Nicholas J. Neill

The spliceosome is a therapeutic vulnerability in MYC-driven cancer

MYC (also known as c-MYC) overexpression or hyperactivation is one of the most common drivers of human cancer. Despite intensive study, the MYC oncogene remains recalcitrant to therapeutic inhibition. MYC is a transcription factor, and many of its pro-tumorigenic functions have been attributed to its ability to regulate gene expression programs. Notably, oncogenic MYC activation has also been shown to increase total RNA and protein production in many tissue and disease contexts. While such increases in RNA and protein production may endow cancer cells with pro-tumour hallmarks, this increase in synthesis may also generate new or heightened burden on MYC-driven cancer cells to process these macromolecules properly. Here we discover that the spliceosome is a new target of oncogenic stress in MYC-driven cancers. We identify BUD31 as a MYC-synthetic lethal gene in human mammary epithelial cells, and demonstrate that BUD31 is a component of the core spliceosome required for its assembly and catalytic activity. Core spliceosomal factors (such as SF3B1 and U2AF1) associated with BUD31 are also required to tolerate oncogenic MYC. Notably, MYC hyperactivation induces an increase in total precursor messenger RNA synthesis, suggesting an increased burden on the core spliceosome to process pre-mRNA. In contrast to normal cells, partial inhibition of the spliceosome in MYC-hyperactivated cells leads to global intron retention, widespread defects in pre-mRNA maturation, and deregulation of many essential cell processes. Notably, genetic or pharmacological inhibition of the spliceosome in vivo impairs survival, tumorigenicity and metastatic proclivity of MYC-dependent breast cancers. Collectively, these data suggest that oncogenic MYC confers a collateral stress on splicing, and that components of the spliceosome may be therapeutic entry points for aggressive MYC-driven cancers.

Copy number variants of schizophrenia susceptibility loci are associated with a spectrum of speech and developmental delays and behavior problems

Purpose: Recently, molecular cytogenetic techniques have identified novel copy number variants in individuals with schizophrenia. However, no large-scale prospective studies have been performed to characterize the broader spectrum of phenotypes associated with such copy number variants in individuals with unexplained physical and intellectual disabilities encountered in a diagnostic setting.Methods: We analyzed 38,779 individuals referred to our diagnostic laboratory for microarray testing for the presence of copy number variants encompassing 20 putative schizophrenia susceptibility loci. We also analyzed the indications for study for individuals with copy number variants overlapping those found in six individuals referred for schizophrenia.Results: After excluding larger gains or losses that encompassed additional genes outside the candidate loci (e.g., whole-arm gains/losses), we identified 1113 individuals with copy number variants encompassing schizophrenia susceptibility loci and 37 individuals with copy number variants overlapping those present in the six individuals referred to our laboratory for schizophrenia. Of these, 1035 had a copy number variant of one of six recurrent loci: 1q21.1, 15q11.2, 15q13.3, 16p11.2, 16p13.11, and 22q11.2. The indications for study for these 1150 individuals were diverse and included developmental delay, intellectual disability, autism spectrum, and multiple congenital anomalies.Conclusion: The results from our study, the largest genotype-first analysis of schizophrenia susceptibility loci to date, suggest that the phenotypic effects of copy number variants associated with schizophrenia are pleiotropic and imply the existence of shared biologic pathways among multiple neurodevelopmental conditions.

Comparative analysis of copy number detection by whole-genome BAC and oligonucleotide array CGH.

BackgroundMicroarray-based comparative genomic hybridization (aCGH) is a powerful diagnostic tool for the detection of DNA copy number gains and losses associated with chromosome abnormalities, many of which are below the resolution of conventional chromosome analysis. It has been presumed that whole-genome oligonucleotide (oligo) arrays identify more clinically significant copy-number abnormalities than whole-genome bacterial artificial chromosome (BAC) arrays, yet this has not been systematically studied in a clinical diagnostic setting.ResultsTo determine the difference in detection rate between similarly designed BAC and oligo arrays, we developed whole-genome BAC and oligonucleotide microarrays and validated them in a side-by-side comparison of 466 consecutive clinical specimens submitted to our laboratory for aCGH. Of the 466 cases studied, 67 (14.3%) had a copy-number imbalance of potential clinical significance detectable by the whole-genome BAC array, and 73 (15.6%) had a copy-number imbalance of potential clinical significance detectable by the whole-genome oligo array. However, because both platforms identified copy number variants of unclear clinical significance, we designed a systematic method for the interpretation of copy number alterations and tested an additional 3,443 cases by BAC array and 3,096 cases by oligo array. Of those cases tested on the BAC array, 17.6% were found to have a copy-number abnormality of potential clinical significance, whereas the detection rate increased to 22.5% for the cases tested by oligo array. In addition, we validated the oligo array for detection of mosaicism and found that it could routinely detect mosaicism at levels of 30% and greater.ConclusionsAlthough BAC arrays have faster turnaround times, the increased detection rate of oligo arrays makes them attractive for clinical cytogenetic testing.

Investigation of NRXN1 deletions: clinical and molecular characterization.

Deletions at 2p16.3 involving exons of NRXN1 are associated with susceptibility for autism and schizophrenia, and similar deletions have been identified in individuals with developmental delay and dysmorphic features. We have identified 34 probands with exonic NRXN1 deletions following referral for clinical microarray‐based comparative genomic hybridization. To more firmly establish the full phenotypic spectrum associated with exonic NRXN1 deletions, we report the clinical features of 27 individuals with NRXN1 deletions, who represent 23 of these 34 families. The frequency of exonic NRXN1 deletions among our postnatally diagnosed patients (0.11%) is significantly higher than the frequency among reported controls (0.02%; P = 6.08 × 10−7), supporting a role for these deletions in the development of abnormal phenotypes. Generally, most individuals with NRXN1 exonic deletions have developmental delay (particularly speech), abnormal behaviors, and mild dysmorphic features. In our cohort, autism spectrum disorders were diagnosed in 43% (10/23), and 16% (4/25) had epilepsy. The presence of NRXN1 deletions in normal parents and siblings suggests reduced penetrance and/or variable expressivity, which may be influenced by genetic, environmental, and/or stochastic factors. The pathogenicity of these deletions may also be affected by the location of the deletion within the gene. Counseling should appropriately represent this spectrum of possibilities when discussing recurrence risks or expectations for a child found to have a deletion in NRXN1.

Haploinsufficiency of SOX5 at 12p12.1 is associated with developmental delays with prominent language delay, behavior problems, and mild dysmorphic features†

SOX5 encodes a transcription factor involved in the regulation of chondrogenesis and the development of the nervous system. Despite its important developmental roles, SOX5 disruption has yet to be associated with human disease. We report one individual with a reciprocal translocation breakpoint within SOX5, eight individuals with intragenic SOX5 deletions (four are apparently de novo and one inherited from an affected parent), and seven individuals with larger 12p12 deletions encompassing SOX5. Common features in these subjects include prominent speech delay, intellectual disability, behavior abnormalities, and dysmorphic features. The phenotypic impact of the deletions may depend on the location of the deletion and, consequently, which of the three major SOX5 protein isoforms are affected. One intragenic deletion, involving only untranslated exons, was present in a more mildly affected subject, was inherited from a healthy parent and grandparent, and is similar to a deletion found in a control cohort. Therefore, some intragenic SOX5 deletions may have minimal phenotypic effect. Based on the location of the deletions in the subjects compared to the controls, the de novo nature of most of these deletions, and the phenotypic similarities among cases, SOX5 appears to be a dosage‐sensitive, developmentally important gene. Hum Mutat 33:728–740, 2012.

Referral patterns for microarray testing in prenatal diagnosis

Objective To understand the prenatal referral patterns from the United States, Canada, and Israel for two whole-genome microarray platforms, each with a different resolution. Method Physicians selected one of the two array designs to be performed on 1483 prenatal specimens for a 1-year period. We retrospectively examined detection rates, indications for study, and physician array selection. Results The lower resolution array (55 K) showed an ~32% decrease in the detection of results of unclear clinical significance while retaining the ability to detect all but one significant abnormality identified by the higher resolution array (135 K). A majority of samples were referred for abnormal ultrasound findings. Whereas the United States and Canada utilized the higher resolution array more often for this indication, Israel preferred the 55 K array. Referral patterns for parental anxiety were similar for the United States and Israel, with most cases being tested on the 55 K array. Few cases were referred for advanced maternal age or family history of a genetic condition from either Canada or Israel. Conclusion Referral patterns varied between the countries and between indications for study. Understanding these differences will provide laboratories the critical information needed to develop array designs to meet the medical needs and patient desires for prenatal testing.

Recurrent HERV‐H‐Mediated 3q13.2–q13.31 Deletions Cause a Syndrome of Hypotonia and Motor, Language, and Cognitive Delays

We describe the molecular and clinical characterization of nine individuals with recurrent, 3.4‐Mb, de novo deletions of 3q13.2–q13.31 detected by chromosomal microarray analysis. All individuals have hypotonia and language and motor delays; they variably express mild to moderate cognitive delays (8/9), abnormal behavior (7/9), and autism spectrum disorders (3/9). Common facial features include downslanting palpebral fissures with epicanthal folds, a slightly bulbous nose, and relative macrocephaly. Twenty‐eight genes map to the deleted region, including four strong candidate genes, DRD3, ZBTB20, GAP43, and BOC, with important roles in neural and/or muscular development. Analysis of the breakpoint regions based on array data revealed directly oriented human endogenous retrovirus (HERV‐H) elements of ∼5 kb in size and of >95% DNA sequence identity flanking the deletion. Subsequent DNA sequencing revealed different deletion breakpoints and suggested nonallelic homologous recombination (NAHR) between HERV‐H elements as a mechanism of deletion formation, analogous to HERV‐I‐flanked and NAHR‐mediated AZFa deletions. We propose that similar HERV elements may also mediate other recurrent deletion and duplication events on a genome‐wide scale. Observation of rare recurrent chromosomal events such as these deletions helps to further the understanding of mechanisms behind naturally occurring variation in the human genome and its contribution to genetic disease.

Referral patterns for microarray testing in prenatal diagnosis: Microarray testing referral patterns

To understand the prenatal referral patterns from the United States, Canada, and Israel for two whole‐genome microarray platforms, each with a different resolution.

Clinical Utility of Chromosomal Microarray Analysis

OBJECTIVE: To test the hypothesis that chromosomal microarray analysis frequently diagnoses conditions that require specific medical follow-up and that referring physicians respond appropriately to abnormal test results. METHODS: A total of 46 298 postnatal patients were tested by chromosomal microarray analysis for a variety of indications, most commonly intellectual disability/developmental delay, congenital anomalies, dysmorphic features, and neurobehavioral problems. The frequency of detection of abnormalities associated with actionable clinical features was tallied, and the rate of physician response to a subset of abnormal tests results was monitored. RESULTS: A total of 2088 diagnoses were made of more than 100 different disorders that have specific clinical features that warrant follow-up. The detection rate for these conditions using high-resolution whole-genome microarrays was 5.4%, which translates to 35% of all clinically significant abnormal test results identified in our laboratory. In a subset of cases monitored for physician response, appropriate clinical action was taken more than 90% of the time as a direct result of the microarray finding. CONCLUSIONS: The disorders diagnosed by chromosomal microarray analysis frequently have clinical features that need medical attention, and physicians respond to the diagnoses with specific clinical actions, thus arguing that microarray testing provides clinical utility for a significant number of patients tested.

Diagnostic utility of microarray testing in pregnancy loss

To determine the frequency of clinically significant chromosomal abnormalities identified by chromosomal microarray in pregnancy losses at any gestational age and to compare microarray performance with that of traditional cytogenetic analysis when testing pregnancy losses.