July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
The Effect of IVMED-80 Eye Drops on Lysinonorleucine (LNL) Amounts In Vivo for treatment of keratoconus.
Author Affiliations & Notes
  • Santosh Kumar Muddana
    OPTHAMOLOGY, Moran Eye Center, , Salt Lake City, Utah, United States
  • Haeli Hauritz
    iVeena Delivery Systems, Salt Lake City, Utah, United States
  • Michael Burr
    iVeena Delivery Systems, Salt Lake City, Utah, United States
  • Balamurali Ambati
    iVeena Delivery Systems, Salt Lake City, Utah, United States
  • Sarah Molokhia
    iVeena Delivery Systems, Salt Lake City, Utah, United States
  • Footnotes
    Commercial Relationships   Santosh Kumar Muddana, None; Haeli Hauritz, None; Michael Burr, None; Balamurali Ambati, None; Sarah Molokhia, None
  • Footnotes
    Support  NIH-NEI R43EY027636
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 3359. doi:
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      Santosh Kumar Muddana, Haeli Hauritz, Michael Burr, Balamurali Ambati, Sarah Molokhia; The Effect of IVMED-80 Eye Drops on Lysinonorleucine (LNL) Amounts In Vivo for treatment of keratoconus.. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3359.

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

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Purpose : Keratoconus (KCN) is a progressive disorder associated with structural changes in corneal collagen organization. Lysinonorleucine (LNL) is a major crosslink of the protein elastin and has been identified in collagen. Lower levels of LNL in the KC cornea suggest that there might be a cross-linking defect. In this study, we investigate the effect of IVMED-80 on (LNL) crosslink in rabbit cornea.

Methods : In this study, 3 groups (n=6, each) were tested using New Zealand white rabbits. IVMED-80 eye drops twice a day were compared to vehicle twice a day and control (no eye drops) for 6 weeks. After six weeks each group of corneas was dissected and used for LNL analysis. Recovered corneas weight was recorded. Samples were reduced with NaBH4 at room temperature then washed with water twice, dried, and hydrolyzed with 6N HCl in vacuo for 18 hours at 110°C. Post hydrolysis, cross-link enrichment was carried out by using a cellulose mini-column method. Further samples were provided for Mass Spec.
For Mass Spectroscopy Method, 1 uL of 100 ug/mL d9-Lysine was added to all samples as an internal standard. A calibration curve was created using serial dilution. Semi-quantitative mass spectral analysis was performed using a Sciex 6500 Q-Trap (Sciex, Farmington, MA). A helicon ihilic-Fusion 2.1 x 100 mm (Umeå, Sweden) was used for chromatography using 10 mM NH4OAc (Buffer A) and ACN (Buffer B). Mass spectrometry was performed in the positive mode with a TurboIon source using optimized source conditions: Curtain Gas = 20 L/min, Collision Gas = high, IonSpray = 5000 V, Temperature = 400 C, Ion Source Gas 1 = 20 L/min, Ion Source Gas 2 = 30. Declustering Potential, and Entrance Potential for all compounds were 51 V, and 10 V, respectively. Transitions for d9-lysine and lysinonorleucine were monitored via MRM.

Results : LC-MS results showed an increase in Lysinonorleucine (LNL) crosslink in treated rabbit cornea compared to Normal and Vehicle group. Lysinonorleucine (LNL) showed statistical difference between treated and control groups.

Conclusions : We have previously demonstrated an increase in Lysyl oxidase (LOX) activity IVMED-80 eye drops in human cadaver eyes. However, this is the first time to show in vivo increase of LNL between IVMED 80 treated corneas and control groups (non-treated and vehicle). This is biochemical evidence of crosslinking being induced by IVMED-80 for treatment of keratoconus.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.


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