Mon. Jun 17th, 2024

Te in intracellular organelles, delivering any payload they carry.3, five, 24, 28?0 The key challenge, although, has been to identify ligands of adequate avidity and selectivity to target cells expressing only the preferred siglec. By far the most prosperous approach to date has been to use sialic acid as a privileged scaffold, with modifications made around the sugar ring, primarily at C9 and C5, to raise affinity and selectivity for the desired siglec.31?1 Despite considerable progress in this arena, efforts have failed to identify ligands of CD22 and CD33 with adequate avidity and selectivity needed for human clinical studies. For hCD33 in particular, you can find no reports describing high affinity ligands of this siglec. In contrast, various groups have generated ligands of CD22 with 100-1000 fold greater affinity than the Sigma 1 Receptor Modulator drug natural ligand, but the greatest of these have not demonstrated adequate selectivity.36, 38, 39, 41 As an example, even though we have shown that doxorubicin-loaded liposomes displaying a high affinity ligand of CD22 (Fig. 1, compound 4) are powerful in prolonging life within a murine model of disseminated human B cell lymphoma, this ligand exhibits a significant cross-reactivity with sialoadhesin (Siglec-1, mSn), expressed on macrophages, which mediate fast clearance of your liposomes.28 Therefore, a extra selective ligand of hCD22 is needed for optimal targeting of B lymphoma cells. Here we report the development of high affinity ligands selective for hCD33 and hCD22. This was accomplished for hCD33 by carrying out iterative cycles of focused library synthesis followed by glycan microarray screening to assess relative avidity and specificity for selected siglecs. In the end this resulted within a ligand exhibiting 350-fold improved affinity more than a organic sialoside, and when displayed on liposomal nanoparticles exhibited higher specificity for hCD33 more than a panel of other human siglecs. Throughout these screens weNIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptChem Sci. Author manuscript; obtainable in PMC 2015 June 01.Rillahan et al.Pagefortuitously identified a sialic acid analog displaying enhanced affinity for hCD22 with no crossreactivity to Siglec-1 (mSn) or hCD33. Additional optimization of this scaffold yielded a ligand with high affinity and selectivity for hCD22. Finally, we show that ligand-bearing liposomes displaying the ligands of hCD33 and hCD22 bind selectively to cells expressing their respective siglec in peripheral human blood.NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptResults and SIRT1 Modulator Gene ID DiscussionIdentification of high-affinity sialoside ligands selective for hCD33 We’ve previously shown that hCD33 binds to 2-6 linked sialoside analogues bearing unnatural hydrophobic substituents appended to C9 or C5.31 Within this earlier perform, screening an extensive library of click-chemistry generated sialoside analogues identified compound 2, using a 4-cyclohexyl-1,two,3-triazole substituent in the C5 position, having a modestly elevated affinity for hCD33 more than the native scaffold (1), and with out crossreactivity to other siglecs in the screen (Fig. 1).31 Even though triazole-containing substituents linked for the C9 position failed to yield affinity gains for hCD33, a previously identified high affinity hCD22/mSn ligand with a benzamide linkage (4) also exhibited an affinity obtain for hCD33, albeit with no selectivity (Fig. 1).31 These observations supplied motivation to extra exhaustively survey C9-substituted b.