Comparative genomic characterization of melanoma of known and unknown primary

BackgroundThis study aims to genomically characterize melanoma of unknown primary (MUP) in comparison to melanomas of cutaneous primary (MCP).MethodsEligible cases were collected from the MSK-IMPACT™ Clinical Sequencing Cohort published in the cBioPortal database. Genomic analysis was performed using a hybridization-capture-based next-generation sequencing assay designed to detect mutations, small insertions and deletions, copy number alterations, and genomic rearrangements.ResultsAmong 462 patients of whom 18.4% had MUP, brain metastasis was more common among patients with MUP (23% vs 7.1%). The differences in genomic profiling between MCP and MUP did not reach statistical significance. The 187 MCP and 44 MUP patients treated with immune checkpoint inhibitors had a median overall survival of 49 and 44 months, respectively (p = 0.705).ConclusionsThe differences in somatic mutation patterns and survival outcomes were not statistically significant. These findings may allude to similar carcinogenic processes but should be considered exploratory and interpreted with caution. (AU)

Identification of a novel nonsense variant in FYCO1 gene associated with infantile cataract and cortical atrophy.

INTRODUCTION: Cataract is a major condition characterized by ocular lens opacification, resulting from alteration in the lens architecture, lens proteins or both. It is responsible for about one-third of infants' blindness worldwide. Variants in the FYCO1 gene have been associated with autosomal recessive infantile cataract. MATERIAL AND METHODS: We conducted whole exome sequencing (WES) in a nine months old male patient who was referred for genetic investigation because of infantile cataract. WES analysis revealed the presence of a homozygous pathogenic variant (c.2365C>T) in exon 8 of the FYCO1 gene. RESULTS AND DISCUSSION: This is the first report on a Lebanese infant with infantile cataract and cortical atrophy which was not previously reported, resulting from a novel homozygous FYCO1 variant; thus expanding the clinical phenotypic spectrum of FYCO1 involvement.

Comparative genomic characterization of melanoma of known and unknown primary.

BACKGROUND: This study aims to genomically characterize melanoma of unknown primary (MUP) in comparison to melanomas of cutaneous primary (MCP). METHODS: Eligible cases were collected from the MSK-IMPACT™ Clinical Sequencing Cohort published in the cBioPortal database. Genomic analysis was performed using a hybridization-capture-based next-generation sequencing assay designed to detect mutations, small insertions and deletions, copy number alterations, and genomic rearrangements. RESULTS: Among 462 patients of whom 18.4% had MUP, brain metastasis was more common among patients with MUP (23% vs 7.1%). The differences in genomic profiling between MCP and MUP did not reach statistical significance. The 187 MCP and 44 MUP patients treated with immune checkpoint inhibitors had a median overall survival of 49 and 44 months, respectively (p = 0.705). CONCLUSIONS: The differences in somatic mutation patterns and survival outcomes were not statistically significant. These findings may allude to similar carcinogenic processes but should be considered exploratory and interpreted with caution.

Deciphering balanced translocations in infertile males by next-generation sequencing to identify candidate genes for spermatogenesis disorders.

Male infertility affects about 7% of the general male population. Balanced structural chromosomal rearrangements are observed in 0.4-1.4% of infertile males and are considered as a well-established cause of infertility. However, underlying pathophysiological mechanisms still need to be clarified. A strategy combining standard and high throughput cytogenetic and molecular technologies was applied in order to identify the candidate genes that might be implicated in the spermatogenesis defect in three male carriers of different balanced translocations. Fluorescence in situ hybridization (FISH) and whole-genome paired-end sequencing were used to characterize translocation breakpoints at the molecular level while exome sequencing was performed in order to exclude the presence of any molecular event independent from the chromosomal rearrangement in the patients. All translocation breakpoints were characterized in the three patients. We identified four variants: a position effect on LACTB2 gene in Patient 1, a heterozygous CTDP1 gene disruption in Patient 2, two single-nucleotide variations (SNVs) in DNAH5 gene and a heterozygous 17q12 deletion in Patient 3. The variants identified in this study need further validation to assess their roles in male infertility. This study shows that beside the mechanical effect of structural rearrangement on meiosis, breakpoints could result in additional alterations such as gene disruption or position effect. Moreover, additional SNVs or copy number variations may be fortuitously present and could explain the variable impact of chromosomal rearrangements on spermatogenesis. In conclusion, this study confirms the relevance of combining different cytogenetic and molecular techniques to investigate patients with spermatogenesis disorders and structural rearrangements on genomic scale.

Char Syndrome a novel mutation and new insights: A clinical report.

Transcription Factor AP-2 Beta (TFAP2B) functions in the differentiation of neural crest cell derivatives and contributes to the embryogenesis of the ductus arteriosus. Mutations of TFAP2B produces Char syndrome. Char syndrome is an autosomal dominant disorder comprising facial dysmorphism, hand anomalies, and patent ductus arteriosus (PDA). In this report, we describe a proband with a de novo TFAP2B frameshift mutation c.650delG p.(Gly217Alafs*32) in the basic domain. The proband presented mainly with musculoskeletal features of Char syndrome. No PDA was identified at presentation suggesting that this syndrome may prove to be phenotypically heterogeneous. This report will help illustrate the genotype/phenotype correlation of TAFB2 mutations and better delineate the clinical features in Char syndrome.

Non-invasive pre-implantation genetic testing of human embryos: an emerging concept.

The accurate genetic screening of pre-implantation embryos currently entails the use of technically challenging and biologically invasive biopsies of the human embryos. Investigating a more conservative sampling approach has emerged as a timely and desired alternative. Circulating cell-free embryonic DNA is present in the blastocoel fluid and spent culture media of blastocysts, and this has lately been sought as an attractive source of genetic information. The genetic analysis of cell-free embryonic DNA has been reported, to be useful in evaluating the genetic constitution of embryos; thus, providing a potential alternative to conventional biopsy-derived pre-implantation genetic testing (PGT). In this review, we have summarized these non-invasive alternative applications of PGT and discussed their current limitations and future clinical implications.

The utilization of pre-implantation genetic testing in the absence of governance: a real-time experience.

To create a diagnostic document describing the utilization of pre-implantation genetic testing (PGT) in the absence of monitoring and regulation. Retrospective cohort study of couples undergoing PGT between 2004 and 2007 in Lebanon. The clinical indications for 192 PGT cycles performed during the study period were gender selection (96.3%), chromosomal aneuploidy (3.1%), and balanced translocation (0.5%). When gender selection was sought, the selection of a son was desired in 94.1% of cases. Of couples undergoing PGT for sex selection, 16.2% were childless, 8.6% had one child of the opposite gender, 28.1% had two same-gender children, 29.7% had three same-gender children, and 11.9% had four or more. Our findings demonstrate the morally questionable consequences of self-regulated systems in which physicians are the sole gatekeepers of norms and ethics.

Idiopathic premature ovarian failure: what is the most suitable ovarian stimulation protocol?

OBJECTIVE: To evaluate the ovarian response to ovarian stimulation in women with idiopathic premature ovarian failure (POF) in a prospective, controlled, and sequential crossover pilot study. MATERIALS AND METHODS: Ten women with idiopathic premature ovarian failure and normal karyotype were included in the study. Phase I was comprised of three consecutive control cycles consisting each of estrogen progestin sequential therapy. Phase II was comprised of three consecutive treatment cycles combining the use of gonadotropin-releasing hormone agonist (GnRHa) in the background of estrogen priming, followed by gonadotropin ovarian stimulation and corticosteroid immunosuppression. RESULTS: Ovulation rates in the treatment cycles (0/10; 0%) did not differ from control cycles (0/10; 0%). CONCLUSIONS: The findings of this pilot study showed that the combination of estrogen priming, corticosteroid immune-suppression, GnRHa pituitary desensitization, and followed by gonadotropin ovarian stimulation is ineffective in restoring ovarian function in women with idiopathic POF.

Genetics of autoimmune thyroid disease in the Lebanese population.

This study aims to investigate the association of human leukocyte antigen (HLA) class II genes and cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) with autoimmune thyroid diseases in the Lebanese population. A total of 128 patients with autoimmune thyroid disease (55 with Graves' disease (GD) and 73 with Hashimoto's thyroiditis (HT)) were typed for HLA DQA1 (0301 and 0501) and DQB1 (0201, 0302, and 0303) and for 49A/G CTLA-4 using PCR-based sequence-specific priming methods. A total of 186 matched controls were typed for the same alleles and compared to the diseased population. Results showed no significant differences in HLA DQB1*0201 or DQB1*0301 allelic frequencies or CTLA-4 polymorphisms between patients and controls. For GD, there was a weak association with HLA DQB1*0302 [34.6% (19 of 55) vs. 21.5% (40 of 186), P = 0.048, odds ratio (OR) = 1.926, confidence interval (CI) = 0.999-3.715] and HLA DQB1*0302-DQA1*0501 haplotype [56.36% (31 of 55) vs. 40.8% (76 of 186), P = 0.042, OR = 1.870, CI = 1.018-3.433]. For HT, the frequencies of DQB1*0302-DQA1*0501 haplotype [28.8% (21of 73) vs. 14.5% (27 of 186), P = 0.008, OR = 2.378, CI = 1.241-4.558] and DQB1*0302-DQA1*0301 haplotype [60.2% (44 of 73) vs. 38.7% (72 of 186), P = 0.002, OR = 2.402, CI = 1.381-4.180] were significantly higher in patients. On the other hand, weak association was found between HT and DQA1*0301 allele [32.9% (24 of 73) vs. 20.9% (39 of 186), P = 0.044, OR = 1.846, CI = 1.011-3.373]. Findings show that DQB1*0302-DQA1*0501 and DQB1*0302-DQA1*0301 haplotypes may play a role in the pathogenesis of HT in the Lebanese population. For the 49A/G CTLA-4 polymorphism, no significant difference was found between patients and controls.

Impact of AQP3 inducer treatment on cultured human keratinocytes, ex vivo human skin and volunteers.

One of the main functions of the skin is to protect the organism against environmental threats, such as thermal stress. Aquaporin-3 (AQP3) facilitates water and glycerol transport across cell membranes and therefore regulates osmotic balance in different situations of stress. This mechanism seems to be particularly important for the resistance of different organisms to cold stress. Consequently, we were interested in investigating the effect of cold and osmotic stress on AQP3 expression in normal human keratinocytes. We developed a new active ingredient to stimulate aquaporins in skin and demonstrated the partial restoration of AQP3 expression in keratinocytes transfected with AQP3 siRNA. Moreover, we examined the effect of cold stress on cell morphology and the impact of a pre-treatment with the active ingredient. Our results indicated that induction of AQP3 helped maintain a correct organization of the actin cytoskeleton, preserving cell morphology and preventing cells from rounding. Immunofluorescent staining revealed cytoplasmic localization of AQP3 and its translocation to the cell membrane following osmotic stress. Histological ex vivo studies of skin under different conditions, such as cold environment and tape-stripping, indicated that increase in AQP3 expression appears to be involved in skin protection and showed that the pattern of AQP3 expression was more enhanced in the active ingredient-treated samples. In vivo confocal microscopy by Vivascope showed a generally healthier appearance of the skin in the treated areas. These results attest to the potential value of the active ingredient in optimizing environmental stress resistance and protecting the skin from stratum corneum damage.

Microarray profiling of gene expression in human keratinocytes suggests a new protective activity against UV-induced DNA damage for a compound previously known to interact with SCF-KIT signalling pathway.

The stem cell factor (SCF) and its protein-tyrosine kinase receptor KIT are together implicated in the regulation of diverse biological processes and particularly in melanogenesis. Indeed, this signalling pathway controls melanoblast migration from the neural crest during embryogenesis and allows the communication between keratinocytes and melanocytes in the adult. In melanocytes, the binding of SCF to its transmembrane receptor leads to the activation of signalling pathways implicating protein kinases which finally control the expression of pigmentation-related genes. We have developed a biological compound called IV09.007, which we previously described as a modulator of the SCF/KIT signalling pathway with a pro-pigmenting effect. In the present work, we have studied the expression and localization of both SCF and KIT mRNAs and proteins in the skin or skin-derived cell lines. Then, we explored with a microarray approach the ability of IV09.007 to modulate the expression of genes in human keratinocytes and melanocytes in culture. Thereby, we observed the regulation of genes implicated in DNA repair, mainly related to base/nucleotides excision pathways. A modulated transcriptional response was also observed for some genes implicated in the response against oxidative stress, in apoptosis inhibition and in lowering inflammatory immune response. These microarray results predicted a conferred protective effect of IV09.007 and we verified this hypothesis by performing comet assays on UVB-irradiated keratinocytes or melanocytes, to demonstrate the efficacy of IV09.007 on preventing DNA damage.

Maintenance of the ubiquitin-proteasome system activity correlates with visible skin benefits.

Researches on longevity and anti-ageing molecules have clearly evidenced the potential to increase lifespan of the cells. These recent scientific data raise interests and questions on the capacity of the cells to live longer and maintain their fundamental mechanisms of protection, reparation or degradation of abnormal proteins to maintain their capital of healthy and functional cellular activity. In this concern, this study was focused on the ubiquitin-proteasome system as an essential cellular tool to maintain the pool of functionally active proteins allowing renewal of proteins and degradation of damaged proteins. As the proteasome keeps the 'cells health capital', it should be particularly interesting to associate the maintenance of the proteasome activity with increasing longevity. Indeed, although oxidative stress damage increases with ageing leading to collagen and cellular membrane alterations, it also leads to a reduction in the proteasome activity which is critical for the cells. The aim of this study was to better understand the cellular role of the proteasome and to provide new data showing the skin beneficial effects in activating the overall system of ubiquitination and proteasomal degradation. For this purpose, in vitro, ex vivo and in vivo experiments were performed to evaluate the effects of maintaining the ubiquitin-proteasome activity in basal and stress conditions on young versus aged cells. Experiments have included evaluation of a newly developed dimerized tripeptide targeting specifically the ubiquitin-proteasome pathway. Our results have demonstrated that maintenance of this essential mechanism that participates in abnormal protein elimination and protein renewal allows maintaining cellular integrity that correlates with visible skin benefits.

Image analysis to quantify histological and immunofluorescent staining of ex vivo skin and skin cell cultures.

Image processing steps and analysis techniques were developed for the quantification of photomicrographs obtained from light and fluorescence microscopy. The substrates examined were either skin cell cultures, such as normal human keratinocytes (NHK) or fibroblasts, or ex vivo skin sections. Examples of the analyses are provided for the comparison of skincare active ingredient treated samples vs. placebo to demonstrate the utility of the methods to quantify and provide numerical data for a procedure that is typically qualitative in nature and based on observations by a histologist. Quantifiable experiments that are discussed include: Fontana Masson staining for melanin expression; Nile red staining to detect cellular lipid droplets; nuclei staining with diamidino-phenylindole (DAPI); and immunofluorescent staining of protein expression with a primary antibody directed against the protein (antigen) and a secondary antibody tagged with a fluorescent dye (Alexa Fluor 488) against the primary antibody.

Molecular analysis of pericentrin gene (PCNT) in a series of 24 Seckel/microcephalic osteodysplastic primordial dwarfism type II (MOPD II) families.

Microcephalic osteodysplastic primordial dwarfism type II (MOPD II, MIM 210720) and Seckel syndrome (SCKL, MIM 210600) belong to the primordial dwarfism group characterised by intrauterine growth retardation, severe proportionate short stature, and pronounced microcephaly. MOPD II is distinct from SCKL by more severe growth retardation, radiological abnormalities, and absent or mild mental retardation. Seckel syndrome is associated with defective ATR dependent DNA damage signalling. In 2008, loss-of-function mutations in the pericentrin gene (PCNT) have been identified in 28 patients, including 3 SCKL and 25 MOPDII cases. This gene encodes a centrosomal protein which plays a key role in the organisation of mitotic spindles. The aim of this study was to analyse PCNT in a large series of SCKL-MOPD II cases to further define the clinical spectrum associated with PCNT mutations. Among 18 consanguineous families (13 SCKL and 5 MOPDII) and 6 isolated cases (3 SCKL and 3 MOPD II), 13 distinct mutations were identified in 5/16 SCKL and 8/8 MOPDII including five stop mutations, five frameshift mutations, two splice site mutations, and one apparent missense mutation affecting the last base of exon 19. Moreover, we demonstrated that this latter mutation leads to an abnormal splicing with a predicted premature termination of translation. The clinical analysis of the 5 SCKL cases with PCNT mutations showed that they all presented minor skeletal changes and clinical features compatible with MOPDII diagnosis. It is therefore concluded that, despite variable severity, MOPDII is a genetically homogeneous condition due to loss-of-function of pericentrin.

Cotton honeydew (Gossypium hirsutum L.) extract offers very interesting properties for hair cosmetics and care products.

Cotton honeydew extract is composed of a unique combination of oligosaccharides, including fructose, glucose, inositol, melezitose, saccharose, trehalose and trehalulose. Studies have shown that these oligosaccharides exhibit a protective effect. Therefore, we were interested in studying the effect of these oligosaccharides on normal and damaged human hair. Both clinical and scanning electron microscopy (SEM) studies were performed. Standardized human hair samples were used to determine the effect of a rinse-off mask with 1% cotton honeydew extract on the ultrastructure of hair. In addition, hair samples were submitted to different aggressions, following various experimental protocols. SEM showed that, without extra aggression, the cuticle scales appeared to lie more smoothly in the hair in cotton honeydew extract-treated samples than in untreated samples. The extract-treated hair samples were also less prone to chipping. In contrast, the control, untreated hair samples retained a dry and damaged appearance and were prone to chipping and progressive splitting. In a clinical study, 15 volunteers had half of their hair treated with a formula with 1% honeydew extract and the other half was left untreated as a control. Pictures and visual evaluation of the hair showed that the honeydew extract formula left the hair with a smoothness that was far superior to the control side and this result was confirmed by SEM. In addition, mRNA studies on epidermal cells were performed and confirmed the stimulating effect of honeydew extract on keratin synthesis. These results demonstrate that cotton honeydew extract can be of great use in hair care products and cosmetics.

Quercus suber cork extract displays a tensor and smoothing effect on human skin: an in vivo study.

Recently, it has become indispensable for anti-aging active ingredients to provide a visible and immediate smoothing antiwrinkle effect. In Quercus suber, suberin is the most important structural component of cork cell walls. Studies have shown that suberin is made up mostly of hydroxycarboxylic acids and that it is endowed with many special mechanical and chemical properties that evoke a possible smoothing effect on the surface of the skin. Therefore, we were interested in investigating the effect of this cork extract on the skin's surface in a double-blind clinical study. The study was conducted in 15 healthy volunteers, aged 22 to 52 years. The volunteers applied a gel formula with 3% of cork extract, or placebo gel, on each forearm. Skin surface roughness was evaluated visually by pictures and by silicone replicas 1 and 2 h after application, followed by statistical analysis using the matched-pairs McNemar statistical test. McNemar analysis of the pictures revealed that application of cork extract on the skin resulted in a highly significant reduction of roughness 1 h after application. This effect was observed in 73.3% of volunteers. Two hours after cork extract application, a highly significant improvement of skin roughness was found in 78.6% of volunteers. Moreover, silicone replica treatment confirmed significant improvement in average of roughness at 2 h. These results demonstrate that cork extract provides a remarkable and highly significant tensor and smoothing effect on the skin, which could be of great use in anti-aging skin care products.

Complex translocation (8;12;21): a new variant of t(8;21) in acute myeloid leukemia.

Variants of the t(8;21)(q22;q22) involving chromosomes 8, 21, and other chromosomes account for approximately 3% of all t(8;21)(q22;q22) in acute myeloid leukemia (AML) patients. In this paper, we report a case of AML-M2 with a t(8;12;21)(q22;p12 approximately p13;q22) associated with chromosomal abnormalities, including loss of the Y chromosome and trisomy 8q22 approximately qter. Using a dual-color fluorescence in situ hybridization (FISH) analysis with ETO and AML1 probes, we demonstrated an ETO/AML1 fusion signal on the derivative chromosome 8. Using whole painting probes for chromosomes 8 and 12, we demonstrated a three-way translocation, t(8;12;21)(q22;p12 approximately p13;q22). Reverse transcription polymerase chain reaction (RT-PCR) analysis showed the presence of AML1/ETO fusion transcript. The present case highlights the importance of the combination of approaches, i.e., standard karyotyping, FISH, and RT-PCR, for the detection of variants of t(8;21)(q22;q22), shedding light on region 8q22 approximately qter which could harbor potential genes responsible for leukemogenesis.