A Brazilian Rare-Disease Center's Experience with Glucosylsphingosine (lyso-Gb1) in Patients with Gaucher Disease: Exploring a Novel Correlation with IgG Levels in Plasma and a Biomarker Measurement in CSF.

Gaucher disease (GD, OMIM 230800) is one of the most common lysosomal disorders, being caused by the deficient activity of the enzyme acid ß-glucocerebrosidase (Gcase). Three clinical forms of Gaucher's disease (GD) are classified based on neurological involvement. Type 1 (GD1) is non-neuronopathic, while types 2 (GD2) and 3 (GD3) are neuronopathic forms. Gcase catalyzes the conversion of glucosylceramide (GlcCer) into ceramide and glucose. As GlcCer accumulates in lysosomal macrophages, it undergoes deacylation to become glycosylsphingosine (lyso-Gb1), which has shown to be a useful and reliable biomarker for the diagnosis and monitoring of treated and untreated patients with GD. Multiple myeloma (MM) is one of the leading causes of cancer-related death among patients with GD and monoclonal gammopathy of undetermined significance (MGUS) is a non-neoplastic condition that can be a telltale sign of a B clonal proliferation caused by the chronic activation of B cells. This study aimed to quantify Lyso-Gb1 levels in dried blood spots (DBS) and cerebrospinal fluid (CSF) as biomarkers for Gaucher disease (GD) and discuss the association of this biomarker with other clinical parameters. This is a mixed-methods study incorporating both cross-sectional and longitudinal elements within a cohort design with a convenience-sampling strategy. Data collection took place from January 2012 to March 2023. Lyso-Gb1 extraction from DBS involved the use of a methanol-acetonitrile-water mixture, followed by incubation and centrifugation. Analysis was performed using UPLC-MS/MS with MassLynx software version 4.2 and the control group for the DBS measurements included general newborns. CSF Lyso-Gb1 was extracted using ethyl acetate, analyzed by UPLC-MS/MS with a calibration curve, and expressed in pmol/L. Lysosomal activity in CSF was assessed by measuring chitotriosidase (Cht), and other lysosomal enzyme activities were assessed as previously described in the literature. Patients with metachromatic leukodystrophy (MLD) were used as controls. Thirty-two treated patients (twenty-nine GD1 and three GD3, all on ERT except for one GD type on SRT with eliglustat) and three untreated patients (one GD1, one GD2, and one GD3) were included. When analyzing only the treated GD1 group, a significant correlation was found between lyso-Gb1 and age (rho = -0.447, p = 0.001), ChT, and IgG levels (rho = 0.73, p < 0.001; and rho = 0.36, p = 0.03, respectively). Five GD1 patients (three females, mean age 40 years) also had their CSF collected and analyzed. The average measurement of lyso-Gb1 in CSF was 94 pmol/L (range: 57.1-157.9 pmol/L) versus <6.2 pmol/L in the control group (MLD). This is the first time, to the best of our knowledge, that lyso-Gb1 has been associated with IgG levels. While this finding reflects a risk for MGUS or MM and not only chronic plasma B-cell activation, it still requires further studies. Moreover, the analysis of CSF lyso-Gb1 levels in GD1 patients was demonstrated to be significantly higher than the control group. This raises the hypothesis that CSF lyso-Gb1 may serve as a valuable indicator for neurological involvement in GD, providing insights into the potential implications for neurological manifestations in GD, including GD1. The correlation between lyso-Gb1 and ChT levels in treated GD1 patients further underscores the interconnectedness of lysosomal markers and their relevance in monitoring.

Clinical and Functional Consequences of C-Terminal Variants in MCT8: A Case Series.

CONTEXT: Genetic variants in SLC16A2, encoding the thyroid hormone transporter MCT8, can cause intellectual and motor disability and abnormal serum thyroid function tests, known as MCT8 deficiency. The C-terminal domain of MCT8 is poorly conserved, which complicates prediction of the deleteriousness of variants in this region. We studied the functional consequences of 5 novel variants within this domain and their relation to the clinical phenotypes. METHODS: We enrolled male subjects with intellectual disability in whom genetic variants were identified in exon 6 of SLC16A2. The impact of identified variants was evaluated in transiently transfected cell lines and patient-derived fibroblasts. RESULTS: Seven individuals from 5 families harbored potentially deleterious variants affecting the C-terminal domain of MCT8. Two boys with clinical features considered atypical for MCT8 deficiency had a missense variant [c.1724A>G;p.(His575Arg) or c.1796A>G;p.(Asn599Ser)] that did not affect MCT8 function in transfected cells or patient-derived fibroblasts, challenging a causal relationship. Two brothers with classical MCT8 deficiency had a truncating c.1695delT;p.(Val566*) variant that completely inactivated MCT8 in vitro. The 3 other boys had relatively less-severe clinical features and harbored frameshift variants that elongate the MCT8 protein [c.1805delT;p.(Leu602HisfsTer680) and c.del1826-1835;p.(Pro609GlnfsTer676)] and retained ~50% residual activity. Additional truncating variants within transmembrane domain 12 were fully inactivating, whereas those within the intracellular C-terminal tail were tolerated. CONCLUSIONS: Variants affecting the intracellular C-terminal tail of MCT8 are likely benign unless they cause frameshifts that elongate the MCT8 protein. These findings provide clinical guidance in the assessment of the pathogenicity of variants within the C-terminal domain of MCT8.

Retraction notice to Plexin-B1 and Semaphorin 4D Cooperate to Promote Perineural Invasion in a RhoA/ROK-Dependent Manner Am J Pathol 180 (2012) 1232-1242.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article is being retracted following correspondence from the Office of Accountability and Compliance at the University of Maryland, Baltimore. An internal investigation into this manuscript by the University of Maryland, Baltimore, found evidence that there are errors with the presentation of the standard deviations and statistical significance shown in Figure 6 which are not supported by the original data, and that these inaccuracies warrant retraction to correct the scientific record. Despite extensive efforts, the journal was unable to contact Dr. Ying-hua Yang and Dr. Hua Zhou with regard to this retraction.

Immunohistochemical analysis of TIMP-3 and MMP-9 in actinic keratosis, squamous cell carcinoma of the skin, and basal cell carcinoma.

The expression of metalloproteinases and their inhibitors has been related to different invasive and metastatic potentials in cancer. This study aims to investigate the immunohistochemical expression of TIMP-3 and MMP-9 in samples of basal cell carcinoma (BCC), squamous cell carcinoma of the skin (SCC), and actinic keratosis (AK). Immunohistochemistry was performed to evaluate the expression of TIMP-3 and MMP-9 in samples of BCC (n=22), SCC (n=10), and AK (n=15). Ten fields of both tumor parenchyma and tumor stroma were photographed and counted in image software. The ratio of positive cells to total cells was used to quantify the staining. A higher expression of MMP-9 was found in tumor stroma of SCC compared to BCC and AK. No significant differences in TIMP-3 expression were observed among the groups. Considering the well-described differences between these neoplasms, these results provide additional evidence of the role of MMP-9 in tumor invasiveness of keratinocyte-derived tumors.

Plexin-B1 and semaphorin 4D cooperate to promote perineural invasion in a RhoA/ROK-dependent manner.

Perineural invasion (PNI) is a tropism of tumor cells for nerve bundles located in the surrounding stroma. It is a pathological feature observed in certain tumors, referred to as neurotropic malignancies, that severely limits the ability to establish local control of disease and results in pain, recurrent growth, and distant metastases. Despite the importance of PNI as a prognostic indicator, its biological mechanisms are poorly understood. The semaphorins and their receptors, the plexins, compose a family of proteins originally shown to be important in nerve cell adhesion, axon migration, and proper central nervous system development. Emerging evidence has demonstrated that these factors are expressed in tissues outside of the nervous system and represent a widespread signal transduction system that is involved in the regulation of motility and adhesion in different cell types. We believe that the plexins and semaphorins, which are strongly expressed in both axons and many carcinomas, play a role in PNI. In this study, we show that plexin-B1 is overexpressed in tissues and cell lines from neurotropic malignancies and is attracted to nerves that express its ligand, semaphorin 4D, in a Rho/Rho kinase-dependent manner. We also demonstrate that nerves are attracted to tumors through this same system of proteins, suggesting that both plexin-B1 and semaphorin 4D are important in the promotion of PNI.