Paper-based electrochemical device for early detection of integrin αvß6 expressing tumors.

Despite progress in the prevention and diagnosis of cancer, current technologies for tumor detection present several limitations including invasiveness, toxicity, inaccuracy, lengthy testing duration and high cost. Therefore, innovative diagnostic techniques that integrate knowledge from biology, oncology, medicinal and analytical chemistry are now quickly emerging in the attempt to address these issues. Following this approach, here we developed a paper-based electrochemical device for detecting cancer-derived Small Extracellular Vesicles (S-EVs) in fluids. S-EVs were obtained from cancer cell lines known to express, at a different level, the αvß6 integrin receptor, a well-established hallmark of numerous epithelial cancer types. The resulting biosensor turned out to recognize αvß6-containing S-EVs down to a limit of 0.7*103 S-EVs/mL with a linear range up to 105 S-EVs /mL, and a relative standard deviation of 11%, thus it may represent a novel opportunity for αvß6 expressing cancers detection.

New Insights into the Identification of Metabolites and Cytokines Predictive of Outcome for Patients with Severe SARS-CoV-2 Infection Showed Similarity with Cancer.

SARS-CoV-2 infection is characterized by several clinical manifestations, ranging from the absence of symptoms to severe forms that necessitate intensive care treatment. It is known that the patients with the highest rate of mortality develop increased levels of proinflammatory cytokines, called the "cytokine storm", which is similar to inflammatory processes that occur in cancer. Additionally, SARS-CoV-2 infection induces modifications in host metabolism leading to metabolic reprogramming, which is closely linked to metabolic changes in cancer. A better understanding of the correlation between perturbed metabolism and inflammatory responses is necessary. We evaluated untargeted plasma metabolomics and cytokine profiling via 1H-NMR (proton nuclear magnetic resonance) and multiplex Luminex assay, respectively, in a training set of a limited number of patients with severe SARS-CoV-2 infection classified on the basis of their outcome. Univariate analysis and Kaplan-Meier curves related to hospitalization time showed that lower levels of several metabolites and cytokines/growth factors, correlated with a good outcome in these patients and these data were confirmed in a validation set of patients with similar characteristics. However, after the multivariate analysis, only the growth factor HGF, lactate and phenylalanine retained a significant prediction of survival. Finally, the combined analysis of lactate and phenylalanine levels correctly predicted the outcome of 83.3% of patients in both the training and the validation set. We highlighted that the cytokines and metabolites involved in COVID-19 patients' poor outcomes are similar to those responsible for cancer development and progression, suggesting the possibility of targeting them by repurposing anticancer drugs as a therapeutic strategy against severe SARS-CoV-2 infection.

Surgical Treatment Following Failed Medical Treatment of an Interstitial Pregnancy.

Interstitial pregnancy (IP) is a type of ectopic pregnancy in which the embryo implants in the interstitial part of the Fallopian tube. It accounts for 2% of all ectopic pregnancies. Signs and symptoms appear later than the other forms of ectopic pregnancies because of its peculiar location. The gold standard for its diagnosis is transvaginal ultrasound. The treatment can be medical or surgical. Medical treatment is based on the systemic or local injection of methotrexate (MTX); a dose of mifepristone can be added with a reported 85-90% success rate. The surgical option is laparoscopic unilateral cornuostomy or unilateral salpingectomy. The therapeutic choice is based on symptoms, serum ß-human chorionic gonadotropin (ß-hCG) values, and sonographic features. Furthermore, the patient's fertility perspectives should be considered. We report a case of IP in a Caucasian woman of 29 years old, with a previous salpingectomy for ectopic pregnancy medically treated by a double dose of intramuscular MTX 50 mg/m2 combined with a single dose of leucovorin 15 mg and a single dose of mifepristone 600 mg orally. Medical therapy failed as suggested by the sudden onset of intense pelvic pain after 10 days. Because of the clinical symptoms and the sonographic suspicious of pregnancy rupture due to the modest amount of fluid in the pouch of Douglas, clinicians decided on an urgent unilateral laparoscopic salpingectomy. The hemoperitoneum was drained. The patient was discharged two days later and ß-hCG serum levels became negative after 45 days. The advantages of fertility sparing should be weighted according to the patient's reproductive perspectives. Appropriate counseling is therefore key in managing the treatment of interstitial pregnancy.

Epilepsy Course and Developmental Trajectories in STXBP1-DEE.

Background and Objectives: Clinical manifestations in STXBP1 developmental and epileptic encephalopathy (DEE) vary in severity and outcome, and the genotypic spectrum is diverse. We aim to trace the neurodevelopmental trajectories in individuals with STXBP1-DEE and dissect the relationship between neurodevelopment and epilepsy. Methods: Retrospective standardized clinical data were collected through international collaboration. A composite neurodevelopmental score system compared the developmental trajectories in STXBP1-DEE. Results: Forty-eight patients with de novo STXBP1 variants and a history of epilepsy were included (age range at the time of the study: 10 months to 35 years, mean 8.5 years). At the time of inclusion, 65% of individuals (31/48) had active epilepsy, whereas 35% (17/48) were seizure free, and 76% of those (13/17) achieved remission within the first year of life. Twenty-two individuals (46%) showed signs of developmental impairment and/or neurologic abnormalities before epilepsy onset. Age at seizure onset correlated with severity of developmental outcome and the developmental milestones achieved, with a later seizure onset associated with better developmental outcome. In contrast, age at seizure remission and epilepsy duration did not affect neurodevelopmental outcomes. Overall, we did not observe a clear genotype-phenotype correlation, but monozygotic twins with de novo STXBP1 variant showed similar phenotype and parallel disease course. Discussion: The disease course in STXBP1-DEE presents with 2 main trajectories, with either early seizure remission or drug-resistant epilepsy, and a range of neurodevelopmental outcomes from mild to profound intellectual disability. Age at seizure onset is the only epilepsy-related feature associated with neurodevelopment outcome. These findings can inform future dedicated natural history studies and trial design.

HDAC class I inhibitor domatinostat sensitizes pancreatic cancer to chemotherapy by targeting cancer stem cell compartment via FOXM1 modulation.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) represents an unmet clinical need due to the very poor prognosis and the lack of effective therapy. Here we investigated the potential of domatinostat (4SC-202), a new class I histone deacetylase (HDAC) inhibitor, currently in clinical development, to sensitize PDAC to first line standard gemcitabine (G)/taxol (T) doublet chemotherapy treatment. METHODS: Synergistic anti-tumor effect of the combined treatment was assessed in PANC1, ASPC1 and PANC28 PDAC cell lines in vitro as well as on tumor spheroids and microtissues, by evaluating combination index (CI), apoptosis, clonogenic capability. The data were confirmed in vivo xenograft models of PANC28 and PANC1 cells in athymic mice. Cancer stem cells (CSC) targeting was studied by mRNA and protein expression of CSC markers, by limiting dilution assay, and by flow cytometric and immunofluorescent evaluation of CSC mitochondrial and cellular oxidative stress. Mechanistic role of forkhead box M1 (FOXM1) and downstream targets was evaluated in FOXM1-overexpressing PDAC cells. RESULTS: We showed that domatinostat sensitized in vitro and in vivo models of PDAC to chemotherapeutics commonly used in PDAC patients management and particularly to GT doublet, by targeting CSC compartment through the induction of mitochondrial and cellular oxidative stress. Mechanistically, we showed that domatinostat hampers the expression and function of FOXM1, a transcription factor playing a crucial role in stemness, oxidative stress modulation and DNA repair. Domatinostat reduced FOXM1 protein levels by downregulating mRNA expression and inducing proteasome-mediated protein degradation thus preventing nuclear translocation correlated with a reduction of FOXM1 target genes. Furthermore, by overexpressing FOXM1 in PDAC cells we significantly reduced domatinostat-inducing oxidative mitochondrial and cellular stress and abolished GT sensitization, both in adherent and spheroid cells, confirming FOXM1 crucial role in the mechanisms described. Finally, we found a correlation of FOXM1 expression with poor progression free survival in PDAC chemotherapy-treated patients. CONCLUSIONS: Overall, we suggest a novel therapeutic strategy based on domatinostat to improve efficacy and to overcome resistance of commonly used chemotherapeutics in PDAC that warrant further clinical evaluation.

Epigenetic Approaches to Overcome Fluoropyrimidines Resistance in Solid Tumors.

Although fluoropyrimidines were introduced as anticancer agents over 60 years ago, they are still the backbone of many combination chemotherapy regimens for the treatment of solid cancers. Like other chemotherapeutic agents, the therapeutic efficacy of fluoropyrimidines can be affected by drug resistance and severe toxicities; thus, novel therapeutic approaches are required to potentiate their efficacy and overcome drug resistance. In the last 20 years, the deregulation of epigenetic mechanisms has been shown to contribute to cancer hallmarks. Histone modifications play an important role in directing the transcriptional machinery and therefore represent interesting druggable targets. In this review, we focused on histone deacetylase inhibitors (HDACis) that can increase antitumor efficacy and overcome resistance to fluoropyrimidines by targeting specific genes or proteins. Our preclinical data showed a strong synergistic interaction between HDACi and fluoropyrimidines in different cancer models, but the clinical studies did not seem to confirm these observations. Most likely, the introduction of increasingly complex preclinical models, both in vitro and in vivo, cannot recapitulate human complexity; however, our analysis of clinical studies revealed that most of them were designed without a mechanistic approach and, importantly, without careful patient selection.

Plasma metabolomics, lipidomics and cytokinomics profiling predict disease recurrence in metastatic colorectal cancer patients undergoing liver resection.

Purpose: In metastatic colorectal cancer (mCRC) patients (pts), treatment strategies integrating liver resection with induction chemotherapy offer better 5-year survival rates than chemotherapy alone. However, liver resection is a complex and costly procedure, and recurrence occurs in almost 2/3rds of pts, suggesting the need to identify those at higher risk. The aim of this work was to evaluate whether the integration of plasma metabolomics and lipidomics combined with the multiplex analysis of a large panel of plasma cytokines can be used to predict the risk of relapse and other patient outcomes after liver surgery, beyond or in combination with clinical morphovolumetric criteria. Experimental design: Peripheral blood metabolomics and lipidomics were performed by 600 MHz NMR spectroscopy on plasma from 30 unresectable mCRC pts treated with bevacizumab plus oxaliplatin-based regimens within the Obelics trial (NCT01718873) and subdivided into responder (R) and non-R (NR) according to 1-year disease-free survival (DFS): ≥ 1-year (R, n = 12) and < 1-year (NR, n = 18). A large panel of cytokines, chemokines, and growth factors was evaluated on the same plasma using Luminex xMAP-based multiplex bead-based immunoassay technology. A multiple biomarkers model was built using a support vector machine (SVM) classifier. Results: Sparse partial least squares discriminant analysis (sPLS-DA) and loading plots obtained by analyzing metabolomics profiles of samples collected at the time of response evaluation when resectability was established showed significantly different levels of metabolites between the two groups. Two metabolites, 3-hydroxybutyrate and histidine, significantly predicted DFS and overall survival. Lipidomics analysis confirmed clear differences between the R and NR pts, indicating a statistically significant increase in lipids (cholesterol, triglycerides and phospholipids) in NR pts, reflecting a nonspecific inflammatory response. Indeed, a significant increase in proinflammatory cytokines was demonstrated in NR pts plasma. Finally, a multiple biomarkers model based on the combination of presurgery plasma levels of 3-hydroxybutyrate, cholesterol, phospholipids, triglycerides and IL-6 was able to correctly classify patients by their DFS with good accuracy. Conclusion: Overall, this exploratory study suggests the potential of these combined biomarker approaches to predict outcomes in mCRC patients who are candidates for liver metastasis resection after induction treatment for defining personalized management and treatment strategies.

Placenta accreta in the first trimester: A case report.

Placenta accreta is a severe complication of pregnancy normally diagnosed during the second trimester. Early detection could reduce the risk of hemorrhage during abortion or miscarriage; however, guidelines on first-trimester diagnosis are lacking. We describe a case of placenta accreta during the first trimester with its sonographic and histological features.

Effect of Bevacizumab in Combination With Standard Oxaliplatin-Based Regimens in Patients With Metastatic Colorectal Cancer: A Randomized Clinical Trial.

Importance: Although bevacizumab is a standard of care in combination treatments for metastatic colorectal cancer (mCRC), its clinical benefit has been limited. Objective: To determine whether sequential scheduling of bevacizumab administration in combination with chemotherapy improves treatment efficacy in patients with mCRC, in keeping with the tumor vascular normalization hypothesis. Design, Setting, and Participants: This open-label, randomized clinical phase 3 trial was conducted from May 8, 2012, to December 9, 2015, at 3 Italian centers. Patients aged 18 to 75 years with unresectable, previously untreated, or single line-treated mCRC were recruited. Follow-up was completed December 31, 2019, and data were analyzed from February 26 to July 24, 2020. Interventions: Patients received 12 biweekly cycles of standard oxaliplatin-based regimens (modified FOLFOX-6 [levo-folinic acid, fluorouracil, and oxaliplatin]/modified CAPOX [capecitabine and oxaliplatin]) plus bevacizumab administered either on the same day as chemotherapy (standard arm) or 4 days before chemotherapy (experimental arm). Main Outcomes and Measures: The primary end point was the objective response rate (ORR) measured with Response Evaluation Criteria in Solid Tumors, version 1.1. Secondary end points included progression-free survival, overall survival, safety, and quality of life (QOL). Results: Overall, 230 patients (136 men [59.1%]; median age, 62.3 [interquartile range, 53.3-67.6] years) were randomly assigned to the standard arm (n = 115) or the experimental arm (n = 115). The median duration of follow-up was 68.3 (95% CI, 61.0-70.0) months. No difference in ORR (57.4% [95% CI, 47.8%-66.6%] in the standard arm and 56.5% [95% CI, 47.0-65.7] in the experimental arm; P = .89) or progression-free survival (10.5 [95% CI, 9.1-12.3] months in the standard arm and 11.7 [95% CI, 9.9-12.9] months in the experimental arm; P = .15) was observed. However, the median overall survival was 29.8 (95% CI, 22.5-41.1) months in the experimental arm compared with 24.1 (95% CI, 18.6-29.8) months in the standard arm (adjusted hazard ratio, 0.73; 95% CI, 0.54-0.99; P = .04). Moreover, the experimental arm was associated with a significant reduction in the rate of severe diarrhea (6 [5.3%] vs 19 [16.5%]; P = .006) and nausea (2 [1.8%] vs 8 [7.0%]; P = .05) and improved physical functioning (mean [SD] change from baseline, 0.65 [1.96] vs -7.41 [2.95] at 24 weeks; P = .02), and constipation scores (mean [SD] change from baseline, -17.2 [3.73] vs -0.62 [4.44]; P = .003). Conclusions and Relevance: In this randomized clinical trial, sequential administration of bevacizumab plus chemotherapy did not improve ORR, the primary end point. However, the overall survival advantage, fewer adverse effects, and better health-related QOL associated with sequential bevacizumab administration might provide the basis for exploring antiangiogenic combination treatments with innovative perspectives. Trial Registration: EudraCT Identifier: 2011-004997-27; ClinicalTrials.gov Identifier: NCT01718873.

Italian cohort of Lafora disease: Clinical features, disease evolution, and genotype-phenotype correlations.

BACKGROUND: Lafora disease (LD) is characterized by progressive myoclonus, refractory epilepsy, and cognitive deterioration. This complex neurodegenerative condition is caused by pathogenic variants in EPM2A/EPM2B genes, encoding two essential glycogen metabolism enzymes known as laforin and malin. Long-term follow-up data are lacking. We describe the clinical features and genetic findings of a cohort of 26 Italian patients with a long clinical follow-up. METHODS: Patients with EPM2A/EPM2B pathogenic variants were identified by direct gene sequencing or gene panels with targeted re-sequencing. Disease progression, motor functions, and mental performance were assessed by a simplified disability scale. Spontaneous/action myoclonus severity was scored by the Magaudda Scale. RESULTS: Age range was 12.2-46.2 years (mean:25.53 ± 9.14). Age at disease onset ranged from 10 to 22 years (mean:14.04 ± 2.62). The mean follow-up period was 11.48 ± 7.8 years. Twelve out of the 26 (46%) patients preserved walking ability and 13 (50%) maintained speech. A slower disease progression with preserved ambulation and speech after ≥4 years of follow-up was observed in 1 (11%) out of the 9 (35%) EPM2A patients and in 6 (35%) out of the 17 (65%) EPM2B patients. Follow-up was >10 years in 7 (41.2%) EPM2B individuals, including two harbouring the homozygous p.(D146N) pathogenic variant. CONCLUSIONS: This study supports an overall worse disease outcome with severe deterioration of ambulation and speech in patients carrying EPM2A mutations. However, the delayed onset of disabling symptoms observed in the EPM2B subjects harbouring the p.(D146N) pathogenic variant suggests that the underlying causative variant may still influence LD severity.

HSP90 identified by a proteomic approach as druggable target to reverse platinum resistance in ovarian cancer.

Acquired resistance to platinum (Pt)-based therapies is an urgent unmet need in the management of epithelial ovarian cancer (EOC) patients. Here, we characterized by an unbiased proteomics method three isogenic EOC models of acquired Pt resistance (TOV-112D, OVSAHO, and MDAH-2774). Using this approach, we identified several differentially expressed proteins in Pt-resistant (Pt-res) compared to parental cells and the chaperone HSP90 as a central hub of these protein networks. Accordingly, up-regulation of HSP90 was observed in all Pt-res cells and heat-shock protein 90 alpha isoform knockout resensitizes Pt-res cells to cisplatin (CDDP) treatment. Moreover, pharmacological HSP90 inhibition using two different inhibitors [17-(allylamino)-17-demethoxygeldanamycin (17AAG) and ganetespib] synergizes with CDDP in killing Pt-res cells in all tested models. Mechanistically, genetic or pharmacological HSP90 inhibition plus CDDP -induced apoptosis and increased DNA damage, particularly in Pt-res cells. Importantly, the antitumor activities of HSP90 inhibitors (HSP90i) were confirmed both ex vivo in primary cultures derived from Pt-res EOC patients ascites and in vivo in a xenograft model. Collectively, our data suggest an innovative antitumor strategy, based on Pt compounds plus HSP90i, to rechallenge Pt-res EOC patients that might warrant further clinical evaluation.

Synergistic antitumor interaction of valproic acid and simvastatin sensitizes prostate cancer to docetaxel by targeting CSCs compartment via YAP inhibition.

BACKGROUND: Despite the introduction of several novel therapeutic approaches that improved survival, metastatic castration-resistant prostate cancer (mCRPC) remains an incurable disease. Herein we report the synergistic antitumor interaction between two well-known drugs used for years in clinical practice, the antiepileptic agent with histone deacetylase inhibitory activity valproic acid and the cholesterol lowering agent simvastatin, in mCRPC models. METHODS: Synergistic anti-tumor effect was assessed on PC3, 22Rv1, DU145, DU145R80, LNCaP prostate cancer cell lines and EPN normal prostate epithelial cells, by calculating combination index (CI), caspase 3/7 activation and colony formation assays as well as on tumor spheroids and microtissues scored with luminescence 3D-cell viability assay. Cancer stem cells (CSC) compartment was studied evaluating specific markers by RT-PCR, western blotting and flow cytometry as well as by limiting dilution assay. Cholesterol content was evaluated by 1H-NMR. Overexpression of wild-type YAP and constitutively active YAP5SA were obtained by lipofectamine-based transfection and evaluated by immunofluorescence, western blotting and RT-PCR. 22Rv1 R_39 docetaxel resistant cells were selected by stepwise exposure to increasing drug concentrations. In vivo experiments were performed on xenograft models of DU145R80, 22Rv1 parental and docetaxel resistant cells, in athymic mice. RESULTS: We demonstrated the capacity of the combined approach to target CSC compartment by a novel molecular mechanism based on the inhibition of YAP oncogene via concurrent modulation of mevalonate pathway and AMPK. Because both CSCs and YAP activation have been associated with chemo-resistance, we tested if the combined approach can potentiate docetaxel, a standard of care in mCRCP treatment. Indeed, we demonstrated, both in vitro and in vivo models, the ability of valproic acid/simvastatin combination to sensitize mCRPC cells to docetaxel and to revert docetaxel-resistance, by mevalonate pathway/YAP axis modulation. CONCLUSION: Overall, mCRPC progression and therapeutic resistance driven by CSCs via YAP, can be tackled by the combined repurposing of two generic and safe drugs, an approach that warrants further clinical development in this disease.

Valproic Acid Synergizes With Cisplatin and Cetuximab in vitro and in vivo in Head and Neck Cancer by Targeting the Mechanisms of Resistance.

Recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC) is a devastating malignancy with a poor prognosis. The combination of cisplatin (CDDP) plus cetuximab (CX) is one of the standard first-line treatments in this disease. However, this therapeutic regimen is often associated with high toxicity and resistance, suggesting that new combinatorial strategies are needed to improve its therapeutic index. In our study, we evaluated the antitumor effects of valproic acid (VPA), a well-known antiepileptic agent with histone deacetylase inhibitory activity, in combination with CDDP/CX doublet in head and neck squamous cell carcinoma (HNSCC) models. We demonstrated, in HNSCC cell lines, but not in normal human fibroblasts, that simultaneous exposure to equitoxic doses of VPA plus CDDP/CX resulted in a clear synergistic antiproliferative and pro-apoptotic effects. The synergistic antitumor effect was confirmed in four different 3D-self-assembled spheroid models, suggesting the ability of the combined approach to affect also the cancer stem cells compartment. Mechanistically, VPA enhanced DNA damage in combination treatment by reducing the mRNA expression of ERCC Excision Repair 1, a critical player in DNA repair, and by increasing CDDP intracellular concentration via upregulation at transcriptional level of CDDP influx channel copper transporter 1 and downregulation of the ATPAse ATP7B involved in CDDP-export. Valproic acid also induced a dose-dependent downregulation of epidermal growth factor receptor (EGFR) expression and of MAPK and AKT downstream signaling pathways and prevent CDDP- and/or CX-induced EGFR nuclear translocation, a well-known mechanism of resistance to chemotherapy. Indeed, VPA impaired the transcription of genes induced by non-canonical activity of nuclear EGFR, such as cyclin D1 and thymidylate synthase. Finally, we confirmed the synergistic antitumor effect also in vivo in both heterotopic and orthotopic models, demonstrating that the combined treatment completely blocked HNSCC xenograft tumors growth in nude mice. Overall, the introduction of a safe and generic drug such as VPA into the conventional treatment for R/M HNSCC represents an innovative and feasible antitumor strategy that warrants further clinical evaluation. A phase II clinical trial exploring the combination of VPA and CDDP/CX in R/M HNSCC patients is currently ongoing in our institute.

Randomized phase II study of valproic acid in combination with bevacizumab and oxaliplatin/fluoropyrimidine regimens in patients with RAS-mutated metastatic colorectal cancer: the REVOLUTION study protocol.

BACKGROUND: Despite effective treatments, metastatic colorectal cancer (mCRC) prognosis is still poor, mostly in RAS-mutated tumors, thus suggesting the need for novel combinatorial therapies. Epigenetic alterations play an important role in initiation and progression of cancers, including CRC. Histone-deacetylase inhibitors (HDACi) have shown activity in combination with chemotherapy in the treatment of solid tumors. Owing to its HDACi activity and its safe use for epileptic disorders, valproic acid (VPA) is a good candidate for anticancer therapy that we have largely explored preclinically translating our findings in currently ongoing clinical studies. We have shown in CRC models that HDACi, including VPA, induces synergistic antitumor effects in combination with fluoropyrimidines. Furthermore, unpublished results from our group demonstrated that VPA induces differentiation and sensitization of CRC stem cells to oxaliplatin. Moreover, preclinical and clinical data suggest that HDACi may prevent/reverse anti-angiogenic resistance. METHODS/DESIGN: A randomized, open-label, two-arm, multicenter phase-II study will be performed to explore whether the addition of VPA to first line bevacizumab/oxaliplatin/fluoropyrimidine regimens (mFOLFOX-6/mOXXEL) might improve progression-free survival (PFS) in RAS-mutated mCRC patients. A sample size of 200 patients was calculated under the hypothesis that the addition of VPA to chemotherapy/bevacizumab can improve PFS from 9 to 12 months, with one-sided alpha of 0.20 and a power of 0.80. Secondary endpoints are overall survival, objective response rate, metastases resection rate, toxicity, and quality of life. Moreover, the study will explore several prognostic and predictive biomarkers on blood samples, primary tumors, and on resected metastases. DISCUSSION: The "Revolution" study aims to improve the treatment efficacy of RAS-mutated mCRC through an attractive strategy evaluating the combination of VPA with standard cancer treatment. Correlative studies could identify novel biomarkers and could add new insight in the mechanism of interaction between VPA, fluoropyrimidine, oxaliplatin, and bevacizumab. TRIAL REGISTRATION: EudraCT: 2018-001414-15; ClinicalTrials.gov identifier: NCT04310176.

Two distinct phenotypes, hemiplegic migraine and episodic Ataxia type 2, caused by a novel common CACNA1A variant.

BACKGROUND: To investigate the genetic and environmental factors responsible for phenotype variability in a family carrying a novel CACNA1A missense mutation. Mutations in the CACNA1A gene were identified as responsible for at least three autosomal dominant disorders: FHM1 (Familial Hemiplegic Migraine), EA2 (Episodic Ataxia type 2), and SCA6 (Spinocerebellar Ataxia type 6). Overlapping clinical features within individuals of some families sharing the same CACNA1A mutation are not infrequent. Conversely, reports with distinct phenotypes within the same family associated with a common CACNA1A mutation are very rare. CASE PRESENTATION: A clinical, molecular, neuroradiological, neuropsychological, and neurophysiological study was carried out in proband and his carrier mother. The new heterozygous missense variant c.4262G > A (p.Arg1421Gln) in the CACNA1A gene was detected in the two affected family members. The proband showed a complex clinical presentation characterized by developmental delay, poor motor coordination, hemiplegic migraine attacks, behavioral dysregulation, and EEG abnormalities. The mother showed typical episodic ataxia attacks during infancy with no other comorbidities and mild cerebellar signs at present neurological evaluation. CONCLUSIONS: The proband and his mother exhibit two distinct clinical phenotypes. It can be hypothesized that other unknown modifying genes and/or environmental factors may cooperate to generate the wide intrafamilial variability.

qPCR Applications for the Determination of the Biological Age.

Individual age is a phenotypic trait that provides useful information not only in forensic investigations but also in the aging research which is becoming an urgent call due to the dramatic growth of the aging population worldwide.TaqMan quantification PCR (qPCR) can be successfully applied to biological age estimation, using method defined in Zubakov et al. (Curr Biol 20:R970-R971, 2010). Since levels of signal joint T-cell receptor rearrangement excision circle (sjTREC) in human lymphocytes are known to decrease with age increasing, the qPCR of sjTREC represents a simple and relatively reproducible technique which offers highly accurate age estimation results.