Galectins are small soluble lectins that bind β-galactosides via their carbohydrate recognition domain (CRD). Their ability to dimerize is critical for the crosslinking of glycoprotein receptors and subsequent cellular signaling. This is particularly important in their immunomodulatory role via the induction of T-cell apoptosis. Because galectins play a central role in many pathologies, including cancer, they represent valuable therapeutic targets. At present, most inhibitors have been directed towards the CRD, a challenging task in terms of specificity given the high structural homology of the CRD among galectins. Such inhibitors are not effective at targeting CRD-independent functions of galectins. Here, we report a new class of galectin inhibitors that specifically binds human galectin-7 (hGal-7), disrupts the formation of homodimers, and inhibits the pro-apoptotic activity of hGal-7 on Jurkat T cells. In addition to representing a new means to achieve specificity when targeting galectins, such inhibitors provide a promising alternative to more conventional galectin inhibitors that target the CRD with soluble glycans or other small molecular weight allosteric inhibitors.

Bacterial resistance to b-lactam antibiotics is a global issue threatening the success of infectious disease treatments worldwide. Mycobacterium tuberculosis has been particularly resilient to blactam treatment, primarily due to the chromosomally encoded BlaC b-lactamase, a broad-spectrum hydrolase that renders ineffective the vast majority of relevant b-lactam compounds currently in use. Recent laboratory and clinical studies have nevertheless shown that specific b-lactam-BlaC inhibitor combinations can be used to inhibit the growth of extensively drug-resistant strains of M. tuberculosis, effectively offering new tools for combined treatment regimens against resistant strains. In the present work, we performed combinatorial active-site replacements in BlaC to demonstrate that specific inhibitor-resistant (IRT) substitutions at positions 69, 130, 220, and/or 234 can act synergistically to yield active-site variants with several thousand fold greater in vitro resistance to clavulanate, the most common clinical b-lactamase inhibitor. While most single and double variants remain sensitive to clavulanate, double mutants R220S-K234R and S130G-K234R are substantially less affected by time-dependent clavulanate inactivation, showing residual b-lactam hydrolytic activities of 46% and 83% after 24 h incubation with a clinically relevant inhibitor concentration (5 lg/ml, 25 mM). These results demonstrate that activesite alterations in BlaC yield resistant variants that remain active and stable over prolonged bacterial generation times compatible with mycobacterial proliferation. These results also emphasize the formidable adaptive potential of inhibitor-resistant substitutions in b-lactamases, potentially casting a shadow on specific b-lactam-BlaC inhibitor combination treatments against M. tuberculosis.

The selective targeting of protein–protein interactions remains a significant determinant for the proper modulation and regulation of cell apoptosis. Prototypic galectins such as human galectin-7 (GAL-7) are characterized by their ability to form homodimers that control the molecular fate of a cell by mediating subtle yet critical glycan-dependent interactions between pro- and anti-apoptotic molecular partners. Altering the structural architecture of GAL-7 can therefore result in resistance to apoptosis in various human cancer cells, further illustrating its importance in cell survival. In this study, we used a combination of biophysical and cellular assays to illustrate that binding of a water-soluble meso-tetraarylporphyrin molecule to GAL-7 induces protein oligomerization and modulation of GAL-7-induced apoptosis in human Jurkat T cells. Our results suggest that the integrity of the GAL-7 homodimer architecture is essential for its molecular function, in addition to providing an interesting porphyrin binding modulator for controlling apoptosis in mammalian cells.

Galectins are small soluble lectins that bind to galactosides via their carbohydrate recognition domain (CRD). They are involved in a variety of biological functions such as cell adhesion and recognition of pathogens. They are also known to modulate cell migration, metastasis and survival. Because their dysregulation is often observed in cancer cells, they are interesting modulators of tumor progression and attractive targets for cancer therapy. Most galectin inhibitors developed so far target the CRD. However, since the CRD is very conserved between the galectins, these inhibitors normally lack specificity and are ineffective against CRD-independent functions. [1,2] Galectin-7 is a prototype galectin that attracts increasing interest due to its implication in various cancers. Galectin-7 ability to form homodimers has been demonstrated essential for its ability to promote tumor formation and progression. The dimer formation in galectin-7 occurs in a "back-to-back" structure ( Figure 1). [1,3] In this study, peptides corresponding to dimer interface segments were synthesized with or without the addition of a polyethylene glycol chain at the N-terminus to improve solubility. The binding of the peptides to galectin-7 was evaluated along with their ability to disrupt galectin-7 dimer formation and to modulate associated biological activities. In parallel, other residues having a role in the dimer stabilization were identified using PyMOL software. Residues R14, R20 and D94 at the dimerization interface were selected and alanine mutants at these positions were produced and purified. Evaluation of the impact of these substitutions on structure, in absence and in presence of ligands, galectin-7 dimer formation as well as affinity to CRD ligands such as lactose, galactose and LacNAc was undertaken. This study will probably highlight key features in the dimerization of galectin-7 and potential targets for the development of galectin-7 specific inhibitors not altering CRD functions or other galectin activities.