| 1. |
|
|
| 2. |
- Abramsson, Alexandra, 1973, et al.
(författare)
-
Defective N-sulfation of heparan sulfate proteoglycans limits PDGF-BB binding and pericyte recruitment in vascular development
- 2007
-
Ingår i: GENES & DEVELOPMENT. - : Cold Spring Harbor Laboratory. - 0890-9369 .- 1549-5477. ; 21:3, s. 316-331
-
Tidskriftsartikel (refereegranskat)abstract
- During vascular development, endothelial platelet-derived growth factor B (PDGF-B) is critical for pericyte recruitment. Deletion of the conserved C-terminal heparin-binding motif impairs PDGF-BB retention and pericyte recruitment in vivo, suggesting a potential role for heparan sulfate (HS) in PDGF-BB function during vascular development. We studied the participation of HS chains in pericyte recruitment using two mouse models with altered HS biosynthesis. Reduction of N-sulfation due to deficiency in N-deacetylase/N-sulfotransferase-1 attenuated PDGF-BB binding in vitro, and led to pericyte detachment and delayed pericyte migration in vivo. Reduced N-sulfation also impaired PDGF-BB signaling and directed cell migration, but not proliferation. In contrast, HS from glucuronyl C5-epimerase mutants, which is extensively N- and 6-O-sulfated, but lacks 2-O-sulfated L-iduronic acid residues, retained PDGF-BB in vitro, and pericyte recruitment in vivo was only transiently delayed. These observations were supported by in vitro characterization of the structural features in HS important for PDGF-BB binding. We conclude that pericyte recruitment requires HS with sufficiently extended and appropriately spaced N-sulfated domains to retain PDGF-BB and activate PDGF receptor β (PDGFRβ) signaling, whereas the detailed sequence of monosaccharide and sulfate residues does not appear to be important for this interaction.
|
|
| 3. |
- Al-Amin, Rasel A., Researcher, 1983-, et al.
(författare)
-
Monitoring drug–target interactions through target engagement-mediated amplification on arrays and in situ
- 2022
-
Ingår i: Nucleic Acids Research. - : Oxford University Press. - 0305-1048 .- 1362-4962. ; 50:22, s. e129-e129
-
Tidskriftsartikel (refereegranskat)abstract
- Drugs are designed to bind their target proteins in physiologically relevant tissues and organs to modulate biological functions and elicit desirable clinical outcomes. Information about target engagement at cellular and subcellular resolution is therefore critical for guiding compound optimization in drug discovery, and for probing resistance mechanisms to targeted therapies in clinical samples. We describe a target engagement-mediated amplification (TEMA) technology, where oligonucleotide-conjugated drugs are used to visualize and measure target engagement in situ, amplified via rolling-circle replication of circularized oligonucleotide probes. We illustrate the TEMA technique using dasatinib and gefitinib, two kinase inhibitors with distinct selectivity profiles. In vitro binding by the dasatinib probe to arrays of displayed proteins accurately reproduced known selectivity profiles, while their differential binding to fixed adherent cells agreed with expectations from expression profiles of the cells. We also introduce a proximity ligation variant of TEMA to selectively investigate binding to specific target proteins of interest. This form of the assay serves to improve resolution of binding to on- and off-target proteins. In conclusion, TEMA has the potential to aid in drug development and clinical routine by conferring valuable insights in drug–target interactions at spatial resolution in protein arrays, cells and in tissues.
|
|
| 4. |
- Al-Amin, Rasel A., 1983-, et al.
(författare)
-
Target Engagement-Mediated Amplification for Monitoring Drug-Target Interactions in Situ
-
Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- It is important to determine the localization of drugs or drug candidates at cellular and subcellular resolution in relevant clinical specimens. This is necessary to evaluate drug candidates from early stages of drug development to clinical evaluation of mutations potentially causing resistance to targeted therapy. We describe a technology where oligonucleotide-conjugated drug molecules are used to visualize and measure target engagement in situ via rolling-circle amplification (RCA) of circularized oligonucleotide probes (padlock probes). We established this target engagement-mediated amplification (TEMA) technique using kinase inhibitor precursor compounds, and we applied the assay to investigate target interactions by microscopy in pathology tissue sections and using flow cytometry for blood samples from patients, as well as in commercial arrays including almost half of all human proteins. In the variant proxTEMAtechnique, in situ proximity ligation assays were performed by combining drug-DNA conjugates with antibody-DNA conjugates to specifically reveal drug binding to particular on- or off-targets in pathological tissues sections. In conclusion, the TEMA methods successfully visualize drug-target interaction by experimental and clinically approved kinase inhibitors in situ and with kinases among a large collection of arrayed proteins.
|
|
| 5. |
- de Oliveira, Felipe, 1984-, et al.
(författare)
-
Autoimmunity detection via proximity assays
-
Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Since autoantibodies are recognized as valuable biomarkers for clinical diagnostics and prognostics in autoimmune diseases such as Stiff Person Syndrome (SPS) and Type 1 diabetes, detection of such markers at improved sensitivity and specificity could be of significant interest. In addition, as proximity assays have been shown to offer highly sensitive and specific detection of multiple proteins, the technique could be expanded to applications for autoimmunity detection. In the present study, we have applied the newly developed proximity ligation assay with rolling circle amplification (PLARCA), and proximity extension assay (PEA) for the detection of GADA, autoantibodies specific for glutamic acid decarboxylase 65 (GAD65). Through the use of oligonucleotide conjugated autoantigen GAD65 and anti-human antibodies, as proximity probes, we were able to apply these proximity assays to detect GADA in a set of SPS patient samples. In summary, we have applied and established both PLARCA and PEA, as a proof of concept, for the use of the specific and sensitive autoimmune detection.
|
|
| 6. |
|
|
| 7. |
- Lagerström-Fermér, Maria, et al.
(författare)
-
Amelogenin signal peptide mutation : correlation between mutations in the amelogenin gene (AMGX) and manifestations of X-linked amelogenesis imperfecta
- 1995
-
Ingår i: Genomics. - : Elsevier BV. - 0888-7543 .- 1089-8646. ; 26:1, s. 159-162
-
Tidskriftsartikel (refereegranskat)abstract
- Formation of tooth enamel is a poorly understood biological process. In this study we describe a 9-bp deletion in exon 2 of the amelogenin gene (AMGX) causing X-linked hypoplastic amelogenesis imperfecta, a disease characterized by defective enamel. The mutation results in the loss of 3 amino acids and exchange of 1 in the signal peptide of the amelogenin protein. This deletion in the signal peptide probably interferes with translocation of the amelogenin protein during synthesis, resulting in the thin enamel observed in affected members of the family. We compare this mutation to a previously reported mutation in the amelogenin gene that causes a different disease phenotype. The study illustrates that molecular analysis can help explain the various manifestations of a tooth disorder and thereby provide insights into the mechanisms of tooth enamel formation.
|
|
| 8. |
- Lagerström-Fermér, Maria, et al.
(författare)
-
X-linked recessive panhypopituitarism associated with a regional duplication in Xq25-q26
- 1997
-
Ingår i: American Journal of Human Genetics. - 0002-9297 .- 1537-6605. ; 60:4, s. 910-916
-
Tidskriftsartikel (refereegranskat)abstract
- We present a linkage analysis and a clinical update on a previously reported family with X-linked recessive panhypopituitarism, now in its fourth generation. Affected members exhibit variable degrees of hypopituitarism and mental retardation. The markers DXS737 and DXS1187 in the q25-q26 region of the X chromosome showed evidence for linkage with a peak LOD score (Zmax) of 4.12 at zero recombination fraction (theta(max) = 0). An apparent extra copy of the marker DXS102, observed in the region of the disease gene in affected males and heterozygous carrier females, suggests that a segment including this marker is duplicated. The gene causing this disorder appears to code for a dosage-sensitive protein central to development of the pituitary.
|
|
| 9. |
- Schallmeiner, Edith, et al.
(författare)
-
Sensitive protein detection via triple-binder proximity ligation assays
- 2007
-
Ingår i: Nature Methods. - : Springer Science and Business Media LLC. - 1548-7091 .- 1548-7105. ; 4:2, s. 135-137
-
Tidskriftsartikel (refereegranskat)abstract
- The detection of weakly expressed proteins and protein complexes in biological samples represents a fundamental challenge. We have developed a new proximity-ligation strategy named 3PLA that uses three recognition events for the highly specific and sensitive detection of as little as a hundred molecules of the vascular endothelial growth factor (VEGF), the biomarkers troponin I, and prostate-specific antigen (PSA) alone or in complex with an inhibitor--demonstrating the versatility of 3PLA.
|
|
| 10. |
|
|
