Anti-c5 antibody for treating patients with complement c5 polymorphism
A polymorphism and antibody technology, applied in the determination/examination of antibodies and microorganisms, anti-animal/human immunoglobulin, etc., can solve the problem of patients not being able to receive treatment
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example 1
[0108] Example 1: Crystallization of Testolumab Fab complexed with human C5.
[0109] Testolumab is a human monoclonal antibody that binds human and macaque (cynomolgus) complement C5 with picomolar affinity, thereby preventing C5 activation and the release of C5a and C5b. A detailed analysis of testolumab complexed with human C5 was performed.
[0110] method:
[0111] Expression and Purification of Tesdolumab Fab
[0112] The testolumab Fab was cloned and expressed in TG1F-E. coli cells (ACE25090). Suspend the frozen cell pellet in 150 ml lysis buffer and homogenize (lysis buffer: 20 mM NaH 2 PO 4 , 10mM Imidazole, 500mM NaCl pH 7.4, 1 tablet of EDTA-free cOmplete per 50ml buffer TM Protease Inhibitor Cocktail (Roche), 450 μl 1.0M MgCl 2 and 15 μl of totipotent nuclease (Novagen). After centrifugation (30 min at 16,000 g, 4° C.), the supernatant was sterile filtered (0.2 μm Stericup filter) and loaded (2.5 ml / min) onto a 5 ml HisTrap HP column (GE Healthcare, Inc....
example 2
[0136] Example 2: MAC Formation of C5 Demonstration that Testolumab, but not Eculizumab, inhibits mutant C5.
[0137] Testolumab and eculizumab were tested in the Wieslab assay using C5-depleted serum spiked with 7ug / ml wt C5 (Arg885) or mutant C5 (His885).
[0138] The results showed that Tesdolizumab, but not Eculizumab, blocked membrane attack complex (MAC) formation in C5-depleted sera spiked with mutant C5. Both antibodies were equally effective in inhibiting MAC formation in sera spiked with wt C5. Tesdolizumab was equally effective in serum spiked with normal or mutant C5. In contrast, eculizumab was inactive in sera spiked with mutant C5 ( image 3 ).
example 3
[0139] Example 3: Testolumab shows anti-hemolytic effect in C5 variant and non-variant PNH.
[0140] An open-label, single-arm study has been conducted to test testolumab (20 mg / kg iv twice a week) in patients with C5 variant and non-variant PNH.
[0141] method:
[0142] To determine the pharmacodynamic response to tesdolizumab, the ability of patient serum to lyse antibody-sensitized chicken erythrocytes in a human serum-complement hemolysis assay can be measured (Hillmen et al., New England Journal of Medicine 2004;350: 552-9). In this assay system, residual hemolysis of less than 20% indicated complete blockage of hemolysis (Nishimura et al., New England Journal of Medicine 2014;370;7).
[0143] result:
[0144] Five patients (two C5 variants) were analyzed after an average treatment duration of 8.5 weeks. No major safety issues were identified (no discontinuation of treatment, no treatment-related safety adverse events). Anti-hemolytic effects in PNH were seen in...
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