Evaluation of Corneal Thickness Measurement in Cynomolgus Monkeys
1. Introduction
Man and nonhuman primates share many ocular similarities, such as retinal cell
anatomy, the blood vessel system and the presence of a fovea. Because of these
similarities, ocular function can be investigated satisfactorily in nonhuman primate
models in toxicity studies (Niggemann, 2000). The cynomolgus monkey (Macaca
fascicularis, Figure 1) has proven to be a highly suitable and well established
animal model for examining the ocular toxicity of drugs, and the spectrum of
ophthalmoscopy techniques available for this animal is large.
Measurement of corneal thickness (CT) is a standard tool in the regulatory driven
safety evaluation of ocular drugs, and also NCEs (new chemical entities) with the
possibility of ophthalmic side effects. New equipment (here: Accutome AccuPach
VI®, Accutome/ADI, The Netherlands) for measuring CT (pachymetry), using
ultrasound technology, needs to undergo an in vivo characterization before being
utilized to ensure scientific and GLP (good laboratory practice) acceptance.
Evaluation of Pachymetry Data for the Measurement of Corneal Thickness in Cynomolgus Monkeys
S. Korte, C.M. Luetjens and B. Niggemann
Covance Laboratories GmbH, Münster, Germany
Discussion
Key elements for successful CT measurements are the accurate positioning and orientation of the probe on the eye surface. Specifically, measurement at
the same location of the cornea (central) and keeping the orientation of the probe perpendicular to the eye globe (in-line with visual axis). Further, it is
also necessary to maintain the animal eyes moist whilst sedated to avoid drying the cornea due to obstructed blinking. A manual blink of the eye before
taking the CT measurements will help support a uniform tear film.
Conclusions
In conclusion, the results for the CT measurements (with Accutome AccuPach VI® and the ultrasound technique) achieved a mean intra-run precision
between 5.3 and 6.9% and an inter-run precision between 1.5 and 1.9%. The mean concordance between staff was 97.9 and 98.3%. The tested system
was considered to be valid. Average results were between 411.3 and 418.7 µm and therefore lower than comparable monkeys measured by Kodamo,
2010 (men 580-616 µm, n=8, 5-7 years) using optical coherence tomography (RTVue-100, Optovue, Inc., Meridianville, AL, USA). The differences
observed related to the 2 technologies routinely being used in pachymetry assessment.
References
Birgit Niggemann (2000): Ocular toxicity investigations in primates and options for improvements. Waxmann Publishing Co., Münster, New York,
München, Berlin, 189-201.
Rinya Kodama, Xiuying Yang, Tomoaki Araki, Masato Sato, Shuichiro Iwashige, Yohei Tanigawa, Mie Uchimura, Keisuke Ishimaru, Yuji Sasaki, Yasuhiro
Kamimura, and Hiroshi Maeda (2010): Keratoconus in a cynomolgus monkey. Toxicol Pathol., 23(1): 53–58.
Figure 1. Group-housed cynomolgus monkeys.
Materials and Methods
In this study, 12 cynomolgus monkeys (Macaca fascicularis, 4.0-8.6 kg, 6-9 years)
underwent multiple readings (Table 1) for CT within a period of 30 minutes in the
late afternoon under light sedation using ketamine hydrochloride and topical
instillation of a local ophthalmic anesthetic (proxymetacain) into both eyes. The
sedated animals were placed in a restrain chair in a horizontal position for
pachymetry measurements. Intra- and inter-run precision (Table 1) were determined
for the right and left eye.
Table 1. Parameters Determined During the Study
Intra-run precision Examination of 12 different animals (both sexes)
with 3 repetitions per animal
Inter-run precision/
Inter-staff-robustness
Examination of 12 different animals (both sexes)
on two different days (2nd technician) with 3
repetitions per animal
Criteria of acceptance The results were accepted if the CV ≤ 25% was
achieved
Schedule 2 assessments (at one time point)
Evaluation for
pachymetry data
Mean value
Standard deviation
Percentage of coefficient of variation
Results
The results of the determination of CT (Tables 1-4) proved to be within the expected normal range
(with ultrasound technique) for cynomolgus monkeys and the equipment used was considered to be
valid (Table 5).
Table 2. Summary of Intra-Run Precision
Corneal Thickness
Left Eye
Average
(µm)
Left Eye
Average
(µm)
Left Eye
Average
(µm)
Right Eye
Average
(µm)
Right Eye
Average
(µm)
Right Eye
Average
(µm)
Technician 1
Mean 425 418 414 420 420 415
SD 43.1 34.2 29.3 34.2 36.9 41.7
N 12 12 12 12 12 12
Precision (% CV) 10.1 8.2 7.1 8.1 8.8 10.0
Mean Precision (% CV) 8.5 9.0
Technician 2
Mean 417 414 404 409 411 411
SD 16.1 27.5 45.9 21.7 23.6 22.4
N 10 10 10 10 10 10
Precision (% CV) 3.9 6.6 11.4 5.3 5.7 5.5
Mean Precision (% CV) 7.3 5.5
Table 3. Determination of Inter-Staff Robustness
Left Eye Right Eye
Animal No. Technician 1 Technician 2 % Concordance
between Staff
Technician 1 Technician 2 % Concordance
between Staff
1 483 - - 492 - -
2 409 447 109 417 430 103
3 432 422 98 424 436 103
4 410 422 103 425 419 99
5 358 384 107 351 366 104
6 408 429 105 417 417 100
7 408 397 97 387 388 100
8 417 358 86 423 426 101
9 412 423 103 410 416 101
10 447 462
11 383 405 106 379 390 103
12 455 429 94 433 415 96
Mean
Concordance
(%)
100.8 101.0
Correlation
(% CC)
37.3 92.2
Table 4. Determination of Inter-Run Precision
Technician
Left Eye
Average
(µm)
Average
(µm)
Average
(µm)
1 425 418 414
2 417 414 404
Mean 421 416 409
SD 5.3 2.3 6.8
N 2 2 2
Precision (% CV) 1.3 0.5 1.7
Mean Precision
(% CV)
1.2
Technician
Right Eye
Average
(µm)
Average
(µm)
Average
(µm)
1 420 420 415
2 409 411 411
Mean 414 416 413
SD 7.9 6.1 3.1
N 2 2 2
Precision (% CV) 1.9 1.5 0.7
Mean Precision
(% CV)
1.4
Table 5. Results of the Pachymetry Validation in
Cynomolgus Monkeys, Using Accutome AccuPach VI®
Test Range of found values (mean) Requested range Valid
Intra-Run Precision 3.9-11.4% CV (7.5% CV) ≤ 25% CV Yes
Inter-Run Precision 0.5-1.8% CV (1.2% CV) ≤ 25% CV Yes
Robustness 100.8-101.0% CV (100.9% CV) ≤ 25% CV Yes
Correlation (% CC) 37.3-92.2% (64.7%) concordance
99.9 % correlation
75-125% concordance
and/or ≤ 90% correlation
Yes
Mean of 3 Results of the Intra-Run Precision
