| 1. |
- 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.
|
|
| 2. |
- 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.
|
|
| 3. |
|
|
| 4. |
|
|
| 5. |
|
|
| 6. |
- Al-Amin, Rasel Abdullah, Researcher, 1983-, et al.
(författare)
-
Sensitive protein detection using site-specifically oligonucleotide-conjugated nanobody reagents
- 2022
-
Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 98:28, s. 10054-10061
-
Tidskriftsartikel (refereegranskat)abstract
- High-quality affinity probes are critical for sensitive and specific protein detection, in particular for detection of protein biomarkers in the early phases of disease development. Proximity extension assays (PEAs) have been used for high-throughput multiplexed protein detection of up to a few thousand different proteins in one or a few microliters of plasma. Clonal affinity reagents can offer advantages over the commonly used polyclonal antibodies (pAbs) in terms of reproducibility and standardization of such assays. Here, we explore nanobodies (Nbs) as an alternative to pAbs as affinity reagents for PEA. We describe an efficient site-specific approach for preparing high-quality oligo-conjugated Nb probes via enzyme coupling using Sortase A (SrtA). The procedure allows convenient removal of unconjugated affinity reagents after conjugation. The purified high-grade Nb probes were used in PEA, and the reactions provided an efficient means to select optimal pairs of binding reagents from a group of affinity reagents. We demonstrate that Nb-based PEA (nano-PEA) for interleukin-6 (IL6) detection can augment assay performance, compared to the use of pAb probes. We identify and validate Nb combinations capable of binding in pairs without competition for IL6 antigen detection by PEA.
|
|
| 7. |
|
|
| 8. |
|
|
| 9. |
|
|
| 10. |
|
|
| 11. |
|
|
| 12. |
- Chen, L, et al.
(författare)
-
Ultra-sensitive monitoring of leukemia patients using superRCA mutation detection assays
- 2022
-
Ingår i: Nature communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 13:1, s. 4033-
-
Tidskriftsartikel (refereegranskat)abstract
- Rare tumor-specific mutations in patient samples serve as excellent markers to monitor the course of malignant disease and responses to therapy in clinical routine, and improved assay techniques are needed for broad adoption. We describe herein a highly sensitive and selective molecule amplification technology - superRCA assays - for rapid and highly specific detection of DNA sequence variants present at very low frequencies in DNA samples. Using a standard flow cytometer we demonstrate precise, ultra-sensitive detection of single-nucleotide mutant sequences from malignant cells against up to a 100,000-fold excess of DNA from normal cells in either bone marrow or peripheral blood, to follow the course of patients treated for acute myeloid leukemia (AML). We also demonstrate that sequence variants located in a high-GC region may be sensitively detected, and we illustrate the potential of the technology for early detection of disease recurrence as a basis for prompt change of therapy.
|
|
| 13. |
|
|
| 14. |
|
|
| 15. |
|
|
| 16. |
|
|
| 17. |
|
|
| 18. |
|
|
| 19. |
- Hagberg, A., et al.
(författare)
-
Beta-globin mRNA increases rapidly during erythropoietin treatment
- 2003
-
Ingår i: Scandinavian Journal of Clinical and Laboratory Investigation. - 0036-5513 .- 1502-7686. ; 63:3, s. 239-245
-
Tidskriftsartikel (refereegranskat)abstract
- Recombinant human erythropoietin (r-HuEpo) has an important role in the treatment of anaemic patients. Because of the high cost of r-HuEpo treatment, an early indicator of whether a patient is responding to the therapy would be valuable. Although measurement of gene expression is a promising new tool, it has not yet been established in clinical practice. The response pattern of a possible new marker, beta-globin mRNA, is compared with reticulocyte count, levels of haemoglobin, transferrin receptor and ferritin after r-HuEpo treatment. Eight healthy volunteers were stimulated with erythropoietin three times a week for four weeks and compared with five untreated control subjects. Blood samples were collected before each erythropoietin injection. Quantitative measurement of beta-globin mRNA was performed by poly(A) selection onto a manifold plastic support, coated with oligo(dT). The mRNA was reverse transcribed, followed by quantitative analysis using PCR via the 5' nuclease assay. The individuals treated with rHuEpo showed a more distinct increase in beta-globin mRNA levels than all other laboratory measurements. Beta-globin mRNA levels are therefore promising as a marker for the response to treatment with Epo.
|
|
| 20. |
|
|
| 21. |
|
|
| 22. |
|
|
| 23. |
|
|
| 24. |
|
|
| 25. |
|
|
| 26. |
|
|
| 27. |
|
|
| 28. |
|
|
| 29. |
|
|
| 30. |
- Lagerkvist, A., et al.
(författare)
-
Manifold sequencing
- 1996
-
Ingår i: Laboratory protocols for mutation detection. - : Oxford University Press, Oxford. ; , s. 119-
-
Bokkapitel (övrigt vetenskapligt/konstnärligt)
|
|
| 31. |
|
|
| 32. |
|
|
| 33. |
|
|
| 34. |
- Landegren, U., et al.
(författare)
-
Detecting genes with ligases.
- 1996
-
Ingår i: Methods: a companion to Methods in Enzymology.. ; , s. 84-
-
Bokkapitel (övrigt vetenskapligt/konstnärligt)
|
|
| 35. |
|
|
| 36. |
|
|
| 37. |
- Landegren, U., et al.
(författare)
-
Ligase-mediated gene detection .
- 1996
-
Ingår i: Encyclopedia of molecular biology: fundamentals and applications.. - : VCH Publisher, Weinheim. ; , s. 391-
-
Bokkapitel (övrigt vetenskapligt/konstnärligt)
|
|
| 38. |
|
|
| 39. |
- Landegren, U.
(författare)
-
Mutation detection now and later
- 1996
-
Ingår i: Laboratory protocols for mutation detection. - : Oxford University Press, Oxford. ; , s. 2-
-
Bokkapitel (övrigt vetenskapligt/konstnärligt)
|
|
| 40. |
|
|
| 41. |
|
|
| 42. |
|
|
| 43. |
|
|
| 44. |
- Ljung, R, et al.
(författare)
-
How do carriers of hemophilia experience prenatal diagnosis by fetal blood sampling?
- 1987
-
Ingår i: Clinical Genetics. - : Wiley. - 0009-9163 .- 1399-0004. ; 31:5, s. 297-302
-
Tidskriftsartikel (refereegranskat)abstract
- A semistructured personal interview was performed with 29 carriers of hemophilia A or B, 1-4 years after a pregnancy in which prenatal diagnosis (PND) of hemophilia was performed by fetal blood sampling. The carriers had received different recommendations regarding future pregnancies, and 14/29 did not know before they became pregnant that PND by fetal blood sampling was possible. One third of the women felt that important information was lacking in the consultations that preceded the PND. The conclusions regarding future genetic counselling are that more attention should be paid to improving education of all female carriers before a pregnancy, to motivating fathers-to-be to attend counselling sessions with the carriers, and to emphasizing the importance of the emotional support given by the family doctor and by other females who have experienced PND.
|
|
| 45. |
|
|
| 46. |
|
|
| 47. |
- Nilsson, M., et al.
(författare)
-
Padlock probes for in situ detection
- 1996
-
Ingår i: Laboratory protocols for mutation detection. - : Oxford University Press, Oxford. ; , s. 135-
-
Bokkapitel (övrigt vetenskapligt/konstnärligt)
|
|
| 48. |
|
|
| 49. |
|
|
| 50. |
|
|
