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Cortical bone adaptation to a moderate level of mechanical loading in male Sost deficient mice
No full textLoss-of-function mutations in the Sost gene lead to high bone mass phenotypes. Pharmacological inhibition of Sost/sclerostin provides a new drug strategy for treating osteoporosis. Questions remain as to how physical activity may affect bone mass under sclerostin inhibition and if that effect differs between males and females. We previously observed in female Sost knockout (KO) mice an enhanced cortical bone formation response to a moderate level of applied loading (900 με at the tibial midshaft). The purpose of the present study was to examine cortical bone adaptation to the same strain level applied to male Sost KO mice. Strain-matched in vivo compressive loading was applied to the tibiae of 10-, 26- and 52-week-old male Sost KO and littermate control (LC) mice. The effect of tibial loading on bone (re)modeling was measured by microCT, 3D time-lapse in vivo morphometry, 2D histomorphometry and gene expression analyses. As expected, Sost deficiency led to high cortical bone mass in 10- and 26-week-old male mice as a result of increased bone formation. However, the enhanced bone formation associated with Sost deficiency did not appear to diminish with skeletal maturation. An increase in bone resorption was observed with skeletal maturation in male LC and Sost KO mice. Two weeks of in vivo loading (900 με at the tibial midshaft) induced only a mild anabolic response in 10- and 26-week-old male mice, independent of Sost deficiency. A decrease in the Wnt inhibitor Dkk1 expression was observed 3 h after loading in 52-week-old Sost KO and LC mice, and an increase in Lef1 expression was observed 8 h after loading in 10-week-old Sost KO mice. The current results suggest that long-term inhibition of sclerostin in male mice does not influence the adaptive response of cortical bone to moderate levels of loading. In contrast with our previous strain-matched study in females showing enhanced bone responses with Sost ablation, these results in males indicate that the influence of Sost deficiency on the cortical bone formation response to a moderate level of loading differs between males and females. Clinical studies examining antibodies to inhibit sclerostin may need to consider that the efficacy of additional physical activity regimens may be sex dependent
Gene therapy: a double-edged sword with great powers.
No full textGene therapy is the treatment of a disease through transferring genetic material into cells of the patients. In the recent several years, gene therapy has experienced rapid progress and achieved huge success. Over two dozens of gene therapies have been approved for clinical use by the drug regulatory agencies from different countries. However, concerns about its efficacy and safety have accompanied gene therapy since its birth. In the present manuscript, we first introduce various strategies employed in gene therapy, which includes ex vivo gene delivery v.s. in vivo gene delivery; gene addition v.s. genome editing; inherited disease v.s. acquired disease; and somatic gene therapy v.s. germline gene therapy. Then we discuss the clinical outcomes of some approved gene therapies. We finish our discussion with the safety issues related to gene therapy. We will see that with the technology improvement, somatic gene therapy has been proved to be efficient and safe enough for clinical practice. However, germline gene therapy has important efficiency and safety issues at present, and should not be put into clinical practice before these issues are solved
Efficacy and Tolerability of Pyrazolo[1,5-a]pyrimidine RET Kinase Inhibitors for the Treatment of Lung Adenocarcinoma
No full textRET (REarranged during Transfection) kinase gain-of-function aberrancies have been identified as potential oncogenic drivers in lung adenocarcinoma, along with several other cancer types, prompting the discovery and assessment of selective inhibitors. Internal mining and analysis of relevant kinase data informed the decision to investigate a pyrazolo[1,5-a]pyrimidine scaffold, where subsequent optimization led to the identification of compound WF-47-JS03 (1), a potent RET kinase inhibitor with >500-fold selectivity against KDR (Kinase insert Domain Receptor) in cellular assays. In subsequent mouse in vivo studies, compound 1 demonstrated effective brain penetration and was found to induce strong regression of RET-driven tumor xenografts at a well-tolerated dose (10 mg/kg, po, qd). Higher doses of 1, however, were poorly tolerated in mice, similar to other pyrazolo[1,5-a]pyrimidine compounds at or near the efficacious dose, and indicative of the narrow therapeutic windows seen with this scaffold
Application of Transition-Metal Catalysis, Biocatalysis and Flow Chemistry as State-of-the-Art Technologies in the Synthesis of LCZ696
No full textLCZ696 is a novel treatment for patients suffering from heart failure that combines the two active pharmaceutical ingredients sacubitril and valsartan in a single chemical compound. While valsartan is an established drug substance, a new manufacturing process suitable for large-scale commercial production had to be developed for sacubitril. The use of chemocatalysis, biocatalysis, and flow chemistry as state-of-the-art technologies allowed to efficiently build up the structure of sacubitril and achieve the defined performance targets
Intravenous immunoglobulin significantly reduces exposure of concomitantly administered anti-C5 monoclonal antibody tesidolumab
No full textAwareness of drug-drug interactions is critical in organ transplant recipient management. However, biologic agents interfering with monoclonal antibodies is not widely considered. We report the effect of high-dose intravenous immunoglobulin (IVIg) on safety, tolerability, pharmacokinetics (PK), and pharmacodynamics (PD) of the human anti-C5 monoclonal antibody tesidolumab (LFG316) in end-stage renal disease patients awaiting kidney transplant. In this single-center, phase 1, open-label, parallel-group study, 8 patients were assigned to receive either single-dose tesidolumab + IVIg or tesidolumab alone, with 56-day follow-up. Within-group PK parameters were consistent. Mean tesidolumab exposure decreased 34%, clearance increased 63%, and half-life decreased 41% comparing tesidolumab + IVIg to tesidolumab alone. IVIg influence on tesidolumab elimination was most evident in the first 3 weeks. Complete suppression of both total and alternative complement activities was maintained for 4 weeks in the tesidolumab alone group and for 2 weeks in the tesidolumab + IVIg group. Tesidolumab was well tolerated. IVIg infused before tesidolumab affected tesidolumab PK and PD, resulting in a shortened period of full complement activity inhibition. These findings suggest a clinically relevant impact of IVIg on monoclonal antibody clearance and indirectly hint at an IVIg mechanism of action in treating autoimmune diseases and allosensitization by accelerating pathogenic IgG antibody degradation. Trial registration number: NCT02878616
Outline for a Perspective Article on Novartis’ approach to compound screening for hit and lead finding and the design of compounds sets for diversity driven screening
No full textDescription of the new NIBR compound screening deck and the underlying design principles. We plan to publish this as a perspective in JMC. It will include our solubility and permeability model, Methods of defining diversity, sub-structure filters and be examplified with structures from our collection, which are non-exclusive and already know in the public Domain (with PubChem numbers).
The actual article has not yet been written and will only be submitted to OAK once we are invited by JMC to submit a perspective
Development of a robust protocol for the synthesis of 6-hydroxybenzofuran-3-carboxylic acid
No full textBenzofuran scaffolds are fundamental moieties found in a variety of biologically active natural products and synthetic drugs. In the course of one of our development programs, we needed to develop a practical and cost-effective manufacturing approach to such a benzofuran scaffold. Here, a 4-step 1-pot process is reported, allowing the access to a 6-hydroxybenzofuran-3-carboxylic acid structure as highly robust. A 1H NMR monitoring study allowed a better understanding of the overall sequence of event and the nature of the detected intermediates. After 6 steps, including the optimized tandem process, the desired hydroxylated benzofuran was obtained with 40% yield and a purity above 99%
Protein phosphatase 1 regulatory subunit 1A regulates cell cycle progression in Ewing sarcoma
No full textIntroduction: We recently identified protein phosphatase 1 regulatory subunit 1A (PPP1R1A) as oneof the EWS/FLI core targets that promotes tumor growth and metastasis in Ewing sarcoma (ES), an aggressive pediatric bone and soft tissue tumor. In the current study, we seek to further define the role of PPP1R1A in ES and identify rational combinatorial therapy with improved and specific efficacy in treating primary and metastatic ES.
Experimental design: We evaluated ES cell proliferation and cell cycle progression in control and PPP1R1A depleted ES cells. PPP1R1A regulation of cell cycle modulators was analyzed to characterize the underlying mechanism of PPP1R1A mediated cell cycle control. The effects of combination of PPP1R1A and IGF-1R inhibition on ES cell viability and migration in vitro as well as tumor growth and metastasis in an orthotopic xenograft mouse model were investigated.
Results: PPP1R1A regulates ES cell cycle in G1/S phase by down-regulating cell cycle inhibitors p21Cip1 and p27Kip1 which results in Rb protein hyperphosphorylation and by promoting normal transcription of replication-dependent histone genes. Furthermore, the combination of PPP1R1A and IGF-1R inhibition induced a synergistic/additive effect on decreasing cell proliferation and migration in vitro and xenograft tumor growth and metastasis in vivo.
Conclusions: Taken together, our findings suggest a role of PPP1R1A as an ES specific cell cycle modulator and that simultaneous targeting of PPP1R1A and IGF-1R pathways is a promising specific and effective strategy to treat both primary and metastatic ES
On the importance of metabolic stability to achieve high oral exposures for cyclic peptides
No full textFollowing up on a previous publication in which we reported a high liver first pass effect in rats for the cyclic peptide (1) [Ala-Leu-NMe-D-Leu-NMe-Leu-Leu-D-Pro], we decided to investigate the type of metabolites formed and to suggest solutions to this problem. As a result of a bile duct cannulation study in rats and subsequent derivatization of this peptide by an isolated Cyp-enzyme, several hydroxylated variants were identified. Cyclopropyl-Ala (Cpa) residues as surrogates for Leu alleviated metabolism at these particular side chains. Significant progress was achieved, when in addition the D-Pro residue was exchanged by 4,4 difluoro-D-Pro (DiF-D-Pro). Albeit the Ala was kept constant in this process, in the corresponding in-vivo studies in rats, peptide (6) [Ala-Cpa-NMe-D-Cpa-NMe-Cpa-Cpa-4,4 difluro-D-Pro] exhibited M exposures at 3mg/kg and an absolute oral bioavailability of > 90%. Thus, we conclude the Cpa- and DiF-D-Pro residues are metabolically stable isosteres for Leu, and D-Pro respectively
Enhanced electrospray in-source fragmentation for higher sensitivity data independent acquisition and autonomous METLIN molecular identification
No full textElectrospray ionization (ESI) in-source fragmentation (ISF) has traditionally been minimized to promote precursor molecular ion formation, and therefore its value in molecular identification underappreciated. Recently a METLIN-guided in-source annotation (MISA) algorithm was introduced to increase confidence in putative identifications by using ubiquitous in-source fragments. However, MISA is limited by ESI sources that are generally designed to minimize ISF. In this study, enhanced ISF with MISA (eMISA) was created by tuning the ISF conditions to generate in-source fragmentation patterns comparable with higher energy fragments generated at higher collision energies as deposited in the METLIN MS/MS library, without compromising the intensity of precursor ions (median loss ≤ 10% in both positive and negative ionization modes). The analysis of 50 molecules was used to validate the approach in comparison to MS/MS spectra produced via data dependent acquisition (DDA) and data independent acquisition mode (DIA) with quadrupole time-of-flight mass spectrometry (QTOF-MS). Enhanced ISF as compared to QTOF DDA, enables for higher peak intensities for the precursor ions (median: 18 times at negative mode and 210 times at positive mode), with the eMISA fragmentation patterns consistent with METLIN for over 90% of the molecules with respect to fragment relative intensity and m/z. eMISA also provides higher peak intensity as opposed to QTOF DIA with a median increase of 20% at negative mode and 80% at positive mode for all precursor ions. Metabolite identification with eMISA was also successfully validated from the analysis of a metabolic extract from macrophages. An interesting side benefit of enhanced ISF is that it significantly improved the compound identification confidence with low resolution single quadrupole mass spectrometry-based untargeted LC/MS experiments. Overall, enhanced ISF allowed for eMISA to be used as a more sensitive alternative to other QTOF DIA and DDA approaches, and further, it enables the acquisition of ESI TOF and ESI single quadrupole mass spectrometry instrumentation spectra with higher sensitivity and improved molecular identification confidence