Affibody molecules are a class of engineered affinity proteins with proven potential for therapeutic, diagnostic and biotechnological applications. Affibody molecules are small (6.5
kDa) single ...domain proteins that can be isolated for high affinity and specificity to any given protein target. Fifteen years after its discovery, the Affibody technology is gaining use in many groups as a tool for creating molecular specificity wherever a small, engineering compatible tool is warranted. Here we summarize recent results using this technology, propose an Affibody nomenclature and give an overview of different HER2-specific Affibody molecules. Cumulative evidence suggests that the three helical scaffold domain used as basis for these molecules is highly suited to create a molecular affinity handle for vastly different applications.
Despite intense interest in expanding chemical space, libraries containing hundreds-of-millions to billions of diverse molecules have remained inaccessible. Here we investigate structure-based ...docking of 170 million make-on-demand compounds from 130 well-characterized reactions. The resulting library is diverse, representing over 10.7 million scaffolds that are otherwise unavailable. For each compound in the library, docking against AmpC β-lactamase (AmpC) and the D
dopamine receptor were simulated. From the top-ranking molecules, 44 and 549 compounds were synthesized and tested for interactions with AmpC and the D
dopamine receptor, respectively. We found a phenolate inhibitor of AmpC, which revealed a group of inhibitors without known precedent. This molecule was optimized to 77 nM, which places it among the most potent non-covalent AmpC inhibitors known. Crystal structures of this and other AmpC inhibitors confirmed the docking predictions. Against the D
dopamine receptor, hit rates fell almost monotonically with docking score, and a hit-rate versus score curve predicted that the library contained 453,000 ligands for the D
dopamine receptor. Of 81 new chemotypes discovered, 30 showed submicromolar activity, including a 180-pM subtype-selective agonist of the D
dopamine receptor.
We uncover the fine structure of a silicon vacancy in isotopically purified silicon carbide (4H-SiC28 ) and reveal not yet considered terms in the spin Hamiltonian, originated from the trigonal ...pyramidal symmetry of this spin-3/2 color center. These terms give rise to additional spin transitions, which would be otherwise forbidden, and lead to a level anticrossing in an external magnetic field. We observe a sharp variation of the photoluminescence intensity in the vicinity of this level anticrossing, which can be used for a purely all-optical sensing of the magnetic field. We achieve dc magnetic field sensitivity better than 100nT/Hz within a volume of 3×10−7mm3 at room temperature and demonstrate that this contactless method is robust at high temperatures up to at least 500 K. As our approach does not require application of radio-frequency fields, it is scalable to much larger volumes. For an optimized light-trapping waveguide of 3mm3 , the projection noise limit is below 100fT/Hz .
The
Pu,
U, and
Am concentrations and
Pu/
Pu,
U/
U, and
U/
U atom ratios were measured in the hair and nail samples using a new method utilized TEVA, UTEVA, and DGA extraction chromatography and ...multi-collector ICP-MS. Samples were collected from individuals who donated their bodies to the United States Transuranium and Uranium Registries. The concentration of
Pu ranged from 0.22 to 15.8 ng/kg. The
Pu/
Pu isotopic ratios ranged from 0.026 to 0.127 which is consistent with weapons-grade plutonium. Concentration of uranium fell between 1.84 μg/kg and 29.5 μg/kg and
U/
U ratios ranged from 4.8 × 10
to 7.6 × 10
. Elevated
U/
U atom ratios were measured in two cases and ranged from 5.0 × 10
- 2.4 × 10
indicating exposure to spent or reprocessed uranium material. The concentration of
Am was measured in four hair samples and ranged from 0.02 to 0.21 ng/kg.
Overbinding of ions is a common and well-known problem in classical molecular dynamics simulations. One of its main causes is the absence of electronic polarizability in the force fields. The current ...approaches for minimizing overbinding typically either retain the original charges and use an ad hoc readjustment of the Lennard-Jones parameters as done in the nonbonded fix (NBFIX) approach or rescale the charges using a theoretical framework. The goal in the latter is to include shielding produced by the missing electronic polarizability as done in the electronic continuum correction (ECC) approach. NBFIX and ECC are the most common corrections, and we compare their performance to the default parameterizations provided by five different commonly used biomolecular force fields, OPLS-AA/L, CHARMM27, CHARMM36m, CHARMM22*, and AMBER99SB-ILDN. As test systems, we use poly-α,l-glutamic and poly-α,l-aspartic amino acid molecules in explicit water together with Na+ and K+ counterions. We demonstrate that the different force fields yield results that are not only quantitatively but also qualitatively different. The resulting structures of the macroions depend strongly on the model for ions. NBFIX corrections alleviate the problem of overbinding, resulting in extended peptides. The ECC corrections depend nontrivially on the original underlying model, and despite being based on a theoretical framework, they cannot always solve the problem.
Abstract
The United States Transuranium and Uranium Registries (USTUR) is a research program that studies actinide biokinetics in occupationally exposed individuals with known intakes of these ...elements. Electron paramagnetic resonance (EPR) in tooth enamel was applied to reconstruct external doses of nine USTUR registrants. Only in two cases there is a reasonable agreement between the EPR-measured dose and the worksite external dose record. For two registrants, high EPR doses can be explained by possible cancer radiotherapy. For the remaining five cases, EPR doses significantly exceed official occupational doses with no plausible explanation for the observed discrepancy. More EPR dose measurements need to be done to explain this anomaly.
The detection of cell-bound proteins that are produced due to aberrant gene expression in malignant tumors can provide important diagnostic information influencing patient management. The use of ...small radiolabeled targeting proteins would enable high-contrast radionuclide imaging of cancers expressing such antigens if adequate binding affinity and specificity could be provided. Here, we describe a HER2-specific 6 kDa Affibody molecule (hereinafter denoted Affibody molecule) with 22 pmol/L affinity that can be used for the visualization of HER2 expression in tumors in vivo using gamma camera. A library for affinity maturation was constructed by re-randomization of relevant positions identified after the alignment of first-generation variants of nanomolar affinity (50 nmol/L). One selected Affibody molecule, Z(HER2:342) showed a >2,200-fold increase in affinity achieved through a single-library affinity maturation step. When radioiodinated, the affinity-matured Affibody molecule showed clear, high-contrast visualization of HER2-expressing xenografts in mice as early as 6 hours post-injection. The tumor uptake at 4 hours post-injection was improved 4-fold (due to increased affinity) with 9% of the injected dose per gram of tissue in the tumor. Affibody molecules represent a new class of affinity molecules that can provide small sized, high affinity cancer-specific ligands, which may be well suited for tumor imaging.
Abstract
Molecular imaging of the gastrin-releasing peptide receptor (GRPR) could improve patient management in prostate cancer. This study aimed to produce gallium-66 (T
½
= 9.5 h) suitable for ...radiolabeling, and investigate the imaging properties of gallium-66 labeled GRPR-antagonist NOTA-PEG
2
-RM26 for later-time point PET-imaging of GRPR expression. Gallium-66 was cyclotron-produced using a liquid target, and enriched
66
ZnZn(NO
3
)
2
. In vitro,
66
GaGa-NOTA-PEG
2
-RM26 was characterized in GRPR-expressing PC-3 prostate cancer cells. In vivo, specificity test and biodistribution studies were performed 3 h and 22 h pi in PC-3 xenografted mice. microPET/MR was performed 3 h and 22 h pi. Biodistribution of
66
GaGa-NOTA-PEG
2
-RM26 was compared with
68
GaGa-NOTA-PEG
2
-RM26 3 h pi.
66
GaGa-NOTA-PEG
2
-RM26 was successfully prepared with preserved binding specificity and high affinity towards GRPR.
66
GaGa-NOTA-PEG
2
-RM26 cleared rapidly from blood via kidneys. Tumor uptake was GRPR-specific and exceeded normal organ uptake. Normal tissue clearance was limited, resulting in no improvement of tumor-to-organ ratios with time. Tumors could be clearly visualized using microPET/MR. Gallium-66 was successfully produced and
66
GaGa-NOTA-PEG
2
-RM26 was able to clearly visualize GRPR-expression both shortly after injection and on the next day using PET. However, delayed imaging did not improve contrast for Ga-labeled NOTA-PEG
2
-RM26.
An ICP-MS method was developed to measure the actinides in autopsy brain tissue of an occupationally exposed individual.
239
Pu,
240
Pu,
241
Am, and
238
U concentrations as well as
240
Pu/
239
Pu,
...235
U/
238
U atom ratios were measured by quadrupole ICP-MS following extraction chromatography. The
239
Pu concentrations measured in the cerebral lobe of the right side of the brain was 0.66 ± 0.08 ng/kg. The
239
Pu/
240
Pu ratio was 0.071 ± 0.025. The
241
Am level was below the LOD. The
238
U concentration was 106.6 ± 0.29 ng/kg and the
235
U/
238
U ratio was 0.00703 ± 0.00087.
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