A protective role for EFTUD2 in the brain.

In this issue of Neuron, Yang et al. report MFDM-like phenotypes in mice with deletion of Eftud2 in their Purkinje cells (PCs), namely cerebellar atrophy alongside motor and social deficits, similar to phenotypes observed in MFDM patients. The absence of Eftud2 caused mis-splicing of Atf4, reduced Scd1 and Gch1, and promoted ferroptosis-regulated PC death.

Sf3b4 regulates chromatin remodeler splicing and Hox expression.

SF3B proteins form a heptameric complex in the U2 small nuclear ribonucleoprotein, essential for pre-mRNA splicing. Heterozygous pathogenic variants in human SF3B4 are associated with head, face, limb, and vertebrae defects. Using the CRISPR/Cas9 system, we generated mice with constitutive heterozygous deletion of Sf3b4 and showed that mutant embryos have abnormal vertebral development.

Craniofacial Defects in Embryos with Homozygous Deletion of in Their Neural Crest Cells Are Not Rescued by Deletion.

Embryos with homozygous mutation of in their neural crest cells () have brain and craniofacial malformations, hyperactivation of the P53-pathway and die before birth. Treatment of embryos with pifithrin-α, a P53-inhibitor, partly improved brain and craniofacial development. To uncover if craniofacial malformations and death were indeed due to P53 hyperactivation we generated embryos with homozygous loss of function mutations in both and in the neural crest cells.

Snrpb is required in murine neural crest cells for proper splicing and craniofacial morphogenesis.

Heterozygous mutations in SNRPB, an essential core component of the five small ribonucleoprotein particles of the spliceosome, are responsible for cerebrocostomandibular syndrome (CCMS). We show that Snrpb heterozygous mouse embryos arrest shortly after implantation. Additionally, heterozygous deletion of Snrpb in the developing brain and neural crest cells models craniofacial malformations found in CCMS, and results in death shortly after birth.

Mutation in Eftud2 causes craniofacial defects in mice via mis-splicing of Mdm2 and increased P53.

EFTUD2 is mutated in patients with mandibulofacial dysostosis with microcephaly (MFDM). We generated a mutant mouse line with conditional mutation in Eftud2 and used Wnt1-Cre2 to delete it in neural crest cells. Homozygous deletion of Eftud2 causes brain and craniofacial malformations, affecting the same precursors as in MFDM patients.

Spliceosomopathies and neurocristopathies: Two sides of the same coin?

Mutations in core components of the spliceosome are responsible for a group of syndromes collectively known as spliceosomopathies. Patients exhibit microcephaly, micrognathia, malar hypoplasia, external ear anomalies, eye anomalies, psychomotor delay, intellectual disability, limb, and heart defects. Craniofacial malformations in these patients are predominantly found in neural crest cells-derived structures of the face and head.

Reproducibility of CRISPR-Cas9 methods for generation of conditional mouse alleles: a multi-center evaluation.

Background: CRISPR-Cas9 gene-editing technology has facilitated the generation of knockout mice, providing an alternative to cumbersome and time-consuming traditional embryonic stem cell-based methods. An earlier study reported up to 16% efficiency in generating conditional knockout (cKO or floxed) alleles by microinjection of 2 single guide RNAs (sgRNA) and 2 single-stranded oligonucleotides as donors (referred herein as "two-donor floxing" method).

Results: We re-evaluate the two-donor method from a consortium of 20 laboratories across the world.

Loss of function mutation of Eftud2, the gene responsible for mandibulofacial dysostosis with microcephaly (MFDM), leads to pre-implantation arrest in mouse.

Mutations in EFTUD2 are responsible for the autosomal dominant syndrome named MFDM (mandibulofacial dysostosis with microcephaly). However, it is not clear how reduced levels of EFTUD2 cause abnormalities associated with this syndrome. To determine if the mouse can serve as a model for uncovering the etiology of abnormalities found in MFDM patients, we used in situ hybridization to characterize expression of Eftud2 during mouse development, and used CRISPR/Cas9 to generate a mutant mouse line with deletion of exon 2 of the mouse gene.

Inhibition of PI3K-AKT-mTOR pathway sensitizes endometrial cancer cell lines to PARP inhibitors.

Background: Phosphatase and Tensin homolog (PTEN) is a tumor suppressor gene. Loss of its function is the most frequent genetic alteration in endometrioid endometrial cancers (70-80%) and high grade tumors (90%). We assessed the sensitivity of endometrial cancer cell lines to PARP inhibitors (olaparib and BMN-673) and a PI3K inhibitor (BKM-120), alone or in combination, in the context of their PTEN mutation status.

Non-alcoholic fatty liver disease in mice with heterozygous mutation in TMED2.

The transmembrane emp24 domain/p24 (TMED) family are essential components of the vesicular transport machinery. Members of the TMED family serve as cargo receptors implicated in selection and packaging of endoplasmic reticulum (ER) luminal proteins into coatomer (COP) II coated vesicles for anterograde transport to the Golgi. Deletion or mutations of Tmed genes in yeast and Drosophila results in ER-stress and activation of the unfolded protein response (UPR).

Metformin Increases E-cadherin in Tumors of Diabetic Patients With Endometrial Cancer and Suppresses Epithelial-Mesenchymal Transition in Endometrial Cancer Cell Lines.

Purpose: Epithelial-mesenchymal transition (EMT) is a critical process for cancer metastasis and recurrence. Metformin, an effective oral antidiabetic drug, has been associated with decreased cancer risk and mortality. In this pilot study, we started to evaluate the effect of metformin on EMT in vivo and in vitro in endometrial cancer (EC).

Suppression of Homologous Recombination by insulin-like growth factor-1 inhibition sensitizes cancer cells to PARP inhibitors.

Background: Impairment of homologous recombination (HR) is found in close to 50 % of ovarian and breast cancer. Tumors with BRCA1 mutations show increased expression of the Insulin-like growth factor type 1 receptor (IGF-1R). We previously have shown that inhibition of IGF-1R results in growth inhibition and apoptosis of ovarian tumor cells.

Chemotherapy reduces PARP1 in cancers of the ovary: implications for future clinical trials involving PARP inhibitors.

Background: PARP inhibitors have shown promising clinical results in cancer patients carrying BRCA1/2 mutations. Their clinical efficacy could logically be influenced by PARP1 protein levels in patient tumors.

Methods: We screened three cohorts of patients with ovarian cancer, totaling 313 samples, and evaluated PARP1 protein expression by immunohistochemistry with further validation by western blotting.

Akt-activated endothelium constitutes the niche for residual disease and resistance to bevacizumab in ovarian cancer.

Ovarian cancer is the second leading cause of cancer-related death in women worldwide. Despite optimal cytoreduction and adequate adjuvant therapies, initial tumor response is often followed by relapse suggesting the existence of a tumor niche. Targeted therapies have been evaluated in ovarian cancer to overcome resistant disease.

Anti-diabetic doses of metformin decrease proliferation markers in tumors of patients with endometrial cancer.

Background: Metformin has been associated with reduced cancer risk. The mechanisms underlying this cancer protective effect remain unknown.

Methods: "Window of opportunity" study of metformin in women with operable endometrial cancer (EC).

Specimen quality evaluation in Canadian biobanks participating in the COEUR repository.

Human biological specimens are important for translational research programs such as the Canadian Ovarian Experimental Unified Resource (COEUR) funded by the Terry Fox Research Institute. Sample quality is an important consideration, as it directly impacts the quality of ensuing research. The aim of the present study was to determine the quality of tissues collected from different sites contributing to the COEUR cohort.

Combination of serum biomarkers to differentiate malignant from benign ovarian tumours.

Objective: To investigate biomarkers and clinical parameters to distinguish ovarian cancers from benign ovarian tumours.

Methods: Serum biomarkers (CA 125, human epididymis protein 4 [HE 4], interleukin-18 [IL-18], leptin, macrophage migration inhibitory factor [MIF], fibroblast growth factor 2 [FGF-2], insulin-like growth factor, osteopontin, prolactin) and the risk of malignancy indexes I and II (RMI-I and RMI-II) scores were obtained prior to surgery in 52 patients with ovarian tumours (37 malignant and 15 benign). ROC curves were built for each individual marker, for logistic regression models using all markers, and for models combining both biomarkers and RMI scores.

Relevance of the OCT1 transporter to the antineoplastic effect of biguanides.

Epidemiologic and laboratory data suggesting that metformin has antineoplastic activity have led to ongoing clinical trials. However, pharmacokinetic issues that may influence metformin activity have not been studied in detail. The organic cation transporter 1 (OCT1) is known to play an important role in cellular uptake of metformin in the liver.

Induction of apoptosis by metformin in epithelial ovarian cancer: involvement of the Bcl-2 family proteins.

Objective: The aims of the study were to evaluate the ability of metformin to induce apoptosis in epithelial ovarian cancer cell lines and to identify the pathways involved in this effect.

Methods: After treatment with metformin and/or cisplatin, OVCAR-3 and OVCAR-4 cellular apoptosis was assessed by flow cytometry and caspase 3/7 activity. Cell cycle analysis was also performed by flow cytometry as well.

Targeting insulin and insulin-like growth factor pathways in epithelial ovarian cancer.

Ovarian cancer is the most lethal of all gynecological malignancies, due in part to the diagnosis at an advanced stage caused by the lack of specific signs and symptoms and the absence of reliable tests for screening and early detection. Most patients will respond initially to treatment but about 70% of them will suffer a recurrence. Therefore, new therapeutic modalities are urgently needed to overcome chemoresistance observed in ovarian cancer patients.

BMS-536924 sensitizes human epithelial ovarian cancer cells to the PARP inhibitor, 3-aminobenzamide.

Objective: To evaluate the anti-neoplastic activity of BMS-536924, an IGF-1R inhibitor, in epithelial ovarian cancer and its capacity to potentiate the effect of a PARP inhibitor, 3-aminobenzamide.

Methods: OVCAR-3, OVCAR-4, SKOV-3 and TOV-81D cell lines were investigated in low-serum tissue culture conditions (1%FBS). Cytotoxicity assays were performed in quadruplicates using the Alamar colorimetric assay in the presence of BMS-536924 and/or 3-aminobenzamide.

In vitro metformin anti-neoplastic activity in epithelial ovarian cancer.

Objective: Metformin, a commonly used drug in the treatment of type II diabetes, may reduce cancer risk and improve cancer prognosis. We evaluated its effect on epithelial ovarian cancer cell lines.

Methods: The OVCAR-3 and OVCAR-4 cell lines were exposed to metformin with and without cisplatin.

Evolutionary conservation of drug action on lipoprotein metabolism-related targets.

Genetic analysis has shown that the slower than normal rhythmic defecation behavior of the clk-1 mutants of Caenorhabditis elegans is the result of altered lipoprotein metabolism. We show here that this phenotype can be suppressed by drugs that affect lipoprotein metabolism, including drugs that affect HMG-CoA reductase activity, reverse cholesterol transport, or HDL levels. These pharmacological effects are highly specific, as these drugs affect defecation only in clk-1 mutants and not in the wild-type and do not affect other behaviors of the mutants.