ABSTRACT
Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase involved in the development of several human cancers and, as a result, is a recognized target for the development of small-molecule inhibitors for the treatment of ALK-positive malignancies. Here, we present the crystal structures of the unphosphorylated human ALK kinase domain in complex with the ATP competitive ligands PHA-E429 and NVP-TAE684. Analysis of these structures provides valuable information concerning the specific characteristics of the ALK active site as well as giving indications about how to obtain selective ALK inhibitors. In addition, the ALK-KD-PHA-E429 structure led to the identification of a potential regulatory mechanism involving a link made between a short helical segment immediately following the DFG motif and an N-terminal two-stranded beta-sheet. Finally, mapping of the activating mutations associated with neuroblastoma onto our structures may explain the roles these residues have in the activation process.
Subject(s)
Models, Molecular , Protein-Tyrosine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/chemistry , Pyrimidines/chemistry , Anaplastic Lymphoma Kinase , Animals , Cell Line , Enzyme Inhibitors/chemistry , Humans , Protein Structure, Tertiary , Protein-Tyrosine Kinases/genetics , Protein-Tyrosine Kinases/metabolism , Receptor Protein-Tyrosine Kinases , SpodopteraABSTRACT
OBJECTIVE: Cholinesterase (ChE) inhibitors are the only medications approved for the treatment of Alzheimer's disease (AD). The features of ChE inhibitors differ considerably. In addition to acetylcholinesterase (AChE) inhibition, rivastigmine also inhibits butrylcholinesterase (BuChE), providing dual AChE and BuChE inhibition. An observational study was performed to determine the response in routine clinical practice to switching AD patients to rivastigmine from a selective AChE inhibitor when that treatment no longer delivered a satisfactory clinical response. RESEARCH DESIGN AND METHODS: A prospective, multicentre, 3-month observational trial in patients with mild to moderately severe AD (adjusted Mini Mental State Examination [MMSE] score 10-26) deteriorating (at least 2 adjusted MMSE points in last 6 months) on selective AChE inhibitor treatment. Adjusted MMSE, activities of daily living (ADL) and instrumental activities of daily living (IADL), the Zarit caregiver burden and global function (short Clinical Global Impression of Change, CGIC) scores were noted before the switch and 3 months after the switch. RESULTS: 225 patients entered the study. The switches made were from donepezil to rivastigmine in (D-R) in 188 patients, galantamine to rivastigmine (G-R) in 33 patients and donepezil to galantamine (D-G) in four patients. Ten patients discontinued due to adverse events and eight for other reasons. More than half of the switches were within 36 hours of a patient's first treatment visit. In the D-R and G-R groups, 67.7% and 66.7% of patients responded (CGIC score < or = 4), respectively. In non-responders, worsening (CGIC score 5-7) was mild in approximately 80% or more of patients. Adjusted MMSE improved after the switch from both donepezil and galantamine to rivastigmine (+0.69 +/- 3.2, p = 0.008 and +0.6 +/- 1.6, p = 0.05, respectively). Mean ADL, IADL, and Zarit scores remained stable. The proportion of patients on concomitant antipsychotic therapy diminished by 30.5% and benzodiazepines were discontinued in all patients, except one. CONCLUSIONS: AD patients deteriorating on selective AChE inhibitor treatment can benefit from switching to a dual AChE-BuChE inhibitor, such as rivastigmine, in terms of stabilization of disease, improvement in cognitive function and reduction in the burden of concomitant psychoactive treatment. The switch was well tolerated. Confirmation of these results is required in a controlled study.
Subject(s)
Alzheimer Disease/drug therapy , Cholinesterase Inhibitors/therapeutic use , Phenylcarbamates/therapeutic use , Acetylcholinesterase/metabolism , Aged , Aged, 80 and over , Alzheimer Disease/physiopathology , Butyrylcholinesterase/metabolism , Female , Humans , Mental Status Schedule , Prospective Studies , Rivastigmine , Treatment OutcomeABSTRACT
We have expressed in Escherichia coli a soluble, truncated form of the human 55 kDa Tumor Necrosis Factor (TNF) receptor. For this purpose a plasmid was constructed which contains the extracellular domain of the 55 kDa TNF receptor fused to the coding sequence of the IgG binding domains of protein A from Staphylococcus aureus. The fusion product (TNFR-PA) obtained in E. coli is a soluble protein which bound human TNF alpha (huTNF alpha) with high affinity. In ligand-blotting experiments huTNF alpha bound to a single 52 kDa protein, a molecular mass corresponding to that expected for the monomeric fusion product. In gel filtration experiments binding activity was recovered from fractions that eluted at a volume corresponding to 140-150 kDa. TNFR-PA neutralized huTNF alpha in an in vitro cytotoxicity assay.
Subject(s)
Escherichia coli/metabolism , Receptors, Tumor Necrosis Factor/biosynthesis , Recombinant Fusion Proteins/biosynthesis , Chromatography, Gel , Cloning, Molecular , Humans , Molecular Weight , Receptors, Tumor Necrosis Factor/genetics , Solubility , Staphylococcal Protein A/biosynthesisABSTRACT
We have isolated and sequenced partial cDNA clones that encode SO-6, a ribosome-inactivating protein from Saponaria officinalis. A cDNA library was constructed from the leaves of this plant and screened with synthetic oligonucleotide probes representing various portions of the protein. The deduced amino acid sequence shows the signal peptide and a coding region virtually accounting for the entire amino acid sequence of SO-6. The sequence reveals regions of similarity to other ribosome-inactivating proteins, especially in a region of the molecule where critical amino acid residues might participate in the active site.
