RESUMEN
Genome-Wide Association Studies (GWAS) have been decisive in elucidating the genetic predisposition of neuroblastoma (NB). The majority of genetic variants identified in GWAS are found in non-coding regions, suggesting that they can be causative of pathogenic dysregulations of gene expression. Nonetheless, pinpointing the potential causal genes within implicated genetic loci remains a major challenge. In this study, we integrated NB GWAS and expression Quantitative Trait Loci (eQTL) data from adrenal gland to identify candidate genes impacting NB susceptibility. We found that ZMYM1, CBL, GSKIP and WDR81 expression was dysregulated by NB predisposing variants. We further investigated the functional role of the identified genes through computational analysis of RNA sequencing (RNA-seq) data from single-cell and whole-tissue samples of NB, neural crest, and adrenal gland tissues, as well as through in vitro differentiation assays in NB cell cultures. Our results indicate that dysregulation of ZMYM1, CBL, GSKIP, WDR81 may lead to malignant transformation by affecting early and late stages of normal program of neuronal differentiation. Our findings enhance the understanding of how specific genes contribute to NB pathogenesis by highlighting their influence on neuronal differentiation and emphasizing the impact of genetic risk variants on the regulation of genes involved in critical biological processes.
RESUMEN
Pharmacogenetics investigates sequence of genes that affect drug response, enabling personalized medication. This approach reduces drug-induced adverse reactions and improves clinical effectiveness, making it a crucial consideration for personalized medical care. Numerous guidelines, drawn by global consortia and scientific organizations, codify genotype-driven administration for over 120 active substances. As the scientific community acknowledges the benefits of genotype-tailored therapy over traditionally agnostic drug administration, the push for its implementation into Italian healthcare system is gaining momentum. This evolution is influenced by several factors, including the improved access to patient genotypes, the sequencing costs decrease, the growing of large-scale genetic studies, the rising popularity of direct-to-consumer pharmacogenetic tests, and the continuous improvement of pharmacogenetic guidelines. Since EMA (European Medicines Agency) and AIFA (Italian Medicines Agency) provide genotype information on drug leaflet without clear and explicit clinical indications for gene testing, the regulation of pharmacogenetic testing is a pressing matter in Italy. In this manuscript, we have reviewed how to overcome the obstacles in implementing pharmacogenetic testing in the clinical practice of the Italian healthcare system. Our particular emphasis has been on germline testing, given the absence of well-defined national directives in contrast to somatic pharmacogenetics.
Asunto(s)
Farmacogenética , Humanos , Italia , Farmacogenética/métodos , Farmacogenética/tendencias , Medicina de Precisión/tendencias , Medicina de Precisión/métodos , Pruebas de Farmacogenómica/métodos , GenotipoRESUMEN
BACKGROUND AND AIM: In surgically altered anatomy (SAA), endoscopic retrograde cholangiopancreatography (ERCP) can be challenging, and it remains debatable the choice of the optimal endoscopic approach within this context. We aim to show our experience and evaluate the technical and clinical success of endoscopic treatment performed in the setting of adverse events (AE) after pancreaticoduodenectomy (PD). METHODS: This study was conducted on a retrospective cohort of patients presenting biliopancreatic complications after PD from 01/01/2012 to 31/12/2022. All patients underwent ERCP at our Endoscopy Unit. Clinical, instrumental data, and characteristics of endoscopic treatments were collected. RESULTS: 133 patients were included (80 M, mean age = 65 y.o.) with a total of 296 endoscopic procedures (median = 2 procedures/treatment). The indications for ERCP were mainly biliary AE (76 cases, 57.1%). Technical success was obtained in 121 patients of 133 (90.9%). 112 out of 133 (84.2%) obtained clinical success. Nine patients out of 112 (8%) experienced AEs. Clinical success rates were statistically different between patients with biliary or pancreatic disease (93.4% vs 73.6%, p < 0.0001). Septic patients were 38 (28.6%) and showed a worse prognosis than non-septic ones (clinical success: 65.7% vs 91.5%, p = 0.0001). During follow-up, 9 patients (8%), experienced recurrence of the index biliopancreatic disease with a median onset at 20 months (IQR 6-40.1). CONCLUSION: Our case series demonstrated that the use of a pediatric colonoscope in ERCP procedures for patients with AEs after PD is both safe and effective in treating the condition, even in a long-term follow-up.
RESUMEN
BACKGROUND: Endoscopic stenting is the mainstay of treatment for benign biliary strictures. There is a not-negligible rate of recurrence and stent migration. Lumen-apposing metal stents (LAMS) have a unique design with short length, large diameter and wide flanges which make them less prone to migration. AIMS: To describe the intraluminal use of LAMS to treat short benign biliary strictures. METHODS: All consecutive patients who underwent bi-flanged LAMS placement for benign biliary strictures, in approximately 6 years, were retrospectively included. Primary outcomes were technical and clinical success; secondary outcomes were number of endoscopic procedures, adverse events evaluation and stricture recurrence during follow-up. RESULTS: Seventy patients (35 male, mean age 67) were enrolled; bilio-enteric anastomotic stricture was the most common etiology. Technical and clinical success were 100 % and 85.7 %, respectively. Patients with post-surgical stricture had a higher success rate than patients with non-surgical stricture or with bilio-enteric anastomotic stricture (90.4 %, 86.3 % and 81.4 %, respectively). Adverse events were 12/70 (17.1 %): stent migration was the most frequent (8/70, 11.4 %). Stricture recurrence was found in 10/54 patients (18.5 %). CONCLUSION: LAMS placement could be safe and effective treatment for short benign biliary strictures in patients in which a significant caliber disproportion between stricture and the duct above was revealed.
RESUMEN
Although a number of susceptibility loci for neuroblastoma (NB) have been identified by genome-wide association studies, it is still unclear whether variants in the HLA region contribute to NB susceptibility. In this study, we conducted a comprehensive genetic analysis of variants in the HLA region among 724 NB patients and 2863 matched controls from different cohorts. We exploited whole-exome sequencing data to accurately type HLA alleles with an ensemble approach on the results from three different typing tools, and carried out rigorous sample quality control to ensure a fine-scale ancestry matching. The frequencies of common HLA alleles were compared between cases and controls by logistic regression under additive and non-additive models. Population stratification was taken into account adjusting for ancestry-informative principal components. We detected significant HLA associations with NB. In particular, HLA-DQB1*05:02 (OR = 1.61; padj = 5.4 × 10-3) and HLA-DRB1*16:01 (OR = 1.60; padj = 2.3 × 10-2) alleles were associated to higher risk of developing NB. Conditional analysis highlighted the HLA-DQB1*05:02 allele and its residue Ser57 as key to this association. DQB1*05:02 allele was not associated to clinical features worse outcomes in the NB cohort. Nevertheless, a risk score derived from the allelic combinations of five HLA variants showed a substantial predictive value for patient survival (HR = 1.53; p = 0.032) that was independent from established NB prognostic factors. Our study leveraged powerful computational methods to explore WES data and HLA variants and to reveal complex genetic associations. Further studies are needed to validate the mechanisms of these interactions that contribute to the multifaceted pattern of factors underlying the disease initiation and progression.
Asunto(s)
Alelos , Secuenciación del Exoma , Predisposición Genética a la Enfermedad , Neuroblastoma , Humanos , Neuroblastoma/genética , Neuroblastoma/mortalidad , Secuenciación del Exoma/métodos , Estudios de Casos y Controles , Masculino , Femenino , Frecuencia de los Genes , Cadenas beta de HLA-DQ/genética , Antígenos HLA/genética , Estudio de Asociación del Genoma Completo , Cadenas HLA-DRB1/genética , Polimorfismo de Nucleótido SimpleRESUMEN
Fluoropyrimidines, crucial in cancer treatment, often cause toxicity concerns even at standard doses. Toxic accumulation of fluoropyrimidine metabolites, culminating in adverse effects, can stem from impaired dihydropyrimidine dehydrogenase (DPYD) enzymatic function. Emerging evidence underscores the role of single nucleotide polymorphisms (SNPs) in DPYD gene, capable of inducing DPYD activity deficiency. Consequently, DPYD genotyping's importance is on the rise in clinical practice before initiating fluoropyrimidine treatment. Although polymerase chain reaction (PCR) followed by Sanger sequencing (SS; PCR-SS) is a prevalent method for DPYD genotyping, it may encounter limitations. In this context, there is reported a case in which a routine PCR-SS approach for genotyping DPYD SNP rs55886062 failed in a proband of African descent. The Clinical Pharmacogenetics Implementation Consortium (CPIC) categorizes the guanine (G) allele of this SNP as non-functional. The enforcement of whole genome sequencing (WGS) approach led to the identification of two adenine (A) insertions near the PCR primers annealing regions in the proband, responsible for a sequence frameshift and a genotyping error for rs55886062. These SNPs (rs145228578, 1-97981199-T-TA and rs141050810, 1-97981622-G-GA) were extremely rare in non-Finnish Europeans (0.05%) but prevalent in African populations (16%). Although limited evidence was available for these SNPs, they were catalogued as benign variants in public databases. Notably, these two SNPs exhibited a high linkage disequilibrium [LD; squared correlation coefficient (R2) = 0.98]. These findings highlighted the importance to consider the prevalence of genetic variants within diverse ethnic populations when designing primers and probes for SNP genotyping in pharmacogenetic testing. This preventive measure is essential to avoid sequence frameshifts or primer misalignments arising from SNP occurrences in the genome, which can compromise PCR-SS and lead to genotyping failures. Furthermore, this case highlights the significance of exploring alternative genotyping approaches, like WGS, when confronted with challenges associated with conventional techniques.
RESUMEN
ATP2B1 is a known regulator of calcium (Ca2+) cellular export and homeostasis. Diminished levels of intracellular Ca2+ content have been suggested to impair SARS-CoV-2 replication. Here, we demonstrate that a nontoxic caloxin-derivative compound (PI-7) reduces intracellular Ca2+ levels and impairs SARS-CoV-2 infection. Furthermore, a rare homozygous intronic variant of ATP2B1 is shown to be associated with the severity of COVID-19. The mechanism of action during SARS-CoV-2 infection involves the PI3K/Akt signaling pathway activation, inactivation of FOXO3 transcription factor function, and subsequent transcriptional inhibition of the membrane and reticulum Ca2+ pumps ATP2B1 and ATP2A1, respectively. The pharmacological action of compound PI-7 on sustaining both ATP2B1 and ATP2A1 expression reduces the intracellular cytoplasmic Ca2+ pool and thus negatively influences SARS-CoV-2 replication and propagation. As compound PI-7 lacks toxicity in vitro, its prophylactic use as a therapeutic agent against COVID-19 is envisioned here.
Asunto(s)
COVID-19 , Calcio , Fosfatidilinositol 3-Quinasas , Proteínas Proto-Oncogénicas c-akt , SARS-CoV-2 , Transducción de Señal , Replicación Viral , Humanos , SARS-CoV-2/efectos de los fármacos , SARS-CoV-2/fisiología , Replicación Viral/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , COVID-19/virología , COVID-19/metabolismo , Transducción de Señal/efectos de los fármacos , Fosfatidilinositol 3-Quinasas/metabolismo , Calcio/metabolismo , Animales , Proteína Forkhead Box O3/metabolismo , Proteína Forkhead Box O3/genética , Chlorocebus aethiops , Tratamiento Farmacológico de COVID-19 , Células Vero , Femenino , ATPasas Transportadoras de Calcio/metabolismo , ATPasas Transportadoras de Calcio/genética , MasculinoRESUMEN
Recently, metabolic associated steatotic liver disease (MASLD) became the leading cause of chronic liver disease worldwide and one of the most frequent causes of hepatocellular carcinoma (HCC). Nonetheless, in this epidemiological trend, viral hepatitis remains the major driver in hepatic carcinogenesis. Globally, hepatitis B virus (HBV) is the leading cause of hepatocellular carcinoma, with an overall attributable risk of approximately 40%, followed by hepatitis C virus (HCV), which accounts for 28-30% of cases, with significant geographic variations between the Eastern and Western world. Considering all the etiologies, HCC risk increases proportionally with the progression of liver disease, but the risk is consistently higher in patients with viral triggers. This evidence indicates that both direct (due to the oncogenic properties of the viruses) and indirect (through the mechanisms of chronic inflammation that lead to cirrhosis) mechanisms are involved, alongside the presence of co-factors contributing to liver damage (smoking, alcohol, and metabolic factors) that synergistically enhance the oncogenic process. The aim of this review is to analyze the oncogenic role of hepatitis viruses in the liver, evaluating epidemiological changes and direct and indirect viral mechanisms that lead to liver cancer.
RESUMEN
The primary treatment for glioblastoma (GBM) is removing the tumor mass as defined by MRI. However, MRI has limited diagnostic and predictive value. Tumor-associated macrophages (TAM) are abundant in GBM tumor microenvironment (TME) and are found in peripheral blood (PB). FKBP51 expression, with its canonical and spliced isoforms, is constitutive in immune cells and aberrant in GBM. Spliced FKBP51s supports M2 polarization. To find an immunologic signature that combined with MRI could advance in diagnosis, we immunophenotyped the macrophages of TME and PB from 37 patients with GBM using FKBP51s and classical M1-M2 markers. We also determined the tumor levels of FKBP51s, PD-L1, and HLA-DR. Tumors expressing FKBP51s showed an increase in various M2 phenotypes and regulatory T cells in PB, indicating immunosuppression. Tumors expressing FKBP51s also activated STAT3 and were associated with reduced survival. Correlative studies with MRI and tumor/macrophages cocultures allowed to interpret TAMs. Tumor volume correlated with M1 infiltration of TME. Cocultures with spheroids produced M1 polarization, suggesting that M1 macrophages may infiltrate alongside cancer stem cells. Cocultures of adherent cells developed the M2 phenotype CD163/FKBP51s expressing pSTAT6, a transcription factor enabling migration and invasion. In patients with recurrences, increased counts of CD163/FKBP51s monocyte/macrophages in PB correlated with callosal infiltration and were accompanied by a concomitant decrease in TME-infiltrating M1 macrophages. PB PD-L1/FKBP51s connoted necrotic tumors. In conclusion, FKBP51s identifies a GBM subtype that significantly impairs the immune system. Moreover, FKBP51s marks PB macrophages associated with MRI features of glioma malignancy that can aid in patient monitoring. SIGNIFICANCE: Our research suggests that by combining imaging with analysis of monocyte/macrophage subsets in patients with GBM, we can enhance our understanding of the disease and assist in its treatment. We discovered a similarity in the macrophage composition between the TME and PB, and through association with imaging, we could interpret macrophages. In addition, we identified a predictive biomarker that drew more attention to immune suppression of patients with GBM.
Asunto(s)
Neoplasias Encefálicas , Glioblastoma , Isoformas de Proteínas , Proteínas de Unión a Tacrolimus , Microambiente Tumoral , Humanos , Glioblastoma/genética , Glioblastoma/patología , Glioblastoma/inmunología , Glioblastoma/metabolismo , Glioblastoma/mortalidad , Glioblastoma/diagnóstico por imagen , Proteínas de Unión a Tacrolimus/genética , Proteínas de Unión a Tacrolimus/metabolismo , Pronóstico , Femenino , Microambiente Tumoral/inmunología , Masculino , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/inmunología , Neoplasias Encefálicas/mortalidad , Persona de Mediana Edad , Macrófagos Asociados a Tumores/inmunología , Macrófagos Asociados a Tumores/metabolismo , Anciano , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/genética , Imagen por Resonancia Magnética , AdultoRESUMEN
BACKGROUND: Neuroblastoma (NB) represents the most frequent and aggressive form of extracranial solid tumor of infants. Although the overall survival of patients with NB has improved in the last years, more than 50% of high-risk patients still undergo a relapse. Thus, in the era of precision/personalized medicine, the need for high-risk NB patient-specific therapies is urgent. METHODS: Within the PeRsonalizEd Medicine (PREME) program, patient-derived NB tumors and bone marrow (BM)-infiltrating NB cells, derived from either iliac crests or tumor bone lesions, underwent to histological and to flow cytometry immunophenotyping, respectively. BM samples containing a NB cells infiltration from 1 to 50 percent, underwent to a subsequent NB cells enrichment using immune-magnetic manipulation. Then, NB samples were used for the identification of actionable targets and for the generation of 3D/tumor-spheres and Patient-Derived Xenografts (PDX) and Cell PDX (CPDX) preclinical models. RESULTS: Eighty-four percent of NB-patients showed potentially therapeutically targetable somatic alterations (including point mutations, copy number variations and mRNA over-expression). Sixty-six percent of samples showed alterations, graded as "very high priority", that are validated to be directly targetable by an approved drug or an investigational agent. A molecular targeted therapy was applied for four patients, while a genetic counseling was suggested to two patients having one pathogenic germline variant in known cancer predisposition genes. Out of eleven samples implanted in mice, five gave rise to (C)PDX, all preserved in a local PDX Bio-bank. Interestingly, comparing all molecular alterations and histological and immunophenotypic features among the original patient's tumors and PDX/CPDX up to second generation, a high grade of similarity was observed. Notably, also 3D models conserved immunophenotypic features and molecular alterations of the original tumors. CONCLUSIONS: PREME confirms the possibility of identifying targetable genomic alterations in NB, indeed, a molecular targeted therapy was applied to four NB patients. PREME paves the way to the creation of clinically relevant repositories of faithful patient-derived (C)PDX and 3D models, on which testing precision, NB standard-of-care and experimental medicines.
Asunto(s)
Variaciones en el Número de Copia de ADN , Neuroblastoma , Lactante , Humanos , Animales , Ratones , Recurrencia Local de Neoplasia , Neuroblastoma/genética , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/patología , Modelos Animales de Enfermedad , Citometría de FlujoRESUMEN
To efficiently tackle certain tumor types, finding new biomarkers for rapid and complete phenotyping of cancer cells is highly demanded. This is especially the case for the most common pediatric solid tumor of the sympathetic nervous system, namely, neuroblastoma (NB). Liquid biopsy is in principle a very promising tool for this purpose, but usually enrichment and isolation of circulating tumor cells in such patients remain difficult due to the unavailability of universal NB cell-specific surface markers. Here, we show that rapid screening and phenotyping of NB cells through stain-free biomarkers supported by artificial intelligence is a viable route for liquid biopsy. We demonstrate the concept through a flow cytometry based on label-free holographic quantitative phase-contrast microscopy empowered by machine learning. In detail, we exploit a hierarchical decision scheme where at first level NB cells are classified from monocytes with 97.9% accuracy. Then we demonstrate that different phenotypes are discriminated within NB class. Indeed, for each cell classified as NB its belonging to one of four NB sub-populations (i.e., CHP212, SKNBE2, SHSY5Y, and SKNSH) is evaluated thus achieving accuracy in the range 73.6%-89.1%. The achieved results solve the realistic problem related to the identification circulating tumor cell, i.e., the possibility to recognize and detect tumor cells morphologically similar to blood cells, which is the core issue in liquid biopsy based on stain-free microscopy. The presented approach operates at lab-on-chip scale and emulates real-world scenarios, thus representing a future route for liquid biopsy by exploiting intelligent biomedical imaging.
RESUMEN
The aim of the study was to evaluate the effects of Active or Sedentary lifestyle on saliva microbiota composition in Italian schoolchildren. Methods: Male (114) and female children (8-10 years) belonging to five primary schools in the neighborhoods of Turin were classified as active (A) or sedentary (S) based on PAQ-C-It questionnaire. PCR amplification of salivary DNA targeted the hypervariable V3-V4 regions of the 16S rRNA bacterial genes. DADA2 workflow was used to infer the Amplicon Sequence Variants and the taxonomic assignments; the beta-diversity was obtained by PCoA with the UniFrac method; LEfSe algorithm, threshold at 5%, and Log LDA cutoff at ±0.5 were used to identify differently abundant species in A compared to S saliva sample. Daily food intake was assessed by 3-Days food record. The metabolic potential of microbial communities was assessed by PICRUSt. Results: No significant differences were found in individual's gender distribution (p = 0.411), anthropometry, BMI (p > 0.05), and all diet composition between A and S groups (p > 0.05). Eight species were differently abundant: Prevotella nigrescens (LDA score = -3.76; FDR = 1.5×10-03), Collinsella aerofaciens (LDA score = -3.17; FDR = 7.45×10-03), Simonsiella muelleri (LDA score = -2.96; FDR = 2.76×10-05), Parabacteroides merdae (LDA score = -2.43; FDR = 1.3×10-02) are enriched in the A group; Gemella parahaemolysans, Prevotella aurantiaca (LDA score = -3.9; FDR = 5.27×10-04), Prevotella pallens (LDA score = 4.23; FDR = 1.93×10-02), Neisseria mucosa (LDA score = 4.43; FDR = 1.31×10-02; LDA score = 2.94; FDR = 7.45×10-03) are enriched in the S group. A prevalence of superpathway of fatty acid biosynthesis initiation (E. coli) and catechol degradation II (meta-cleavage pathway) was found in saliva from A compared to S children. Conclusion: Our results showed that active children had an enrichment of species and genera mainly associated with a healthier profile. By contrast, the genera and the species enriched in the sedentary group could be linked to human diseases.
Asunto(s)
Divertículo , Endoscopía , Humanos , Divertículo/diagnóstico por imagen , Divertículo/cirugíaRESUMEN
Loss of dopaminergic midbrain neurons perturbs l-serine and d-serine homeostasis in the post-mortem caudate putamen (CPu) of Parkinson's disease (PD) patients. However, it is unclear whether the severity of dopaminergic nigrostriatal degeneration plays a role in deregulating serine enantiomers' metabolism. Here, through high-performance liquid chromatography (HPLC), we measured the levels of these amino acids in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys and MPTP-plus-probenecid (MPTPp)-treated mice to determine whether and how dopaminergic midbrain degeneration affects the levels of serine enantiomers in various basal ganglia subregions. In addition, in the same brain regions, we measured the levels of key neuroactive amino acids modulating glutamatergic neurotransmission, including l-glutamate, glycine, l-aspartate, d-aspartate, and their precursors l-glutamine, l-asparagine. In monkeys, MPTP treatment produced severe denervation of nigrostriatal dopaminergic fibers (â75%) and increased the levels of serine enantiomers in the rostral putamen (rPut), but not in the subthalamic nucleus, and the lateral and medial portion of the globus pallidus. Moreover, this neurotoxin significantly reduced the protein expression of the astrocytic serine transporter ASCT1 and the glycolytic enzyme GAPDH in the rPut of monkeys. Conversely, concentrations of d-serine and l-serine, as well as ASCT1 and GAPDH expression were unaffected in the striatum of MPTPp-treated mice, which showed only mild dopaminergic degeneration (â30%). These findings unveil a link between the severity of dopaminergic nigrostriatal degeneration and striatal serine enantiomers concentration, ASCT1 and GAPDH expression. We hypothesize that the up-regulation of d-serine and l-serine levels occurs as a secondary response within a homeostatic loop to support the metabolic and neurotransmission demands imposed by the degeneration of dopaminergic neurons.
Asunto(s)
1-Metil-4-fenil-1,2,3,6-Tetrahidropiridina , Serina , Ratones , Animales , Serina/metabolismo , 1-Metil-4-fenil-1,2,3,6-Tetrahidropiridina/farmacología , Dopamina/metabolismo , Cuerpo Estriado/metabolismo , Mesencéfalo/metabolismo , Aminoácidos/metabolismo , Putamen/metabolismo , HomeostasisRESUMEN
Cholangiocarcinoma (CCA), a highly heterogeneous cancer, is the second most common type of primary liver cancer. It is characterized by resistance to therapy and poor prognosis, with a 5-year survival rate lower than 20%. The pathogenesis of CCA is complex and multifactorial, and in recent years, bile acids (BAs) have been implicated in CCA development and prognosis. BAs belong to a category of amphipathic compounds that hold significant importance as signaling molecules and inflammatory agents. They possess the ability to activate transcriptional factors and cellular signaling pathways, thereby governing the regulation of lipid, glucose, and energy metabolism in diverse human disorders. These disorders encompass chronic liver diseases among other conditions. In this review, we provided an update on the current knowledge on the molecular mechanisms involving BAs in cholangiocarcinogenesis. Additionally, we analyzed the role of gut and biliary microbiota in CCA pathogenesis. Future research is required to better understand how to modulate BA activity and, possibly, identify new therapeutic strategies.
Asunto(s)
Neoplasias de los Conductos Biliares , Sistema Biliar , Colangiocarcinoma , Humanos , Ácidos y Sales Biliares , Neoplasias de los Conductos Biliares/patología , Colangiocarcinoma/patología , Sistema Biliar/metabolismo , Conductos Biliares Intrahepáticos/patologíaRESUMEN
Image-based identification of circulating tumor cells in microfluidic cytometry condition is one of the most challenging perspectives in the Liquid Biopsy scenario. Here we show a machine learning-powered tomographic phase imaging flow cytometry system capable to provide high-throughput 3D phase-contrast tomograms of each single cell. In fact, we show that discrimination of tumor cells against white blood cells is potentially achievable with the aid of artificial intelligence in a label-free flow-cyto-tomography method. We propose a hierarchical machine learning decision-maker, working on a set of features calculated from the 3D tomograms of the cells' refractive index. We prove that 3D morphological features are adequately distinctive to identify tumor cells versus the white blood cell background in the first stage and, moreover, in recognizing the tumor type at the second decision step. Proof-of-concept experiments are shown, in which two different tumor cell lines, namely neuroblastoma cancer cells and ovarian cancer cells, are used against monocytes. The reported results allow claiming the identification of tumor cells with a success rate higher than 97% and with an accuracy over 97% in discriminating between the two cancer cell types, thus opening in a near future the route to a new Liquid Biopsy tool for detecting and classifying circulating tumor cells in blood by stain-free method.
Asunto(s)
Inteligencia Artificial , Células Neoplásicas Circulantes , Humanos , Citometría de Flujo/métodos , Aprendizaje Automático , Biopsia Líquida , TomografíaRESUMEN
The low response to vaccines is a well-known problem in cirrhosis. We evaluated the safety and immunogenicity of booster doses in patients with chronic liver disease (CLD), comparing the humoral response in cirrhotic vs. non-cirrhotic patients, and the impact of different factors on immune response. From September 2021 to April 2022, outpatients with CLD who completed the primary vaccination course and the booster dose against SARS-CoV-2 were enrolled. Blood samples were collected after second and third doses for detecting anti-spike protein IgG. We enrolled 340 patients; among them, 91 subjects were cirrhotic. After primary vaccination course, 60 (17.6%) patients did not develop a positive antibody titer, without significant differences between cirrhotic and non-cirrhotic patients (p = 0.076); most of them (88.3%) developed it after booster dose. At multivariable analysis, factors associated with higher humoral response after booster dose were only porto-sinusoidal vascular disorder (p = 0.007) as an etiology of CLD and the use of the mRNA-1273 vaccine (p = 0.001). In conclusion, in patients with CLD, a booster dose against SARS-CoV-2 induces an excellent immunogenicity and leads to an adequate antibody response. Cirrhosis is not associated with a worse humoral response, compared to patients with non-cirrhotic CLD.
RESUMEN
Neuroblastoma (NB) is the most common extracranial solid tumor in childhood. Patients with relapsed/refractory disease have a poor prognosis, and additional therapeutic options are needed. Mutations and amplifications in the ALK (Anaplastic Lymphoma Kinase) gene constitute a key target for treatment. Our goal, within the Italian project of PeRsonalizEdMEdicine (PREME), was to evaluate the genomic status of patients with relapsed/refractory NB and to implement targeted therapies in those with targetable mutations. From November 2018 to November 2021, we performed Whole Exome Sequencing or Targeted Gene Panel Sequencing in relapsed/refractory NB patients in order to identify druggable variants. Activating mutations of ALK were identified in 8(28.57%) of 28 relapsed/refractory NB patients. The mutation p.F1174L was found in six patients, whereas p.R1275Q was found in one and the unknown mutation p.S104R in another. Three patients died before treatment could be started, while five patients received crizotinib: two in monotherapy (one with p.F1174L and the other with p.S104R) and three (with p.F1174L variant) in combination with chemotherapy. All treated patients showed a clinical improvement, and one had complete remission after two cycles of combined treatment. The most common treatment-related toxicities were hematological. ALK inhibitors may play an important role in the treatment of ALK-mutated NB patients.
RESUMEN
Cornelia de Lange Syndrome (CdLS) is a rare genetic disorder that affects many organs. The diagnosis of this condition is primarily clinical and it can be confirmed by molecular analysis of the genes known to cause this disease, although about 30% of CdLS patients are without a genetic diagnosis. Here we report clinical and genetic findings of a patient with CdLS type 4, a syndrome of which the clinical features of only 30 patients have been previously described in the literature. The index patient presented with clinical characteristics previously associated with CdLS type 4 (short nose, thick eyebrow, global development delay, synophrys, microcephaly, weight < 2DS, small hands, height < 2DS). She also presented cardiac anomalies, cleft palate and laryngomalacia, which was never described before. The index patient was diagnosed with a novel de novo RAD21 variant (c.1722_1723delTG, p.Gly575SerfsTer2): segregation analysis, bioinformatic analysis, population data and in silico structural modelling indicate the pathogenicity of the novel variant. This report summarizes previously reported clinical manifestations of CdLS type 4 but also highlights new clinical symptoms, which will aid correct counselling of future CdLS type 4 cases.
Asunto(s)
Fisura del Paladar , Síndrome de Cornelia de Lange , Hipertricosis , Femenino , Humanos , Síndrome de Cornelia de Lange/diagnóstico , Síndrome de Cornelia de Lange/genética , Proteínas de Ciclo Celular/genética , Fenotipo , Proteínas de Unión al ADN/genéticaRESUMEN
Pleiotropic genetic factors (e.g., DNA polymorphisms) may be involved in the initiation of neuroblastoma (NB) and coronary artery disease (CAD) given their common origin from defects in neural crest development. To discover novel NB susceptibility genes, we conducted a three-stage survey including a meta-analysis of NB and CAD genome-wide association data, prioritization of NB causal variants, and validation in an independent cohort of affected individuals-control subjects. The lead SNP, rs13337397 at the 16q23.1 locus, associated with both diseases in the meta-analysis and with NB in the validation study. All the SNPs in linkage disequilibrium with rs13337397 were annotated using the H3K27ac epigenetic marker of neural crest cells (NCC) and NB cell lines. Indeed, we identified the functional SNP rs13337017, mapping within an enhancer of NCCs and NB cell lines and showing long-range interactions with CFDP1 by Hi-C analysis. Luciferase assays indicated that the risk allele of rs13337017 increased CFDP1 expression in NB cell lines. Of note, CFDP1 high expression associated with unfavorable prognostic markers in an analysis including 498 NB transcriptomes. Moreover, depletion of CFDP1 markedly decreased viability and migration and increased apoptotic rates in NB cell lines. Finally, transcriptome and qPCR analyses revealed that the depletion of CFDP1 may affect noradrenergic neuron differentiation by downregulating master regulators of sympathetic noradrenergic identity, including PHOX2B, HAND2, and GATA3. Our data strongly suggest that CFDP1 acts as oncogene in NB. In addition, we provide evidence that genetic predisposition to NB can be mediated by the alteration of noradrenergic lineage-specific gene expression.