Katy Phelan

22q13.2q13.32 genomic regions associated with severity of speech delay, developmental delay, and physical features in Phelan-McDermid syndrome

Purpose:Phelan–McDermid syndrome is a developmental disability syndrome with varying deletions of 22q13 and varying clinical severity. We tested the hypothesis that, in addition to loss of the telomeric gene SHANK3, specific genomic regions within 22q13 are associated with important clinical features.Methods:We used a customized oligo array comparative genomic hybridization of 22q12.3-terminus to obtain deletion breakpoints in a cohort of 70 patients with terminal 22q13 deletions. We used association and receiver operating characteristic statistical methods in a novel manner and also incorporated protein interaction networks to identify 22q13 genomic locations and genes associated with clinical features.Results:Specific genomic regions and candidate genes within 22q13.2q13.32 were associated with severity of speech/language delay, neonatal hypotonia, delayed age at walking, hair-pulling behaviors, male genital anomalies, dysplastic toenails, large/fleshy hands, macrocephaly, short and tall stature, facial asymmetry, and atypical reflexes. We also found regions suggestive of a negative association with autism spectrum disorders.Conclusion:This work advances the field of research beyond the observation of a correlation between deletion size and phenotype and identifies candidate 22q13 loci, and in some cases specific genes, associated with singular clinical features observed in Phelan–McDermid syndrome. Our statistical approach may be useful in genotype–phenotype analyses for other microdeletion or microduplication syndromes.Genet Med 2014:16(4):318–328.

Growth in Phelan-McDermid syndrome.

Phelan–McDermid syndrome (or 22q13.3 deletion syndrome, OMIM #606232) results from a deletion in the terminal region of the long arm of chromosome 22 [Phelan et al., 2001]. Characteristic traits include hypotonia, developmental delay, and severely delayed or absent speech. Minor dysmorphic features are often present, and include long eyelashes, abnormal external ears, large fleshy hands, and hypoor dysplastic toenails [Phelan, 2008]. The growth aspect of the clinical picture of this syndrome is not welldefined. We report growth data on a cohort of 55 patients with terminal 22q13 deletions. Our results help delineate the clinical picture ofPhelan–McDermid syndromeanddemonstrate thenovel observation of occasional undergrowth. The postulation that ‘‘accelerated’’ growth may be a feature of Phelan–McDermid syndrome can be traced to an early report of seven patients with 22q13.3 deletions [Nesslinger et al., 1994]. This assertion was partially based on the observation that while none of the seven patients was undergrown, one had tall stature, one had large hands and feet, and three hadmacrocephaly. Of the three who had macrocephaly, two of their fathers did as well, suggesting the macrocephaly may have been inherited [Weaver and Christian, 1980] independently of the de novo 22q13.3 deletions. One of the two patients with familial macrocephaly was also the patient with tall stature, and head circumference is a known correlate of stature [Bushby et al., 1992], raising furtherquestionof a true associationof overgrowth with the deletion. On the other hand, most previous reports of short stature occurred in patients with ring chromosome 22, so growth failure was dismissed as a phenomenon of any autosomal ring structure rather than deletion of 22q13 [Luciani et al., 2003; Dhar et al., 2010]. Thus, ambiguous and subjective references to advanced or accelerated growth have often been repeated among the list of features of Phelan–McDermid syndrome [Phelan et al., 2001; Jones, 2006; Cusmano-Ozog et al., 2007; Phelan, 2008], although objective evidence has been weak. Wehave collected the largest set of clinical and laboratory data to date on patients with Phelan–McDermid syndrome. Most growth measurements were gathered from physical examinations at Phelan–McDermid Syndrome Foundation conferences in 1998, 2000, 2004, 2006, and 2008; measurements for two additional patients were derived from medical records. This study and the consent forms were approved by the Institutional Review Board of Self Regional Healthcare in Greenwood, South Carolina. After excluding patients with all but terminal deletions of 22q13 (i.e., rings, translocations, interstitial deletions, etc.), the study included growth data on 55 patients (26 males, 29 females) between 10 months and 40 years of age (median age 5.17 years). In five cases where multiple measurements were available from the same patient, only the most recent measurements were used. A total of 45 length/height measurements (19 males, 26 females) and 53 head circumference measurements (25 males, 28 females) were compared by deriving their centiles from the most recent growth references for the United States population [Kuczmarski et al., 2002; Rollins et al., 2010]. One-sample t-tests performed using Stata SE/10 (StataCorp, College Station, TX) revealed that average length-for-age, staturefor-age, andhead circumference-for-age centiles of our patients did not differ significantly from the expected value of 50 (Table I). However, the data appeared to be heavily weighted in the tails; the Chi-square goodness of fit test, also performed using Stata, revealed that the proportion of measurements falling above and below the respective cutoffs for tall and short stature (>95th centile and<5th centile, respectively) was significantly different than expected (P1⁄4 0.0239). The number of patients with short stature equalled

APPENDIX 3B Basic Techniques in Mammalian Cell Tissue Culture

Cultured mammalian cells are used extensively in cell biology studies. It requires a number of special skills in order to be able to preserve the structure, function, behavior, and biology of the cells in culture. This unit describes the basic skills required to maintain and preserve cell cultures: maintaining aseptic technique, preparing media with the appropriate characteristics, passaging, freezing and storage, recovering frozen stocks, and counting viable cells.

Mammalian Cell Tissue Culture Techniques

Cultured tissues and cells are used extensively in physiological and pharmacological studies. In vitro cultures provide a means of examining cells and tissues without the complex interactions that would be present if the whole organism were studied. A number of special skills are required in order to preserve the structure, function, behavior, and biology of cells in culture. This unit describes the basic skills required to maintain and preserve cell cultures: maintaining aseptic technique, preparing media with the appropriate characteristics, passaging, freezing and storage, recovering frozen stocks, and counting viable cells.

Letter to the editor regarding Disciglio et al.: Interstitial 22q13 deletions not involving SHANK3 gene: A new contiguous gene syndrome

Letter to the Editor Regarding Disciglio et al.: Interstitial 22q13 Deletions not Involving SHANK3 Gene: A New Contiguous Gene Syndrome Katy Phelan,* Luigi Boccuto, R. Curtis Rogers, Sara M. Sarasua, and Heather E. McDermid Department of Pediatrics, Hayward Genetics Center, Tulane University School of Medicine, New Orleans, Louisiana Greenwood Genetic Center, Greenwood, South Carolina Biological Sciences, University of Alberta, Edmonton, Alberta, Canada

Variability in Phelan-McDermid syndrome: The impact of the PNPLA3 p.Ile148M polymorphism

The PNPLA3 gene maps in the 22q13 region and can have modifying effects on the phenotype of patients with Phelan-McDermid syndrome (PMS). The PNPLA3 p.I148M variant was detected in two PMS patients presenting with refractory seizures, gastrointestinal issues, and liver dysfunction. The p.I148M variant leads to macrovescicular steaosis and predisposes to liver disorders from steatohepatitis to fibrosis. Accumulation of lipid macrovescicles in the hepatocytes affects several pathways, including the metabolismof anti-epileptics, possibly leading to the lack of response to anti-epileptic treatments reported in the two cases. Screening for the p.I148M variant can identify PMS patients at higher risk for liver dyfunction and help designing personalized therapeutic protocols.

Mammalian Cell Tissue Culture

Cultured mammalian cells are used extensively in the field of human genetics. It requires a number of special skills in order to be able to preserve the structure, function, behavior, and biology of the cells in culture. This unit describes the basic skills required to maintain and preserve cell cultures: maintaining aseptic technique, preparing media with the appropriate characteristics, passaging, freezing and storage, recovering frozen stocks, and counting viable cells.