A complement C4-derived glycopeptide is a biomarker for PMM2-CDG.

BACKGROUNDDiagnosis of PMM2-CDG, the most common congenital disorder of glycosylation (CDG), relies on measuring carbohydrate-deficient transferrin (CDT) and genetic testing. CDT tests have false negatives and may normalize with age. Site-specific changes in protein N-glycosylation have not been reported in sera in PMM2-CDG.METHODSUsing multistep mass spectrometry-based N-glycoproteomics, we analyzed sera from 72 individuals to discover and validate glycopeptide alterations. We performed comprehensive tandem mass tag-based discovery experiments in well-characterized patients and controls. Next, we developed a method for rapid profiling of additional samples. Finally, targeted mass spectrometry was used for validation in an independent set of samples in a blinded fashion.RESULTSOf the 3,342 N-glycopeptides identified, patients exhibited decrease in complex-type N-glycans and increase in truncated, mannose-rich, and hybrid species. We identified a glycopeptide from complement C4 carrying the glycan Man5GlcNAc2, which was not detected in controls, in 5 patients with normal CDT results, including 1 after liver transplant and 2 with a known genetic variant associated with mild disease, indicating greater sensitivity than CDT. It was detected by targeted analysis in 2 individuals with variants of uncertain significance in PMM2.CONCLUSIONComplement C4-derived Man5GlcNAc2 glycopeptide could be a biomarker for accurate diagnosis and therapeutic monitoring of patients with PMM2-CDG and other CDGs.FUNDINGU54NS115198 (Frontiers in Congenital Disorders of Glycosylation: NINDS; NCATS; Eunice Kennedy Shriver NICHD; Rare Disorders Consortium Disease Network); K08NS118119 (NINDS); Minnesota Partnership for Biotechnology and Medical Genomics; Rocket Fund; R01DK099551 (NIDDK); Mayo Clinic DERIVE Office; Mayo Clinic Center for Biomedical Discovery; IA/CRC/20/1/600002 (Center for Rare Disease Diagnosis, Research and Training; DBT/Wellcome Trust India Alliance).

Older Age, Male Sex, and Early Start Time Lengthen the Recovery Room Stay Following Total Joint Arthroplasty in an Ambulatory Surgical Center.

Background: Total joint arthroplasty (TJA) performed in the ambulatory surgical center (ASC) has been shown to be safe and cost-effective for an expanding cohort of patients. As criteria for TJA in the ASC become less restrictive, data guiding the efficient use of ASC resources are crucial. Purpose: We sought to identify factors associated with length of stay in the recovery room after primary total hip arthroplasty (THA) and total knee arthroplasty (TKA) performed in the ASC. Methods: We conducted a retrospective review of 411 patients who underwent primary THA or TKA at our institution's ASC between November 2020 and March 2022. We collected patient demographics, perioperative factors, success of same-day discharge (SDD), and length of time in the recovery room. Results: Of 411 patients, 100% had successful SDD. The average length of time spent in recovery was 207 minutes (SD: 73.9 minutes). Predictors of longer time in recovery were increased age, male sex, and operative start time before 9:59 am. Body mass index, preoperative opioid use, Charlson Comorbidity Index, type of surgery (THA vs TKA), urinary retention risk, and type of anesthesia (spinal vs general) were not significant predictors of length of time in the recovery room. Conclusion: In this retrospective study, factors associated with increased length of time in the recovery room included older age, male sex, and operative start time before 9:59 am. Such factors may guide surgeons in determining the optimal order of cases for each day at the ASC, but further prospective studies should seek to confirm these observations.

Neurological manifestations in PMM2-congenital disorders of glycosylation (PMM2-CDG): Insights into clinico-radiological characteristics, recommendations for follow-up, and future directions.

PURPOSE: In the absence of prospective data on neurological symptoms, disease outcome, or guidelines for system specific management in phosphomannomutase 2-congenital disorders of glycosylation (PMM2-CDG), we aimed to collect and review natural history data. METHODS: Fifty-one molecularly confirmed individuals with PMM2-CDG enrolled in the Frontiers of Congenital Disorders of Glycosylation natural history study were reviewed. In addition, we prospectively reviewed a smaller cohort of these individuals with PMM2-CDG on off-label acetazolamide treatment. RESULTS: Mean age at diagnosis was 28.04 months. Developmental delay is a constant phenotype. Neurological manifestation included ataxia (90.2%), myopathy (82.4%), seizures (56.9%), neuropathy (52.9%), microcephaly (19.1%), extrapyramidal symptoms (27.5%), stroke-like episodes (SLE) (15.7%), and spasticity (13.7%). Progressive cerebellar atrophy is the characteristic neuroimaging finding. Additionally, supratentorial white matter changes were noted in adult age. No correlation was observed between the seizure severity and SLE risk, although all patients with SLE have had seizures in the past. "Off-label" acetazolamide therapy in a smaller sub-cohort resulted in improvement in speech fluency but did not show statistically significant improvement in objective ataxia scores. CONCLUSION: Clinical and radiological findings suggest both neurodevelopmental and neurodegenerative pathophysiology. Seizures may manifest at any age and are responsive to levetiracetam monotherapy in most cases. Febrile seizure is the most common trigger for SLEs. Acetazolamide is well tolerated.

PIGO-CDG: A case study with a new genotype, expansion of the phenotype, literature review, and nosological considerations.

The phosphatidylinositol glycan anchor biosynthesis class O protein (PIGO) enzyme is an important step in the biosynthesis of glycosylphosphatidylinositol (GPI), which is essential for the membrane anchoring of several proteins. Bi-allelic pathogenic variants in PIGO lead to a congenital disorder of glycosylation (CDG) characterized by global developmental delay, an increase in serum alkaline phosphatase levels, congenital anomalies including anorectal, genitourinary, and limb malformations in most patients; this phenotype has been alternately called "Mabry syndrome" or "hyperphosphatasia with impaired intellectual development syndrome 2." We report a 22-month-old female with PIGO deficiency caused by novel PIGO variants. In addition to the Mabry syndrome phenotype, our patient's clinical picture was complicated by intermittent hypoglycemia with signs of functional hyperinsulinism, severe secretory diarrhea, and osteopenia with a pathological fracture, thus, potentially expanding the known phenotype of this disorder, although more studies are necessary to confirm these associations. We also provide an updated review of the literature, and propose unifying the nomenclature of PIGO deficiency as "PIGO-CDG," which reflects its pathophysiology and position in the broad scope of metabolic disorders and congenital disorders of glycosylation.

The Effect of Preoperative Exposure to Benzodiazepines on Opioid Consumption After One and Two-level Anterior Cervical Discectomy and Fusion.

STUDY DESIGN: Retrospective cohort. OBJECTIVE: Investigate the relationship between preoperative benzodiazepine exposure and postoperative opioid use in patients undergoing primary 1 or 2-level anterior cervical discectomy and fusion (ACDF). BACKGROUND: Little is known about the effect of preoperative benzodiazepine exposure on postoperative opioid use in spine surgery. PATIENTS AND METHODS: Patients undergoing primary 1 or 2-level ACDF at a single institution from February 2020 to November 2021 were identified through electronic medical records. The prescription drug monitoring program was utilized to record the name, dosage, and quantity of preoperative benzodiazepines/opioids filled within 60 days before surgery and postoperative opioids 6 months after surgery. Patients were classified as benzodiazepine naïve or exposed according to preoperative usage, and postoperative opioid dose and duration were compared between groups. Regression analysis was performed for outcomes that demonstrated statistical significance, adjusting for preoperative opioid use, age, sex, and body mass index. RESULTS: Sixty-seven patients comprised the benzodiazepine-exposed group whereas 90 comprised the benzodiazepine-naïve group. There was no significant difference in average daily morphine milligram equivalents between groups (median: 96.0 vs 65.0, P = 0.11). The benzodiazepine-exposed group received postoperative opioids for a longer duration (median: 32.0 d vs 12.0 d, P = 0.004) with more prescriptions (median: 2.0 vs 1.0, P = 0.004) and a greater number of pills (median: 110.0 vs 59.0, P = 0.007). On regression analysis, preoperative benzodiazepine use was not significantly associated with postoperative opioid duration [incidence rate ratio (IRR): 0.93, P = 0.74], number of prescriptions (IRR: 1.21, P = 0.16), or number of pills (IRR: 0.89, P = 0.58). CONCLUSIONS: While preoperative benzodiazepine users undergoing primary 1 or 2-level ACDF received postoperative opioids for a longer duration compared with a benzodiazepine naïve cohort, preoperative benzodiazepine use did not independently contribute to this observation. These findings provide insight into the relationship between preoperative benzodiazepine use and postoperative opioid consumption. LEVEL OF EVIDENCE: Level III.

The Complication of a Retained Broken Distal Interlocking Screw Within a Cephalomedullary Nail During Conversion Hip Arthroplasty: A Case Report.

In patients who undergo femoral fracture fixation with a cephalomedullary nail, the breakage of one or more of the distal interlocking screws is a well-described phenomenon. The presence of a broken interlocking screw in patients who require the removal of their cephalomedullary nail presents a unique challenge. The broken interlocking screw may be retrieved, or the screw may be retained if it is not engaged within the nail and the nail can safely be removed while leaving the broken screw fragment behind. We report a hip conversion arthroplasty case with a broken interlocking screw where the nail was removed with ease and the broken screw was assumed to have been left behind. Cerclage wires were placed for an apparent proximal femoral fracture. Postoperative X-rays demonstrated a large lucency tracking from the prior location of the distal interlocking screw to the calcar region. This finding made it evident that the broken screw had been retained in the nail and was dragged up the femur upon nail removal, causing a large gouge spanning the entire femur.

Fractionated plasma N-glycan profiling of novel cohort of ATP6AP1-CDG subjects identifies phenotypic association.

ATP6AP1-CDG is an X-linked disorder typically characterized by hepatopathy, immunodeficiency, and an abnormal type II transferrin glycosylation pattern. Here, we present 11 new patients and clinical updates with biochemical characterization on one previously reported patient. We also document intrafamilial phenotypic variability and atypical presentations, expanding the symptomatology of ATP6AP1-CDG to include dystonia, hepatocellular carcinoma, and lysosomal abnormalities on hepatic histology. Three of our subjects received successful liver transplantation. We performed N-glycan profiling of total and fractionated plasma proteins for six patients and show associations with varying phenotypes, demonstrating potential diagnostic and prognostic value of fractionated N-glycan profiles. The aberrant N-linked glycosylation in purified transferrin and remaining plasma glycoprotein fractions normalized in one patient post hepatic transplant, while the increases of Man4GlcNAc2 and Man5GlcNAc2 in purified immunoglobulins persisted. Interestingly, in the single patient with isolated immune deficiency phenotype, elevated high-mannose glycans were detected on purified immunoglobulins without glycosylation abnormalities on transferrin or the remaining plasma glycoprotein fractions. Given the diverse and often tissue specific clinical presentations and the need of clinical management post hepatic transplant in ATP6AP1-CDG patients, these results demonstrate that fractionated plasma N-glycan profiling could be a valuable tool in diagnosis and disease monitoring.

Utilizing augmented artificial intelligence for aminoacidopathies using collaborative laboratory integrated reporting- A cross-sectional study.

Introduction: Plasma amino acids profiling can aid in the screening and diagnosis of aminoacidopathies. The goal of the current study was to analyze and report the metabolic profiles of plasma amino acid (PAA) and additionally to compare PAA-reference intervals (RI) from Pakistan with more countries utilizing Clinical Laboratory Integrated Reports (CLIR). Methods: This was a cross sectional prospective single center study. Twenty-two amino acids were analyzed in each sample received for one year at the clinical laboratory. Data was divided into reference and case data files after interpretation by a team of pathologists and technologists. All PAA samples were analyzed using ion-exchange high-performance chromatography. The CLIR application of Amino Acid in Plasma (AAQP) was used for statistical analysis for both data sets and post-analytical interpretive tools using a single condition tool was applied. Result: The majority of 92% (n = 1913) of PAA profiles out of the total 2081 tests run were non-diagnostic; the PAA values were within the age-specific RI. The PAA median was in close comparison close to the 50th percentile of reference data available in CLIR software. Out of the total 2081 tests run, one hundred and sixty-eight had abnormal PAA levels; 27.38% were labeled as non-fasting samples, and the main aminoacidopathies identified were Phenylketonuria and Maple Syrup Urine Disorder. Conclusion: An agreement of >95% was observed between the reporting done by the pathologists and technologists' team and then after the application of CLIR. Augmented artificial intelligence using CLIR can improve the accuracy of reporting rare aminoacidopathies in a developing country like ours.

The effect of long-term opioid use on back-specific disability and health-related quality of life in patients with chronic low back pain.

CONTEXT: Opioids are commonly utilized for the treatment of chronic pain. However, research regarding the long-term (≥12 months) outcomes of opioid therapy remains sparse. OBJECTIVES: This study aims to evaluate the effects of long-term opioid therapy on measures of back-specific disability and health-related quality of life in patients with chronic low back pain. METHODS: In this retrospective cohort study, patients with chronic low back pain who reported consistent opioid use or abstinence for at least 12 months while enrolled in the Pain Registry for Epidemiological, Clinical, and Interventional Studies and Innovation Pain Research Registry were classified as long-term opioid users or nonusers, respectively. For comparison, intermediate-term and short-term opioid users and nonusers were also identified. Multiple linear regression analysis was performed to compare back-specific disability (Roland-Morris Disability Questionnaire [RMDQ]) and health-related quality of life (29-item Patient-Reported Outcomes Measurement Information System [PROMIS]) between opioid users and nonusers while controlling for pain intensity, depression, age, body mass index (BMI), and eight common comorbid conditions (herniated disc, sciatica, osteoporosis, osteoarthritis, heart disease, hypertension, diabetes, and asthma). Statistically significant findings were assessed for clinical relevance. RESULTS: There were 96 long-term opioid users and 204 long-term opioid nonusers. After controlling for potential confounders, long-term opioid use was a predictor of worse back-specific disability (adjusted mean difference=2.85, p<0.001), physical function (adjusted mean difference=-2.90, p=0.001), fatigue (adjusted mean difference=4.32, p=0.001), participation in social roles (adjusted mean difference=-4.10, p<0.001), and pain interference (adjusted mean difference=3.88, p<0.001) outcomes. Intermediate-term opioid use was a predictor of worse back-specific disability (adjusted mean difference=2.41, p<0.001), physical function (adjusted mean difference=-2.26, p=0.003), fatigue (adjusted mean difference=3.70, p=0.002), and sleep disturbance outcomes (adjusted mean difference=3.03, p=0.004), whereas short-term opioid use was a predictor of worse back-specific disability (adjusted mean difference=2.42, p<0.001) and physical function outcomes (adjusted mean difference=-1.90, p<0.001). CONCLUSIONS: The findings of this study are largely consistent with existing literature regarding the outcomes of long-term opioid therapy. Taken in conjunction with the well-established risks of opioid medications, these findings draw into question the utility of long-term opioid therapy for chronic low back pain.

Medial Clavicle Fracture Fixation Including the Sternum: A Case Report.

CASE: A 29-year-old man presented with a displaced medial clavicle fracture. Surgical repair was performed using a precontoured plate designed for the contralateral distal clavicle, and medial fixation was accomplished at the sternum. The patient had no complications and demonstrated full strength and range of motion at the 8-month follow-up. CONCLUSION: Medial clavicle fractures with a small medial fragment can be immobilized using a plate designed for the contralateral distal clavicle that crosses the sternoclavicular joint to obtain medial fixation in the sternum. This technique may provide a viable treatment modality for this unique fracture pattern.

De novo and bi-allelic variants in AP1G1 cause neurodevelopmental disorder with developmental delay, intellectual disability, and epilepsy.

Adaptor protein (AP) complexes mediate selective intracellular vesicular trafficking and polarized localization of somatodendritic proteins in neurons. Disease-causing alleles of various subunits of AP complexes have been implicated in several heritable human disorders, including intellectual disabilities (IDs). Here, we report two bi-allelic (c.737C>A [p.Pro246His] and c.1105A>G [p.Met369Val]) and eight de novo heterozygous variants (c.44G>A [p.Arg15Gln], c.103C>T [p.Arg35Trp], c.104G>A [p.Arg35Gln], c.229delC [p.Gln77Lys∗11], c.399_400del [p.Glu133Aspfs∗37], c.747G>T [p.Gln249His], c.928-2A>C [p.?], and c.2459C>G [p.Pro820Arg]) in AP1G1, encoding gamma-1 subunit of adaptor-related protein complex 1 (AP1γ1), associated with a neurodevelopmental disorder (NDD) characterized by mild to severe ID, epilepsy, and developmental delay in eleven families from different ethnicities. The AP1γ1-mediated adaptor complex is essential for the formation of clathrin-coated intracellular vesicles. In silico analysis and 3D protein modeling simulation predicted alteration of AP1γ1 protein folding for missense variants, which was consistent with the observed altered AP1γ1 levels in heterologous cells. Functional studies of the recessively inherited missense variants revealed no apparent impact on the interaction of AP1γ1 with other subunits of the AP-1 complex but rather showed to affect the endosome recycling pathway. Knocking out ap1g1 in zebrafish leads to severe morphological defect and lethality, which was significantly rescued by injection of wild-type AP1G1 mRNA and not by transcripts encoding the missense variants. Furthermore, microinjection of mRNAs with de novo missense variants in wild-type zebrafish resulted in severe developmental abnormalities and increased lethality. We conclude that de novo and bi-allelic variants in AP1G1 are associated with neurodevelopmental disorder in diverse populations.

UBA2 variants underlie a recognizable syndrome with variable aplasia cutis congenita and ectrodactyly.

PURPOSE: The human chromosome 19q13.11 deletion syndrome is associated with a variable phenotype that includes aplasia cutis congenita (ACC) and ectrodactyly as specific features. UBA2 (ubiquitin-like modifier-activating enzyme 2) lies adjacent to the minimal deletion overlap region. We aimed to define the UBA2-related phenotypic spectrum in humans and zebrafish due to sequence variants and to establish the mechanism of disease. METHODS: Exome sequencing was used to detect UBA2 sequence variants in 16 subjects in 7 unrelated families. uba2 loss of function was modeled in zebrafish. Effects of human missense variants were assessed in zebrafish rescue experiments. RESULTS: Seven human UBA2 loss-of-function and missense sequence variants were detected. UBA2-phenotypes included ACC, ectrodactyly, neurodevelopmental abnormalities, ectodermal, skeletal, craniofacial, cardiac, renal, and genital anomalies. uba2 was expressed in zebrafish eye, brain, and pectoral fins; uba2-null fish showed deficient growth, microcephaly, microphthalmia, mandibular hypoplasia, and abnormal fins. uba2-mRNAs with human missense variants failed to rescue nullizygous zebrafish phenotypes. CONCLUSION: UBA2 variants cause a recognizable syndrome with a wide phenotypic spectrum. Our data suggest that loss of UBA2 function underlies the human UBA2 monogenic disorder and highlights the importance of SUMOylation in the development of affected tissues.

Immune dysfunction in MGAT2-CDG: A clinical report and review of the literature.

Glycosylation is a critical post/peri-translational modification required for the appropriate development and function of the immune system. As an example, abnormalities in glycosylation can cause antibody deficiency and reduced lymphocyte signaling, although the phenotype can be complex given the diverse roles of glycosylation. Human MGAT2 encodes N-acetylglucosaminyltransferase II, which is a critical enzyme in the processing of oligomannose to complex N-glycans. Complex N-glycans are essential for immune system functionality, but only one individual with MGAT2-CDG has been described to have an abnormal immunologic evaluation. MGAT2-CDG (CDG-IIa) is a congenital disorder of glycosylation (CDG) associated with profound global developmental disability, hypotonia, early onset epilepsy, and other multisystem manifestations. Here, we report a 4-year old female with MGAT2-CDG due to a novel homozygous pathogenic variant in MGAT2, a 4-base pair deletion, c.1006_1009delGACA. In addition to clinical features previously described in MGAT2-CDG, she experienced episodic asystole, persistent hypogammaglobulinemia, and defective ex vivo mitogen and antigen proliferative responses, but intact specific vaccine antibody titers. Her infection history has been mild despite the testing abnormalities. We compare this patient to the 15 previously reported patients in the literature, thus expanding both the genotypic and phenotypic spectrum for MGAT2-CDG.

Osteopathic Manipulation in the Management of Chronic Pain: Current Perspectives.

Chronic pain is a common condition that often interferes with work or other activities. Guidelines support the use of non-pharmacological treatments, such as spinal manipulation, in patients with chronic pain. Osteopathic physicians in the United States are uniquely positioned to manage chronic pain because their professional philosophy embraces the biopsychosocial model and they are trained in the use of osteopathic manipulative treatment (OMT) to complement conventional medical care. This narrative review provides current perspectives on the osteopathic approach to chronic pain management, including evidence for the efficacy of OMT based on systematic searches of the biomedical literature and the ClinicalTrials.gov database. Men, persons with low levels of education, and non-White and Hispanic patients are significantly less likely to have received OMT during their lifetime. Patients with low back and neck pain are most likely to be treated with OMT, and osteopathic manipulative medicine specialty physicians and family medicine physicians most often use OMT. However, many osteopathic physicians report using OMT infrequently. Although OMT is considered safe, based on millions of patient encounters over more than a century, there is limited evidence on its efficacy in treating chronic pain. The lone exception involves chronic low back pain, wherein there is evidence from systematic reviews, a large clinical trial, and observational studies. There is lesser evidence to support cost effectiveness and patient satisfaction associated with OMT for chronic pain. The only clinical practice guideline established by the American Osteopathic Association recommends that OMT should be used to treat chronic low back pain in patients with somatic dysfunction. Given the philosophy of osteopathic medicine, universal training of osteopathic physicians to use OMT, and national guidelines supporting non-pharmacological treatments for chronic pain, it is unclear why OMT use is reported to be remarkably low in physician surveys.

Laboratory monitoring of patients with hereditary tyrosinemia type I.

BACKGROUND: The prognosis of patients with Hereditary Tyrosinemia Type 1 (HT-1) has greatly improved with early detection through newborn screening and the introduction of nitisinone (NTBC) therapy. A recent guideline calls for periodic monitoring of biochemical markers and NTBC levels to tailor treatment; however, this is currently only achieved through a combination of clinical laboratory tests. We developed a multiplexed assay measuring relevant amino acids, succinylacetone (SUAC), and NTBC in dried blood spots (DBS) to facilitate treatment monitoring. METHODS: Tyrosine, phenylalanine, methionine, NTBC and SUAC were eluted from DBS with methanol containing internal standards for each analyte and analyzed by liquid chromatography tandem mass spectrometry over 6.5 min in the multiple reaction monitoring positive mode. RESULTS: Pre-analytical and analytical factors were studied and demonstrated a reliable assay. Chromatography resolved an unknown substance that falsely elevates SUAC concentrations and was present in all samples. To establish control and disease ranges, the method was applied to DBS collected from controls (n = 284) and affected patients before (n = 2) and after initiation of treatment (n = 29). In the treated patients SUAC concentrations were within the normal range over a wide range of NTBC levels. CONCLUSIONS: This assay enables combined, accurate measurement of revelevant metabolites and NTBC in order to simplify treatment monitoring of patients with HT-1. In addition, the use of DBS allows for specimen collection at home to facilitate more standardization in relation to drug and dietary treatment.

Expanding the clinical and phenotypic heterogeneity associated with biallelic variants in ACO2.

OBJECTIVE: We describe the clinical characteristics and genetic etiology of several new cases within the ACO2-related disease spectrum. Mitochondrial aconitase (ACO2) is a nuclear-encoded tricarboxylic acid cycle enzyme. Homozygous pathogenic missense variants in the ACO2 gene were initially associated with infantile degeneration of the cerebrum, cerebellum, and retina, resulting in profound intellectual and developmental disability and early death. Subsequent studies have identified a range of homozygous and compound heterozygous pathogenic missense, nonsense, frameshift, and splice-site ACO2 variants in patients with a spectrum of clinical manifestations and disease severities. METHODS: We describe a cohort of five novel patients with biallelic pathogenic variants in ACO2. We review the clinical histories of these patients as well as the molecular and functional characterization of the associated ACO2 variants and compare with those described previously in the literature. RESULTS: Two siblings with relatively mild symptoms presented with episodic ataxia, mild developmental delays, severe dysarthria, and behavioral abnormalities including hyperactivity and depressive symptoms with generalized anxiety. One patient presented with the classic form with cerebellar hypoplasia, ataxia, seizures, optic atrophy, and retinitis pigmentosa. Another unrelated patient presented with ataxia but developed severe progressive spastic quadriplegia. Another patient demonstrated a spinal muscular atrophy-like presentation with severe neonatal hypotonia, diminished reflexes, and poor respiratory drive, leading to ventilator dependence until death at the age of 9 months. INTERPRETATION: In this study, we highlight the importance of recognizing milder forms of the disorder, which may escape detection due to atypical disease presentation.

Detection of cryptic CCND1 rearrangements in mantle cell lymphoma by next generation sequencing.

The accurate detection of recurrent genetic abnormalities for most hematologic neoplasms is critical for diagnosis, prognosis and/or treatment. Rearrangements involving CCND1 are observed in a subset of mature B-cell neoplasms and can be reliably detected by fluorescence in situ hybridization (FISH) in most cases. However, cryptic and complex chromosomal rearrangements may pose a technical challenge for accurate diagnosis. Herein, we describe two patients with suspected mantle cell lymphoma that lacked obvious CCND1 rearrangements by FISH studies. A next generation sequencing (NGS) based assay, mate-pair sequencing (MPseq), was utilized in each case to investigate potential cryptic CCND1 rearrangements and revealed cryptic insertional events resulting in CCND1/IGH and CCND1/IGK rearrangements. These cases demonstrate that NGS-based assays, including MPseq, are a powerful approach to identify cryptic rearrangements of clinical importance that are not detected by current clinical genomics evaluation.

Targeted gene approach with biochemical assay confirms ABCD1 mutation of X-linked adrenoleukodystrophy in a 62-year-old man with gait imbalance.

X-linked adrenoleukodystrophy is a peroxisomal disorder caused by a mutation in ABCD1 gene. The three main phenotypes of X-linked adrenoleukodystrophy include cerebral adrenoleukodystrophy, adrenomyeloneuropathy, and isolated primary adrenal insufficiency. More than 750 non-recurrent mutations exist throughout the coding region of the ABCD1 gene. We report a 62-year-old man with a 17-year history of progressive gait imbalance and numb feet. He had noted difficulty rising from a chair for 3 years. Examination revealed proximal lower limb weakness and length-dependent sensory loss with preservation of reflexes and unilateral Babinski sign. Electrodiagnostic evaluation confirmed a length-dependent sensorimotor peripheral neuropathy and proximal myopathy. Family history was remarkable for similar symptoms in 6 siblings. A targeted gene approach for 102 known peripheral neuropathy genes led to discovery of ABCD1 mutation confirmed by kindred evaluation and biochemical assay. This case highlights the importance of combining targeted gene approaches with functional assay confirmation especially for atypical clinical presentations.

The Tumor-Associated Glycosyltransferase ST6Gal-I Regulates Stem Cell Transcription Factors and Confers a Cancer Stem Cell Phenotype.

The glycosyltransferase ST6Gal-I, which adds α2-6-linked sialic acids to substrate glycoproteins, has been implicated in carcinogenesis; however, the nature of its pathogenic role remains poorly understood. Here we show that ST6Gal-I is upregulated in ovarian and pancreatic carcinomas, enriched in metastatic tumors, and associated with reduced patient survival. Notably, ST6Gal-I upregulation in cancer cells conferred hallmark cancer stem-like cell (CSC) characteristics. Modulating ST6Gal-I expression in pancreatic and ovarian cancer cells directly altered CSC spheroid growth, and clonal variants with high ST6Gal-I activity preferentially survived in CSC culture. Primary ovarian cancer cells from patient ascites or solid tumors sorted for α2-6 sialylation grew as spheroids, while cells lacking α2-6 sialylation remained as single cells and lost viability. ST6Gal-I also promoted resistance to gemcitabine and enabled the formation of stably resistant colonies. Gemcitabine treatment of patient-derived xenograft tumors enriched for ST6Gal-I-expressing cells relative to pair-matched untreated tumors. ST6Gal-I also augmented tumor-initiating potential. In limiting dilution assays, subcutaneous tumor formation was inhibited by ST6Gal-I knockdown, whereas in a chemically induced tumor initiation model, mice with conditional ST6Gal-I overexpression exhibited enhanced tumorigenesis. Finally, we found that ST6Gal-I induced expression of the key tumor-promoting transcription factors, Sox9 and Slug. Collectively, this work highlighted a previously unrecognized role for a specific glycosyltransferase in driving a CSC state. Cancer Res; 76(13); 3978-88. ©2016 AACR.