Anti-CCR5 agents and methods of treatment to block cancer metastasis or enhance cell death induced by DNA damage chemotherapy
A cancer and drug technology, applied in chemical instruments and methods, receptors/cell surface antigens/cell surface determinants, anti-tumor drugs, etc., can solve problems such as unmet needs and unproven clinical treatment plans
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Embodiment 1
[0161] Example 1. Binding of lelimumab to CCR5 expressed in breast cancer cells
[0162] Figure 1A and Figure 1B : Lelimumab binds to CCR5 in human breast cancer cells.
[0163] like Figure 1A As shown, to determine the binding of lelimumab to human CCR5 in breast cancer cells, the MDA-MB-231 human breast cancer cell line was transfected with the human CCR5 expression vector as a model system. CCR5 positive cells were assessed using a previously tested commercial APC-conjugated mouse anti-human / mouse / rat CCR5 antibody (FAB1802A) (APC-αCCR5) from R&D as a positive control. MDA-MB-231-CCR5 cells were stained with APC-αCCR5 and lelimumab using concentrations of 1-140 g / ml. Mouse anti-human IgG conjugated to Alexa Fluor 488 was used as the secondary antibody to measure lelimumab binding to cells. Figure 1A Analysis of lelimumab binding to CCR5 by FACS is shown in . Binding of lelimumab to human CCR5 was demonstrated.
[0164] like Figure 1B As shown, the efficiency of ...
Embodiment 2
[0165] Example 2. Effect of PRO140 on CCL5-induced Ca in MDA-MB-231-CCR5 cells 2+ Effects of ionic reactions
[0166] Figure 2A , Figure 2B , Figure 2C and Figure 2D : PRO140 (lelimumab) blocks human CCR5-mediated signaling in human breast cancer cells. CCR5 activation induces calcium flux (Mueller et al., 2002; Petkovic et al., 2004). To assess the effect of tetralimumab on CCR5 function, we analyzed live cell images ( Figure 2A , Figure 2B and Figure 2C ) measured the calcium response induced by CCL5 in MDA-MB-231-CCR5 cells in the presence or absence of lelimumab. Fluo-4 was used as a calcium concentration indicator. The CCR5 antagonist velivero was used as a positive control ( Figure 2A and Figure 2D ). The results showed that lelimumab was able to block CCL5-induced calcium responses in MDA-MB-231-CCR5 cells (1.23±0.10 for control cells, N=10, and 0.54±0.13 for PRO140-treated cells, N=12. CCL5 induced calcium peak (P<0.001).
Embodiment 3
[0167] Example 3. Lelimumab blocks 3D Matrigel invasion of breast cancer cells
[0168] Figure 3A , Figure 3B , Figure 3C and Figure 3D : Lelimumab blocks CCR5-mediated invasion of human breast cancer cells into the extracellular matrix. The ability of breast cancer cells to invade the extracellular matrix differs from, but is an important step in, tumor metastasis (Zetter, 1990). To test the ability of PRO140 to block the 3D Matrigel invasion assay, MDA-MB-231 cells were used. CCL5 acts as a chemokine to induce invasion. Velivero, a small molecule inhibitor of CCR5, was used as a form of positive control. Lelimumab reduces CCL5-induced invasion of MDA-MB-231 breast cancer cells with similar efficacy to veliverol ( Figure 3A , Figure 3B ) (855±9 for control, N=8, 855±9 for lelimumab, N=9, P Figure 3C , Figure 3D ).
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