ABSTRACT
PDAC (also termed Matthew Wood) syndrome is a rare, autosomal recessive disorder characterized by pulmonary hypoplasia/aplasia, diaphragmatic defects, bilateral anophthalmia, and cardiac malformations. The disorder is caused by mutations in STRA6, an important regulator of vitamin A and retinoic acid metabolism. We describe six cases from four families of Hmong ancestry, seen over a 30 years period in California. These include: (i) consanguineous siblings with a combination of bilateral anophthalmia, diaphragmatic abnormalities, truncus arteriosus, and/or pulmonary agenesis/hypoplasia; (ii) a singleton fetus with bilateral anophthalmia, pulmonary agenesis, cardiac malformation, and renal hypoplasia; (iii) a sibling pair with a combination of antenatal contractures, camptodactyly, fused palpebral fissures, pulmonary agenesis, and/or truncus arteriosus; (iv) a fetus with bilateral anophthalmia, bushy eyebrows, pulmonary agenesis, heart malformation, and abnormal hand positioning. The phenotypic spectrum of PDAC syndrome has until now not included contractures or camptodactyly. Sequencing of STRA6 in unrelated members of families three and four identified a novel, shared homozygous splice site alteration (c.113 + 3_4delAA) that is predicted to be pathogenic. We hypothesize this may represent a unique disease allele in the Hmong. We also provide a focused review of all published PDAC syndrome cases with confirmed or inferred STRA6 mutations, illustrating the phenotypic and molecular variability that characterizes this disorder.
Subject(s)
Abnormalities, Multiple/genetics , Alternative Splicing/genetics , Anophthalmos/genetics , Contracture/genetics , Hand Deformities, Congenital/genetics , Lung Diseases/genetics , Lung/abnormalities , Membrane Proteins/genetics , Microphthalmos/genetics , Mutation/genetics , Abnormalities, Multiple/pathology , Anophthalmos/pathology , California , Consanguinity , Contracture/pathology , Female , Gestational Age , Hand Deformities, Congenital/pathology , Heart Defects, Congenital/genetics , Heart Defects, Congenital/pathology , Homozygote , Humans , Infant, Newborn , Lung/pathology , Lung Diseases/pathology , Male , Microphthalmos/pathology , Pedigree , Pregnancy , Prognosis , SyndromeABSTRACT
The RASopathies are a family of developmental disorders caused by heritable defects of the RAS/MAPK signaling pathway. While the postnatal presentation of this group of disorders is well known, the prenatal and neonatal findings are less widely recognized. We report on the perinatal presentation of 10 patients with Noonan syndrome (NS), nine with Cardiofaciocutaneous syndrome (CFCS) and three with Costello syndrome (CS), in conjunction with the results of a comprehensive literature review. The majority of perinatal findings in NS, CS, and CFCS are shared: polyhydramnios; prematurity; lymphatic dysplasia; macrosomia; relative macrocephaly; respiratory distress; hypotonia, as well as cardiac and renal anomalies. In contrast, fetal arrhythmia and neonatal hypoglycemia are relatively specific to CS. NS, CS, and CFCS should all be considered as a possible diagnosis in pregnancies with a normal karyotype and ultrasound findings of a RASopathy. Recognition of the common perinatal findings of these disorders should facilitate both their prenatal and neonatal diagnosis.
Subject(s)
Costello Syndrome/diagnosis , Ectodermal Dysplasia/diagnosis , Failure to Thrive/diagnosis , Heart Defects, Congenital/diagnosis , Noonan Syndrome/diagnosis , Phenotype , Abnormalities, Multiple , Costello Syndrome/genetics , DNA Mutational Analysis , Diagnosis, Differential , Ectodermal Dysplasia/genetics , Facies , Failure to Thrive/genetics , Female , Heart Defects, Congenital/genetics , Humans , Infant, Newborn , Male , Mutation , Neonatal Screening , Noonan Syndrome/genetics , Prenatal DiagnosisABSTRACT
Patients with physical findings suggestive of Treacher Collins syndrome (TCS) or mandibulofacial dysostosis (MFD) and macrocytic anemia diagnostic of Diamond-Blackfan anemia (DBA) have been reported. Disease-causing genes have been identified for TCS and other MFDs. Mutations in several ribosomal protein genes and the transcription factor GATA1 result in DBA. However, no disease-causing mutation had been identified in the reported patients with the combination of TCS/MFD and DBA phenotype, and we hypothesized that pathogenic mutations in a single gene could be identified using whole exome analysis. We studied probands from six unrelated families. Combining exome analysis and Sanger sequencing, we identified likely pathogenic mutations in 5/6 families. Two mutations in unrelated families were seen in RPS26, the known DBA10 gene. One variant was predicted to affect mRNA splicing, and the other to lead to protein truncation. In another family a likely pathogenic X-linked mutation affecting a highly conserved residue was found in TSR2, which encodes a direct binding partner of RPS26. De novo mutations affecting the RPS28 start codon were found in two unrelated probands, identifying RPS28 as a novel disease gene. We conclude that the phenotype combining features of TCS with DBA is genetically heterogeneous. Each of the pathogenic variants identified is predicted to impede ribosome biogenesis, which in turn could result in altered cell growth and proliferation, causing abnormal embryologic development, defective erythropoiesis and reduced growth. The phenotype combining TCS/MFD and DBA is highly variable, overlaps with DBA and lies within the phenotypic spectrum of ribosomopathies. © 2014 Wiley Periodicals, Inc.
Subject(s)
Anemia, Diamond-Blackfan/complications , Anemia, Diamond-Blackfan/genetics , Apoptosis Regulatory Proteins/genetics , Genetic Heterogeneity , Mandibulofacial Dysostosis/complications , Mandibulofacial Dysostosis/genetics , Ribosomal Proteins/genetics , Adult , Child, Preschool , Exome/genetics , Family , Female , Humans , Infant , Infant, Newborn , Male , Pedigree , Phenotype , Pregnancy , Young AdultABSTRACT
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.
Subject(s)
Cell Adhesion Molecules, Neuronal/genetics , Gene Deletion , Nerve Tissue Proteins/genetics , Abnormalities, Multiple/diagnosis , Abnormalities, Multiple/genetics , Adolescent , Adult , Autistic Disorder/genetics , Calcium-Binding Proteins , Child , Child, Preschool , Comparative Genomic Hybridization , Developmental Disabilities/genetics , Exons , Facies , Female , Gene-Environment Interaction , Genome-Wide Association Study , Humans , Infant , Intellectual Disability/genetics , Male , Middle Aged , Neural Cell Adhesion Molecules , Penetrance , Phenotype , Schizophrenia/genetics , Young AdultABSTRACT
Interstitial deletions of 6q are associated with variable phenotypes, including growth retardation, dysmorphic features, upper limb malformations, and Prader-Willi (PW)-like features. Only a minority of cases in the literature have been characterized with high resolution techniques, making genotype-phenotype correlations difficult. We report 12 individuals with overlapping, 200-kb to 16.4-Mb interstitial deletions within 6q15q22.33 characterized by microarray-based comparative genomic hybridization to better correlate deletion regions with specific phenotypes. Four individuals have a PW-like phenotype, though only two have deletion of SIM1, the candidate gene for this feature. Therefore, other genes on 6q may contribute to this phenotype including multiple genes on 6q16 and our newly proposed candidate, the transcription cofactor gene VGLL2 on 6q22.2. Two individuals present with movement disorders as a major feature, and ataxia is present in a third. The 4.1-Mb 6q22.1q22.2 critical region for movement disorders includes the cerebellar-expressed candidate gene GOPC. Observed brain malformations include thick corpus callosum in two subjects, cerebellar vermal hypoplasia in two subjects, and cerebellar atrophy in one subject. Seven subjects' deletions overlap a ~250-kb cluster of four genes on 6q22.1 including MARCKS, HDAC2, and HS3ST5, which are involved in neural development. Two subjects have only this gene cluster deleted, and one deletion was apparently de novo, suggesting at least one of these genes plays an important role in development. Although the phenotypes associated with 6q deletions can vary, using overlapping deletions to delineate critical regions improves genotype-phenotype correlation for interstitial 6q deletions.
Subject(s)
Genetic Association Studies , Abnormalities, Multiple/genetics , Adult , Child , Child, Preschool , Chromosome Deletion , Chromosomes, Human, Pair 6/genetics , Computational Biology , Developmental Disabilities/genetics , Female , Humans , In Situ Hybridization, Fluorescence , Infant , Male , Microarray Analysis , Young AdultABSTRACT
Toll-like receptor 3 (TLR3) binds and signals in response to dsRNA and poly(I:C), a synthetic double stranded RNA analog. Activation of TLR3 triggers innate responses that may play a protective or detrimental role in viral infections or in immune-mediated inflammatory diseases through amplification of inflammation. Two monoclonal antibodies, CNTO4685 (rat anti-mouse TLR3) and CNTO5429 (CDRs from CNTO4685 grafted onto a mouse IgG1 scaffold) were generated and characterized. These mAbs bind the extracellular domain of mouse TLR3, inhibit poly(I:C)-induced activation of HEK293T cells transfected with mTLR3, and reduce poly(I:C)-induced production of CCL2 and CXCL10 by primary mouse embryonic fibroblasts. CNTO5429 decreased serum IL-6 and TNFα levels post-intraperitoneal poly(I:C) administration, demonstrating in vivo activity. In summary, specific anti-mTLR3 mAbs have been generated to assess TLR3 antagonism in mouse models of inflammation.
Subject(s)
Antibodies, Monoclonal/immunology , Poly I-C/immunology , Toll-Like Receptor 3/immunology , Animals , Cell Line , Cells, Cultured , Humans , Inflammation/immunology , Intracellular Space/immunology , Macrophages/immunology , Mice , Mice, Inbred C57BL , Toll-Like Receptor 3/geneticsABSTRACT
The Na+/K+- ATPase acts as an ion pump maintaining the essential plasma membrane potential in all mammalian cell types, and is essential for many cellular functions. There are four α isoforms (α1, α2, α3 and α4) with distinct expression patterns, kinetic properties and substrate affinity. The α2-isoform is encoded by ATP1A2 and evidence supports its utmost importance in Cl- homeostasis in neurons, and in the function of respiratory neurons at birth. Monallelic pathogenic variants in ATP1A2 are associated with familial hemiplegic migraine type 2 (FHM2) and on rare occasions with alternating hemiplegia of childhood 1 (AHC1). To date, no instances of biallelic loss of function variants have been reported in humans. However, Atp1a2 homozygous loss of function knockout mice (α2-/- mice) show severe motor deficits, with lack of spontaneous movements, and are perinatally lethal due to absent respiratory activity. In this report we describe three newborns from two unrelated families, who died neonatally, presenting in utero with an unusual form of fetal hydrops, seizures and polyhydramnios. At birth they had multiple joint contractures (e.g. arthrogryposis), microcephaly, malformations of cortical development, dysmorphic features and severe respiratory insufficiency. Biallelic loss of function variants in ATP1A2, predicted to be pathogenic were found on whole exome sequencing. We propose that this is a distinctive new syndrome caused by complete absence of Na+/K+- ATPase α2-isoform expression.
Subject(s)
Arthrogryposis/genetics , Hydrops Fetalis/genetics , Microcephaly/genetics , Migraine with Aura/genetics , Sodium-Potassium-Exchanging ATPase/genetics , Alleles , Animals , Arthrogryposis/pathology , Child , Female , Genetic Predisposition to Disease , Humans , Hydrops Fetalis/pathology , Infant, Newborn , Loss of Function Mutation/genetics , Male , Mice , Microcephaly/pathology , Migraine with Aura/pathology , Phenotype , Pregnancy , Protein Isoforms/genetics , Exome SequencingABSTRACT
The generation of anti-variable region monoclonal antibodies (mAbs) against therapeutic antibodies is essential in the pharmacokinetic/pharmacodynamic (PK/PD) assessments of the drugs in clinical study samples. Sandwich EIA and other methods are typically employed to achieve sensitivity and selectivity for the PK/PD analyses. These assays usually require generation of mAb reagents that bind specifically to the drug in non-competing pair combinations. Thus, large panels of anti-variable region mAbs must be generated in an expeditious manner to increase the probability of success. Herein we describe a novel immunization method that utilizes type 1 interferons (IFNs) as immunomodulators coupled with an agonistic anti-CD40 mAb as a B cell proliferative agent to drive immune responses. This novel protocol allows for rapid and robust mAb reagent generation without the use of conventional protein-denaturing adjuvants. The use of IFNs allowed for the generation of comparable and in some cases, increased numbers of anti-variable region mAbs in a dramatically shorter timeframe. This IFN based, immunostimulatory approach utilizing a non-denaturing adjuvant, likely presents conformational epitopes and may optimize the humoral response for the rapid generation of anti-variable region reagents to therapeutic mAb candidates.
Subject(s)
Adjuvants, Immunologic/pharmacology , Antibodies, Monoclonal/biosynthesis , Interferons/immunology , Animals , Antibodies, Monoclonal/immunology , B-Lymphocytes/immunology , Enzyme-Linked Immunosorbent Assay , Female , Mice , Mice, Inbred BALB C , Sensitivity and Specificity , Spleen/immunologyABSTRACT
Defective clearance of apoptotic cells can result in sustained inflammation and subsequent autoimmunity. Macrophages, the "professional phagocyte" of the body, are responsible for efficient, non-phlogistic, apoptotic cell clearance. Controlling phagocytosis of apoptotic cells by macrophages is an attractive therapeutic opportunity to ameliorate inflammation. Using high content imaging, we have developed a system for evaluating the effects of antibody treatment on apoptotic cell uptake in primary human macrophages by comparing the Phagocytic Index (PI) for each antibody. Herein we demonstrate the feasibility of evaluating a panel of antibodies of unknown specificities obtained by immunization of mice with primary human macrophages and show that they can be distinguished based on individual PI measurements. In this study ~50% of antibodies obtained enhance phagocytosis of apoptotic cells while approximately 5% of the antibodies in the panel exhibit some inhibition. Though the specificities of the majority of antibodies are unknown, two of the antibodies that improved apoptotic cell uptake recognize recombinant MerTK; a receptor known to function in this capacity in vivo. The agonistic impact of these antibodies on efferocytosis could be demonstrated without addition of either of the MerTK ligands, Gas6 or ProS. These results validate applying the mechanism of this fundamental biological process as a means for identification of modulators that could potentially serve as therapeutics. This strategy for interrogating macrophages to discover molecules regulating apoptotic cell uptake is not limited by access to purified protein thereby increasing the possibility of finding novel apoptotic cell uptake pathways.
Subject(s)
Antibodies/immunology , Apoptosis , Macrophages/immunology , Phagocytosis , Animals , Antibodies/classification , Cells, Cultured , Humans , Mice , Mice, Inbred BALB C , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/immunology , Receptor Protein-Tyrosine Kinases/genetics , Receptor Protein-Tyrosine Kinases/immunology , c-Mer Tyrosine KinaseABSTRACT
Most antigen-specific mouse antibodies have been derived by hybridoma technology, predominantly through use of the Balb/c strain. Much of the Balb/c germline repertoire of variable genes (V regions) is known. However, there is little information about the background expressed repertoire of IgG antibodies in mice, which reflects the baseline against which antigen-specific antibodies are generated through immunization. To assess this baseline repertoire, RNA was isolated from splenic B-cells enriched for expression of IgG from three mice. The RNA was individually amplified with three distinct PCR primer sets for comprehensive recovery of the heavy and light chain variable regions. Each PCR product was independently subjected to deep sequencing using 454 pyro-sequencing technology and analysed for redundancy, open reading frame, germline representation, and CDR3 sequence of the heavy chain variable region (VH CDR3) within and across the primer sets and mice. A highly skewed abundance of heavy and light chain variable gene usage was observed for all three primers in all three mice. While showing considerable overlap, there were differences among these profiles indicative of primer bias and animal-to-animal variation. VH CDR3 sequences were likewise highly skewed indicating that the heavy chain genes profiles substantially reflected individual antibodies. This observation was confirmed through analysis of randomly selected complete heavy chain variable sequences. However, there was very little redundancy in VH CDR3 sequences across the different mice. We conclude that the background IgG repertoire in young, unimmunized mice is highly skewed within individual mice and is diverse among them, a pattern similar to that observed in highly immunized mice.
Subject(s)
B-Lymphocytes/immunology , Immunoglobulin G/genetics , Immunoglobulin Heavy Chains/genetics , Immunoglobulin Variable Region/genetics , Single-Domain Antibodies/genetics , Animals , B-Lymphocytes/cytology , Base Sequence , High-Throughput Nucleotide Sequencing , Immunization , Immunoglobulin G/immunology , Immunoglobulin Heavy Chains/immunology , Immunoglobulin J-Chains/genetics , Immunoglobulin Variable Region/immunology , Mice , Mice, Inbred BALB C , RNA, Messenger/analysis , RNA, Messenger/genetics , Sequence Analysis, RNAABSTRACT
The chemokines CCL17 (TARC) and CCL22 (MDC) function through the same receptor, CCR4, but have been proposed to differentially affect the immune response. To better understand the role of the individual ligands, a panel of rat anti-mouse CCL17 surrogate antibodies was generated that can be used to differentiate CCL17 and CCL22 function in vitro and in vivo. We have successfully identified a panel of neutralizing antibodies by screening hybridomas for the ability to inhibit CCL17-mediated calcium mobilization. Chemotaxis in response to CCL17 is also inhibited, providing further evidence that the antibodies in this panel are antagonistic. Using a recombinant cell line expressing human CCR4, we show that the antibodies block ß-arrestin recruitment as evidence that the antibodies are specifically blocking CCL17 signaling through CCR4. The antibodies within this panel inhibit calcium mobilization with varying potency in the calcium flux assay, having apparent IC50 ranging from approximately 1 to >400 ng/mL. Although both CCL17 and CCL22 function through CCR4, only a single antibody was identified as having detectable binding to CCL22. This panel of CCL17-specific antibodies provides tools that can be used to differentiate CCL17 and CCL22 function through CCR4 interaction in vitro and in vivo.
Subject(s)
Antibodies, Neutralizing/immunology , Chemokine CCL17/immunology , Chemokine CCL22/immunology , Receptors, CCR4/immunology , Animals , Antibody Affinity/immunology , Arrestins/immunology , Cell Line , Chemotaxis/immunology , Humans , Rats , beta-ArrestinsABSTRACT
PURPOSE: In 1963, Charles Van Riper published "My Grandfather," a short reading passage that has evolved into a ubiquitous metric of reading ability and speech intelligibility. In this historical note, we describe several heretofore unacknowledged similarities between "The Grandfather Passage" (Darley, Aronson, & Brown, 1975) and a portion of The Valley of Fear (Conan Doyle, 1915/2006), the final novel of the Sherlock Holmes series. We also describe overlap between "My Grandfather" and "The Grandfather Passage." METHOD: We contrasted propositions within The Valley of Fear to "My Grandfather" and "The Grandfather Passage." We also compared the respective text strings using the Turnitin antiplagiarism software application (iParadigms, 2011). RESULTS: "My Grandfather" and "The Grandfather Passage" are nearly identical passages with 88% string overlap. In addition, both passages show similarities with text from The Valley of Fear. CONCLUSIONS: Darley et al. (1975) did not acknowledge Van Riper (1963) as the original author of "The Grandfather Passage." In addition to this citation oversight, neither Darley et al. nor Van Riper attributed Conan Doyle as original source material. We describe the colorful history of this passage that has seen a remarkable breadth of utility in speech and language sciences.
Subject(s)
Aphasia/diagnosis , Language Tests/history , Plagiarism , Speech Articulation Tests/history , Speech Intelligibility , Algorithms , History, 20th Century , Humans , Pattern Recognition, Automated , Psychometrics/history , Reading , SemanticsABSTRACT
ST2L is a transmembrane receptor that belongs to the IL-1 receptor family. The receptor is expressed on various cell types including Th2 cells, mast cells, basophils, growth-activated fibroblasts, and vascular endothelial cells. ST2L activation by its ligand IL-33 has been implicated in Th2-mediated immunity, inflammation, and allergic responses in vivo. Inhibition of ST2L activity can attenuate Th2-dominated immune responses such as lung eosinophilia, airway hyper-responsiveness, and arthritis in animal models. Here we report the generation and in vitro characterization of a panel of rat anti-mouse ST2L monoclonal antibodies. We demonstrate that the antibodies specifically bind to recombinant receptor protein and that a subset of the binders inhibits mouse ST2L activity in multiple in vitro assays. Four of the identified anti-mouse ST2L antibodies were shown to prevent IL-33 from binding to ST2L, down-regulate IL-33-induced NF-κB signaling, and neutralize the ability of IL-33 to stimulate mouse Th2 cell proliferation. The characterized monoclonal antibodies are important tools that will be used to study mouse ST2L receptor functionality in vivo.
Subject(s)
Antibodies, Monoclonal , Interleukins/immunology , Receptors, Interleukin-1/immunology , Recombinant Proteins/immunology , Signal Transduction/drug effects , Th2 Cells/drug effects , Animals , Antibodies, Monoclonal/biosynthesis , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/isolation & purification , Antibodies, Monoclonal/pharmacology , Antibody Specificity , Biotin/chemistry , Biotin/metabolism , Cell Proliferation/drug effects , Down-Regulation , HEK293 Cells , Humans , Hybridomas/immunology , Hybridomas/metabolism , Hypersensitivity/drug therapy , Hypersensitivity/immunology , Immunoconjugates/chemistry , Immunoconjugates/metabolism , Inflammation/drug therapy , Inflammation/immunology , Interleukin-1/immunology , Interleukin-33 , Interleukins/metabolism , Mice , Mice, Inbred BALB C , NF-kappa B/metabolism , Plasmids/genetics , Plasmids/metabolism , Rats , Receptors, Interleukin-1/genetics , Receptors, Interleukin-1/metabolism , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Signal Transduction/immunology , Th2 Cells/immunology , TransfectionABSTRACT
ABSTRACT Herein we describe the use of an agonistic anti-murine CD40 MAb as a B cell proliferative agent to enhance the generation of monoclonal antibodies (MAbs) in Balb/c mice. While hybridoma technology has been validated repeatedly over the decades, little work has been described to improve upon the overall numbers of in vivo B cells and specific antibodies obtained from a fusion. To begin to address this situation, strategies to boost B lymphocyte yields for hybridoma production were employed. Anti-CD40 agonist antibodies have been reported to activate and amplify human resting B lymphocytes in vitro, resulting in increased cell numbers available for the generation of human hybridomas or B cell clones. An agonistic anti-murine CD40 MAb was administered to immunized mice 3 days prior to splenic harvest, and B lymphocyte yields were found to be approximately 2-fold higher in treated animals when compared to untreated animals. Moreover, the resulting hybridoma fusions using lymphocytes from treated animals yielded 5- to 10-fold more antigen reactive hybrids when compared to untreated animals. This novel addition to conventional approaches utilizes the proliferative effects of agonistic anti-CD40 MAbs to markedly enhance monoclonal antibody generation.
Subject(s)
Antibodies, Monoclonal/immunology , CD40 Antigens/agonists , Hybridomas/immunology , Receptors, Antigen, B-Cell/immunology , Animals , B-Lymphocytes/cytology , B-Lymphocytes/immunology , CD40 Antigens/immunology , Cell Fusion , Female , Humans , Hybridomas/cytology , Immunization , Mice , Mice, Inbred BALB C , Spleen/cytology , Spleen/immunologyABSTRACT
We describe the cytogenetic diagnosis using BAC- and oligonucleotide microarrays of a 16-year-old Laotian-American female, who first presented at 2 1/2 years of age with microcephaly, developmental retardation, and skeletal abnormalities of the upper limb including mild syndactyly of the second and third and the third and fourth fingers, short middle phalanges and clinodactyly of the fifth digit at the distal interphalangel joint on both hands, and symphalangism of the metacarpal-phalangeal joints of the second and fifth digits bilaterally. Her lower limbs displayed symphalangism of the metatarsal-phalangeal joint of the second, third, and fourth digits on both feet, with fusion of the middle and distal phalanges of the second and fifth digits and hallux valgus bilaterally. G-banded chromosomal study at age 4 was normal. However, comparative genomic hybridization at age 15 with the Spectral Genomics 1 Mb Hu BAC array platform indicated a microdeletion involving two BAC clones, RP11-451F14 --> RP11-12N7 at 2q31.1. The maximal deletion on initial analysis comprised the HOXD cluster, which is implicated in limb development. Fluorescence in situ hybridization (FISH) using the RP11-451F14 probe confirmed the deletion. Both parents were negative for the deletion. Additional FISH using BAC RP11-387A1, covering the HOXD cluster, limited the maximal deletion to approximately 2.518 Mb, and excluded involvement of the HOXD cluster. The Agilent 44K and 244K platforms demonstrated a deletion of approximately 2,011,000 bp, which did not include the HOXD cluster. The malformations in our patient may be caused by deletion of a regulatory element far upstream of the HOXD cluster.