July 2018
Volume 59, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2018
Evaluation of the Potential of AR-13324 and AR-13503 to Induce Phospholipidosis
Author Affiliations & Notes
  • Cheng-Wen Lin
    Aerie Pharmaceuticals, Inc., Durahm, North Carolina, United States
  • Briana E Foley
    Aerie Pharmaceuticals, Inc., Durahm, North Carolina, United States
  • Kevin Carbajal
    Aerie Pharmaceuticals, Inc., Durahm, North Carolina, United States
  • Casey Kopczynski
    Aerie Pharmaceuticals, Inc., Durahm, North Carolina, United States
  • Footnotes
    Commercial Relationships   Cheng-Wen Lin, Aerie Pharmaceuticals, Inc. (E); Briana Foley, Aerie Pharmaceuticals, Inc. (E); Kevin Carbajal, Aerie Pharmaceuticals, Inc. (E); Casey Kopczynski, Aerie Pharmaceuticals, Inc. (E)
  • Footnotes
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Investigative Ophthalmology & Visual Science July 2018, Vol.59, 6125. doi:
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      Cheng-Wen Lin, Briana E Foley, Kevin Carbajal, Casey Kopczynski; Evaluation of the Potential of AR-13324 and AR-13503 to Induce Phospholipidosis
      . Invest. Ophthalmol. Vis. Sci. 2018;59(9):6125.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose : Drug-induced phospholipidosis (PLD) is characterized by excessive lysosomal accumulation of phospholipids with the appearance of multilamellar bodies detectable by transmission electron microscopy (TEM). The purpose of this study was to assess the potential of AR-13324, an inhibitor of Rho kinase, and its primary metabolite, AR-13503, to induce PLD in Chinese hamster ovary (CHO-K1), human corneal epithelial (HCEpiC), and human retinal pigment epithelial (ARPE-19) cells.

Methods : We evaluated the potential of AR-13324 and AR-13503 to induce PLD using a validated cell-based fluorescent phospholipid assay (LipidTOXTM). The lysosome marker (LysoTracker®) was used to determine if the fluorescent phospholipid accumulation co-localized with lysosomes. Transmission electron microscopy (TEM) was used to assess the presence of multilamellar bodies in the lysosomes. In a separate pharmacokinetic (PK) study in New Zealand white (NZW) rabbits, the concentrations of AR-13324 and AR-13503 in aqueous humor (AH) following single topical administration of 0.02% AR-13324 were determined using an LC/MS/MS bioanalytical assay with a lower limit of quantification (LLOQ) of 0.100 ng/mL.

Results : The results showed that AR-13324 at micro-molar concentrations induced fluorescent phospholipid accumulation in a dose-dependent manner that co-localized with lysosomes in CHO-K1, HCEpiC, and ARPE-19 cells. TEM images revealed that AR-13324 induced the formation of lamellar bodies, confirming that the increase in phospholipid accumulation was due to PLD. In contrast, AR-13503 did not induce measurable phospholipid accumulation at any concentration tested in the cell-based assay. In the rabbit ocular PK study, only one of the 24 AH samples had AR-13324 levels above the LLOQ (1.82 ng/mL), whereas AR-13503 was measurable in all AH samples. These results indicate that AR-13324 was effectively converted to AR-13503 in rabbit AH following topical administration.

Conclusions : High concentrations of AR-13324 induce phospholipid accumulation in lysosomes in the same manner as other cationic amphiphilic drugs, such as amiodarone. AR-13503, the active metabolite of AR-13324, does not induce PLD and is the predominant form of the drug in AH following topical ocular dosing.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.


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