DCC mutation update: Congenital mirror movements, isolated agenesis of the corpus callosum, and developmental split brain syndrome

The deleted in colorectal cancer (DCC) gene encodes the netrin‐1 (NTN1) receptor DCC, a transmembrane protein required for the guidance of commissural axons. Germline DCC mutations disrupt the development of predominantly commissural tracts in the central nervous system (CNS) and cause a spectrum of neurological disorders. Monoallelic, missense, and predicted loss‐of‐function DCC mutations cause congenital mirror movements, isolated agenesis of the corpus callosum (ACC), or both. Biallelic, predicted loss‐of‐function DCC mutations cause developmental split brain syndrome (DSBS). Although the underlying molecular mechanisms leading to disease remain poorly understood, they are thought to stem from reduced or perturbed NTN1 signaling. Here, we review the 26 reported DCC mutations associated with abnormal CNS development in humans, including 14 missense and 12 predicted loss‐of‐function mutations, and discuss their associated clinical characteristics and diagnostic features. We provide an update on the observed genotype–phenotype relationships of congenital mirror movements, isolated ACC and DSBS, and correlate this to our current understanding of the biological function of DCC in the development of the CNS. All mutations and their associated phenotypes were deposited into a locus‐specific LOVD (https://databases.lovd.nl/shared/genes/DCC).

The development of commissural tracts is dependent on neurons forming synaptic connections with their target cells located on the opposite side of the CNS. To achieve this, neurons extend commissural axons tipped with a specialized, motile sensing device called a growth cone (Tessier-Lavigne & Goodman, 1996). Growth cones express an array of axon guidance receptors such as DCC. The binding of a chemotactic (diffusible) or haptotactic (substrate-bound) guidance cue to an axon guidance receptor generates a permissive or non-permissive axonal outgrowth signal, or an attractive or repulsive directional response (Raper & Mason, 2010;Tessier-Lavigne & Goodman, 1996).
The signals transduced by axon guidance receptors converge to modulate the assembly of growth cone filamentous actin and cytoskeletal microtubules required for directing axon outgrowth and guidance toward or away from a guiding cell population (Kahn & Baas, 2016;Luo, 2002).

VARIANT DATABASE
We created a new repository for all reported disease-associated DCC sequence variants using the Leiden Open Variation Database (LOVD 3.0: https://databases.lovd.nl/shared/genes/DCC) (Fokkema et al., 2011). The International Research Consortium for the Corpus Callosum and Cerebral Connectivity (IRC 5 , https://www.irc5.org) encourages users to register and submit data, including clinical and neuroimaging phenotypic data.

VARIANT NOMENCLATURE
The nomenclature for DNA and protein sequence variants adheres to the guidelines of the Human Genome Variation Society (den Dunnen & Antonarakis, 2000). Variant descriptions were based on the following Genbank reference sequences: NG_013341.2, NM_005215.3, and NP_005206.2. We utilized the Mutalyzer program

VARIANTS
To date, 26 unique DCC mutations have been associated with MMs, iACC, and DSBS in 66 individuals from 25 unrelated families ( Figure 1).
There are 29 reported unaffected heterozygous mutation carriers; 11 confirmed carriers from seven unrelated families assessed for MMs and brain abnormalities using magnetic resonance imaging (MRI) or computed tomography (CT), and 18 carriers from nine unrelated families that appear clinically unaffected but have not been assessed for MMs and brain abnormalities. There are no reported unaffected individuals with biallelic, predicted LoF DCC mutations.
Monoallelic or biallelic missense (14 out of 24; 58%) and monoallelic, predicted LoF (10 out of 24; 42%) DCC mutations may cause MMs, iACC, or both (Table 1). These include one case each where two heterozygous missense mutations were inherited in cis p.(Met1217Val;p.Ala1250Thr) or in trans p.(Gly470Asp);(p.(Gly803Arg) and a biallelic p.(Gln691Lys) mutation observed in conjunction with an interhemispheric cyst and modest learning difficulties (Jamuar et al., 2017;Marsh et al., 2017;Méneret et al., 2014a). Alternatively, biallelic, predicted LoF (two out of two; 100%) DCC mutations cause DSBS. Germline transmission of the mutant allele, except for a p.(Gly803Arg) mutation that arose de novo, was confirmed in all pedigrees where parental genotypes were available (Méneret et al., 2014a). DCC-MMs and DCC-iACC are both associated with reduced penetrance: MMs ≈42% penetrance and iACC ≈26% penetrance (Marsh et al., 2017). In contrast, DSBS appears to be fully penetrant. Monoallelic DCC mutations are also associated with variable expressivity and affected individuals within one family may present with MMs, iACC, or both (Marsh et al., 2017). All DCC residues altered by a missense mutation are conserved throughout vertebrate evolution, although it should be noted that Dcc has been lost in passeriformes and galliformes during bird evolution, and are either novel (eight out of 14; 57%) or found at a minor allele frequency of less than 0.5% (six out of 14; 43%) in the Exome Aggregation Consortium (ExAC) (Supp . Table S1) (Friocourt et al., 2017). Predicted LoF DCC mutations include nine frameshift mutations, two truncating mutations and an intragenic deletion encompassing exons 4 and 5 (Table 1).    Figure S1) (Kolodziej et al., 1996). Each region of DCC appears to have a unique functional role required for the transduction of NTN1 signaling.
This extracellular binding brings two DCC proteins into close proximity, enabling their intracellular domains to homodimerize and initiate the recruitment of multi-protein complexes required to transduce NTN1 signaling (Mille et al., 2009;Stein, Zou, Poo, & Tessier-Lavigne, 2001).
The DCC ligand, NTN1, is an extracellular protein belonging to the laminin superfamily and is composed of three structurally distinct regions: a laminin-like domain (LN), three laminin-type epidermal growth factor-like domains (LE) and a netrin-like domain (NTR) . NTN1 has three distinct DCC-binding sites (binding site 1, 2, and 0; BS1, BS2, and BS0) ( Figure 2). BS1 is located on the third LE of NTN1 and interacts exclusively with the 5th FN3 domain of DCC. This site functions as a DCC-specific binding site and is stabilized by several hydrophobic interactions supported by a surrounding network of hydrogen bonds ( Figure 3A) (Finci et al., 2014). BS2 is located on the first and second LE domains of NTN1 and interacts with a distinct region of the 5th FN3 domain as well as the N-terminal region of the 6th FN3 domain of DCC. This site functions as a generic binding site that can interact with receptors other than DCC (Finci et al., 2014). BS2 is predominantly stabilized by a group of sulfate and chloride anions that neutralize positively charged patches on NTN1 and DCC residues located at the binding interface ( Figure 3B) (Finci et al., 2014). The adaptable nature of BS2 allows binding to the 1st and 2nd Ig-like domains of members of the unc-5 netrin receptor (UNC5; MIM# 603610) family of repulsive axon guidance proteins, which appear to have a higher affinity for BS2 relative to DCC (Finci et al., 2014;Geisbrecht et al., 2003;Hong et al., 1999;Leonardo et al., 1997). As a result, UNC5 are able to outcompete DCC at BS2 and switch an attractive growth cone response (mediated via NTN1induced DCC homodimerization) to a repulsive response (mediated via NTN1-induced DCC and UNC5 heterodimerization) (Finci et al., 2014;Grandin et al., 2016;Hong et al., 1999). In contrast to BS1 and BS2, BS0 is located on the LN of NTN1 and interacts with the 4th FN3 domain of DCC (Xu et al., 2014). This NTN1 protein is different to the one simultaneously engaged at BS1 and BS2, but binds the same DCC protein occupied at BS1 ( Figure 3C). The binding of DCC by a second NTN1 protein at BS0 is thought to generate a signaling cluster important for the transduction of a directional response toward or away from a guiding cell population (Finci, Zhang, Meijers, & Wang, 2015).
Taken together, these studies indicate that NTN1 forms two distinct binding clusters through BS0, BS1, and BS2 and that these clusters underpin its bifunctionality as an attractive and repulsive guidance cue. The first cluster involves the 4th and 5th FN3 domains of one DCC protein at BS0 and BS1, while the second cluster involves the 5th and 6th FN3 domains of a second DCC protein or the 1st and 2nd Ig-like domains of an UNC5 family member protein at BS2.
However, it should be noted that the actual in vivo binding structure may differ as these studies utilized a recombinant NTN1 protein lacking its NTR domain and different truncated DCC fragments, both lacking the NTN1 binding region in its entirety (Finci et al., 2014;Xu et al., 2014).
The importance of the 4th, 5th, and 6th FN3 domains are highlighted by the significant enrichment of DCC missense mutations linked to MMs and/or iACC within these NTN1 binding regions compared with missense variants located in these domains in the ExAC ( Figure 4A and B) (Marsh et al., 2017 (Li et al., 2002;Li et al., 2004;Liu et al., 2004;Meriane et al., 2004;Ren et al., 2004;Ren et al., 2008;

GENOTYPE-PHENOTYPE CORRELATION
Monoallelic frameshift and nonsense DCC mutations are predicted to result in haploinsufficiency via nonsense mediated mRNA decay or rapid turnover of the truncated protein, resulting in reduced NTN1 binding (Srour et al., 2010). This is in contrast to DCC missense mutations that likely generate a full-length protein within the cell (Finci et al., 2014;Meriane et al., 2004 Monoallelic, missense mutations located within the NTN1 binding sites of DCC appear to be strongly associated with iACC (Marsh et al., 2017). Indeed, the majority of DCC mutations located within the 4th and 5th FN3 domains are associated with iACC (five out of six; 83%).
Interestingly, the p.(Val793Gly) and p.(Gly805Glu) missense mutations located directly within the NTN1 binding interface are associated with a more severe phenotype that features both iACC and MMs (Marsh et al., 2017). Previous studies have shown a differential disruption to NTN1-induced axon attraction and repulsion dependent on the location of the mutated DCC residue within the NTN1 binding interface ( Figure 4C) (Finci et al., 2014). Consequently, DCC mutations located within the NTN1 binding interface may cause more severe disruptions to commissural axon outgrowth and guidance that result in a phenotype that features both iACC and MMs. Overall, it appears that the majority of missense iACC mutations localize to the FN3 domains (seven out of nine; 78%), highlighting the importance of these domains to DCC function and callosal development.
Sex differences in callosal morphology have previously been linked with testosterone levels during prenatal brain development (Chura et al., 2010;Moffat, Hampson, Wickett, Vernon, & Lee, 1997). Consistent with these observations, RNAseq and RT-qPCR analyses have detected a significant dose-dependent increase in DCC expression in testosterone-treated neural stem cells derived from human embryonic stem cells (Marsh et al., 2017). These findings suggest that iACC may occur when the expression of DCC falls below a threshold level during CC development, as would occur more commonly in females. However, given the incomplete penetrance observed in both sexes, the pheno-type must also be influenced by additional genetic, epigenetic, and/or environment factors (Marsh et al., 2017). Furthermore, the sex-specific nature of this phenotype imbalance remains to be verified as reciprocal analyses utilizing estrogens have not been reported. Additional in vivo investigations are required to evaluate the potential impact the hormonal context during brain development has on DCC expression and whether it influences the expressivity of the MMs and iACC phenotype.

BIOLOGICAL SIGNIFICANCE
Commissural axons form connections between the left and right sides of the brain that are required for the transfer and integration of information generated by sensory, motor, and associative neurons. The tightly regulated, spatiotemporal expression of axon guidance receptors and their ligands control the development of tracts such as the CC and CST (Chedotal & Richards, 2010). As highlighted below, our understanding of the development of these tracts and the associated role of DCC and NTN1 signaling in humans has been predominantly informed by the study of these biological processes in mice.
The formation of the CC is complex and dependent on the execution of several developmental steps. Briefly, these include midline patterning and remodeling of the interhemispheric fissure by astroglia to create a permissible substrate for commissural axons to cross the midline (Gobius et al., 2016;Hayhurst, Gore, Tessier-Lavigne, & McConnell, 2008;Okada, et al., 2008;Silver, Lorenz, Wahlsten, & Coughlin, 1982); secretion of guidance cues by populations of glia and neurons situated around the midline (Shu & Richards, 2001;Shu, Li, Keller, & Richards, 2003;Unni et al., 2012); pioneering of the callosal tract by axons extending from neurons in the cingulate cortex (Rash & Richards, 2001); and fasciculation of callosal axons with pioneer axons as they grow toward and across the midline (Koester & O'Leary, 1994;Rash & Richards, 2001). Dcc and Ntn1 are required for the normal development of the CC and their loss or dysfunction in mice is associated with complete ACC (Fazeli et al., 1997;Finger et al., 2002;Serafini et al., 1996). While both pre-crossing pioneer and callosal axons express Dcc, only the former axonal population appears to be attracted toward the midline by Ntn1 (Fothergill et al., 2014;Shu et al., 2000). Instead, it has been shown that pre-crossing callosal axons utilize Dcc and Ntn1 signaling to attenuate the repulsive signaling of Robo1 and Slit2 (another axon guidance receptor-ligand pair) to approach and cross the midline (Fothergill et al., 2014;. The expression of Dcc in post-crossing callosal axons is subsequently downregulated, thereby restoring the repulsive effect of Robo1 and Slit2 signaling to direct axons away from the midline and toward their targets in the contralateral cerebral hemisphere (Fothergill et al., 2014;Shu et al., 2000).
The subcerebrally projecting neurons that comprise the CST mainly originate from the primary motor and premotor areas of the cortex and follow a stereotyped route to innervate their targets in the spinal cord (Dum & Strick, 1991). During development, axons from these projection neurons converge and descend through the internal capsule and cerebral peduncles of the midbrain before entering the ventral brainstem where they form the medullary pyramids. Most CST projections then cross the midline in the caudal region of the medulla to form the pyramidal decussation, before projecting inferiorly in the spinal cord to synapse with lower motor neurons or interneurons in the ventral spinal cord to facilitate voluntary movement of the limbs (Welniarz et al., 2017a). Dcc is required for the normal development of the CST in mice and Dcc dysfunction is associated with a failure of the CST to cross the midline at the level of the pyramidal decussation (Finger et al., 2002;Welniarz et al., 2017b). However, Dcc does not appear to be expressed in the brainstem CST of mice as it is significantly downregulated in the distal portion of the axon that grows beyond the internal capsule (Finger et al., 2002;Shu et al., 2000). Interestingly, the role of Dcc in the development of the CST at the midline was reported to be non-cell autonomous as conditional knockout of Dcc in the cortex (and therefore the CST) caused ACC but not a failure of the CST to cross the midline (Welniarz et al., 2017b).
In the brainstem and spinal cord, Dcc is present on commissural axons, which are guided circumferentially from the dorsal roof plate toward the ventral floor plate (where midline crossing occurs) in response to Ntn1 (Dominici et al., 2017;Holley, 1982;Tessier-Lavigne, Placzek, Lumsden, Dodd, & Jessell, 1988;Yee, Simon, Tessier-Lavigne, & O'Leary, 1999). The attractive response of these commissural axons to Ntn1 is regulated by Dcc in partnership with Robo3, another axon guidance receptor required for normal development of the hindbrain and spinal cord (Chen, Gore, Long, Ma, & Tessier-Lavigne, 2008;Marillat et al., 2004;Sabatier et al., 2004;Zelina et al., 2014). Pioneering studies utilizing embryonic chick spinal cord demonstrated that this axonal navigation is reliant on a ventral-dorsal gradient of Ntn1 diffused from the floor plate .
Subsequent studies in mice also detected a similar graded distribution of Ntn1 extending from the floor plate, supporting the observation that Ntn1 functions as a long-range diffusible chemoattractant (Kennedy, Wang, Marshall, & Tessier-Lavigne, 2006;Serafini et al., 1996). However, recent reports have revised this model by demonstrating that Ntn1 produced by neural progenitors in the ventral ventricular zone of hindbrain and spinal cord neuroepithelium are essential for commissural axon extension toward the ventral midline in embryonic mice (Dominici et al., 2017;Yamauchi et al., 2017).
These reports propose that, in mice, commissural axons are directed via Dcc-dependent haptotaxis toward the ventral midline by Ntn1 produced and deposited at the pial surface of the hindbrain and spinal cord, not via a diffusible gradient of Ntn1 emanating from the floor plate (Dominici et al., 2017;Yamauchi et al., 2017). Therefore, regardless of how Ntn1 is presented to axons, it is clear that it functions to direct Dccexpressing commissural axons toward the ventral midline of the hindbrain and spinal cord during development.

CLINICAL RELEVANCE
MMs are involuntary movements on one side of the body that mirror voluntary movements made on the opposite side (Cincotta & Ziemann, 2008). DCC-MMs commonly present in infancy or early childhood and persist stably into adulthood (Depienne et al., 2011;Franz et al., 2015;Marsh et al., 2017;Méneret et al., 2014a;Srour et al., 2009;Srour et al., 2010). DCC-MMs manifest in the fingers and hands but may also be present in the forearms, toes, and feet in a subset of affected individuals (Supp . Table S2) (Marsh et al., 2017;Méneret et al., 2014a;Srour et al., 2010). With effort, some affected individuals can partly suppress these involuntary movements (Srour et al., 2009). Individuals with DCC-MMs exhibit a range of functional disabilities besides difficulties in fine bimanual activities. These include fatigue, spontaneous muscle contractions, and pain in the upper limbs during extended manual activities such as writing, as well as general clumsiness and compensatory maneuvers to inhibit involuntary movements (Depienne et al., 2011;Marsh et al., 2017;Méneret et al., 2014a;Meneret, Trouillard, Brochard, & Roze, 2015). As a result, DCC-MMs may preclude affected individuals from professions and social activities that demand sustained or complex bimanual coordination. Individuals with DCC-MMs do not appear to exhibit any additional clinical manifestations and have a normal developmental outcome (Depienne et al., 2011;Franz et al., 2015;Marsh et al., 2017;Méneret et al., 2014a;Meneret, et al., 2015;Sharafaddinzadeh et al., 2008;Srour et al., 2009;Srour et al., 2010;Welniarz et al., 2017b). However, only a subset of individuals diagnosed with DCC-MMs are reported to have undergone brain imaging (12 out of 42; 29%) and formal neuropsychological evaluation (two out 42; 5%) and therefore the phenotypic spectrum of DCC-MMs remains to be defined (Marsh et al., 2017). Individuals with monoallelic, missense DCC mutations may also present with iACC, with or without MMs (Marsh et al., 2017). ACC describes the partial or complete absence of the CC and is characterized by the failure of callosal axons to cross the midline. Apart from the abnormalities typically expected to be associated with ACC, such as the absence of the hippocampal commissure and cingulate gyrus and dysmorphic lateral ventricles or colpocephaly, individuals with iACC do not present with additional brain abnormalities (Cesaretti et al., 2016;Kuker, Mayrhofer, Mader, Nagele, & Krageloh-Mann, 2003).
Approximately half of these individuals also present with MMs (nine out of 19; 47%), which suggests that DCC-iACC may frequently present as part of a global disorder of midline crossing (Marsh et al., 2017;Parrish, Roessmann, & Levinsohn, 1979;Paul et al., 2007). Other callosal abnormalities such as hypoplasia (a uniformly thin CC) and/or dysplasia (abnormal CC shape) have not been observed in individuals with DCC-iACC (Marsh et al., 2017). Individuals with DCC-iACC show no consistent additional gross brain abnormalities in the posterior commissure, ventricular system, cerebral cortex, white matter, hippocampi, brainstem, basal ganglia, cerebral and cerebellar peduncles, cerebellar vermis and hemispheres, optic chiasm, and pituitary gland ( Figure 5) (Marsh et al., 2017). However, the anterior commissure of some DCC-iACC individuals may be enlarged (Marsh et al., 2017). Enlargement of the anterior commissure is also observed in a minority of individuals with ACC and has been suggested to represent a compensatory mechanism to maintain connectivity between the cerebral hemispheres (Barr and Corballis, 2002;Hetts, Sherr, Chao, Gobuty, & Barkovich, 2006 with DCC-iACC do commonly exhibit typical neurobehavioral consequences associated with ACC, such as impairments in emotional and social functioning in addition to attention, language, visuospatial, and literacy and numeracy deficits (Marsh et al., 2017;Paul et al., 2007;Sotiriadis & Makrydimas, 2012).
Biallelic, DCC mutations leading to predicted LoF are associated with DSBS, a complex syndrome associated with a broad disorganization of white-matter tracts throughout the CNS (Jamuar et al., 2017).
The features of DSBS include absence of all commissures (including the CC, anterior, and posterior), brainstem defects (including hypoplasia of the pons and midbrain), horizontal gaze palsy and progressive scoliosis with a variable age of onset ( Figure 5).
Several features of DSBS overlap with HGPPS1 (MIM# 607313), a congenital syndrome caused by biallelic mutations in the axon guidance receptor ROBO3 (MIM# 608630) (Chan et al., 2006;Jen et al., 2004;Sicotte et al., 2006). Robo3 is a functional, intracellular binding partner of Dcc, expressed by commissural axons in the brainstem and spinal cord (Marillat et al., 2004;Sabatier et al., 2004;Zelina et al., 2014). The overlapping feature of horizontal gaze palsy in DSBS and HGPPS1 affected individuals appears to originate from hindbrain midline axon guidance defects in tracts that control conjugate horizontal eye movement (Chan et al., 2006;Jamuar et al., 2017;Jen et al., 2004;Renier et al., 2010;Sicotte et al., 2006). The pathogenesis of progressive scoliosis in both these disorders is unknown, but may stem from defective spinal commissural interneurons or abnormal development of extrapyramidal projections (Jamuar et al., 2017;Jen et al., 2004;Rabe Bernhardt et al., 2012). Diffusion MRI of one DSBS individual with a biallelic p.(Val263Alafs*36) mutation revealed several tract defects, including absence of the decussation of the superior cerebellar peduncles and transverse pontocerebellar projections (Jamuar et al., 2017). These commissural defects are also observed in individuals with HGPPS1 (Sicotte et al., 2006). Individuals with DSBS may also present with MMs, which are associated with the reduced midline crossing of descending CST projections at the pyramidal decussation (Jamuar et al., 2017). This is in contrast to individuals with HGPPS1 who do not display MMs, a feature attributed to an uncrossed CST leading to reversed lateralization of motor control (Jen et al., 2004). It is currently unknown why some individuals with DSBS and complete loss of DCC do not present with MMs. However, like HGPPS1, this may be due to a complete, rather than partial, failure of the descending CST to cross the midline. The other major clinical manifestations of DSBS include intellectual disability, global developmental delay, and hypotonia (Supp . Table S2) (Jamuar et al., 2017). Individuals with DSBS have a poor developmental outcome compared to individuals with DCC-iACC, likely attributed to additional brain abnormalities affecting the formation of other commissural tracts (Jamuar et al., 2017;Marsh et al., 2017).

DIAGNOSTIC STRATEGIES
The clinical assessment and diagnosis of MMs is commonly based on the Woods and Teuber severity scale developed in 1978 (Woods & Teuber, 1978). The assessment typically consists of three different on a scale between 1-barely discernible repetitive movement and 4movement equal to that expected for the intended hand (Woods & Teuber, 1978). The majority of individuals with DCC-MMs score between 2 (slight but unsustained or stronger, but briefer, repetitive movement) and 3 (strong and sustained repetitive movement). Recently, accelerometer gloves were utilized to quantitatively evaluate MMs in a multiplex MMs family with a monoallelic, p.(Pro551Leufs*26) DCC mutation (Franz et al., 2015). These gloves detect subtle movement, in contrast to electromyography that measures the electrical activity produced by skeletal muscle (Franz et al., 2015).  (Chan et al., 2006;Jen et al., 2004;Sicotte et al., 2006). However, the low diagnostic yield of genetic screening studies indicates that additional MMs and iACC disease genes remain to be identified and therefore mutations in DCC, NTN1, RAD51, and CDK5RAP2 will not always be identified in individuals with these phenotypes (Franz et al., 2015;Jamuar et al., 2017;Marsh et al., 2017;Méneret et al., 2014a;Méneret et al., 2017;Méneret, et al., 2014b).

VARIANTS OF UNKNOWN SIGNIFICANCE
The classification and clinical interpretation of a DCC sequence variant is challenged by the variable expressivity of monoallelic DCC mutations and the incomplete penetrance associated with DCC-MMs and DCC-iACC. It may also be complicated by phenotypic heterogeneity and the identification of other potential pathogenic variants in an individual.
In this context and in the absence of sufficient evidence to classify a DCC variant as pathogenic or not, it is recommended to report such a genetic alteration as a variant of unknown significance (VUS).
To illustrate these issues, we describe individual N-0083-01 with two novel, monoallelic VUS predicted to be damaging by most in silico tools: a maternally inherited NM_005215.3:c.916G > A:p.(Gly306Arg) DCC alteration and a de novo NM_001270399.1:c.
1246G > A:p.(Gly416Ser) TUBA1A (MIM# 602529) alteration (Supp .   Table S3) (Jamuar et al., 2017). The individual has complete ACC with a small pons and small inferior aspect of the midbrain, but no definite cortical malformation as would be expected to result from a pathogenic TUBA1A mutation (Keays et al., 2007). However, the individual does not exhibit the mild clinical manifestations associated with DCC-iACC. Instead, she has global developmental delay and severe cognitive impairments with significant emotional and behavioral problems in addition to spastic diplegia, strabismus and cortical visual impairment (Supp.

FUTURE PROSPECTS
Mutations in DCC disrupt the development of predominantly commissural tracts in the CNS and cause a spectrum of neurological disorders ranging from MMs and iACC with a normal or favorable developmental outcome, to DSBS with a poor developmental outcome. These findings are consistent with previously reported animal models and support the notion that DCC, in partnership with NTN1, functions as a master regulator of commissural axon guidance at the midline (Fazeli et al., 1997;Finger et al., 2002;Welniarz et al., 2017b).
Future investigations into the underlying molecular mechanisms leading to these disorders will strengthen current genotype-phenotype correlations. However, insightful correlations will be dependent on the multidisciplinary phenotyping of affected individuals, employing standardized neurological assessment, formal neuropsychological evaluation, and brain imaging studies. Utilization of advanced neuroimaging modalities, such as diffusion MRI-based tractography and functional MRI, will also aid our understanding of how the brains of these affected individuals is wired in the context of DCC dysfunction or LoF during development. We anticipate the DCC locus-specific LOVD will function as a central data repository that will assist researchers to establish these important genotype-phenotype correlations.

ACKNOWLEDGMENTS
The authors gratefully acknowledge the participation of the patients and their families in these studies and the support of the Australian Disorders of the Corpus Callosum (AusDoCC) organization, as well as the generous financial support of the Lefroy and Handbury families.
The authors also acknowledge and thank Ivo Fokkema and Johan T. den Dunnen from the Leiden University Medical Center for their assistance in establishing the DCC LOVD.

DISCLOSURE STATEMENT
A.M. received travel funding from Zambon Company and AbbVie Inc. E.R. has received research support from Merz-Pharma, Orkyn,