Anti-mct1 antibodies and uses thereof
A technology of antibodies and antigens, applied to fusion or multispecific proteins, compositions containing such antibodies and antigen-binding fragments thereof, multispecific and bispecific antibodies, therapeutic and diagnostic uses, antagonistic anti-MCT1 antibodies and In the field of antigen-binding fragments, it can solve problems such as antibodies that have not yet been reported
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Embodiment 1
[0637] Example 1. Differential expression of MCTs on T cells.
[0638] Materials and methods
[0639] The commercially available SM compound AZ3965 (MedChem Express, NJ) and related analogs were used to help uncover the unique biology of the MCT1 pathway in immune cells. For clarity, we will collectively refer to AZ3965 and its analogs with the same binding affinity, PK and MCT1 / 2 selectivity as "AZ3965".
[0640] Expression of MCT1, MCT2, MCT4 and BSG (CD147) was measured in unstimulated and stimulated leukocytes from two different donors. For "stimulated" conditions, cells were activated with CD3 / CD28 for 3 days. AZ3965 was tested for inhibition of stimulated cell proliferation.
[0641] result
[0642] MCT1 promotes the transfer of metabolites, including products of glycolysis, which are more important in activated T / B cells ( figure 1 ). figure 2 The expression levels of MCT1, MCT2, MCT4 and BSG (CD147) are shown in the two donors, demonstrating that activated T ...
Embodiment 2
[0644] Example 2: Validation of the feasibility of targeting MCT1 in vitro and in vivo for inflammatory / autoimmune disorders using the AZ3965 inhibitor.
[0645] in vitro
[0646] Effects on lactate transport. AZ3965 inhibited human T cells (CD4 + and CD8 + both), B-cell lymphoma (Daudi) and lactate transport in PBMCs, but not in monocytes ( image 3). AZ3965 inhibits lactate transport by up to 80% in affected cells, but does not affect transport in monocytes, which is important for protecting the innate immune response in treated individuals.
[0647] Human T cell proliferation. Administration of MCT1 inhibitor with an IC of 0.54 nM in a human T cell proliferation assay 50 Reduce T cell proliferation ( Figure 4 ).
[0648] Human mixed lymphocyte reaction (MLR). Administration of the MCT1 inhibitor with an IC of 1.34 nM in the human MLR assay 50 Reduce T cell proliferation ( Figure 5 ).
[0649] T cell cytokine secretion. T cells were activated by CD3 / CD28 in ...
Embodiment 3
[0658] Example 3: Development and binding characterization of anti-human MCT1 antibodies
[0659] Selection of MCT1 Ab1 mAbs. MCT1 Ab1 is a rat anti-human MCT1 monoclonal antibody selected after cell-based rodent immunization and binding screening using MCT1 expressing and MCT1 knockout (KO) cell lines.
[0660] Binding affinity and MCT1 cross-reactivity. Kinetic exclusion assay (KinExA) analysis showed that MCT1 Ab1 binds to human MCT1 with a Kd of 6.3 nM. MCT1 Ab1 is also highly cross-reactive with cynomolgus monkey (cyno) and rabbit MCT1, but not rodent MCT1 ( Figure 12A -D).
[0661] specificity of binding. HEK-293WT cells only expressed MCT1 / CD147, but no other MCTs, as determined by RT-PCR. To measure the binding specificity of the antibodies of the invention, the HEK-293MCT1 / CD147 double knockout cell line can be used as a negative control. In addition, this double knockout cell line was engineered to express individual transporters (MCT1, MCT2, MCT3, MCT4, CD147...
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