Developmental Toxicity Assessment in Zebrafish - Zebrafish: Methods for Assessing Drug Safety and Toxicity - page 20

8.9% lethality. One hundred percent lethality was observed at 400 M; however,

precipitation was observed in the solution. Therefore, the actual LC 50 concentration
was not determined. Valproic acid has an estimated LC 50 of 59.8

m

M. 9-cis-Retinoic
acid did not cause lethality at the lowest testing concentration, 0.1

m

m M; therefore,

LC 50 was not determined.
Figure 2.2a shows the organ structure of treated zebrafish at 8

magnification.
Figure 2.2b shows the same images with organs outlined to highlight dysmorphogen-
esis: heart (blue), liver (red), and intestine (green). SSR101010-treated zebrafish
exhibited similar morphology as untreated and DMSO controls at all test concentra-
tions, although precipitation was observed at concentrations M. M100907-

treated zebrafish exhibited abnormal jaw (blue arrow in the left panel), and dark
opaque color in liver (black arrow in the left panel) and in intestine (yellow arrow in
the left panel), although organ morphology was not affected (outlined in the right
panel). In addition, arrhythmia was observed (not shown). Valproic acid-treated
zebrafish exhibited pericardial edema (red arrow in the left panel); therefore, the heart
chambers were elongated to form a thin tube (blue area in the right panel). In addition,
small liver and intestine were observed (red and green areas, respectively, in the right
panel). 9-cis-Retinoic acid-treated zebrafish exhibited pericardial edemawith a small,
elongated heart chamber (blue area in the right panel); small intestine (green area in
the right panel) and absence of liver tissue were also observed (lack of red area in the
right panel).
Using the TI definition, the values for SSR101010 could not be determined.
However, since no abnormalitywas observed at any test concentration, we determined
that SSR101010 was not teratogenic. Although LC 50 was not determined for
M100907, based on low lethality observed at the highest soluble concentration
(100

5 m

1.04; therefore, M100907 was considered teratogenic.
Valproic acid exhibited a TI value of 2.08, and was therefore teratogenic. Although TI
was not determined for 9-cis-retinoic acid, 100% of treated zebrafish exhibited
abnormalities and severe body deformation at the lowest tested nonlethal concen-
tration (0.1

m M), estimated TI was >

M), indicating that this drug is a potent teratogen. Results are summa-
rized in Table 2.3.

m

2.3.2 Results Based on Alternative TI Definition

In a recent comprehensive study, 12 blinded compounds, including valproic acid and
9-cis-retinoic acid (Bristol-Myers Squibb), were evaluated for developmental toxicity
in zebrafish embryos using TI value defined as visual assessment at maximal nonlethal
concentration. During the first phase of the study (LC 50 determination), compounds
were ranked according to LC 50 values prior to performing the time-consuming
developmental toxicity screen (Fig. 2.3). Teratogenicity properties of these com-
pounds were further evaluated by visual assessment in vivo (Table 2.4).
The data from this study were evaluated for strength in predictivity and accuracy
based on teratogenicity properties identified in vivo (Table 2.4). The assay was good
( > 70%, but < 80%) for specificity and excellent ( > 80%) for sensitivity. That is, it