| 1. |
- Mattsson, Jenny, et al.
(författare)
-
Accelerating target deconvolution for therapeutic antibody candidates using highly parallelized genome editing
- 2021
-
Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 12, s. 1-8
-
Tidskriftsartikel (refereegranskat)abstract
- Therapeutic antibodies are transforming the treatment of cancer and autoimmune diseases. Today, a key challenge is finding antibodies against new targets. Phenotypic discovery promises to achieve this by enabling discovery of antibodies with therapeutic potential without specifying the molecular target a priori. Yet, deconvoluting the targets of phenotypically discovered antibodies remains a bottleneck; efficient deconvolution methods are needed for phenotypic discovery to reach its full potential. Here, we report a comprehensive investigation of a target deconvolution approach based on pooled CRISPR/Cas9. Applying this approach within three real-world phenotypic discovery programs, we rapidly deconvolute the targets of 38 of 39 test antibodies (97%), a success rate far higher than with existing approaches. Moreover, the approach scales well, requires much less work, and robustly identifies antibodies against the major histocompatibility complex. Our data establish CRISPR/Cas9 as a highly efficient target deconvolution approach, with immediate implications for the development of antibody-based drugs.
|
|
| 2. |
- Mattsson, Jenny, et al.
(författare)
-
Sequence enrichment profiles enable target-agnostic antibody generation for a broad range of antigens
- 2023
-
Ingår i: Cell reports methods. - 2667-2375. ; 3:5
-
Tidskriftsartikel (refereegranskat)abstract
- Phenotypic drug discovery (PDD) enables the target-agnostic generation of therapeutic drugs with novel mechanisms of action. However, realizing its full potential for biologics discovery requires new technologies to produce antibodies to all, a priori unknown, disease-associated biomolecules. We present a methodology that helps achieve this by integrating computational modeling, differential antibody display selection, and massive parallel sequencing. The method uses the law of mass action-based computational modeling to optimize antibody display selection and, by matching computationally modeled and experimentally selected sequence enrichment profiles, predict which antibody sequences encode specificity for disease-associated biomolecules. Applied to a phage display antibody library and cell-based antibody selection, ∼105 antibody sequences encoding specificity for tumor cell surface receptors expressed at 103–106 receptors/cell were discovered. We anticipate that this approach will be broadly applicable to molecular libraries coupling genotype to phenotype and to the screening of complex antigen populations for identification of antibodies to unknown disease-associated targets.
|
|
| 3. |
- Roghanian, Ali, et al.
(författare)
-
Antagonistic Human FcγRIIB (CD32B) Antibodies Have Anti-Tumor Activity and Overcome Resistance to Antibody Therapy In Vivo.
- 2015
-
Ingår i: Cancer Cell. - : Elsevier BV. - 1878-3686 .- 1535-6108. ; 27:4, s. 473-488
-
Tidskriftsartikel (refereegranskat)abstract
- Therapeutic antibodies have transformed cancer therapy, unlocking mechanisms of action by engaging the immune system. Unfortunately, cures rarely occur and patients display intrinsic or acquired resistance. Here, we demonstrate the therapeutic potential of targeting human (h) FcγRIIB (CD32B), a receptor implicated in immune cell desensitization and tumor cell resistance. FcγRIIB-blocking antibodies prevented internalization of the CD20-specific antibody rituximab, thereby maximizing cell surface accessibility and immune effector cell mediated antitumor activity. In hFcγRIIB-transgenic (Tg) mice, FcγRIIB-blocking antibodies effectively deleted target cells in combination with rituximab, and other therapeutic antibodies, from resistance-prone stromal compartments. Similar efficacy was seen in primary human tumor xenografts, including with cells from patients with relapsed/refractory disease. These data support the further development of hFcγRIIB antibodies for clinical assessment.
|
|
| 4. |
- Dexlin Mellby, Linda, et al.
(författare)
-
Design of recombinant antibody microarrays for cell surface membrane proteomics
- 2008
-
Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 7:1, s. 319-327
-
Tidskriftsartikel (refereegranskat)abstract
- Generating proteomic maps of membrane proteins, common targets for therapeutic interventions and disease diagnostics, has turned out to be a major challenge. Antibody-based microarrays are among the novel rapidly evolving proteomic technologies that may enable global proteome analysis to be performed. Here, we have designed the first generation of a scaleable human recombinant scFv antibody microarray technology platform for cell surface membrane proteomics as well as glycomics targeting intact cells. The results showed that rapid and multiplexed profiling of the cell surface proteome (and glycome) could be performed in a highly specific and sensitive manner and that differential expression patterns due to external stimuli could be monitored.
|
|
| 5. |
|
|
| 6. |
- Frendeus, Björn, et al.
(författare)
-
Escherichia coli P fimbriae utilize the Toll-like receptor 4 pathway for cell activation
- 2001
-
Ingår i: Molecular Microbiology. - : Wiley. - 1365-2958 .- 0950-382X. ; 40:1, s. 37-51
-
Tidskriftsartikel (refereegranskat)abstract
- Fimbriae mediate bacterial attachment to host cells and provide a mechanism for tissue attack. They activate a host response by delivery of microbial products such as lipopolysaccharide (LPS) or through direct fimbriae-dependent signalling mechanisms. By coupling to glycosphingolipid (GSL) receptors, P fimbriae trigger cytokine responses in CD14 negative host cells. Here we show that P fimbriae utilize the Toll-like receptor 4 (TLR4)-dependent pathway to trigger mucosal inflammation. Escherichia coli strains expressing P fimbriae as their only virulence factor stimulated chemokine and neutrophil responses in the urinary tract of TLR4 proficient mice, but TLR4 defective mice failed to respond to infection. Mucosal cells were CD14 negative but expressed several TLR species including TLR4, and TLR4 protein was detected. Infection with P fimbriated bacteria stimulated an increase in TLR4 mRNA levels. The activation signal did not involve the LPS-CD14 pathway and was independent of lipid A myristoylation, as shown by mutational inactivation of the msbB gene. Co-staining experiments revealed that TLR4 and the GSL receptors for P fimbriae co-localized in the cell membrane. The results demonstrate that P fimbriae activate epithelial cells by means of a TLR4-dependent signalling pathway, and suggest that GSL receptors for P fimbriae can recruit TLR4 as co-receptors.
|
|
| 7. |
- Frendeus, Björn, et al.
(författare)
-
Interleukin 8 receptor deficiency confers susceptibility to acute experimental pyelonephritis and may have a human counterpart
- 2000
-
Ingår i: Journal of Experimental Medicine. - 1540-9538. ; 192:6, s. 881-890
-
Tidskriftsartikel (refereegranskat)abstract
- Neutrophils migrate to infected mucosal sites that they protect against invading pathogens. Their interaction with the epithelial barrier is controlled by CXC chemokines and by their receptors. This study examined the change in susceptibility to urinary tract infection (UTI) after deletion of the murine interleukin 8 receptor homologue (mIL-8Rh). Experimental UTIs in control mice stimulated an epithelial chemokine response and increased chemokine receptor expression. Neutrophils migrated through the tissues to the epithelial barrier that they crossed into the lumen, and the mice developed pyuria. In mIL-8Rh knockout (KO) mice, the chemokine response was intact, but the epithelial cells failed to express IL-8R, and neutrophils accumulated in the tissues. The KO mice were unable to clear bacteria from kidneys and bladders and developed bacteremia and symptoms of systemic disease, but control mice were fully resistant to infection. The experimental UTI model demonstrated that IL-8R-dependent mechanisms control the urinary tract defense, and that neutrophils are essential host effector cells. Patients prone to acute pyelonephritis also showed low CXC chemokine receptor 1 expression compared with age-matched controls, suggesting that chemokine receptor expression may also influence the susceptibility to UTIs in humans. The results provide a first molecular clue to disease susceptibility of patients prone to acute pyelonephritis.
|
|
| 8. |
|
|
| 9. |
- Frendéus, Björn
(författare)
-
Role of Neutrophils and Innate Immune Mechanisms in Urinary Tract Infections
- 2001
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Urinary tract infections (UTI) are among the most common infections in man. Despite their prevalence, information on the molecular mechanisms defects that explain the increased susceptibility to disease are lacking. This study focused on mucosal inflammation and the molecular mechanisms by which uropathogenic Escherichia coli activate this response. P fimbriae are expressed by most E. coli strains that cause acute pyelonephritis. By binding to Gala1-4Galb containing motifs in the globoseries of glycosphingolipids P fimbriae trigger inflammatory responses that underlie disease pathology. P fimbriae were shown here to utilize the Toll-like receptor 4 (TLR4) pathway to activate inflammation. P fimbriated E. coli induced a strong inflammatory response in TLR4+ mice, but not in TLR4- mice. In contrast to previously described activation of TLR4, the P fimbriae induced signal was independent of the LPS/LBP/CD14 pathway. The TLR4 deficiency was shown to impair resistance to UTI through an effect on neutrophil recruitment. Uropathogenic E. coli induced a rapid and strong neutrophil recruitment in TLR4+ mice, but only a weak response in TLR4- mice. TLR4+ mice cleared infection, while in TLR4- mice bacterial numbers increased with time. Neutrophil depletion converted the TLR4+ mice to the TLR4- susceptible phenotype. Neutrophil recruitment was shown to depend on IL-8 and IL-8 receptors. In vitro neutrophil transepithelial migration was blocked by antibodies to the interleukin-8 receptor CXCR1, and neutrophils of IL-8R knockout mice failed to cross the urinary tract epithelium and accumulated in subepithelial tissue compartments. mIL-8Rh knock-out mice failed to clear infection, developed symptoms of disease, and showed evidence of renal scarring. Subsequent studies showed that children prone to acute pyelonephritis have decreased neutrophil CXCR1 expression. The results suggest that TLR and CXCR expression determine neutrophil recruitment to the infected urinary tract, and suggest that a CXCR1 deficiency might underlie increased susceptibility to acute pyelonephritis.
|
|
| 10. |
|
|
