Genetic counselors outside of the genetics clinic: Roles, practices, and ethico-legal implications in light of lagging legal recognition across Canada.

Advances in medical genetics have led to a significant increase in demand for genetic services and expertise across almost all medical specialties. Genetic counselors (GCs) in Canada play key roles in genetic services both within and outside of the Genetics Clinic, while not being regulated or legally recognized as healthcare professionals (HCPs) in most provinces. Understanding whether GCs outside of the "traditional" Genetics Clinic influence patient care, their level of professional autonomy and supervisory structure is, therefore, important. In this study, we explore the current landscape of GC practice outside of the Genetics Clinic by describing positions, determining the professional scope of practice, as defined by the Canadian Association of Genetic Counselors (CAGC) and Canadian Board of Genetic Counseling (CBGC) core competencies, and by elucidating associated ethico-legal implications. An online survey was developed and distributed to GCs working with patient-related data in Canada in positions outside of the Genetics Clinic through the CAGC ListServ and accessed between March 5 and April 9, 2021. Thirty GCs were included in the study, with 16/30 in public healthcare system positions. Most respondents held roles with direct (11/30) and indirect (14/30) impact on patient care and management, and the majority reported performing their primary roles with minimal supervision (56%) or complete independence (36%). Most roles (22/25) elicited by respondents were considered to be within the GC scope of practice, except for administrative tasks and special projects. GCs were the only genetics-trained professional(s) in 8/30 of respondents' workplaces. The results of the current study support the value of GCs translatable skillset in positions beyond the Genetics Clinic, and outline ethico-legal implications for GCs, regulated HCPs, patients, and health institutions in the absence of legal recognition, including medical-legal liability and title protection. This study provides evidence in support of regulation of GCs as HCPs.

Evaluation of the diagnostic accuracy of exome sequencing and its impact on diagnostic thinking for patients with rare disease in a publicly funded health care system: A prospective cohort study.

PURPOSE: To evaluate the diagnostic utility of publicly funded clinical exome sequencing (ES) for patients with suspected rare genetic diseases. METHODS: We prospectively enrolled 297 probands who met eligibility criteria and received ES across 5 sites in Ontario, Canada, and extracted data from medical records and clinician surveys. Using the Fryback and Thornbury Efficacy Framework, we assessed diagnostic accuracy by examining laboratory interpretation of results and assessed diagnostic thinking by examining the clinical interpretation of results and whether clinical-molecular diagnoses would have been achieved via alternative hypothetical molecular tests. RESULTS: Laboratories reported 105 molecular diagnoses and 165 uncertain results in known and novel genes. Of these, clinicians interpreted 102 of 105 (97%) molecular diagnoses and 6 of 165 (4%) uncertain results as clinical-molecular diagnoses. The 108 clinical-molecular diagnoses were in 104 families (35% diagnostic yield). Each eligibility criteria resulted in diagnostic yields of 30% to 40%, and higher yields were achieved when >2 eligibility criteria were met (up to 45%). Hypothetical tests would have identified 61% of clinical-molecular diagnoses. CONCLUSION: We demonstrate robustness in eligibility criteria and high clinical validity of laboratory results from ES testing. The importance of ES was highlighted by the potential 40% of patients that would have gone undiagnosed without this test.

Persistent chylothorax associated with lymphatic malformation type 6 due to biallelic pathogenic variants in PIEZO1.

PIEZO1 is required for lymphatic valve formation, and several lymphatic abnormalities have been reported to be associated with autosomal recessive PIEZO1 pathogenic variants including neonatal hydrops, lymphedema involving various body regions, and chylothorax. Persistent or recurrent chylothorax has been infrequently described in association with pathogenic variants in the PIEZO1 gene. We present a 4-year-old female with bilateral pleural effusions detected prenatally, who was diagnosed with bilateral chylothoraces post-partum. She subsequently had recurrent pleural effusions involving both pleural cavities, which tended to improve with restriction of her fat intake, and, one occasion, subcutaneous octreotide. She also had bilateral calf, and intermittent cheek swelling. Genetic testing revealed two deleterious variants in PIEZO1: c.2330-2_2330-1del and c.3860G > A (p.Trp1287*), both of which were classified as likely pathogenic. This supported a diagnosis of Lymphatic Malformation Type 6 (OMIM 616843), also known as Hereditary Lymphedema Type III. Hereditary Lymphedema type III can be associated with persistent chylothorax that can vary in size over time.

The implementation of an enhanced clinical model to improve the diagnostic yield of exome sequencing for patients with a rare genetic disease: A Canadian experience.

The introduction of clinical exome sequencing (ES) has provided a unique opportunity to decrease the diagnostic odyssey for patients living with a rare genetic disease (RGD). ES has been shown to provide a diagnosis in 29%-57% of patients with a suspected RGD, with as many as 70% remaining undiagnosed. There is a need to advance the clinical model of care by more formally integrating approaches that were previously considered research into an enhanced diagnostic workflow. We developed an Exome Clinic, which set out to evaluate a workflow for improving the diagnostic yield of ES for patients with an undiagnosed RGD. Here, we report the outcomes of 47 families who underwent clinical ES in the first year of the clinic. The diagnostic yield from clinical ES was 40% (19/47). Families who remained undiagnosed after ES had the opportunity for follow-up studies that included phenotyping and candidate variant segregation in relatives, genomic matchmaking, and ES reanalysis. This enhanced diagnostic workflow increased the diagnostic yield to 55% (26/47), predominantly through the resolution of variants and genes of uncertain significance. We advocate that this approach be integrated into mainstream clinical practice and highlight the importance of a coordinated translational approach for patients with RGD.

Biallelic NLRP7 variants in patients with recurrent hydatidiform mole: A review and expert consensus.

Hydatidiform mole (HM) is an abnormal human pregnancy characterized by excessive growth of placental trophoblasts and abnormal early embryonic development. Following a first such abnormal pregnancy, the risk for women of successive molar pregnancies significantly increases. To date variants in seven maternal-effect genes have been shown to cause recurrent HMs (RHM). NLRP7 is the major causative gene for RHM and codes for NOD-like receptor (NLR) family pyrin domain containing 7, which belongs to a family of proteins involved in inflammatory disorders. Since its identification, all NLRP7 variants have been recorded in Infevers, an online registry dedicated to autoinflammatory diseases (https://infevers.umai-montpellier.fr/web/). Here, we reviewed published and unpublished recessive NLRP7 variants associated with RHM, scored their pathogenicity according to the American College of Medical Genetics classification, and recapitulated all functional studies at the level of both the patients and the conceptions. We also provided data on further variant analyses of 32 patients and genotypes of 36 additional molar pregnancies. This comprehensive review integrates published and unpublished data on NLRP7 and aims at guiding geneticists and clinicians in variant interpretation, genetic counseling, and management of patients with this rare condition.

Heterozygous loss-of-function variants significantly expand the phenotypes associated with loss of GDF11.

PURPOSE: Growth differentiation factor 11 (GDF11) is a key signaling protein required for proper development of many organ systems. Only one prior study has associated an inherited GDF11 variant with a dominant human disease in a family with variable craniofacial and vertebral abnormalities. Here, we expand the phenotypic spectrum associated with GDF11 variants and document the nature of the variants. METHODS: We present a cohort of six probands with de novo and inherited nonsense/frameshift (4/6 patients) and missense (2/6) variants in GDF11. We generated gdf11 mutant zebrafish to model loss of gdf11 phenotypes and used an overexpression screen in Drosophila to test variant functionality. RESULTS: Patients with variants in GDF11 presented with craniofacial (5/6), vertebral (5/6), neurological (6/6), visual (4/6), cardiac (3/6), auditory (3/6), and connective tissue abnormalities (3/6). gdf11 mutant zebrafish show craniofacial abnormalities and body segmentation defects that match some patient phenotypes. Expression of the patients' variants in the fly showed that one nonsense variant in GDF11 is a severe loss-of-function (LOF) allele whereas the missense variants in our cohort are partial LOF variants. CONCLUSION: GDF11 is needed for human development, particularly neuronal development, and LOF GDF11 alleles can affect the development of numerous organs and tissues.

Channelopathies Are a Frequent Cause of Genetic Ataxias Associated with Cerebellar Atrophy.

BACKGROUND: Cerebellar atrophy is a nonspecific imaging finding observed in a number of neurological disorders. Genetic ataxias associated with cerebellar atrophy are a heterogeneous group of conditions, rendering the approach to diagnosis challenging. OBJECTIVES: To define the spectrum of genetic ataxias associated with cerebellar atrophy in a Canadian cohort and the diagnostic yield of exome sequencing for this group of conditions. METHODS: A total of 92 participants from 66 families with cerebellar atrophy were recruited for this multicenter prospective cohort study. Exome sequencing was performed for all participants between 2011 and 2017 as part of 1 of 2 national research programs, Finding of Rare Genetic Disease Genes or Enhanced Care for Rare Genetic Diseases in Canada. RESULTS: A genetic diagnosis was established in 53% of families (35/66). Pathogenic variants were found in 21 known genes, providing a diagnosis for 31/35 families (89%), and in 4 novel genes, accounting for 4/35 families (11%). Of the families, 31/66 (47%) remained without a genetic diagnosis. The most common diagnoses were channelopathies, which were established in 9/35 families (26%). Additional clinical findings provided useful clues to specific diagnoses. CONCLUSIONS: We report on the high frequency of channelopathies as a cause of genetic ataxias associated with cerebellar atrophy and the utility of exome sequencing for this group of conditions.

A report of congenital adrenal hyperplasia due to 17α-hydroxylase deficiency in two 46,XX sisters.

Congenital adrenal hyperplasia (CAH) is a group of rare orphan disorders caused by mutations in seven different enzymes that impair cortisol biosynthesis. The 17α-hydroxylase deficiency (17OHD) is one of the less common forms of CAH, corresponding to approximately 1% of the cases, with an estimated annual incidence of 1 in 50,000 newborns. Cases description - two phenotypically female Ecuadorian sisters, both with primary amenorrhea, absence of secondary sexual characteristics, and osteoporosis. High blood pressure was present in the older sister. Hypergonadotropic hypogonadism profile was observed: decreased cortisol and dehydroepiandrosterone sulfate (DHEAS), increased adrenocorticotropic hormone (ACTH) and normal levels of 17-hydroxyprogesterone, extremely high deoxycorticosterone (DOC) levels, and a tomography showed bilateral adrenal hyperplasia in both sisters. Consanguinity was evident in their ancestors. Furthermore, in the exon 7, the variant c.1216T > C, p.Trp406Arg was detected in homozygosis in the CYP17A1 gene of both sisters. We report a homozygous missense mutation in the CYP17A1 gene causing 17OHD in two sisters from Loja, Ecuador. According to the authors, this is the first time such deficiency and mutation are described in two members of the same family in Ecuador.

Phenotype and mutation expansion of the PTPN23 associated disorder characterized by neurodevelopmental delay and structural brain abnormalities.

PTPN23 is a His-domain protein-tyrosine phosphatase implicated in ciliogenesis, the endosomal sorting complex required for transport (ESCRT) pathway, and RNA splicing. Until recently, no defined human phenotype had been associated with alterations in this gene. We identified and report a cohort of seven patients with either homozygous or compound heterozygous rare deleterious variants in PTPN23. Combined with four patients previously reported, a total of 11 patients with this disorder have now been identified. We expand the phenotypic and variation spectrum associated with defects in this gene. Patients have strong phenotypic overlap, suggesting a defined autosomal recessive syndrome caused by reduced function of PTPN23. Shared characteristics of affected individuals include developmental delay, brain abnormalities (mainly ventriculomegaly and/or brain atrophy), intellectual disability, spasticity, language disorder, microcephaly, optic atrophy, and seizures. We observe a broad range of variants across patients that are likely strongly reducing the expression or disrupting the function of the protein. However, we do not observe any patients with an allele combination predicted to result in complete loss of function of PTPN23, as this is likely incompatible with life, consistent with reported embryonic lethality in the mouse. None of the observed or reported variants are recurrent, although some have been identified in homozygosis in patients from consanguineous populations. This study expands the phenotypic and molecular spectrum of PTPN23 associated disease and identifies major shared features among patients affected with this disorder, while providing additional support to the important role of PTPN23 in human nervous and visual system development and function.

p21 protein-activated kinase 1 is associated with severe regressive autism, and epilepsy.

The p21-activated kinase (PAK) family of proteins function as key effectors of RHO family GTPases in mammalian cells to regulate many pathways including Ras/Raf/MEK/ERK and Wnt/ß-catenin, amongst others. Here we report an individual with a novel autosomal dominant disorder characterized by severe regressive autism, intellectual disability, and epilepsy. Exome sequencing of the proband and her parents revealed a de novo variant in the PAK1 gene ([NM_001128620] c.362C>T/p.Pro121Leu). Studies in patient cells showed a clear effect on PAK1 protein function, including altered phosphorylation of targets (JNK and ERK), decreased abundance of ß-catenin, and concomitant altered expression downstream of these key regulators. Our findings add PAK1 to the list of PAK proteins and kinases which when mutated cause rare genetic diseases.

MRI-based methodology to monitor the impact of positional changes on the airway caliber in obstructive sleep apnea patients.

PURPOSE: To develop a non-invasive MRI-based methodology to visually and quantitatively assess the impact of head and chest rotations on the airway caliber. METHODS: An MRI table set-up was developed for independent rotations of the head and chest along B0 field and tested for feasibility using phantom scans. The accuracy of the head and chest rotations was validated with ten volunteer scans. A 3T MRI protocol was optimized to image the regions of interest (ROIs) that were the retropalatal (RP) and retroglossal (RG) sections of the upper airway. A workflow for data analysis was developed to assess the changes of the airway caliber following the independent head and chest rotations. RESULTS: A prototype MRI table setup was established with two separate plates each supporting and rotating the head or chest independently. Subject positioning and image acquisition were finished within seven minutes for each position. Thus, each subject MRI was set up with seven positions and completed for less than one hour. The implemented angles were within 0.3-degree deviation from the targeted angles. The data analysis workflow provided 2D and 3D visualization and quantification with the measurements of cross-sectional area, lateral and anterior-posterior distances of the ROIs. Sharp contrast of the airway and its surrounding tissues facilitated an automatic approach to ROI placement to minimize subjectivity. CONCLUSIONS: The 3T MRI data acquisition and analysis methodology could reliably assess the impact of head and chest rotations on the upper airway caliber to identify the optimal position for obstructive sleep apnea patients.

The unsolved rare genetic disease atlas? An analysis of the unexplained phenotypic descriptions in OMIM®.

For years, the genetics community has estimated the number of individual rare genetic diseases to be approximately 6,000-8,000. A commonly quoted derivation of this estimate is based on the simple addition of the number of phenotypic entries with and without confirmed molecular etiologies in the Online Mendelian Inheritance in Man (OMIM®). Here, we examine the validity of this estimation by mining the phenotypic entries in OMIM that are of likely or suspected Mendelian inheritance without a molecular cause (MIM number prefix "%" or "null"). Of the 3,204 unsolved phenotypic entries in OMIM, only two-thirds (2,034 entries) represented rare diseases. Of these, 8% were considered "well-established" based on their description in commonly used reference textbooks. We hypothesize based on the large proportion of entries that represent single families reported prior to 2011, that a number of the unsolved entries represent pathogenic variants in known genes. The novel gene discovery potential of these entries is therefore likely lower than originally thought. Given that the majority of the ~300 new disease-gene associations curated each year by OMIM were never associated with a "%" or "null" sign, the true scope of the rare disease atlas is likely much larger than previously anticipated.

Stabilizing proteins from sequence statistics: the interplay of conservation and correlation in triosephosphate isomerase stability.

Understanding the determinants of protein stability remains one of protein science's greatest challenges. There are still no computational solutions that calculate the stability effects of even point mutations with sufficient reliability for practical use. Amino acid substitutions rarely increase the stability of native proteins; hence, large libraries and high-throughput screens or selections are needed to stabilize proteins using directed evolution. Consensus mutations have proven effective for increasing stability, but these mutations are successful only about half the time. We set out to understand why some consensus mutations fail to stabilize, and what criteria might be useful to predict stabilization more accurately. Overall, consensus mutations at more conserved positions were more likely to be stabilizing in our model, triosephosphate isomerase (TIM) from Saccharomyces cerevisiae. However, positions coupled to other sites were more likely not to stabilize upon mutation. Destabilizing mutations could be removed both by removing sites with high statistical correlations to other positions and by removing nearly invariant positions at which "hidden correlations" can occur. Application of these rules resulted in identification of stabilizing mutations in 9 out of 10 positions, and amalgamation of all predicted stabilizing positions resulted in the most stable yeast TIM variant we produced (+8 °C). In contrast, a multimutant with 14 mutations each found to stabilize TIM independently was destabilized by 2 °C. Our results are a practical extension to the consensus concept of protein stabilization, and they further suggest the importance of positional independence in the mechanism of consensus stabilization.