HT(1B/1D) agonists sumatriptan, and rizatriptan. Moreover, the observed inhibitory activity of the compounds used in this study at 20 microM showed a strong correlation with their in silico docking energies, r(2)=0.64, which was consistent with that found in previous studies. The AQP4 inhibitory IC(50) values of three compounds, 2-(nicotinamido)-1,3,4-thiadiazole, sumatriptan and rizatriptan, were subsequently found to be 3, 11, and 2 microM, respectively.

Activity-dependent glial swelling is impaired in aquaporin-4 knockout mice.

We investigated the role of aquaporin-4 (AQP4), a water channel expressed in glial cells, in neural activity mediated morphological changes observed in brain slice preparation. Changes in flavoprotein fluorescence (FF) and infrared light scattering (LS) signals were measured before and after repetitive stimulation of layer VI in rostral somatosensory cortical slices taken from AQP4 knockout (KO) and wild-type (WT) mice. Changes in FF, which reflect neural aerobic activities, were comparable for the two groups in all cortical layers. However, changes in LS signals, which are indicative of cell swelling, were significantly decreased in layer I of AQP4 KO mice compared to that of WT mice. We conclude that AQP4 likely plays a significant role in neural activity-dependent glial swelling.

Development of a Novel Ligand, [11C]TGN-020, for Aquaporin 4 Positron Emission Tomography Imaging

Aquaporin 4 (AQP4), the most abundant isozyme of the water specific membrane transporter aquaporin family, has now been implicated to play a significant role in the pathogenesis of various disease processes of the nervous system from epilepsy to Alzheimer’s disease. Considering its clinical relevance, it is highly desirable to develop a noninvasive method for the quantitative analysis of AQP distribution in humans under clinical settings. Currently, the method of choice for such diagnostic examinations continues to be positron emission tomography (PET). Here, we report the successful development of a PET ligand for AQP4 imaging based on TGN-020, a potent AQP4 inhibitor developed previously in our laboratory. Utilizing [11C]-TGN-020, PET images were successfully generated in wild type and AQP4 null mice, providing a basis for future evaluation regarding its suitability for clinical studies.

Aquaporin-4 Positron Emission Tomography Imaging of the Human Brain: First Report

Aquaporin 4 (AQP‐4) is the most abundant aquaporin isoform in the brain. Alterations in its expression and distribution have been correlated with the progression of several clinical disorders; however, the specific roles of AQP‐4 in those disorders are not well understood. Visualizing AQP‐4 in vivo is expected to provide fresh insights into its roles in disease pathology, as well as aiding the clinical assessment of those disorders.

N-acetylaspartate Decrease in Acute Stage of Ischemic Stroke:A Perspective from Experimental and Clinical Studies

N-acetylaspartate (NAA) appears in a prominent peak in proton magnetic resonance spectroscopy ((1)H-MRS) of the brain. Exhibition by NAA of time-dependent attenuation that reflects energy metabolism during the acute stage of cerebral ischemia makes this metabolite a unique biomarker for assessing ischemic stroke. Although magnetic resonance (MR) imaging is a powerful technique for inspecting the pathological changes that occur during ischemic stroke, biomarkers that directly reflect the drastic metabolic changes associated with acute-stage ischemia are strongly warranted for appropriate therapeutic decision-making in daily clinical settings. In this review, we provide a brief overview of NAA metabolism and focus on the use of attenuation in NAA as a means for assessing the pathophysiological changes that occur during the acute stage of ischemic stroke.

Ligand-based molecular MRI: O-17 JJVCPE amyloid imaging in transgenic mice.

Development of molecular MR imaging (MRI) similar to PET imaging using contrast agents such as gadolinium as probe have been inherently hampered by incompatibility between potential probe (charged molecules) and membrane permeability. Nevertheless, considering the inherent spatial resolution limit for PET of 700μ, the superior microscopic resolution of MRI of 4 μ presents a strong incentive for research into ligand‐based molecular MRI.

Aquaporin-4 facilitator TGN-073 demonstrates novel analgesic activity

During pre-clinical development, we tested the novel, internally developed AQP-4 facilitator TGN-073 for its effect in a rodent pain model. Therein, TGN-073 was found to exert a strong analgesic effect. Following a single 200 mg/kg (i.p.) administration of TGN-073, a virtually complete block in the acetic acid writhing test was observed. Subsequent in vitro tests demonstrated that TGN-073 had no binding affinity for the μ-opioid or NK-1 receptors. Accordingly, we suspect TGN-073 or other AQP-4 facilitators may be developed into potent non-opioid analgesic agents. Given the potential significance of this discovery, we feel it should be openly shared with the scientific community.

Aquaporin Positron Emission Tomography Differentiates Between Grade III and IV Human Astrocytoma

BACKGROUND Aquaporin (AQP) water channels play a significant role in mesenchymal microvascular proliferation and infiltrative growth. AQPs are highly expressed in malignant astrocytomas, and a positive correlation is observed between their expression levels and histological tumor grade. OBJECTIVE To examine the utility of aquaporin positron emission tomography (PET) for differentiating between astrocytoma grade III and grade IV using the AQP radioligand [11C]TGN‐020. METHODS Fifteen astrocytoma patients, grade III (n = 7) and grade IV (n = 8) and 10 healthy volunteers underwent [11C]TGN‐020 aquaporin PET imaging. Surgical tissues of astrocytoma patients were examined for histopathological grading using the WHO classification standard and expression of AQP1 and AQP4 immunohistochemically. RESULTS Mean standardized uptake values of astrocytoma grade III and IV (0.51 ± 0.11 vs 1.50 ± 0.44, respectively) were higher than normal white matter (0.17 ± 0.02, P < .001) for both tumor grades. Importantly, mean standardized uptake values of astrocytoma grade IV were significantly higher than grade III (P < .01). CONCLUSION Our study demonstrated that [11C]TGN‐020 aquaporin PET imaging differentiated between astrocytoma grades III and IV. We suggest its clinical application as a noninvasive diagnostic tool would lead to advancements in the management of these malignant brain tumors.