Mutations in KIF21A and PHOX2A are absent in 16 patients with congenital vertical incomitant strabismus

To the Editor:

Genetic studies of families with rare forms of familial congenital incomitant strabismus (eg, congenital fibrosis of the extraocular muscles [CFEOM]) have led to identification of genes responsible for ocular cranial nerve development and to the unifying concept of “congenital cranial dysinnervation disorder (CCDD)” to describe such phenotypes.¹ CFEOM1 (On-line Mendelian Inheritance in Man [MIM] #135700) is caused by heterozygous mutation in KIF21A (MIM *608283), which codes a developmental motor kinesin responsible for anterograde axonal transport of cargo along neurons such as that of the superior division of cranial nerve III. The CFEOM1 phenotype is characterized by bilateral ptotic hypotropia, an inability to supraduct above the horizontal midline, exotropia, and a variable degree of ophthalmoplegia with abnormal synkinesis. CFEOM2 (MIM #602078) is caused by homozygous mutation in PHOX2A (MIM *602753), a hindbrain transcription factor responsible for development of cranial nerves 3 and 4. The CFEOM2 phenotype is characterized by bilateral large-angle exotropia, ptosis, miosis, and a variable degree of ophthalmoplegia with abnormal synkineis. Commoner forms of isolated congenital incomitant strabismus (eg, Brown syndrome, superior oblique palsy, monocular elevation deficiency) are usually not familial but could also be related to abnormal ocular cranial nerve innervation. To assess this hypothesis, following institutional review board approval and informed consent we performed KIF21A and PHOX2A analysis over a 2-year period on 16 consecutive Saudi patients with common forms of congenital incomitant vertical strabismus. Only patients with unequivocal clinical diagnoses and without additional medical problems were recruited. Patients with classic clinical CFEOM1 and CFEOM2 were excluded, as were patients with incomitant strabismus that could be attributed to trauma or related to abnormal development of cranial nerve VI (eg, Duane syndrome, Moebius syndrome). All patients were from the referral pediatric ophthalmology practice of one of the authors (AOK), who performed a comprehensive ophthalmic examination for each patient. Venous blood samples were collected; polymerase chain reaction (PCR) was used to amplify the opening reading frame and exons for KIF21A (38 exons) and PHOX2A (3 exons) [primers and conditions available upon request].² Ensemble reference sequences for KIF21A and PHOX2A were ENSF00000000538 and ENSG0000165462, respectively.

For the 16 patients there were 6 diagnoses (5 Brown syndrome, 5 superior oblique weakness, 2 third nerve weakness, 2 monocular elevation deficiency, one “A” pattern, and one “Y” pattern). All patients had strabismus noted at birth, and no patient had a family history of strabismus. Clinical features are summarized in Table 1. No pathogenic mutations in KIF21A or PHOX2A were found in any of the patients. Only one variant was identified, which was heterozygous and did not alter the predicted amino acid sequence (c. 4771 C>T p.L1591 in KIF21A in patient #3).

TABLE 1  Clinical summary of the 16 patients

ID	Age	Gender	Dx	Laterality	AHP	Forced Ductions	Comments
1	11y	M	Brown	OU	Mod Chinup	NA
2	8y	F	Brown	OD	Mod L face turn	NA
3	10y	M	Brown	OD	None	NA
4	8y	F	Brown	OS	Mild R face turn	NA
5	5y	M	Brown	OD	Mild L face turn	NA
6	9y	F	SOP	OS	none	lax SO OS
7	4y	F	SOP	OS	Large R tilt	lax SO OS	Anomalous SO insertion OS
8	6y	F	SOP	OS	Large R tilt	lax SO OS
9	25y	F	SOP	OS	Large R tilt	lax SO OS
10	5y	M	SOP	OU	Mod Chin-up	lax SO OU	V -ET
11	5y	F	3^rd	OS	none	NA	pupil not involved; multiple MRIs normal
12	4y	F	3^rd	OS	none	NA	pupil not involved; multiple MRIs normal
13	8m	F	MED	OD	Large chin up	normal OU
14	7y	M	MED	OD	Large chin up	Mild RIR tightness
15	12y	F	A-ET	OU	None	tight SO OU	Fundus incyclorison OU
16	8y	M	Y-XT	OU	None	NA	XT only on upgaze

ID: identification number; Dx: diagnosis; AHP: Anomalous head posture; y: years; m: months; M: male; F: female; SOP: superior oblique weakness; 3rd: 3rd nerve weakness; MED: monocular elevation deficiency; OU: both eyes; OD: right eye; OS: left eye; Mod: moderate; R: right; L: left; NA: not available; IR: inferior rectus; V-ET: V-pattern esotropia; Y–XT: Y pattern exotropia.

Prior published studies of KIF21A and PHOX2A in relevant forms of congenital incomitant strabismus were in patients with superior oblique palsy and Brown syndrome and did not clearly demonstrate an association^2–5. Jiang et al.^3,4 suggested that PHOX2A polymorphism was associated with congenital superior oblique palsy; 11 out of 31 patients had 1 of 5 nucleotide variations in exon 1 and/or the promoter of PHOX2A that were not found in 54 normal controls. Imai et al.⁵ suggested that PHOX2A 153G>A predisposed to congenital superior oblique palsy based on analysis of 6 patients from 3 families; however, the PHOX2A 153G>A variant was also found in normal controls. In the same study Imai et al.⁵ assessed exons 8, 20, and 21 of KIF21A for mutations; although they did not find sequence variation in these exons, the other exons of KIF21A were not assessed. In a study of 3 siblings with late-onset Brown syndrome, Iannaccone et al.² found no abnormalities in PHOX2A.

Our 16-patient sample suggests that KIF21A and PHOX2A sequence variation does not have a role in common forms of congenital incomitant vertical strabismus.