The Fate of Water in Hydrogen-Based Iron Oxide Reduction.

Gas-solid reactions are important for many redox processes that underpin the energy and sustainability transition. The specific case of hydrogen-based iron oxide reduction is the foundation to render the global steel industry fossil-free, an essential target as iron production is the largest single industrial emitter of carbon dioxide. This perception of gas-solid reactions has not only been limited by the availability of state-of-the-art techniques which can delve into the structure and chemistry of reacted solids, but one continues to miss an important reaction partner that defines the thermodynamics and kinetics of gas phase reactions: the gas molecules. In this investigation, cryogenic-atom probe tomography is used to study the quasi in situ evolution of iron oxide in the solid and gas phases of the direct reduction of iron oxide by deuterium gas at 700°C. So far several unknown atomic-scale characteristics are observed, including, D2 accumulation at the reaction interface; formation of a core (wüstite)-shell (iron) structure; inbound diffusion of D through the iron layer and partitioning of D among phases and defects; outbound diffusion of oxygen through the wüstite and/or through the iron to the next free available inner/outer surface; and the internal formation of heavy nano-water droplets at nano-pores.

Increased PRL-1 in BM-derived MSCs triggers anaerobic metabolism via mitochondria in a cholestatic rat model.

Mesenchymal stem cell (MSC) therapy in chronic liver disease is associated with mitochondrial anaerobic metabolism. Phosphatase of regenerating liver-1 (PRL-1), known as protein tyrosine phosphatase type 4A, member 1 (PTP4A1), plays a critical role in liver regeneration. However, its therapeutic mechanism remains obscure. The aim of this study was to establish genetically modified bone marrow (BM)-MSCs overexpressing PRL-1 (BM-MSCsPRL-1) and to investigate their therapeutic effects on mitochondrial anaerobic metabolism in a bile duct ligation (BDL)-injured cholestatic rat model. BM-MSCsPRL-1 were generated with lentiviral and nonviral gene delivery systems and characterized. Compared with naive cells, BM-MSCsPRL-1 showed an improved antioxidant capacity and mitochondrial dynamics and decreased cellular senescence. In particular, mitochondrial respiration in BM-MSCsPRL-1 generated using the nonviral system was significantly increased as well as mtDNA copy number and total ATP production. Moreover, transplantation of BM-MSCsPRL-1 generated using the nonviral system had predominantly antifibrotic effects and restored hepatic function in a BDL rat model. Decreased cytoplasmic lactate and increased mitochondrial lactate upon the administration of BM-MSCsPRL-1 indicated significant alterations in mtDNA copy number and ATP production, activating anaerobic metabolism. In conclusion, BM-MSCsPRL-1 generated by a nonviral gene delivery system enhanced anaerobic mitochondrial metabolism in a cholestatic rat model, improving hepatic function.

Reducing Iron Oxide with Ammonia: A Sustainable Path to Green Steel.

Iron making is the biggest single cause of global warming. The reduction of iron ores with carbon generates about 7% of the global carbon dioxide emissions to produce ≈1.85 billion tons of steel per year. This dramatic scenario fuels efforts to re-invent this sector by using renewable and carbon-free reductants and electricity. Here, the authors show how to make sustainable steel by reducing solid iron oxides with hydrogen released from ammonia. Ammonia is an annually 180 million ton traded chemical energy carrier, with established transcontinental logistics and low liquefaction costs. It can be synthesized with green hydrogen and release hydrogen again through the reduction reaction. This advantage connects it with green iron making, for replacing fossil reductants. the authors show that ammonia-based reduction of iron oxide proceeds through an autocatalytic reaction, is kinetically as effective as hydrogen-based direct reduction, yields the same metallization, and can be industrially realized with existing technologies. The produced iron/iron nitride mixture can be subsequently melted in an electric arc furnace (or co-charged into a converter) to adjust the chemical composition to the target steel grades. A novel approach is thus presented to deploying intermittent renewable energy, mediated by green ammonia, for a disruptive technology transition toward sustainable iron making.

Laser-equipped gas reaction chamber for probing environmentally sensitive materials at near atomic scale.

Numerous metallurgical and materials science applications depend on quantitative atomic-scale characterizations of environmentally-sensitive materials and their transient states. Studying the effect upon materials subjected to thermochemical treatments in specific gaseous atmospheres is of central importance for specifically studying a material's resistance to certain oxidative or hydrogen environments. It is also important for investigating catalytic materials, direct reduction of an oxide, particular surface science reactions or nanoparticle fabrication routes. This manuscript realizes such experimental protocols upon a thermochemical reaction chamber called the "Reacthub" and allows for transferring treated materials under cryogenic & ultrahigh vacuum (UHV) workflow conditions for characterisation by either atom probe or scanning Xe+/electron microscopies. Two examples are discussed in the present study. One protocol was in the deuterium gas charging (25 kPa D2 at 200°C) of a high-manganese twinning-induced-plasticity (TWIP) steel and characterization of the ingress and trapping of hydrogen at various features (grain boundaries in particular) in efforts to relate this to the steel's hydrogen embrittlement susceptibility. Deuterium was successfully detected after gas charging but most contrast originated from the complex ion FeOD+ signal and the feature may be an artefact. The second example considered the direct deuterium reduction (5 kPa D2 at 700°C) of a single crystal wüstite (FeO) sample, demonstrating that under a standard thermochemical treatment causes rapid reduction upon the nanoscale. In each case, further studies are required for complete confidence about these phenomena, but these experiments successfully demonstrate that how an ex-situ thermochemical treatment can be realised that captures environmentally-sensitive transient states that can be analysed by atomic-scale by atom probe microscope.

Activation of the EGFR-PI3K-CaM pathway by PRL-1-overexpressing placenta-derived mesenchymal stem cells ameliorates liver cirrhosis via ER stress-dependent calcium.

BACKGROUND: Cholesterol accumulation and calcium depletion induce hepatic injury via the endoplasmic reticulum (ER) stress response. ER stress regulates the calcium imbalance between the ER and mitochondria. We previously reported that phosphatase of regenerating liver-1 (PRL-1)-overexpressing placenta-derived mesenchymal stem cells (PD-MSCsPRL-1) promoted liver regeneration via mitochondrial dynamics in a cirrhotic rat model. However, the role of PRL-1 in ER stress-dependent calcium is not clear. Therefore, we demonstrated that PD-MSCsPRL-1 improved hepatic functions by regulating ER stress and calcium channels in a rat model of bile duct ligation (BDL). METHODS: Liver cirrhosis was induced in Sprague-Dawley (SD) rats using surgically induced BDL for 10 days. PD-MSCs and PD-MSCsPRL-1 (2 × 106 cells) were intravenously administered to animals, and their therapeutic effects were analyzed. WB-F344 cells exposed to thapsigargin (TG) were cocultured with PD-MSCs or PD-MSCsPRL-1. RESULTS: ER stress markers, e.g., eukaryotic translation initiation factor 2α (eIF2α), activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP), were increased in the nontransplantation group (NTx) compared to the control group. PD-MSCsPRL-1 significantly decreased ER stress markers compared to NTx and induced dynamic changes in calcium channel markers, e.g., sarco/endoplasmic reticulum Ca2+ -ATPase 2b (SERCA2b), inositol 1,4,5-trisphosphate receptor (IP3R), mitochondrial calcium uniporter (MCU), and voltage-dependent anion channel 1 (VDAC1) (*p < 0.05). Cocultivation of TG-treated WB-F344 cells with PD-MSCsPRL-1 decreased cytosolic calmodulin (CaM) expression and cytosolic and mitochondrial Ca2+ concentrations. However, the ER Ca2+ concentration was increased compared to PD-MSCs (*p < 0.05). PRL-1 activated phosphatidylinositol-3-kinase (PI3K) signaling via epidermal growth factor receptor (EGFR), which resulted in calcium increase via CaM expression. CONCLUSIONS: These findings suggest that PD-MSCsPRL-1 improved hepatic functions via calcium changes and attenuated ER stress in a BDL-injured rat model. Therefore, these results provide useful data for the development of next-generation MSC-based stem cell therapy for regenerative medicine in chronic liver disease.

PEDF-Mediated Mitophagy Triggers the Visual Cycle by Enhancing Mitochondrial Functions in a H2O2-Injured Rat Model.

Retinal degenerative diseases result from oxidative stress and mitochondrial dysfunction, leading to the loss of visual acuity. Damaged retinal pigment epithelial (RPE) and photoreceptor cells undergo mitophagy. Pigment epithelium-derived factor (PEDF) protects from oxidative stress in RPE and improves mitochondrial functions. Overexpression of PEDF in placenta-derived mesenchymal stem cells (PD-MSCs; PD-MSCsPEDF) provides therapeutic effects in retinal degenerative diseases. Here, we investigated whether PD-MSCsPEDF restored the visual cycle through a mitophagic mechanism in RPE cells in hydrogen peroxide (H2O2)-injured rat retinas. Compared with naïve PD-MSCs, PD-MSCsPEDF augmented mitochondrial biogenesis and translation markers as well as mitochondrial respiratory states. In the H2O2-injured rat model, intravitreal administration of PD-MSCsPEDF restored total retinal layer thickness compared to that of naïve PD-MSCs. In particular, PTEN-induced kinase 1 (PINK1), which is the major mitophagy marker, exhibited increased expression in retinal layers and RPE cells after PD-MSCPEDF transplantation. Similarly, expression of the visual cycle enzyme retinol dehydrogenase 11 (RDH11) showed the same patterns as PINK1 levels, resulting in improved visual activity. Taken together, these findings suggest that PD-MSCsPEDF facilitate mitophagy and restore the loss of visual cycles in H2O2-injured rat retinas and RPE cells. These data indicate a new strategy for next-generation MSC-based treatment of retinal degenerative diseases.

5-O-Demethylnobiletin Alleviates CCl4-Induced Acute Liver Injury by Equilibrating ROS-Mediated Apoptosis and Autophagy Induction.

Polymethoxyflavanoids (PMFs) have exhibited a vast array of therapeutic biological properties. 5-O-Demethylnobiletin (5-DN) is one such PMF having anti-inflammatory activity, yet its role in hepatoprotection has not been studied before. Results from in vitro study revealed that 5-DN did not exert a high level of cytotoxicity on HepG2 cells at 40 µM, and it was able to rescue HepG2 cell death induced by carbon tetrachloride (CCl4). Subsequently, we investigated acute liver injury on BALB/c mice induced by CCl4 through the intraperitoneal injection of 1 mL/kg CCl4 and co-administration of 5-DN at (1 and 2 mg/kg) by oral gavage for 15 days. The results illustrated that treatment with 5-DN attenuated CCl4-induced elevated serum aminotransferase (AST)/alanine aminotransferase (ALT) ratio and significantly ameliorated severe hepatic damage such as inflammation and fibrosis evidenced through lesser aberrations in the liver histology of 5-DN dose groups. Additionally, 5-DN efficiently counteracted and equilibrated the production of ROS accelerated by CCl4 and dramatically downregulated the expression of CYP2E1 vitally involved in converting CCl4 to toxic free radicals and also enhanced the antioxidant enzymes. 5-DN treatment also inhibited cell proliferation and inflammatory pathway abnormally regulated by CCl4 treatment. Furthermore, the apoptotic response induced by CCl4 treatment was remarkably reduced by enhanced Bcl-2 expression and noticeable reduction in Bax, Bid, cleaved caspase 3, caspase 9, and apaf-1 expression. 5-DN treatment also induced the conversion of LC3 and promoted the autophagic flux. Conclusively, 5-DN exhibited hepatoprotective effects in vitro and in vivo and prevented liver fibrosis induced by CCl4.

Efficacy of Gene Modification in Placenta-Derived Mesenchymal Stem Cells Based on Nonviral Electroporation.

Mesenchymal stem cell (MSC)-based therapy using gene delivery systems has been suggested for degenerative diseases. Although MSC-based clinical applications are effective and safe, the mode of action remains unclear. Researchers have commonly applied viral-based gene modification because this system has efficient vehicles. While viral transfection carries many risks, such as oncogenes and chromosomal integration, nonviral gene delivery techniques are less expensive, easier to handle, and safe, although they are less efficient. The electroporation method, which uses Nucleofection technology, provides critical opportunities for hard-to-transfect primary cell lines, including MSCs. Therefore, to improve the therapeutic efficacy using genetically modified MSCs, researchers must determine the optimal conditions for the introduction of the Nucleofection technique in MSCs. Here, we suggest optimal methods for gene modification in PD-MSCs using an electroporation gene delivery system for clinical application.

Controlling the Oxidation of Magnetic and Electrically Conductive Solid-Solution Iron-Rhodium Nanoparticles Synthesized by Laser Ablation in Liquids.

This study focuses on the synthesis of FeRh nanoparticles via pulsed laser ablation in liquid and on controlling the oxidation of the synthesized nanoparticles. Formation of monomodal γ-FeRh nanoparticles was confirmed by transmission electron microscopy (TEM) and their composition confirmed by atom probe tomography (APT). For these particles, three major contributors to oxidation were analysed: (1) dissolved oxygen in the organic solvents, (2) the bound oxygen in the solvent and (3) oxygen in the atmosphere above the solvent. The decrease of oxidation for optimized ablation conditions was confirmed through energy-dispersive X-ray (EDX) and Mössbauer spectroscopy. Furthermore, the time dependence of oxidation was monitored for dried FeRh nanoparticles powders using ferromagnetic resonance spectroscopy (FMR). By magnetophoretic separation, B2-FeRh nanoparticles could be extracted from the solution and characteristic differences of nanostrand formation between γ-FeRh and B2-FeRh nanoparticles were observed.

BLI-Based Functional Assay in Phage Display Benefits the Development of a PD-L1-Targeting Therapeutic Antibody.

The therapeutic functionality of the antibodies from phage display is verified after an initial screening. Several immunological assays such as ELISA, flow cytometry, the western blot, and surface plasmon resonance (SPR) assay are commonly used; the IgG-format antibody is usually preferred to verify the functionality of antibodies, which need elaborative mammalian expression and purification work. Here, we describe a biolayer interferometry (BLI)-based assay that can evaluate the inhibitory functions of antibodies at an earlier stage of screening. To develop a PD-L1-targeting antibody from phage display, we applied the BLI assay to the initial scFv antibody screening, in addition to common ELISA and fluorescence-activated cell sorting (FACS) assays, which showed high advantages and relevance with the in vitro cell-based PD-1/PD-L1 inhibition assay. The same assays for IgG-format antibodies showed high efficiency of the BLI assay in the functional characterization of antibodies, and one candidate selected from the BLI assay resulted in highly efficacious antitumor activity in an in vivo syngeneic mouse study. The BLI assay was also beneficial when searching for antibodies with diverse epitopes. These results demonstrated that the BLI-based inhibition assay is an excellent technique for high-throughput scFv antibody screening in earlier stages and can make phage-display antibody screening more efficient to develop therapeutic candidates.

Enhanced Congo red dye removal from aqueous solutions using iron nanoparticles: adsorption, kinetics, and equilibrium studies.

We report on the Congo red dye removal properties of body centred cubic and amorphous iron nanoparticles, synthesized by a facile borohydride reduction method under ambient conditions. We have analyzed the adsorption of Congo red as a function of dye concentration, time, and temperature and measured a Congo red adsorption capacity of 1735 mg g-1 for the amorphous iron nanoparticles. To our knowledge, this is the highest value reported so far for Congo red adsorption. The acquired data have been evaluated applying various models for adsorption kinetics and thermodynamic studies. The isotherm models as well as acquired Fourier transform infrared spectra suggest that both chemi- and physisorption occur for Congo red adsorption on iron nanoparticles, where chemisorption appears to be dominant. The kinetics of adsorption of Congo red on both bcc-structured and amorphous iron follow a pseudo-second order equation and are characterized by high initial adsorption rates. Diffusion studies indicate that adsorption occurs in two stages, namely film diffusion followed by intraparticle diffusion. Our studies show that amorphous iron nanoparticles are highly promising for dye adsorption and wastewater treatment applications.

Geriatric risk in the surgical management of infectious spondylitis.

AIM: The purpose of the present study was to evaluate and compare clinical outcomes in patients aged >65 years and <65 years who have undergone surgery for infectious spondylitis, and to identify any prognostic factors. METHODS: We reviewed 60 consecutive patients treated surgically for infectious spondylitis. We reviewed patients' medical records especially focusing on comorbidities, preoperative serum albumin levels, postoperative complications and control of infection on follow up. Statistical analysis was carried out to determine whether these various factors affected clinical outcomes. RESULTS: Postoperative complication rates (P = 0.764) and infection control rates (P = 0.275) were not significantly different between the two groups. Univariate analysis did not show a correlation between age and clinical outcome, whereas body mass index (BMI; P = 0.04), Charlson Comorbidity Index (CCI; P = 0.017), American Society of Anesthesiologists (ASA) grade (P = 0.006) and serum albumin (P = 0.003) were associated with overall postoperative complications. BMI (P = 0.002) and CCI (P = 0.000) were also associated with postoperative fatalities. The χ2 -test for trend also showed that CCI (P = 0.018), ASA grade (P = 0.007) and low serum albumin (<3.5 mg/dL; P = 0.004) were associated with postoperative complications. Logistic regression analysis showed that ASA grade (P = 0.034) and BMI (P = 0.044) were related to overall postoperative complications. Receiver operating characteristic curve analysis using ASA grade and BMI to predict major postoperative complications and fatality showed an area under the curve value of 0.793 (P = 0.001) and 0.942 (p=0.002), respectively. CONCLUSIONS: BMI, ASA grade, CCI scores and serum albumin levels, rather than age, might be useful in predicting clinical outcome in surgery for infectious spondylitis in elderly patients. Geriatr Gerontol Int 2017; 17: 984-990.

ER2, a novel human anti-EGFR monoclonal antibody inhibit tumor activity in non-small cell lung cancer models.

OBJECTIVES: The epidermal growth factor receptor (EGFR) abnormalities including amplification, mutation, and overexpression are frequent in non-small cell lung cancer (NSCLC). We investigated in vitro and in vivo antitumor activity of ER2, a novel human anti-EGFR monoclonal antibody, in NSCLC. METHODS: A panel of NSCLC cell lines (A549, H460, H322, H358, H1299, HCC827, PC9, H1975, and PC9-GR) was used to evaluate in vitro antitumor activity of ER2 and cetuximab. The inhibitory effects of ER2 and cetuximab on downstream signaling were assessed by western blot. Secreted VEGF was measured by Human VEGF Quantikine ELISA kit. Antitumor effects of ER2 and cetuximab as single agents and in combination with cisplatin were evaluated in H322, HCC827 and A549 xenograft models. RESULTS: ER2 efficiently inhibits EGFR and its downstream signaling molecules including Akt and Erk1/2 in NSCLC cell lines with wild-type or mutant EGFR. ER2 inhibited cell viability of H322, HCC827 and A549 cells in a dose-dependent manner by inducing cell cycle arrest and apoptosis. Also, ER2 suppressed EGF-stimulated VEGF production as efficiently as cetuximab in H322, HCC827 and A549 cells. Moreover, ER2 alone and in combination with cisplatin showed a significant anti-tumor efficacy in xenograft mouse models. CONCLUSION: Taken together, ER2 has significant anti-tumor activity in in vitro and in vivo NSCLC models, suggesting a rationale for clinical development of ER2 in NSCLC.

GC1118, an Anti-EGFR Antibody with a Distinct Binding Epitope and Superior Inhibitory Activity against High-Affinity EGFR Ligands.

The EGFR-targeted monoclonal antibodies are a valid therapeutic strategy for patients with metastatic colorectal cancer (mCRC). However, only a small subset of mCRC patients has therapeutic benefits and there are high demands for EGFR therapeutics with a broader patient pool and more potent efficacy. In this study, we report GC1118 exhibiting a different character in terms of binding epitope, affinity, mode of action, and efficacy from other anti-EGFR antibodies. Structural analysis of the EGFR-GC1118 crystal complex revealed that GC1118 recognizes linear, discrete N-terminal epitopes of domain III of EGFR, critical for EGF binding but not overlapping with those of other EGFR-targeted antibodies. GC1118 exhibited superior inhibitory activity against high-affinity EGFR ligands in terms of EGFR binding, triggering EGFR signaling, and proliferation compared with cetuximab and panitumumab. EGFR signaling driven by low-affinity ligands, on the contrary, was well inhibited by all the antibodies tested. GC1118 demonstrated robust antitumor activity in tumor xenografts with elevated expression of high-affinity ligands in vivo, whereas cetuximab did not. Considering the significant role of high-affinity EGFR ligands in modulating tumor microenvironment and inducing resistance to various cancer therapeutics, our study suggests a potential therapeutic advantage of GC1118 in terms of efficacy and a range of benefited patient pool. Mol Cancer Ther; 15(2); 251-63. ©2015 AACR.

Discovery of an L-alanine ester prodrug of the Hsp90 inhibitor, MPC-3100.

Various types of Hsp90 inhibitors have been and continue to undergo clinical investigation. One development candidate is the purine-based, synthetic Hsp90 inhibitor 1 (MPC-3100), which successfully completed a phase I clinical study. However, further clinical development of 1 was hindered by poor solubility and consequent formulation issues and promoted development of a more water soluble prodrug. Towards this end, numerous pro-moieties were explored in vitro and in vivo. These studies resulted in identification of L-alanine ester mesylate, 2i (MPC-0767), which exhibited improved aqueous solubility, adequate chemical stability, and rapid bioconversion without the need for solubilizing excipients. Based on improved physical characteristics and favorable PK and PD profiles, 2i mesylate was selected for further development. A convergent, scalable, chromatography-free synthesis for 2i mesylate was developed to support further clinical evaluation.

Co-transplantation of human fetal thymus, bone and CD34(+) cells into young adult immunodeficient NOD/SCID IL2Rγ(null) mice optimizes humanized mice that mount adaptive antibody responses.

Both the thymus (T) and bone (B) are necessary hematopoietic niches in adult humans. We previously showed that co-transplantation of human fetal T and B tissues into neonatal immunodeficient NOD/SCID IL2Rγ(null) (NSG, N) mice facilitated hematopoiesis. However, transplantation into neonatal mice resulted in high frequency of early death, making it unrealistic for repetitive experiments. In this study, young adult N mice were pre-engrafted with T and B, T alone, B alone or no tissues. The animals were irradiated and injected with autologous fetal liver (FL)-derived CD34(+) cells (34). The resultant mice were TB34N, T34N, B34N and 34N, respectively, and challenged with T cell dependent antigens (Ags). The humanized TB34N mice showed best performance of these mouse models in many aspects resembling the adult human Ag-experienced spleen. The TB34N mice exhibited better hematopoietic reconstitution; balanced development of T- and B-cell, and common progenitor cells; follicular lymphoid structures with a functional germinal center (GC) enriched with follicular dendritic cells (FDCs) and plasma cells (PCs); secretion of hIgG in the sera in response to Ags at comparable levels to those of human; derivations of hIgG mAb-secreting hybridoma clones. Collectively, the humanized TB34N mice could develop an adaptive immunity that was capable of producing Ag-specific hIgG at a significant level via class switching. This unprecedented TB34N platform in humanized mice would be useful in dissecting human immunity, for generating human Abs and clinical applications.

Enhancement of the tumor penetration of monoclonal antibody by fusion of a neuropilin-targeting peptide improves the antitumor efficacy.

The limited localization and penetration of monoclonal antibodies (mAb) into solid tumors restricts their antitumor efficacy. Here, we describe a solid tumor-targeting antibody with enhanced tumor penetration activity. We designed a 22-residue peptide (A22p), which was extracted from the C-terminal basic region of semaphorin 3A (Sema3A) but modified to have higher affinity with neuropilin receptors (NRP), and genetically fused it to the C-terminus of Fc of human immunoglobulin G1 via a 15-residue (G4S)3 linker, generating Fc-A22p, for the bivalent binding to NRPs. In contrast to Fc or the monovalent A22p peptide alone, Fc-A22p homed to tumor vessels and induced vascular permeability through VE-cadherin downregulation and penetrated tumor tissues by interacting with NRPs in mice bearing human tumor xenografts. We extended the Fc-A22p platform by generating mAb-A22p antibodies of two clinically approved solid tumor-targeting mAbs, the anti-EGF receptor mAb cetuximab (erbitux), and the anti-Her2 mAb trastuzumab (herceptin). The mAb-A22p antibodies retained the intrinsic antigen binding, natural Fc-like biophysical properties, and productivity in mammalian cell cultures, comparable with those of the parent mAbs. In mouse xenograft tumor models, the mAb-A22p antibodies more efficiently homed to tumor vessels and spread into the extravascular tumor parenchyma, which significantly enhanced antitumor efficacy compared with the parent mAbs. Our results suggest that mAb-A22p is a superior format for solid tumor-targeting antibodies due to its enhanced tumor tissue penetration and greater antitumor efficacy compared with conventional mAbs.

Affinity Maturation of an Epidermal Growth Factor Receptor Targeting Human Monoclonal Antibody ER414 by CDR Mutation.

It is well established that blocking the interaction of EGFR with growth factors leads to the arrest of tumor growth, resulting in tumor cell death. ER414 is a human monoclonal antibody (mAb) derived by guided selection of the mouse mAb A13. The ER414 exhibited a ~17-fold lower affinity and, as a result, lower efficacy of inhibition of the EGF-mediated tyrosine phosphorylation of EGFR when compared with mAb A13 and cetuximab. We performed a stepwise in vitro affinity maturation to improve the affinity of ER414. We obtained a 3D model of ER414 to identify the amino acids in the CDRs that needed to be mutated. Clones were selected from the phage library with randomized amino acids in the CDRs and substitution of amino acids in the HCDR3 and LCDR1 of ER414 led to improved affinity. A clone, H3-14, with a ~20-fold increased affinity, was selected from the HCDR3 randomized library. Then three clones, ER2, ER78 and ER79, were selected from the LCDR1 randomized library based on the H3-14 but did not show further increased affinities compared to that of H3-14. Of the three, ER2 was chosen for further characterization due to its better expression than others. We successfully performed affinity maturation of ER414 and obtained antibodies with a similar affinity as cetuximab. And antibody from an affinity maturation inhibits the EGF-mediated tyrosine phosphorylation of EGFR in a manner similar to cetuximab.

Discovery of (2S)-1-[4-(2-{6-amino-8-[(6-bromo-1,3-benzodioxol-5-yl)sulfanyl]-9H-purin-9-yl}ethyl)piperidin-1-yl]-2-hydroxypropan-1-one (MPC-3100), a purine-based Hsp90 inhibitor.

Modulation of Hsp90 (heat shock protein 90) function has been recognized as an attractive approach for cancer treatment, since many cancer cells depend on Hsp90 to maintain cellular homeostasis. This has spurred the search for small-molecule Hsp90 inhibitors. Here we describe our lead optimization studies centered on the purine-based Hsp90 inhibitor 28a containing a piperidine moiety at the purine N9 position. In this study, key SAR was established for the piperidine N-substituent and for the congeners of the 1,3-benzodioxole at C8. These efforts led to the identification of orally bioavailable 28g that exhibits good in vitro profiles and a characteristic molecular biomarker signature of Hsp90 inhibition both in vitro and in vivo. Favorable pharmacokinetic properties along with significant antitumor effects in multiple human cancer xenograft models led to the selection of 28g (MPC-3100) as a clinical candidate.

Triple disruption of the superior shoulder suspensory complex.

The superior shoulder suspensory complex (SSSC) is an extremely important structure composed of a ring of bone and soft tissues at the superior aspect of the shoulder. Double disruption leads to instability of the construct and usually requires operative treatment. Triple disruption of the SSSC is extremely rare and is encountered in high-energy trauma cases often in association with other injuries. The authors experienced a case of triple disruption involving the acromion, coracoid process, and acromioclavicular separation. This type of SSSC disruption is unlikely to have been caused by a single impact and is rather caused by multiple impacts during one traumatic event.