Synthesis and Structure-Activity Relationships of 2,5-Dimethoxy-4-Substituted Phenethylamines and the Discovery of CYB210010: A Potent, Orally Bioavailable and Long-Acting Serotonin 5-HT Receptor Agonist.

Structure-activity studies of 4-substituted-2,5-dimethoxyphenethylamines led to the discovery of 2,5-dimethoxy-4-thiotrifluoromethylphenethylamines, including CYB210010, a potent and long-acting serotonin 5-HT receptor agonist. CYB210010 exhibited high agonist potency at 5-HT and 5-HT receptors, modest selectivity over 5-HT, 5-HT, 5-HT, and adrenergic α receptors, and lacked activity at monoamine transporters and over 70 other proteins. CYB210010 (0.

Long-Term Follow-Up of Pediatric Excimer Laser-Assisted Penetrating Keratoplasty for Congenital Stromal Corneal Dystrophy.

Purpose: The purpose of this study was to highlight characteristic clinical and microscopic findings and report the long-term follow-up of pediatric excimer laser-assisted penetrating keratoplasty (excimer-PKP) for congenital stromal corneal dystrophy (CSCD).

Methods: A 2-year-old Greek child presented with CSCD at our department. Clinical examination showed bilateral flake-like whitish corneal opacities affecting the entire corneal stroma up to the limbus.

Intercellular transfer of cancer cell invasiveness via endosome-mediated protease shedding.

Overexpression of the transmembrane matrix metalloproteinase MT1-MMP/MMP14 promotes cancer cell invasion. Here we show that MT1-MMP-positive cancer cells turn MT1-MMP-negative cells invasive by transferring a soluble catalytic ectodomain of MT1-MMP. Surprisingly, this effect depends on the presence of TKS4 and TKS5 in the donor cell, adaptor proteins previously implicated in invadopodia formation.

Next-generation sequencing reveals mitogenome diversity in plasma extracellular vesicles from colorectal cancer patients.

Background: Recent reports have demonstrated that the entire mitochondrial genome can be secreted in extracellular vesicles (EVs), but the biological attributes of this cell-free mitochondrial DNA (mtDNA) remain insufficiently understood. We used next-generation sequencing to compare plasma EV-derived mtDNA to that of whole blood (WB), peripheral blood mononuclear cells (PBMCs), and formalin-fixed paraffin-embedded (FFPE) tumor tissue from eight rectal cancer patients and WB and fresh-frozen (FF) tumor tissue from eight colon cancer patients.

Methods: Total DNA was isolated before the mtDNA was enriched by PCR with either two primer sets generating two long products or multiple primer sets (for the FFPE tumors), prior to the sequencing.

siRNA screening reveals that SNAP29 contributes to exosome release.

Cells release extracellular vesicles (EVs) of different sizes. Small EVs (< 200 nm) can originate from the fusion of multivesicular bodies with the plasma membrane, i.e.

Hsp90 inhibition and co-incubation with pertuzumab induce internalization and degradation of trastuzumab: Implications for use of T-DM1.

The receptor tyrosine kinase HER2 is associated with a number of human malignancies and is an important therapeutic target. The antibody-drug conjugate trastuzumab emtansine (T-DM1; Kadcyla ) is recommended as a first-line treatment for patients with HER2-positive metastatic breast cancer. T-DM1 combines the antibody-induced effects of the anti-HER2 antibody trastuzumab (Herceptin ) with the cytotoxic effect of the tubulin inhibitor mertansine (DM1).

Protein kinase C regulates ErbB3 turnover.

ErbB3, which belongs to the epidermal growth factor receptor (EGFR) or ErbB family of receptor tyrosine kinases, is involved in progression of several human cancers and a tight regulation of its expression is crucial. An important mechanism for regulation of ErbB proteins is endocytosis and we recently showed that ErbB3, contrary to other ErbB proteins, like EGFR and ErbB2, is constitutively internalized and degraded. Several studies show that protein kinase C (PKC) can regulate the activation, localization and stability of EGFR and ErbB2.

Protein kinase C mediated internalization of ErbB2 is independent of clathrin, ubiquitination and Hsp90 dissociation.

Overexpression of ErbB2 is frequent in cancer and understanding the mechanisms which regulate its expression is important. ErbB2 is considered endocytosis resistant. It has no identified ligand, but upon heterodimerization it is a potent mediator of proliferative signaling.

Macrocyclic pyrrolobenzodiazepine dimers as antibody-drug conjugate payloads.

Macrocyclic pyrrolobenzodiazepine dimers were designed and evaluated for use as antibody-drug conjugate payloads. Initial structure-activity exploration established that macrocyclization could increase the potency of PBD dimers compared with non-macrocyclic analogs. Further optimization overcame activity-limiting solubility issues, leading to compounds with highly potent (picomolar) activity against several cancer cell lines.

ErbB3 interacts with Hrs and is sorted to lysosomes for degradation.

The ErbB family of receptor tyrosine kinases mediates activation of a wide network of signaling pathways. ErbB3 has weak kinase activity, but its six docking sites for the p85 subunit of phosphoinositide 3-kinase make it an important contributor to proliferative signaling. ErbB3 has a relatively short half-life but the exact mechanisms controlling its turnover are unclear as contradictory reports exist.

A combination of two antibodies recognizing non-overlapping epitopes of HER2 induces kinase activity-dependent internalization of HER2.

The human epidermal growth factor receptor 2 (HER2/ErbB2) is overexpressed in a number of human cancers. HER2 is the preferred heterodimerization partner for other epidermal growth factor receptor (EGFR) family members and is considered to be resistant to endocytic down-regulation, properties which both contribute to the high oncogenic potential of HER2. Antibodies targeting members of the EGFR family are powerful tools in cancer treatment and can function by blocking ligand binding, preventing receptor dimerization, inhibiting receptor activation and/or inducing receptor internalization and degradation.

Interaction with epsin 1 regulates the constitutive clathrin-dependent internalization of ErbB3.

Background: In contrast to other members of the EGF receptor family, ErbB3 is constitutively internalized in a clathrin-dependent manner. Previous studies have shown that ErbB3 does not interact with the coated pit localized adaptor complex 2 (AP-2), and that ErbB3 lacks two AP-2 interacting internalization signals identified in the EGF receptor. Several other clathrin-associated sorting proteins which may recruit cargo into coated pits have, however, been identified, and the study was performed to identify adaptors needed for constitutive internalization of ErbB3.

CTGF/CCN2 Postconditioning Increases Tolerance of Murine Hearts towards Ischemia-Reperfusion Injury.

Background And Purpose: Previous studies of ischemia-reperfusion injury (IRI) in hearts from mice with cardiac-restricted overexpression of CCN2 have shown that CCN2 increases tolerance towards IRI. The objectives of this study were to investigate to what extent post-ischemic administration of recombinant human CCN2 (rhCCN2) would limit infarct size and improve functional recovery and what signaling pathways are involved.

Experimental Approach: Isolated mice hearts were perfused ad modum Langendorff, subjected to no-flow, global ischemia, and subsequently, exposed to mammalian cell derived, full-length (38-40kDa) rhCCN2 (250 nM) or vehicle during the first 15 min of a 60 min reperfusion period.

Syndecan-4 Regulates Muscle Differentiation and Is Internalized from the Plasma Membrane during Myogenesis.

The cell surface proteoglycan syndecan-4 has been reported to be crucial for muscle differentiation, but the molecular mechanisms still remain to be fully understood. During in vitro differentiation of bovine muscle cells immunocytochemical analyses showed strong labelling of syndecan-4 intracellularly, in close proximity with Golgi structures, in membranes of intracellular vesicles and finally, in the nuclear area including the nuclear envelope. Chase experiments showed that syndecan-4 was internalized from the plasma membrane during this process.

Neonatal Abstinence Syndrome: Influence of a Combined Inpatient/Outpatient Methadone Treatment Regimen on the Average Length of Stay of a Medicaid NICU Population.

Prescription opioid and heroin abuse have been increasing steadily year after year, and continue to be a serious national problem. A sequela of the increase in opioid abuse has been an increase in the number of infants born with opioid dependence. These infants often require costly, prolonged stays in the neonatal intensive care unit (NICU) for drug withdrawal treatment.

Discovery of tetrahydroisoquinoline-based bivalent heterodimeric IAP antagonists.

Bivalent heterodimeric IAP antagonists that incorporate (R)-tetrahydroisoquinoline in the P3' subunit show high affinity for the BIR2 domain and demonstrated potent IAP inhibitory activities in biochemical and cellular assays. Potent in vivo efficacy was observed in a variety of human tumor xenograft models. The bivalent heterodimeric molecule 3 with a P3-P3' benzamide linker induced pharmacodynamic markers of apoptosis and was efficacious when administered intravenously at a dose of 1mg/kg to mice harboring A875 human melanoma tumors.

Tartrate-resistant acid phosphatase (TRAP) co-localizes with receptor activator of NF-KB ligand (RANKL) and osteoprotegerin (OPG) in lysosomal-associated membrane protein 1 (LAMP1)-positive vesicles in rat osteoblasts and osteocytes.

Tartrate-resistant acid phosphatase (TRAP) is well known as an osteoclast marker; however, a recent study from our group demonstrated enhanced number of TRAP + osteocytes as well as enhanced levels of TRAP located to intracellular vesicles in osteoblasts and osteocytes in experimental osteoporosis in rats. Such vesicles were especially abundant in osteoblasts and osteocytes in cancellous bone as well as close to bone surface and intracortical remodeling sites. To further investigate TRAP in osteoblasts and osteocytes, long bones from young, growing rats were examined.

The Mysterious Ways of ErbB2/HER2 Trafficking.

The EGFR- or ErbB-family of receptor tyrosine kinases consists of EGFR/ErbB1, ErbB2/HER2, ErbB3/HER3 and ErbB4/HER4. Receptor activation and downstream signaling are generally initiated upon ligand-induced receptor homo- or heterodimerization at the plasma membrane, and endocytosis and intracellular membrane transport are crucial for regulation of the signaling outcome. Among the receptors, ErbB2 is special in several ways.

Pertuzumab counteracts the inhibitory effect of ErbB2 on degradation of ErbB3.

Overexpression of ErbB2 and ErbB3 is found in several human cancers, and ErbB2-ErbB3 heterodimers are known as the most potent signaling units among ErbB dimers. While ErbB2 probably undergoes weak endocytosis, ErbB3 is readily internalized even in the absence of added ligand and without requirement for kinase activity. Overexpression of ErbB2 has been demonstrated to inhibit epidermal growth factor-induced internalization and degradation of epidermal growth factor receptor.

Preubiquitinated chimeric ErbB2 is constitutively endocytosed and subsequently degraded in lysosomes.

The oncoprotein ErbB2 is endocytosis-deficient, probably due to its interaction with Heat shock protein 90. We previously demonstrated that clathrin-dependent endocytosis of ErbB2 is induced upon incubation of cells with Ansamycin derivatives, such as geldanamycin and its derivative 17-AAG. Furthermore, we have previously demonstrated that a preubiquitinated chimeric EGFR (EGFR-Ub(4)) is constitutively endocytosed in a clathrin-dependent manner.

Cetuximab in combination with anti-human IgG antibodies efficiently down-regulates the EGF receptor by macropinocytosis.

The monoclonal antibody C225 (Cetuximab) blocks binding of ligand to the epidermal growth factor receptor (EGFR). In addition, it is known that incubation with C225 induces endocytosis of the EGFR. This endocytosis has previously been shown to be increased when C225 is combined with an additional monoclonal anti-EGFR antibody.

Statins affect the presentation of endothelial chemokines by targeting to multivesicular bodies.

Background: In addition to lowering cholesterol, statins are thought to beneficially modulate inflammation. Several chemokines including CXCL1/growth-related oncogene (GRO)-α, CXCL8/interleukin (IL)-8 and CCL2/monocyte chemoattractant protein (MCP)-1 are important in the pathogenesis of atherosclerosis and can be influenced by statin-treatment. Recently, we observed that atorvastatin-treatment alters the intracellular content and subcellular distribution of GRO-α in cultured human umbilical vein endothelial cells (HUVECs).

The oncoprotein ErbB3 is endocytosed in the absence of added ligand in a clathrin-dependent manner.

The oncoprotein ErbB3 is overexpressed in several human cancers, for example in pancreatic adenocarcinoma and in ovarian cancers, and ErbB3-containing heterodimers have been demonstrated to be potent signaling units in carcinogenesis. This especially applies to ErbB2-ErbB3 and epidermal growth factor receptor (EGFR)-ErbB3 heterodimers providing anti-apoptotic signaling. Relatively little is understood about the signaling of EGFR-ErbB3 heterodimers and especially about mechanisms involved in downregulation of ErbB3 from the plasma membrane.

Pertuzumab Increases 17-AAG-Induced Degradation of ErbB2, and This Effect Is Further Increased by Combining Pertuzumab with Trastuzumab.

ErbB2 is an important oncogenic protein involved in carcinogenesis of, among others, breast, gastric, and ovarian carcinoma. Over-expression of ErbB2 is found in almost 20% of breast cancers, and this results in proliferative and anti-apoptotic signalling. ErbB2 is therefore an important treatment target.

Molecular complexity via C-H activation: a dehydrogenative Diels-Alder reaction.

Traditionally, C-H oxidation reactions install oxidized functionality onto a preformed molecular skeleton, resulting in a local molecular change. The use of C-H activation chemistry to construct complex molecular scaffolds is a new area with tremendous potential in synthesis. We report a Pd(II)/bis-sulfoxide-catalyzed dehydrogenative Diels-Alder reaction that converts simple terminal olefins into complex cycloadducts in a single operation.