S‐Adenosylmethionine (SAM) is an enzyme cofactor involved in methylation, aminopropyl transfer, and radical reactions. This versatility renders SAM‐dependent enzymes of great interest in ...biocatalysis. The usage of SAM analogues adds to this diversity. However, high cost and instability of the cofactor impedes the investigation and usage of these enzymes. While SAM regeneration protocols from the methyltransferase (MT) byproduct S‐adenosylhomocysteine are available, aminopropyl transferases and radical SAM enzymes are not covered. Here, we report a set of efficient one‐pot systems to supply or regenerate SAM and SAM analogues for all three enzyme classes. The systems’ flexibility is showcased by the transfer of an ethyl group with a cobalamin‐dependent radical SAM MT using S‐adenosylethionine as a cofactor. This shows the potential of SAM (analogue) supply and regeneration for the application of diverse chemistry, as well as for mechanistic studies using cofactor analogues.
The biomimetic regeneration system for S‐adenosylmethionine (SAM) and SAM analogues presented is based on the salvage of the adenine moiety and in situ supply of d‐ribose and polyphosphate. It is compatible with a broad range of SAM‐dependent enzymes including aminopropyl transferases, and is shown to support ethylation reactions with both conventional and radical SAM methyltransferases.
Corticosteroid receptors in the mammalian brain mediate genomic as well as non-genomic actions. Although receptors mediating genomic actions were already cloned 35 years ago, it remains unclear ...whether the same molecules are responsible for the non-genomic actions or that the latter involve a separate class of receptors.
Here we focus on one type of corticosteroid receptors, i.e. the mineralocorticoid receptor (MR). We summarize some of the known properties and the current insight in the localization of the MR in peripheral cells and neurons, especially in relation to non-genomic signaling. Previous studies from our own and other labs provided evidence that MRs mediating non-genomic actions are identical to the ones involved in genomic signaling, but may be translocated to the plasma cell membrane instead of the nucleus. With fixed cell imaging and live cell imaging techniques we tried to visualize these presumed membrane-associated MRs, using antibodies or overexpression of MR-GFP in COS7 and hippocampal cultured neurons. Despite the physiological evidence for MR location in or close to the cell membrane, we could not convincingly visualize membrane localization of endogenous MRs or GFP-MR molecules. However, we did find punctae of labeled antibodies intracellularly, which might indicate transactivating spots of MR near the membrane. We also found some evidence for trafficking of MR via beta-arrestins. In beta-arrestin knockout mice, we didn't observe metaplasticity in the basolateral amygdala anymore, indicating that internalization of MRs could play a role during corticosterone activation. Furthermore, we speculate that membrane-associated MRs could act indirectly via activating other membrane located structures like e.g. GPER and/or receptor tyrosine kinases.
Platinum-based chemotherapeutics exhibit excellent antitumor properties. However, these drugs cause severe side effects including toxicity, drug resistance, and lack of tumor selectivity. ...Tumor-targeted drug delivery has demonstrated great potential to overcome these drawbacks. Herein, we aimed to design radioactive bisphosphonate-functionalized platinum (
Pt-BP) complexes to confirm preferential accumulation of these Pt-based drugs in metabolically active bone. In vitro NMR studies revealed that release of Pt from Pt BP complexes increased with decreasing pH. Upon systemic administration to mice, Pt-BP exhibited a 4.5-fold higher affinity to bone compared to platinum complexes lacking the bone-seeking bisphosphonate moiety. These Pt-BP complexes formed less Pt-DNA adducts compared to bisphosphonate-free platinum complexes, indicating that in vivo release of Pt from Pt-BP complexes proceeded relatively slow. Subsequently, radioactive
Pt-BP complexes were synthesized using
Pt(NO
)
(en) as precursor and injected intravenously into mice. Specific accumulation of
Pt-BP was observed at skeletal sites with high metabolic activity using micro-SPECT/CT imaging. Furthermore, laser ablation-ICP-MS imaging of proximal tibia sections confirmed that
Pt BP co-localized with calcium in the trabeculae of mice tibia.
Hydroxyapatite (HA) nanoparticles are commonly used as building blocks in the design of bone-substituting biomaterials. Recently, these nanoparticles have been considered for the treatment of ...metastasis disease, since their pH-dependent dissolution behavior allows for precise tuning of release kinetics of loaded cargo. Herein we show that the capacity of drug-loaded nanoparticles stabilized with citrate ions reduce cancer cell survival in an embryonic zebrafish xenograft model. In particular,
in vitro
studies demonstrate that PtPP-loaded HA nanoparticles exhibit anti-proliferative activity against breast cancer cells at reduced pH.
In vivo
studies using an embryonic zebrafish xenograft model reveal that PtPP co-delivered with human breast cancer cells strongly reduce cancer cell survival. Similarly, co-injection of breast cancer cells with citrate-functionalized and PtPP-loaded HA nanoparticles into zebrafish significantly reduces survival of cancer cells due to release of chemotherapeutically active kiteplatin species. These results demonstrate the preclinical efficacy of drug-loaded nanoparticles against human breast cancer cells in a xenogenic embryonic
in vivo
model.
Kiteplatin-pyrophosphate (PtPP) loaded hydroxyapatite nanoparticles effectively reduce breast cancer cell
in vivo
, which opens up new opportunities for controlled local treatment of bone metastases.
This study was designed to assess the efficacy of perioperative administration of celecoxib (Celebrex; Pharmacia GmbH, Erlangen, Germany) in reducing pain and opioid requirements after single-level ...lumbar microdiscectomy.
We studied 34 patients (mean age, 44.26 yr; standard deviation SD, 13.09 yr) allocated randomly to receive celecoxib 200 mg twice a day for 72 hours starting on the evening before surgery or placebo capsules in a double-blind study. Fourteen patients received 20 to 80 mg dexamethasone intravenously during surgery (mean, 40 mg; SD, 19.22 mg) because of visible signs of compression of the affected nerve root. After lumbar disc surgery, patients were monitored for visual analog scores for pain at rest and on movement, patient-controlled analgesia (PCA) piritramide requirements, and von Frey thresholds in the wound area.
Pain scores decreased and wound von Frey thresholds increased continuously until discharge, with no intergroup differences. Mean 24-hour PCA piritramide requirements were 22.63 mg (SD, 23.72 mg) and 26.14 mg (SD, 22.57 mg) in the celecoxib and placebo groups, respectively (P = not significant). However, patients with intraoperative dexamethasone (n = 14) required only 10.29 mg (SD, 8.55 mg) 24-hour PCA piritramide, in contrast to the 34.25 mg (SD, 24.69 mg) needed in those who did not receive intraoperative dexamethasone (P = 0.001). In addition, 24 hours after the operation, pain scores on movement were significantly lower in the dexamethasone subgroup (P = 0.003).
Celecoxib has no effect on postoperative pain scores and PCA piritramide requirements. The intraoperative use of 20 to 80 mg dexamethasone is able to significantly decrease postoperative piritramide consumption and pain scores on the first day after surgery.
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