TNXB-Related Disorders via the TNXB Gene, Exons 32-44

Ehlers-Danlos syndrome (EDS), due to complete tenascin-X deficiency, is an autosomal recessive disorder of connective tissues characterized by hyperextensible skin, hypermobile joints, and tissue fragility (Burch et al. 1997. PubMed ID: 9288108; Schalkwijk et al. 2001. PubMed ID: 11642233; Lindor et al. 2005. PubMed ID: 15793839; Bristow et al. 2005. PubMed ID: 16278880; Demirdas et al. 2017. PubMed ID: 27582382). The major clinical features of this condition resemble the classic type of Ehlers-Danlos syndrome, except for lack of poor wound healing and atrophic scarring. Large joint dislocations are the most frequent debilitating finding, and easy bruising is a prominent feature. Some documented patients were also affected by congenital adrenal hyperplasia due to 21-hydroxylase deficiency because these patients were homozygous for a contiguous 30kb deletion disrupting both the CYP21A2 and TNXB genes. Patients also have features that are not typically found in Ehlers-Danlos syndrome, such as spina bifida occulta, mitral valve prolapse, gastrointestinal bleeding, severe diverticular intestinal disease with ruptured diverticula, pancolonic diverticulitis, rectal prolapse, premature arteriosclerosis, stroke, and obstructive airway disease (Schalkwijk et al. 2001. PubMed ID: 11642233; Lindor et al. 2005. PubMed ID: 15793839). These patients are rare in the literature, and the exact incidence is unknown. Symptoms could be apparent during childhood while age of onset varies into adulthood.

Heterozygous TNXB deficiency has also been associated with hypermobility type of Ehlers-Danlos syndrome, suggesting an autosomal dominant inheritance. It is characterized by joint hypermobility often in association with joint subluxations and chronic musculoskeletal pain (Levy. 2013. PubMed ID: 20301456; Zweers et al. 2003. PubMed ID: 12865992; Merke et al. 2013. PubMed ID: 23284009). Compared with other types of Ehlers-Danlos syndrome, the hypermobility type is more common, but much less severe. Its clinical presentations are extremely variable, and its molecular etiology is largely unknown. Tenascin-X haploinsufficiency has been reported to explain the causes in some patients (Levy. 2013. PubMed ID: 20301456; Zweers et al. 2003. PubMed ID: 12865992; Zweers et al. 2005. PubMed ID: 15733269). Of note, the contiguous 30kb deletion disrupting both CYP21A2 and TNXB genes is common in patients with congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency, and tenascin-X haploinsufficiency has been associated with Ehlers-Danlos syndrome features in those patients who are heterozygous for this contiguous deletion (Chen et al. 2009. PubMed ID: 19921645; Merke et al. 2013. PubMed ID: 23284009; Morissette et al. 2015. PubMed ID: 26075496; Chen et al. 2016. PubMed ID: 27297501). In addition, cardiac abnormalities such as quadricuspid aortic valve have been reported in this group of CAH patients. Average age of diagnosis is 20-30 years old (Miller and Merke. 2018. PubMed ID: 29734195).

In addition, rare heterozygous TNXB variants have been implicated in hereditary vesicoureteral reflux (VUR) in association with joint hypermobility (Gbadegesin et al. 2013. PubMed ID: 23620400; Elahi et al. 2015. PubMed ID: 26408188). VUR is the retrograde flow of urine from the bladder to the ureter and the kidney, representing the most common type of congenital anomaly of the kidney and the urinary tract (CAKUT).

Defects in TNXB were originally found to cause EDS in an autosomal recessive manner (Burch et al. 1997. PubMed ID: 9288108; Schalkwijk et al. 2001. PubMed ID: 11642233). Later, heterozygous pathogenic variants in TNXB were implicated in hypermobility type EDS, suggesting an autosomal dominant inheritance (Zweers et al. 2003. PubMed ID: 12865992; Levy. 2013. PubMed ID: 20301456).

Tenascin-X encoded by TNXB is an extracellular matrix protein acting as an important regulator of collagen deposition in vivo (Bristow et al. 2005. PubMed ID: 16278880; Mao et al. 2002. PubMed ID: 11925569). Defects in TNXB have been implicated in autosomal dominant or recessive type of Ehlers-Danlos syndrome as well as hereditary vesicoureteral reflux (VUR) (Schalkwijk et al. 2001. PubMed ID: 11642233; Zweers et al. 2003. PubMed ID: 12865992; Gbadegesin et al. 2013. PubMed ID: 23620400; Elahi et al. 2015. PubMed ID: 26408188).

TNXB is one of the constitutional genes of the RCCX module (RP-C4-CYP21-TNX) in the human leukocyte antigen histocompatibility complex on chromosome 6. The RCCX module represents a complicated genomic region characterized by frequent homologous recombination events due to existence of highly homologous pseudogenes in tandem (Lee. 2005. PubMed ID: 15639189; Chen et al. 2012. PubMed ID: 22156666). The pseudogene TNXA is a partially duplicated segment that corresponds to intron 31 to exon 44 of TNXB. A contiguous 30kb deletion disrupting both CYP21A2 and TNXB genes, resulting in a nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35, is commonly found in patients (Schalkwijk et al. 2001. PubMed ID: 11642233; Koppens et al. 2002. PubMed ID: 12354783; Lindor et al. 2005. PubMed ID: 15793839; Merke et al. 2013. PubMed ID: 23284009). In addition, two types of chimera TNXA/TNXB genes (with different junction sites) that are not involving the 120 bp deletion have been reported in CAH patients with EDS phenotype (Morissette et al. 2015. PubMed ID: 26075496; Chen et al. 2016. PubMed ID: 27297501). Other documented genetic defects in TNXB are missense substitutions and truncating variants (small indels and splicing variants) (Human Gene Mutation Database). Pathogenic exon-level copy number changes have not been reported within the unique portion (exons 1 to 31) of TNXB.

De novo variants are rare in this gene. Both missense and loss-of-function variants can be pathogenic. TNXB has been cited as a nonessential gene for growth of human tissue culture cells (Online Gene Essentiality, ogee.medgenius.info)

In a group of 151 patients with the classic, hypermobility, or vascular types of the Ehlers-Danlos syndrome, five patients (3.3%) were found to have the common contiguous 30kb deletion allele (i.e. nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35) or truncating variants (Schalkwijk et al. 2001. PubMed ID: 11642233).

In a cohort of 80 Dutch patients with hypermobility type Ehlers-Danlos syndrome, two patients (2.5%) were found to have a common contiguous 30kb deletion allele (nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35) and a truncating 2bp deletion, respectively (Zweers et al. 2003. PubMed ID: 12865992).

In a study of 192 patients with congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency, the common contiguous 30kb deletion allele (nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35) was found in a heterozygous state in 12 (6.25%) of these CAH patients (Merke et al. 2013. PubMed ID: 23284009). In addition, a heterozygous frameshift variant (c.6903insC) was also found in a patient (0.5%).

In a study of 246 unrelated CAH patients screened for TNXB defects, 14 CAH probands carried the TNXA/TNXB chimera involving the 120bp deletion (TNXA/TNXB CH-1) and 7 unrelated patients carried novel chimeras (TNXA/TNXB CH-2), leading to a CAH-X prevalence of 8.5% (Morissette et al. 2015. PubMed ID: 26075496).

In a study of 110 families with primary vesicoureteral reflux (PVUR), rare missense variants in TNXB were found in 5/55 (9%) of families with familial PVUR (FPVUR) and 2/55 (4%) of non-familial PVUR (NFPVUR) (Elahi et al. 2015. PubMed ID: 26408188).

This test is performed using Sanger sequencing.

Due to the homologous sequence at the region of intron 31 to exon 44, we utilize a long-range PCR strategy to specifically amplify and sequence these exons, which is able to overcome the limitation of current short-read NGS technology in dealing with this type of paralogous sequence (Mandelker et al. 2016. PubMed ID: 27228465).

Our test strategy is able to detect those TNXA/TNXB chimera alleles documented in the literature (Merke et al. 2013. PubMed ID: 23284009; Morissette et al. 2015. PubMed ID: 26075496; Chen et al. 2016. PubMed ID: 27297501).