Nonsyndromic Holoprosencephaly via the CRIPTO/TDGF1 Gene

Nonsyndromic holoprosencephaly (HPE) is a structural anomaly of the brain in humans resulting from incomplete cleavage of the prosencephalon (forebrain) into right and left hemispheres between the third to fourth weeks of gestation (Martinez et al. 2018. PubMed ID: 29761634). Nonsyndromic HPE can be divided into four types according to the degree of brain division. Alobar HPE is the most severe form, with a single monoventricle and no interhemispheric fissure. Partial separation of the lobes is observed in semilobar HPE, with the interhemispheric fissure only present posteriorly. In lobar HPE, the right and left ventricles are separated, but incomplete separation of the frontal lobes occurs. Middle interhemispheric fusion variant (MIHF/MIHV or syntelencephaly) is a milder subtype, characterized by abnormal midline union of the posterior frontal and parietal lobes with variable fusion of thalami (Dubourg et al. 2007. PubMed ID: 17274816; Tekendo-Ngongang et al. 2000. PubMed ID: 20301702). In addition to the structural brain abnormality, patients with nonsyndromic HPE may have a distinctive pattern of facial features. The most severe affected individuals often have one central eye (cyclopia) and a tubular nasal structure (proboscis) located above the eye. In the less severe HPE forms, a small head (microcephaly), abnormally small eyes (microphthalmia), absent eyes (anophthalmia), a close set of eyes (hypotelorism), and cleft lip or palate may be observed (Solomon et al. 2010. PubMed ID: 20104594).

Nonsyndromic HPE accounts for approximately 25 to 50 percent of all cases of HPE, which affects an estimated 1 in 10,000 newborns and 1 in 250 fetuses (Martinez et al. 2018. PubMed ID: 29761634). Nonsyndromic HPE is inherited in an autosomal dominant pattern with incomplete penetrance. Therefore, the phenotype of affected individuals is extremely variable, ranging from alobar HPE with cyclopia to clinically normal (Solomon et al. 2010. PubMed ID: 19955556; Solomon et al. 2010. PubMed ID: 20104608; Dubourg et al. 2018. PubMed ID: 29785796). The most severely affected newborns usually do not survive beyond the first week of life, while a significant proportion of more mildly affected individuals survive the first year of life. Some individuals with various HPE subtypes, including severe subtypes, can survive until adulthood (Croen et al. 1996. PubMed ID: 8862623; Levey et al. 2010. PubMed ID: 20104615; Weiss et al. 2018. PubMed ID: 30182446). Variants in CRIPTO/TDGF1 are also associated with tumorigenesis and congenital heart defects (Wang et al. 2011. PubMed ID: 19853938; Geng et al. 2014. PubMed ID: 25516202).

Currently, there are no treatments for nonsyndromic HPE. However, genetic testing may provide advantages, including prognostic information, such as reported genotype-phenotype correlations, which assist in predicting the expected severity of the disorder for a given patient. Early identification and treatment of symptoms such as seizures and feeding difficulties and prenatal testing or pre-implantation genetic diagnosis for future pregnancies may also be possible. Knowledge of a de novo variant may decrease anxiety for future reproductive planning due to reduced recurrence risk.

Nonsyndromic HPE is inherited as an autosomal dominant manner with incomplete penetrance and intrafamilial variable expression. Fourteen genes have been associated with nonsyndromic HPE: CDON, CRIPTO, DISP1, DLL1, FGF8, FOXH1, GAS1, GLI2, NODAL, PTCH1, SHH, SIX3, TGIF1, and ZIC2. Of those, SHH, SIX3, ZIC2, and TGIF1 are common monogenic causes, representing approximately 24% of all cases with known etiology. Pathogenic variants in the SHH gene are the most common cause of nonsyndromic HPE; the frequency of pathogenic variants in the other genes, including CRIPTO, are unknown or rare (Tekendo-Ngongang et al. 2020. PubMed ID: 20301702; Martinez et al. 2018. PubMed ID: 29761634).

CRIPTO (crypto growth factor (CRGF)) is part of the EGF-CFC gene family that encodes a highly conserved epidermal growth factor-like domain (EGF) and a second cysteine-rich region called the CFC domain. Recent findings indicate that members of this gene family act as essential cofactors for Nodal, a member of the transforming growth factor β (TGF-β) family. They also play a crucial role in regional and cellular differentiation during embryonic development, especially in the specification of the endomesoderm, whose derivatives participate in the division of the eye field and brain (Minchiotti et al. 2002. PubMed ID: 11992720; de la Cruz et al. 2002. PubMed ID: 12073012).

CRIPTO is relatively tolerant to loss of function variants (Genome Aggregation Database). Gene function studies in zebrafish and mouse models indicate a role for CRIPTO in human midline and forebrain development (Minchiotti et al. 2002. PubMed ID: 11992720; de la Cruz et al. 2002. PubMed ID: 12073012; Shen and Schier. 2000. PubMed ID: 10858660). Genetic analysis of this gene is complicated by its pseudogenes found on chromosomes 2, 3, 6, 19, and X (Dono et al. 1991. PubMed ID: 1882841; Scognamiglio et al. 1999. PubMed ID: 10393436). Overall, CRIPTO has been cited as a non-essential gene for growth of human tissue culture cells (Online Gene Essentiality).

The clinical sensitivity in CRIPTO is expected to be very low since variants in CRIPTO have rarely been reported in individuals with nonsyndromic HPE (Tekendo-Ngongang et al. 2020. PubMed ID: 20301702).

This test is performed using Next-Gen sequencing with additional Sanger sequencing as necessary.

This test provides full coverage of all coding exons of the CRIPTO gene plus 10 bases of flanking noncoding DNA in all available transcripts along with other non-coding regions in which pathogenic variants have been identified at PreventionGenetics or reported elsewhere. We define coverage as ≥20X NGS reads or Sanger reads.

Dependent on the sequencing backbone selected for this testing, discounted reflex testing to any other similar backbone-based test is available (i.e., PGxome panel to whole PGxome; PGnome panel to whole PGnome).