Clinical presentations
With the exception of one asymptomatic female patient, the age at onset of 68 patients was shown in Fig.1. Fifteen cases (21.7%), 13 males and 2 females, suffered first attack during neonatal period, and the median onset age was 3 days (range, 1 day - 27 days). The remaining 53 cases (76.8%), 21 males and 32 females, had late-onset OTCD, and the median onset age was 2 years (range, 3 months - 56 years). Over all, three quarters of patients underwent the first attack of OTCD within 3 years of age, and the peak age was neonatal period in males and 1-2 years in females, respectively.
Clinical symptoms of these patients were summarized in Fig.2. Neurological symptoms were the most common clinical manifestations in both early-onset and late-onset groups. The acute severe neurological symptoms (including coma, lethargy and seizures) occurred in 14 cases (93.3%) in the early-onset and 33 cases (62.3%) in the late-onset group, indicating clinical symptoms were more serious in early-onset group. The gastrointestinal symptoms ranked second in both groups; vomiting was the most prominent symptom.
Biochemical results
The results of the biochemical detection were showed in Table 1. The levels of blood ammonia, and urine orotic acid were all elevated in patients tested. Nevertheless, elevated serum glutamine were found in 22.2% (12/54) of patients, decreased serum citrulline in 47.6% (30/63) of patients, and elevated uracil in 94.4% (51/54) of patients, respectively. In addition, 73.1% (38/52) of patients had elevated plasma alanine aminotransferase (ALT), among whom 73.7% (28/38) showed severe elevation of ALT. International normalized ratio was elevated in 11 patients tested.
Table 1
Clinical biochemical findings in 69 patients with OTCD
Patient No.
|
Sex
|
Onset age
|
Current/ Death age
|
Type
|
CITa μmol/L
|
GLNa μmol/L
|
Orotic acida mmol/molCr
|
Uracila mmol/molCr
|
Peak AMONa μmol/L
|
ALTa U/L
|
1
|
M
|
7d
|
8m
|
EO
|
1.71
|
3.25
|
425.42
|
8.60
|
300
|
32
|
2
|
M
|
3d
|
7d
|
EO
|
ND
|
ND
|
ND
|
ND
|
346
|
18
|
3
|
M
|
20d
|
24d
|
EO
|
ND
|
ND
|
ND
|
ND
|
ND
|
ND
|
4
|
M
|
3d
|
7d
|
EO
|
3.09
|
33.54
|
231.20
|
151.20
|
637
|
ND
|
5
|
M
|
9d
|
44d
|
EO
|
4.05
|
ND
|
ND
|
ND
|
815
|
ND
|
6
|
M
|
3d
|
7d
|
EO
|
2.48
|
80.60
|
267.60
|
53.26
|
700
|
18
|
7
|
M
|
5d
|
3.1y
|
EO
|
3.24
|
15.15
|
22.28
|
8.70
|
50
|
172
|
8
|
M
|
1d
|
5d
|
EO
|
2.18
|
24.79
|
127.33
|
90.78
|
>500
|
15
|
9
|
M
|
3d
|
7d
|
EO
|
4.80
|
43.73
|
1638.32
|
61.91
|
780
|
ND
|
10
|
M
|
3d
|
7d
|
EO
|
4.96
|
ND
|
214.77
|
9.89
|
1015
|
ND
|
11
|
M
|
2d
|
6d
|
EO
|
3.25
|
45.78
|
423.71
|
256.54
|
>500
|
ND
|
12
|
M
|
2d
|
2d
|
EO
|
3.40
|
ND
|
ND
|
ND
|
657
|
648
|
13
|
M
|
1d
|
7d
|
EO
|
ND
|
ND
|
ND
|
ND
|
1340
|
ND
|
14
|
M
|
56y
|
56y
|
LO
|
8.83
|
5.84
|
827.28
|
28.69
|
1152
|
23
|
15
|
M
|
1.8y
|
2.3y
|
LO
|
3.07
|
27.20
|
ND
|
17.93
|
180
|
42
|
16
|
M
|
8m
|
2.5y
|
LO
|
ND
|
ND
|
ND
|
ND
|
300
|
ND
|
17
|
M
|
2y
|
*
|
LO
|
5.74
|
10.83
|
65.68
|
29.17
|
218
|
ND
|
18
|
M
|
4m
|
9.7y
|
LO
|
6.83
|
44.39
|
228.79
|
311.67
|
350
|
28
|
19
|
M
|
36y
|
39y
|
LO
|
3.87
|
17.93
|
ND
|
ND
|
427
|
36
|
20
|
M
|
4.3y
|
4.3y
|
LO
|
21.74
|
16.37
|
312.35
|
248.40
|
114
|
244
|
21
|
M
|
1.4y
|
4.7y
|
LO
|
5.62
|
9.92
|
413.21
|
143.06
|
351
|
55
|
22
|
M
|
2.6y
|
4.4y
|
LO
|
ND
|
ND
|
ND
|
ND
|
244
|
ND
|
23
|
M
|
1.4y
|
2.4y
|
LO
|
25.43
|
5.61
|
137.00
|
6.07
|
114
|
40
|
24
|
M
|
9.7y
|
9.9y
|
LO
|
8.58
|
11.23
|
96.00
|
42.90
|
120
|
69
|
25
|
M
|
10m
|
1.4y
|
LO
|
2.28
|
ND
|
ND
|
ND
|
131
|
327
|
26
|
M
|
3y
|
10.8y
|
LO
|
7.48
|
22.23
|
48.31
|
165.31
|
289
|
119
|
27
|
M
|
7m
|
2.2y
|
LO
|
1.10
|
14.28
|
543.54
|
130.02
|
392
|
462
|
28
|
M
|
1y
|
8.7y
|
LO
|
7.09
|
14.21
|
1.85
|
19.79
|
45
|
26
|
29
|
M
|
1.2y
|
8.1y
|
LO
|
3.86
|
15.48
|
455.63
|
133.00
|
187
|
48
|
30
|
M
|
8m
|
1.8y
|
LO
|
13.60
|
ND
|
82.82
|
339.11
|
335
|
1782
|
31
|
M
|
2.1y
|
3y
|
LO
|
12.77
|
13.53
|
11.60
|
17.53
|
257
|
116
|
32
|
M
|
5y
|
5y
|
LO
|
2.80
|
ND
|
172.80
|
27.80
|
2500
|
19.7
|
33
|
M
|
3.4y
|
4y
|
LO
|
18.62
|
14.04
|
77.30
|
62.90
|
447
|
228.9
|
34
|
M
|
3m
|
4.5y
|
LO
|
2.43
|
12.81
|
1.79
|
5.72
|
265
|
105
|
35
|
F
|
1d
|
1m
|
EO
|
2.88
|
21.31
|
282.12
|
34.56
|
2000
|
ND
|
36
|
F
|
27d
|
8.8y
|
EO
|
14.83
|
29.73
|
197.00
|
51.80
|
410
|
2602
|
37
|
F
|
6y
|
9.6y
|
LO
|
19.24
|
14.22
|
198.45
|
205.32
|
194
|
509
|
38
|
F
|
2.5y
|
4.5y
|
LO
|
15.75
|
13.50
|
202.00
|
67.50
|
190
|
665
|
39
|
F
|
5.6y
|
5.6y
|
LO
|
12.60
|
11.65
|
189.20
|
50.04
|
500
|
759
|
40
|
F
|
2y
|
*
|
LO
|
14.07
|
18.96
|
141.39
|
35.73
|
ND
|
ND
|
41
|
F
|
1.5y
|
2y
|
LO
|
8.81
|
9.87
|
48.92
|
71.86
|
103
|
508
|
42
|
F
|
2.3y
|
5.1y
|
LO
|
9.70
|
12.78
|
43.50
|
251.00
|
1300
|
1330
|
43
|
F
|
No
|
4m
|
AS
|
6.61
|
ND
|
224.31
|
ND
|
59
|
28
|
44
|
F
|
2.3y
|
3.8y
|
LO
|
11.84
|
4.93
|
43.83
|
131.88
|
120
|
215
|
45
|
F
|
2y
|
2.8y
|
LO
|
7.68
|
32.68
|
ND
|
ND
|
256
|
ND
|
46
|
F
|
2.4y
|
3.9y
|
LO
|
13.44
|
61.66
|
673.28
|
161.53
|
385
|
380
|
47
|
F
|
6m
|
3.8y
|
LO
|
12.68
|
20.81
|
1435.31
|
153.89
|
387
|
214
|
48
|
F
|
29y
|
29y
|
LO
|
8.41
|
18.16
|
38.75
|
51.55
|
421
|
ND
|
49
|
F
|
1.5y
|
8.4y
|
LO
|
15.90
|
23.87
|
457.51
|
112.32
|
133
|
ND
|
50
|
F
|
2.3y
|
2.8y
|
LO
|
15.66
|
37.02
|
251.06
|
42.70
|
300
|
ND
|
51
|
F
|
3y
|
13.2y
|
LO
|
10.91
|
31.70
|
356.42
|
65.03
|
286
|
422
|
52
|
F
|
1.5y
|
3.3y
|
LO
|
1.77
|
ND
|
ND
|
ND
|
350
|
1455.00
|
53
|
F
|
1.1y
|
3.2y
|
LO
|
4.27
|
ND
|
469.38
|
261.67
|
260
|
72
|
54
|
F
|
1.8y
|
9.3y
|
LO
|
5.94
|
15.18
|
204.05
|
83.30
|
216
|
415
|
55
|
F
|
10m
|
2.6y
|
LO
|
ND
|
ND
|
ND
|
ND
|
315
|
885
|
56
|
F
|
3.5y
|
5.1y
|
LO
|
9.53
|
17.52
|
38.21
|
6.75
|
234
|
687
|
57
|
F
|
1.3y
|
7y
|
LO
|
8.21
|
26.24
|
227.12
|
441.98
|
189
|
139
|
58
|
F
|
1y
|
2.8y
|
LO
|
4.21
|
21.26
|
4.53
|
ND
|
480
|
97
|
59
|
F
|
6y
|
6.7y
|
LO
|
10.40
|
42.54
|
199.30
|
ND
|
170
|
338.2
|
60
|
F
|
2y
|
8.4y
|
LO
|
15.56
|
17.72
|
305.73
|
48.85
|
278
|
964
|
61
|
F
|
4y
|
9.9y
|
LO
|
5.69
|
23.48
|
19.93
|
338.64
|
>500
|
839
|
62
|
F
|
1.1y
|
2.6y
|
LO
|
3.17
|
14.11
|
77.65
|
29.97
|
300
|
303
|
63
|
F
|
1.4y
|
7.3y
|
LO
|
8.82
|
49.73
|
691.94
|
559.21
|
197
|
1639
|
64
|
F
|
1y
|
7.3y
|
LO
|
3.70
|
5.18
|
55.46
|
156.57
|
383
|
49
|
65
|
F
|
9.3y
|
*
|
LO
|
7.44
|
56.51
|
106.88
|
61.87
|
268
|
ND
|
66
|
F
|
1.7y
|
3.3y
|
LO
|
11.54
|
20.67
|
129.10
|
107.17
|
>500
|
623
|
67
|
F
|
5.5y
|
5.7y
|
LO
|
7.57
|
19.38
|
308.70
|
78.95
|
231
|
311
|
68
|
F
|
2y
|
*
|
LO
|
6.94
|
29.11
|
213.13
|
164.51
|
256
|
388
|
69
|
F
|
1.5y
|
5.7y
|
LO
|
12.37
|
24.27
|
2.72
|
71.62
|
320
|
111
|
Abbreviations: M: Male; F: Female; d: days; m: months; y: years; EO: Early-onset; LO: Late-onset; AS: Asymptomatic; CIT: Blood citrulline; GLN: Blood glutamine; AMON: Blood ammonia; ND: not detected; “*” donates loss to follow-up.
a Normal reference values: CIT 7-35 umol/L; GLN 6-30 umol/L; Urine orotic acid 0-1.5 mmol/molCr; Uracil: 0-7 mmol/molCr; AMON 9-30 μmol/L; ALT 9-52U/L; severe elevation of ALT was defined as >175 U/L in females, >200 U/L in males.
|
We also compared the biochemical data between male and female patients (Table 2). Among the patients with detected citrulline concentrations, 69.0% (20/29) of male patients and 29.4% (10/34) of female patients had decreased serum citrulline concentrations, with significantly lower levels of serum citrulline concentrations in the male group (P<0.01). However, there were no significant differences in other metabolic characteristics between male and female groups (P > 0.05).
Table 2
Comparison of biochemical data between different gender groups and between different phenotypes
|
CIT
μmol/L
|
GLN
μmol/L
|
Orotic acid
mmol/molCr
|
Uracil
mmol/molCr
|
AMON
μmol/L
|
Reference values
|
7~35
|
6~30
|
0~1.5
|
0~7
|
9~30
|
Male group
(n=34)
|
4.1 (29)
1.1~25.4
|
15.1 (23)
3.3~80.6
|
154.9 (24)
1.7~1638.3
|
53.3 (25)
5.7~339.1
|
335 (33)
45~2500
|
Female group
(n=34)
|
9.2 (34)
1.8~19.2
|
20.8 (31)
4.9~61.7
|
198.9 (32)
2.7~1435.3
|
79.0 (29)
6.8~559.2
|
273 (34)
59~2000
|
Z
|
-3.061
|
-1.268
|
-0.199
|
-1.795
|
-1.117
|
P
|
0.002
|
0.205
|
0.843
|
0.073
|
0.264
|
EO group
(n=15)
|
3.3 (12)
1.7~14.8
|
29.7 (9)
3.3~80.6
|
223.0 (10)
1.7~1638.3
|
52.5 (10)
8.6~256.5
|
679 (14)
50~2000
|
LO group
(n=53)
|
8.4 (51)
1.1~25.4
|
17.7 (45)
4.9~61.7
|
181.0 (46)
1.8~1435.3
|
75.4 (44)
5.7~559.2
|
265 (53)
45~2500
|
Z
|
-3.326
|
-1.938
|
-0.642
|
-1.336
|
-3.726
|
P
|
0.001
|
0.053
|
0.521
|
0.182
|
0.000
|
Quantitative data were expressed as the median (number of patients tested) and the range (minimum~ maximum).
|
In addition, the biochemical data were compared between early-onset and late-onset groups (Table 2). The peak ammonia levels in the early-onset group were much higher than those in the late-onset group (P < 0.01), which indicated that it may be associated with disease severity. Approximately 91.7% (11/12) of the patients with early-onset and 37.3% (19/51) of patients with late-onset showed decreased citrulline concentrations. The citrulline concentration was significantly lower in patients with early-onset (P < 0.01). Nevertheless, blood glutamine, urine orotic acid and uracil concentration were similar between the two groups (P > 0.05).
Mutation analysis of the OTC gene
We identified 54 different mutations in 63 unrelated patients including 28 (51.9%) missense mutations, 5 (9.3%) nonsense mutations, 8 (14.8%) splicing mutations, 7 (13.0%) gross deletions, 3 (5.6%) small deletions, 2 (3.7%) gross duplications, and 1(1.9%) small insertion (Table 3). In 3 patients (P67-P69), the results of WES were negative, but no MPLA was performed due to parental refusal. De novo mutations had been found in 36.2% (25/69) of the patients in this study. Mutations observed in male patients were mainly distributed in exon 6, 8, and 9, while mutations observed in females were highly dispersed. The most common mutations in our cohort were R277W (7/66, 10.6%), followed by G195R (3/66; 4.6%) and A209V (2/66; 3.0%). Of all mutations, 18 were novel: c.77+1G>C (IVS1+1G>C), c.78-2 A>G (IVS1-2A>G), c.122A>C (p.D41A), c.124_126delCTT (p.42delL), c.207-226del20, c.552insGAAC (p.S185Efs*41), c.703C>T (p.Q235X), c.704A>C (p.Q235P), c.860C>T (p.T287I), c.868-1 G>C (IVS8-1G>C), c.913C>T(p.P305S), E1-4 deletion, E1-4 duplication, E2-6 duplication, E5-8 deletion, E7-10 deletion, E9-10 deletion, and 7.8 Mb deletion of Xp11.4p21.2, which have not been previously reported in the literature or registered in the HGMD, ClinVarMiner, and ExAC database.
Table 3
Molecular detective results, family history, and clinical outcomes identified in 69 patients with OTCD
NO.a
|
Type
|
Exon/
Intron
|
Nucleotide change
|
Amino acid change
|
Nature of mutation
|
Family history
|
Therapy receivedb
|
Outcome/ Neurologic damage
|
1
|
EO
|
E4
|
c.386G>A
|
p.R129H
|
NA
|
No
|
LPD, Drug
|
Alive/No
|
2
|
EO
|
E5
|
c.482A>G
|
p.N161S
|
Inherited
|
Yes
|
ND
|
Deceased
|
3
|
EO
|
E6
|
c.579G>A
|
p.W193X
|
Inherited
|
Yes
|
ND
|
Deceased
|
4
|
EO
|
E6
|
c.583G>A
|
p.G195R
|
Inherited
|
Yes
|
ND
|
Deceased
|
5
|
EO
|
E8
|
c.725C>T
|
p.T242I
|
Inherited
|
No
|
LPD, Drug
|
Deceased
|
6
|
EO
|
E8
|
c.829C>T
|
p.R277W
|
De novo
|
No
|
MV,TPN, PD
|
Deceased
|
7
|
EO
|
E8
|
c.860C>T
|
p.T287>I
|
Inherited
|
Yes
|
Drug
|
Alive/No
|
8
|
EO
|
E9
|
c.904C>T
|
p.H302Y
|
Inherited
|
No
|
Drug
|
Deceased
|
9
|
EO
|
E2-6
|
E2-6 duplication
|
Inherited
|
No
|
ND
|
Deceased
|
10
|
EO
|
E7-10
|
E7-10 deletion
|
Inherited
|
Yes
|
ND
|
Deceased
|
11
|
EO
|
NA
|
NA
|
NA
|
NA
|
No
|
ND
|
Deceased
|
12
|
EO
|
NA
|
NA
|
NA
|
NA
|
No
|
ND
|
Deceased
|
13
|
EO
|
NA
|
NA
|
NA
|
NA
|
No
|
Drug
|
Deceased
|
14
|
LO
|
E2
|
c.119G>A
|
p.P40H
|
NA
|
Yes
|
Drug
|
Deceased
|
15
|
LO
|
E2
|
c.122A>C
|
p.D41A
|
Inherited
|
Yes
|
Drug
|
Alive/Yes
|
16
|
LO
|
E4
|
c.386G>T
|
p.R129C
|
Inherited
|
Yes
|
LT
|
Alive/Yes
|
17
|
LO
|
E6
|
c.562G>C
|
p.G188R
|
Inherited
|
Yes
|
Withdraw
|
Withdraw
|
18
|
LO
|
E6
|
c.604C>T
|
p.H202Y
|
Inherited
|
No
|
LT
|
Alive/Yes
|
19
|
LO
|
E6
|
c.622G>A
|
p.A208T
|
Inherited
|
No
|
Drug
|
Alive/Yes
|
20
|
LO
|
E8
|
c.794G>A
|
p.W265X
|
De novo
|
No
|
Drug
|
Deceased
|
21
|
LO
|
E8
|
c.829C>T
|
p.R277W
|
Inherited
|
No
|
LPD, Drug
|
Alive/No
|
22
|
LO
|
E8
|
c.829C>T
|
p.R277W
|
Inherited
|
No
|
LT
|
Alive/Yes
|
23
|
LO
|
E8
|
c.829C>T
|
P.R277W
|
Inherited
|
No
|
LPD, Drug
|
Alive/No
|
24
|
LO
|
E8
|
c.829C>T
|
P.R277W
|
Inherited
|
Yes
|
Drug
|
Alive/Yes
|
25
|
LO
|
E8
|
c.829C>T
|
p.R277W
|
Inherited
|
No
|
LPD, Drug
|
Deceased
|
26
|
LO
|
E8
|
c.829C>T
|
p.R277W
|
Inherited
|
No
|
LPD, Drug
|
Alive/Yes
|
27
|
LO
|
E9
|
c.913C>T
|
p.P305S
|
De novo
|
No
|
LPD, Drug
|
Alive/Yes
|
28
|
LO
|
E9
|
c.919A>G
|
p.K307E
|
Inherited
|
No
|
LPD
|
Alive/Yes
|
29
|
LO
|
E9
|
c.931G>A
|
p.V311M
|
Inherited
|
No
|
LPD, Drug
|
Alive/No
|
30
|
LO
|
E10
|
c.1019C>T
|
p.S340F
|
Inherited
|
No
|
LPD, Drug
|
Alive/Yes
|
31
|
LO
|
E9
|
c.929_931delAAG
|
p.309delE
|
De novo
|
No
|
Drug
|
Alive/ No
|
32
|
LO
|
E1-4
|
E1-4 duplication
|
Inherited
|
Yes
|
ND
|
Deceased
|
33
|
LO
|
-
|
7.8Mb deletion of Xp11.4p21.2
|
De novo
|
No
|
CVVH, Drug
|
Alive/Yes
|
34
|
LO
|
E2
|
c.78-2 A>G
|
IVS1-2A>G
|
Inherited
|
Yes
|
Drug
|
Deceased
|
35
|
EO
|
I1
|
c.77+1G>C
|
IVS1+1G>C
|
Inherited
|
No
|
ND
|
Deceased
|
36
|
EO
|
E9-10
|
E9-10 deletion
|
De novo
|
No
|
LT
|
Alive/Yes
|
37
|
LO
|
E1
|
c.3G>A
|
p.M1I
|
De novo
|
No
|
LPD, Drug
|
Alive/Yes
|
38
|
LO
|
E1
|
c.67C>T
|
p.R23X
|
Inherited
|
No
|
LPD, Drug
|
Alive/No
|
39
|
LO
|
E2
|
c.140delA
|
p.N47Tfs*17
|
De novo
|
No
|
Drug
|
Deceased
|
40
|
LO
|
E2
|
c.174G>A
|
p.W58X
|
De novo
|
No
|
Withdraw
|
Withdraw
|
41
|
LO
|
E3
|
c.275G>A
|
p.R92Q
|
De novo
|
No
|
LPD, Drug
|
Deceased
|
42
|
LO
|
E4
|
c.317G>T
|
p.G106V
|
Inherited
|
No
|
LT
|
Alive/No
|
43
|
AS
|
E5
|
c.421C>G
|
p.R141G
|
Inherited
|
Yes
|
Drug
|
Alive/No
|
44
|
LO
|
E5
|
c.422G>A
|
p.R141Q
|
Inherited
|
No
|
Drug
|
Deceased
|
45
|
LO
|
E5
|
c.540G>C
|
p.Q180H
|
De novo
|
No
|
LT
|
Alive/Yes
|
46
|
LO
|
E6
|
c.552insGAAC
|
p.S185Efs*41
|
De novo
|
No
|
Drug
|
Deceased
|
47
|
LO
|
E6
|
c.583G>A
|
p.G195R
|
Inherited
|
No
|
LT
|
Alive/Yes
|
48
|
LO
|
E6
|
c.583G>A
|
p.G195R
|
Inherited
|
No
|
ND
|
Deceased
|
49
|
LO
|
E6
|
c.626C>T
|
p.A209V
|
Inherited
|
No
|
LPD, Drug
|
Alive/Yes
|
50
|
LO
|
E6
|
c.626C>T
|
p.A209V
|
De novo
|
No
|
Drug
|
Alive/Yes
|
51
|
LO
|
E7
|
c.704A>C
|
p.Q235P
|
Inherited
|
No
|
LT
|
Alive/Yes
|
52
|
LO
|
E7
|
c.703C>T
|
p.Q235X
|
Inherited
|
Yes
|
CVVH, LPD, Drug
|
Alive/Yes
|
53
|
LO
|
E8
|
c.779T>C
|
p.L260S
|
Inherited
|
No
|
ND
|
Deceased
|
54
|
LO
|
E9
|
c.914C>G
|
p.P305R
|
De novo
|
No
|
LT
|
Alive/Yes
|
55
|
LO
|
E9
|
c.944T>G
|
p.V315G
|
De novo
|
No
|
LT
|
Alive/No
|
56
|
LO
|
I3
|
c.298+2T>G
|
IVS3+2 T>G
|
Inherited
|
No
|
LT
|
Alive/Yes
|
57
|
LO
|
I3
|
c.298+5G>C
|
IVS3+5G>C
|
De novo
|
No
|
LPD, Drug
|
Alive/Yes
|
58
|
LO
|
I5
|
c.540+265G>A
|
IVS5+265 G>A
|
De novo
|
No
|
LPD, Drug
|
Alive/Yes
|
59
|
LO
|
I6
|
c.664-1G>A
|
IVS6-1 G>A
|
De novo
|
No
|
Drug
|
Alive/No
|
60
|
LO
|
I7
|
c.718-1G>A
|
IVS7-1G>A
|
De novo
|
No
|
Drug
|
Alive/Yes
|
61
|
LO
|
I8
|
c.868-1G>C
|
IVS8-1G>C
|
De novo
|
No
|
Drug
|
Alive/Yes
|
62
|
LO
|
E2
|
c.124_126delCTT
|
p.42delL
|
De novo
|
No
|
LT
|
Alive/No
|
63
|
LO
|
E2-3
|
c.207-226del20
|
De novo
|
No
|
LPD, Drug
|
Alive/No
|
64
|
LO
|
E1-4
|
E1-4 deletion
|
Inherited
|
Yes
|
LPD, Drug
|
Alive/Yes
|
65
|
LO
|
E2-4
|
E2-4 deletion
|
De novo
|
No
|
Withdraw
|
Withdraw
|
66
|
LO
|
E5-8
|
E5-8 deletion
|
De novo
|
No
|
LPD, Drug
|
Alive/No
|
67
|
LO
|
ND
|
ND
|
ND
|
ND
|
Yes
|
LPD, Drug
|
Deceased
|
68
|
LO
|
ND
|
ND
|
ND
|
ND
|
No
|
Withdraw
|
Withdraw
|
69
|
LO
|
ND
|
ND
|
ND
|
ND
|
No
|
LT
|
Alive/Yes
|
Abbreviations: E: Exon, I: Intron; LPD: Low-protien diet; MV: mechanical ventilation; TPN: total parenteral nutrition; PD: peritoneal dialysis; LT: Liver transplantation; CVVH: continuous veno venous hemodiafltration; NA: not analysed; ND: not detected.
a Patients 1-34 are males, and patients 35-69 are females. bDrug: Referred to L-arginine, L-Citrulline, sodium benzoate, and sodium phenylbutyrate.
|
The bioinformatic characteristics of the 4 novel missense variants (D41A, Q235P, T287I, and P305S) are shown in Table 4. D41 interlinks with carbamoyl phosphate domain through hydrogen bonds. The known mutant D41G has been predicted to decrease the stability of carbamoyl phosphate domain by structure based analysis [17]. Therefore, the mutant D41A may also affect the function of this domain and is predicted to be damaging. Q235 is located at the α-helix in the equatorial domain [18]. The mutant P235 may affect the folding of the equatorial domain and is predicted to be damaging by SIFT and Mutation Taster, but benign by PROVEAN and PolyPhen-2. T287 is located at the surface of the enzyme, which is close to the conserved amino acid motif Ser-Met-Gly (SMG loop) that swings towards the active site to help catalysis [19]. The mutant I287 may affect this local neighborhood structure and is predicted to be deleterious by all servers. P305 is located at the C-terminal ornithine binding domain, which contains the constant motif Leu-His-Cys-Leu-Pro. The nonpolar amino acid proline forms a rare cis-peptide bond that is found exclusively at this location, and largely determines the shape of the ornithine site. As such, P305R mutations would be highly damaging, as observed in a male patient with neonatal-onset, which has complete enzyme deficiency [18]. Consequently, the mutant residue S305, a polar and neutral amino acid, may affect the interaction of ornithine binding and is predicted to be pathogenic by all servers.
Table 4
Prediction of the potential pathogenic effect of novel missense variants of OTC gene
Mutation
|
Domain
|
PROVEANa
|
PolyPhen-2b
|
SIFTc
|
Mutation Taster d
|
Conservation
|
D41A
|
β-sheet in polar domain
|
Deleterious
|
Benign
|
Damaging
|
126
|
Conserved
|
Q235P
|
α-helix in equatorial domain
|
Neutral
|
Benign
|
Damaging
|
76
|
Conserved
|
T287I
|
Surface of the enzyme
|
Deleterious
|
Probably damaging
|
Damaging
|
89
|
Conserved
|
P305S
|
Ornithine binding domain
|
Deleterious
|
Probably damaging
|
Damaging
|
74
|
Conserved
|
a PROVEAN prediction: default threshold is -2.5, that is variants with a score equal to or below -2.5 are considered “deleterious”, whereas variants with a score above -2.5 are considered “neutral”.
b PolyPhen-2 prediction: probably damaging with a score of 1, in contrast, possibly damaging with a score under 1.
c SIFT prediction: amino acids with probabilities <0.05 are predicted to be deleterious, whereas variants with a score above 0.05 are considered “neutral”.
d Mutation taster prediction: scores range from 0.0 to 215. The more they score, the more deleterious protein mutations.
|
Clinical outcomes
During 15 years of follow-up, 24 cases (16 males, 8 females) had died, 41 cases (17 males, 24 females) survived, and 4 cases (1 male, 3 females) withdrawn from this study. The mortality was 48.5% (16/33) in male patients and 25.0% (8/32) in female patients. The median age at death was 7 days (range, 2 days–56 years) in males and 3.9 years (range, 1 month–29 years) in females. In the early-onset group, 12 cases died following their first hyperammonemic encephalopathy at the median age of 7 days (range, 2 days-44 days), resulting in a high mortality of 80% (12/15). In the late-onset group, except for that 4 cases lose to follow-up, 12 patients deceased at median age of 4.4 years (range, 1.4 years–56 years), with a mortality of 24.5% (12/49). Estimated survival rates of these groups were shown in Fig.3. For the 41 survivors, they survived during the initial hyperammonemic episode through nutritional support, administration of intravenous ammonia scavengers, and intensive care. Subsequent clinical progress showed neurological impairments including developmental delay, mental retardation, learning disorder, behavior disorders, motor disorders and epilepsy, were common (65.8%, 27/41).
In this study, 13 patients (3 males, 10 females) received liver transplantation (LT). The median age at operation was 3 years (ranged from 19 months to 11.4 years). The overall median peak blood ammonia before LT was 300 µmol/L (range 216–1300) in 13 patients. Prior to transplantation, the median number of hospital admissions for hyperammonemic crises was 3 (range, 1-12). At an average follow-up period of 1.25 years (range, 0–2.7 years) following LT, the children could have a protein-unrestricted diet, with the blood ammonia being maintained in the normal range and no hyperammonemia episodes happened. Postoperative complications occurred in 2 patients, one (P18) having hepatapostema and the other (P36) having biliary fistula. Three patients (P42, P55, and P62) had good prognosis and underwent normal development. For the other 10 patients, neurological impairments were found before transplantation, and remained stable condition in 8 of them (P18, P22, P36, P45, P47, P51, P54, and P56) and got improved in 2 cases (P16, P69) after transplantation.