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Abstract: Background context: Intradural disc herniation is relatively rare complication of the spinal degenerative process that occurs most frequently in the lumbar part of the spine. Both myelographic and magnetic resonance features of this entity have been described, and the mechanism of intradural herniation has already been proposed and generally accepted. In this article, we present a case of spontaneous resorption of an intradural, fragmented intervertebral disc. Spontaneous resorption of intradural disc fragments has not been previously reported. Purpose: To discuss a possible mechanism of spontaneous resorption of the subdural disc fragments. Study design: Case report and literature review. Methods: Radiological follow-up of a 46-year-old man with the intradural herniation of disc fragments. Conclusion: The reaction generated by the meninges might lead to the complete resorption of intrathecally localized disc fragments. [Copyright &y& Elsevier]

Investigates structural perturbations caused by mutagenesis in the refolding of human carbonic anhydrase II. Intrinsic tryptophan fluorescence; Formation of tertiary structures at various tryptophan side chains; Equilibrium unfolding intermediates and kinetic intermediates.

Presents the experiment conducted by the authors on mapping the folding intermediate of human carbonic anhydrase II by probing the substructure by chemical reactivity and spin and fluorescence labeling of engineered Cysteine residues. Introduction; Materials used and methodology adopted; Results; Discussion.

Measures the fluorescence spectra produced by pseudo-wild-type human carbonic anhydrase II and mutants in which the tryptophan residues had been replaced by phenylalanine or cysteine residues. Indication of no long range effect of mutation on structure; Deduction of individual contributions of seven tryptophan residues from measurements on native proteins; Materials, methods, results, discussion and conclusion.

The kinetics for trypsin cleavage of different fusion proteins, consisting of human proinsulin and two IgG-binding domains (ZZ), were investigated. To achieve simultaneous removal of the fusion tag and processing of proinsulin to insulin and free C peptide. three versions of the ZZ-proinsulin fusion protein were generated, having different trypsin-sensitive cleavage sites, Mg. Lys-Arg or Lys. The ZZ-proinsulin fusion proteins which accumulated as inclusion bodies in Escherichia coli cells were solubilized, refolded and purified by IgG affinity chromatography. The yield of ZZ-proinsulin monomers exceeded 90%. The kinetics for the trypsin cleavage revealed unexpected differences when comparing the three linkers amid it was found that the single arginine linker was most efficiently processed. Characterization of the cleavage products by reverse-phase chromatography, mass spectrometry and N-terminal sequencing verified that human insulin and C peptide were generated. The results demonstrate that high yields of native insulin. C peptide and affinity tag can be achieved by simultaneous cleavage of a fusion protein at three different trypsin-sensitive sites in a single step. The implications for production and recovery of various recombinant proteins arc discussed. [ABSTRACT FROM AUTHOR]

Site-specific mutagenesis has been used to replace amino acid residues in the active site of human carbonic anhydrase II with residues characterizing carbonic anhydrases I. Previous studies of [Thr200→ His]isoenzyme II [Behravan, G., Jonsson, B.-H. & Lindskog, S. (1990) Eur. J. Biochem. 190, 351–357] showed that His200 is important for the specific catalytic properties of isoenzymes I. In this paper some properties of two single mutants, Asn62 → Val and Asn67 → His, as well as a double mutant, Asn67 → His/Thr200 → His, are described. The results show that neither Va162 nor His67 give rise to isoenzyme-I-like properties, while the double mutant behaves like the single mutant with His200. At pH 8.9, the variant with Va162 has a higher value of kcat/Km for CO2 hydration than unmodified isoenzyme II, whereas the variant with His67 has an enhanced kcat value. The replacement of Asn62 with Val results in a 20% increase of the 4-nitrophenyl acetate hydrolase activity. For the double mutant, the esterase activity is quite close to that calculated on the assumption that the effects of the two single mutations on the free energy of activation are additive. [ABSTRACT FROM AUTHOR]

Objective: To study: (i) acute computed tomography (CT) characteristics and clinical outcome; (ii) clinical course and (iii) Corticosteroid Randomisation after Significant Head Injury acute calculator protocol (CRASH) model and clinical outcome in patients with severe traumatic brain injury (sTBI). Methods: Initial CT (CTi) and CT 24 hours post-trauma (CT24) were evaluated according to Marshall and Rotterdam classifications. Rancho Los Amigos Cognitive Scale-Revised (RLAS-R) and Glasgow Outcome Scale Extended (GOSE) were assessed at three months and one year post-trauma. The prognostic value of the CRASH model was evaluated. Results: Thirty-seven patients were included. Marshall CTi and CT24 were significantly correlated with RLAS-R at three months. Rotterdam CT24 was significantly correlated with GOSE at three months. RLAS-R and the GOSE improved significantly from three months to one year. CRASH predicted unfavourable outcome at six months for 81% of patients with bad outcome and for 85% of patients with favourable outcome according to GOSE at one year. Conclusion: Neither CT nor CRASH yielded clinically useful predictions of outcome at one year postinjury. The study showed encouragingly many instances of significant recovery in this population of sTBI. The combination of lack of reliable prognostic indicators and favourable outcomes supports the case for intensive acute management and rehabilitation as the default protocol in the cases of sTBI. [ABSTRACT FROM AUTHOR]

Introduction This is the first report on the simultaneous successful treatment of a large ruptured saccular aneurysm and stenotic parent artery with a single flow-diverting stent. Case presentation We report the case of a 68-year-old Caucasian man with occlusion of the right vertebral artery and a ruptured aneurysm at the junction of the left posterior inferior cerebellar arteryleft vertebral artery that was successfully treated by the deployment of a single flow-diverting stent in the stenotic left vertebral artery. Stent deployment was complicated by thrombotic occlusion of the basilar artery, which was successfully reopened. The patient recovered completely, and follow-up angiography at 4 months and 1 year showed patent vertebral artery with gradual shrinkage of the aneurysm. Conclusions This report contributes to the literature on treatment of large ruptured aneurysms localized in stenotic arteries and in areas of the endocranium where a mass of embolic material in the aneurysm (coils) might compromise the circulation in the parent blood vessel or compress vital brain structures. [ABSTRACT FROM AUTHOR]

Introduction: This is the first report on the simultaneous successful treatment of a large ruptured saccular aneurysm and stenotic parent artery with a single flow-diverting stent.Case Presentation: We report the case of a 68-year-old Caucasian man with occlusion of the right vertebral artery and a ruptured aneurysm at the junction of the left posterior inferior cerebellar artery-left vertebral artery that was successfully treated by the deployment of a single flow-diverting stent in the stenotic left vertebral artery. Stent deployment was complicated by thrombotic occlusion of the basilar artery, which was successfully reopened. The patient recovered completely, and follow-up angiography at 4 months and 1 year showed patent vertebral artery with gradual shrinkage of the aneurysm.Conclusions: This report contributes to the literature on treatment of large ruptured aneurysms localized in stenotic arteries and in areas of the endocranium where a mass of embolic material in the aneurysm (coils) might compromise the circulation in the parent blood vessel or compress vital brain structures. [ABSTRACT FROM AUTHOR]

Background: Our aim was to evaluate the long-term treatment results in patients with intracranial aneurysms treated with endovascular techniques. Methods: Forty-four patients treated due to intracranial aneurysms between 1996 and 2002 were investigated with a time-of-flight sequence magnetic resonance angiography (TOF MRA). Results: Depending on the assessment, 47% to 51% of the treated aneurysms had a residual neck at the last digital subtraction angiography follow-up. There was filling of the aneurysm base (2%) in only 1 patient, whereas the remaining aneurysms were totally occluded. A TOF MRA performed 6 to 14 (mean 9.68) years after the last procedure showed a stable result in 93.9% of the treated aneurysms. There were no de novo aneurysms and previously untreated aneurysms were unchanged in size. Conclusion: Our long-term follow-up showed a stable result in previously coiled intracranial aneurysms. [ABSTRACT FROM PUBLISHER]

The purpose of the study is to evaluate patients with wide-necked or complex aneurysms of the anterior circulation who underwent Solitaire™ AB Neurovascular Remodeling Device-assisted coil embolization. From February 2008 to March 2009, consecutive data were collected from 45 patients with anterior circulation aneurysms. Eighteen of the patients presented with acute subarachnoid hemorrhage. Forty-six aneurysms were treated with the aid of different applications ( n = 49) of the Solitaire™ AB Remodeling Device followed by standard coiling procedure ( n = 43) using bioactive coils or/and bare coils. Successful positioning of the remodeling device was obtained in 95.9% of the cases. There were two thromboembolic complications (4.1%) and one severe vasospasm requiring retrieval of the device. Permanent procedural morbidity was observed in one patient (2%). The proportion of patients in whom Raymond class 1 occlusion was obtained was 53.5% ( n = 23). Raymond class 2 occlusion was achieved in 42% ( n = 18) and Raymond class 3 occlusion in 4.7% ( n = 2). Thirty-nine patients left the hospital with a good clinical status. The initial technical and clinical results of Solitaire™ AB device-assisted coiling of aneurysms in the anterior circulation are highly encouraging. This technique may enhance the possibilities of the endovascular treatment of these aneurysms in clinical routine. [ABSTRACT FROM AUTHOR]

Abstract: Affibody molecules are non-immunoglobulin-derived affinity proteins based on a three-helical bundle protein domain. Here, we describe the design process of an optimized Affibody molecule scaffold with improved properties and a surface distinctly different from that of the parental scaffold. The improvement was achieved by applying an iterative process of amino acid substitutions in the context of the human epidermal growth factor receptor 2 (HER2)-specific Affibody molecule ZHER2:342. Replacements in the N-terminal region, loop 1, helix 2 and helix 3 were guided by extensive structural modeling using the available structures of the parent Z domain and Affibody molecules. The effect of several single substitutions was analyzed followed by combination of up to 11 different substitutions. The two amino acid substitutions N23T and S33K accounted for the most dramatic improvements, including increased thermal stability with elevated melting temperatures of up to +12 °C. The optimized scaffold contains 11 amino acid substitutions in the nonbinding surface and is characterized by improved thermal and chemical stability, as well as increased hydrophilicity, and enables generation of identical Affibody molecules both by chemical peptide synthesis and by recombinant bacterial expression. A HER2-specific Affibody tracer, [MMA-DOTA-Cys61]-ZHER2:2891-Cys (ABY-025), was produced by conjugating MMA-DOTA (maleimide-monoamide-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) to the peptide produced either chemically or in Escherichia coli. ABY-025 showed high affinity and specificity for HER2 (equilibrium dissociation constant, K D, of 76 pM) and detected HER2 in tissue sections of SKOV-3 xenograft and human breast tumors. The HER2-binding capacity was fully retained after three cycles of heating to 90 °C followed by cooling to room temperature. Furthermore, the binding surfaces of five Affibody molecules targeting other proteins (tumor necrosis factor α, insulin, Taq polymerase, epidermal growth factor receptor or platelet-derived growth factor receptor β) were grafted onto the optimized scaffold, resulting in molecules with improved thermal stability and a more hydrophilic nonbinding surface. [Copyright &y& Elsevier]

Radiolabeled Affibody-based targeting agent 177Lu-ABY-027, a fusion of an anti-HER2 Affibody molecule with albumin binding domain (ABD) site-specifically labeled at the C-terminus, has demonstrated a promising biodistribution profile in mice; binding of the construct to albumin prevents glomerular filtration and significantly reduces renal uptake. In this study, we tested the hypothesis that site-specific positioning of the chelator at helix 1 of ABD, at a maximum distance from the albumin binding site, would further increase the strength of binding to albumin and decrease the renal uptake. The new construct, ABY-271 with DOTA conjugated at the back of ABD, has been labelled with 177Lu. Targeting properties of 177Lu-ABY-271 and 177Lu-ABY-027 were compared directly. 177Lu-ABY-271 specifically accumulated in SKOV-3 xenografts in mice. The tumor uptake of 177Lu-ABY-271 exceeded uptake in any other organ 24 h and later after injection. However, the renal uptake of 177Lu-ABY-271 was two-fold higher than the uptake of 177Lu-ABY-027. Thus, the placement of chelator on helix 1 of ABD does not provide desirable reduction of renal uptake. To conclude, minimal modification of the design of Affibody molecules has a strong effect on biodistribution, which cannot be predicted a priori. This necessitates extensive structure-properties relationship studies to find an optimal design of Affibody-based targeting agents for therapy. [ABSTRACT FROM AUTHOR]

Radionuclide imaging of HER2 expression in tumours may enable stratification of patients with breast, ovarian, and gastroesophageal cancers for HER2-targeting therapies. A first-generation HER2-binding affibody molecule [99mTc]Tc-ZHER2:V2 demonstrated favorable imaging properties in preclinical studies. Thereafter, the affibody scaffold has been extensively modified, which increased its melting point, improved storage stability, and increased hydrophilicity of the surface. In this study, a second-generation affibody molecule (designated ZHER2:41071) with a new improved scaffold has been prepared and characterized. HER2-binding, biodistribution, and tumour-targeting properties of [99mTc]Tc-labelled ZHER2:41071 were investigated. These properties were compared with properties of the first-generation affibody molecules, [99mTc]Tc-ZHER2:V2 and [99mTc]Tc-ZHER2:2395. [99mTc]Tc-ZHER2:41071 bound specifically to HER2 expressing cells with an affinity of 58 ± 2 pM. The renal uptake for [99mTc]Tc-ZHER2:41071 and [99mTc]Tc-ZHER2:V2 was 25–30 fold lower when compared with [99mTc]Tc-ZHER2:2395. The uptake in tumour and kidney for [99mTc]Tc-ZHER2:41071 and [99mTc]Tc-ZHER2:V2 in SKOV-3 xenografts was similar. In conclusion, an extensive re-engineering of the scaffold did not compromise imaging properties of the affibody molecule labelled with 99mTc using a GGGC chelator. The new probe, [99mTc]Tc-ZHER2:41071 provided the best tumour-to-blood ratio compared to HER2-imaging probes for single photon emission computed tomography (SPECT) described in the literature so far. [99mTc]Tc-ZHER2:41071 is a promising candidate for further clinical translation studies. [ABSTRACT FROM AUTHOR]

Human epidermal growth factor receptor 3 (HER3) has been increasingly scrutinized as a potential drug target since the elucidation of its role in mediating tumor growth and acquired therapy resistance. Affibody molecules are so-called scaffold proteins with favorable biophysical properties, such as a small size for improved tissue penetration and extravasation, thermal and chemical stability, and a high tolerance to modifications. Additionally, affibody molecules are efficiently produced in prokaryotic hosts or by chemical peptide synthesis. We have previously evaluated the biodistribution profiles of five mono- and bivalent anti-HER3 affibody molecules (designated as 3) fused to an albumin-binding domain (designated as A), 3A, 33A, 3A3, A33, and A3, that inhibit ligand-dependent phosphorylation. In the present study, we examined the therapeutic efficacy of the three most promising variants, 3A, 33A, and 3A3, in a direct comparison with the HER3-targeting monoclonal antibody seribantumab (MM-121) in a preclinical BxPC-3 pancreatic cancer model. Xenografted mice were treated with either an affibody construct or MM-121 and the tumor growth was compared to a vehicle group. Receptor occupancy was estimated by positron emission tomography/computed tomography (PET/CT) imaging using a HER3-targeting affibody imaging agent [68Ga]Ga-(HE)3-Z08698-NODAGA. The affibody molecules could inhibit ligand-dependent phosphorylation and cell proliferation in vitro and demonstrated tumor growth inhibition in vivo comparable to that of MM-121. PET/CT imaging showed full receptor occupancy for all tested drug candidates. Treatment with 3A and 3A3 affibody constructs was more efficient than with 33A and similar to the anti-HER3 antibody seribantumab, showing that the molecular design of affibody-based therapeutics targeting HER3 in terms of the relative position of functional domains and valency has an impact on therapeutic effect. [ABSTRACT FROM AUTHOR]