Physiological recognition

To explore the activity of the synthesized and selected compounds 1 to 6 and 9 to 11 on the TLR2 activity, reporter cells overexpressing hTLR2 were used. 7, 10a, 11 Except for 1, all selected compounds decreased TLR2-mediated NF- actividad de citas?B activation in a primary screen (Figure 3A). Additionally, cuatro, 6, 9 and 10 reduced Pam₃CSK₄-induced TLR2/1 responses to less than 40 % at fifty ?M and 2–6, 9 and 10 diminished Pam₂CSK₄-induced TLR2/6 responses to less than 50 % at 50 ?Mpound 6 and 11 abrogated the TLR2 response of both heterodimers almost completely and therefore showed a similar TLR2 inhibition as CU-CPT22 (Figure 3A). None of the tested compounds appeared to have any agonistic TLR2 activity (Figure S1A in the Supporting Information).

Inhibition of TLR2-dependent NF-?B activation and IL-8 secretion. A) HEK-Blue hTLR2 cells were pre-incubated with CU-CPT22 or compounds 1–6 and 9–11 for 1 h and then stimulated additionally with TLR2/1 agonist Pam₃CSK₄ (10 ng/mL) or TLR2/6 agonist Pam₂CSK₄ (1 ng/mL) for 24 h. B) HEK-Blue hTLR2 cells were pre-incubated with increasing concentrations of 4, 6, 9, 10 and then stimulated additionally with Pam₃CSK₄ or Pam₂CSK₄ for 24 h. Supernatants were tested for TLR2-mediated NF-?B/AP-1 activity by quantification of SEAP release (OD₆₄₀). C) THP-1 macrophages were incubated with CU-CPT22 (25 ?M) or compound 6 for 1 h and afterwards stimulated with Pam₃CSK₄ (10 ng/mL), Pam₂CSK₄ (1ng/mL), LPS (10 ng/mL), flagellin (1 ?g/mL), CL075 (8 ?M) or ODN2006 (5 ?M) for 24 h. Supernatants were analyzed for IL-8 secretion by ELISA. Mean+SD or ±SD (n=3).

No cytotoxic effects were observed within the activity range except for 11 which reduced cell viability starting at 50 ?M (Figure S1B). Hence, the apparent inhibition of TLR2-mediated responses by 11 might be due its cytotoxicity. Consequently, compound 11 was excluded from further investigation.