Crystal Packing Reveals a Potential Autoinhibited KRAS Dimer Interface and a Strategy for Small-Molecule Inhibition of RAS Signaling.

KRAS GTPases harbor oncogenic mutations in more than 25% of human tumors. KRAS is considered to be largely undruggable due to the lack of a suitable small-molecule binding site. Here, we report a unique crystal structure of His-tagged KRASG12D that reveals a remarkable conformational change. The Switch I loop of one His-KRASG12D structure extends into the Switch I/II pocket of another His-KRASG12D in an adjacent unit cell to create an elaborate interface that is reminiscent of high-affinity protein-protein complexes. We explore the contributions of amino acids at this interface using alanine-scanning studies with alchemical free energy perturbation calculations based on explicit-solvent molecular dynamics simulations. Several interface amino acids were found to be hot spots as they contributed more than 1.5 kcal/mol to the protein-protein interaction. Computational analysis of the complex revealed the presence of two large binding pockets that possess physicochemical features typically found in pockets considered druggable. Small-molecule binding to these pockets may stabilize this autoinhibited structure of KRAS if it exists in cells to provide a new strategy to inhibit RAS signaling.

Chloroacetamide fragment library screening identifies new scaffolds for covalent inhibition of the TEAD·YAP1 interaction.

Transcriptional enhanced associate domain (TEAD) binding to co-activator yes-associated protein (YAP1) leads to a transcription factor of the Hippo pathway. TEADs are regulated by S-palmitoylation of a conserved cysteine located in a deep well-defined hydrophobic pocket outside the TEAD·YAP1 interaction interface. Previously, we reported the discovery of a small molecule based on the structure of flufenamic acid that binds to the palmitate pocket, forms a covalent bond with the conserved cysteine, and inhibits TEAD4 binding to YAP1. Here, we screen a fragment library of chloroacetamide electrophiles to identify new scaffolds that bind to the palmitate pocket of TEADs and disrupt their interaction with YAP1. Time- and concentration-dependent studies with wild-type and mutant TEAD1-4 provided insight into their reaction rates and binding constants and established the compounds as covalent inhibitors of TEAD binding to YAP1. Binding pose hypotheses were generated by covalent docking revealing that the fragments and compounds engage lower, middle, and upper sub-sites of the palmitate pocket. Our fragments and compounds provide new scaffolds and starting points for the design of derivatives with improved inhibition potency of TEAD palmitoylation and binding to YAP1.

Exploring Covalent Bond Formation at Tyr-82 for Inhibition of Ral GTPase Activation.

Ral RAS GTPases are directly activated by KRAS through a trimeric complex with a guanine exchange factor. Ral is considered undruggable and lacks an accessible cysteine for covalent drug development. Previously we had reported an aryl sulfonyl fluoride fragment that formed a covalent bond at Tyr-82 on Ral and created a deep and well-defined pocket. Here, we explore this pocket further through design and synthesis of several fragment derivatives. The fragment core is modified by introducing tetrahydronaphthalene or benzodioxane rings to enhance affinity and stability of the sulfonyl fluoride reactive group. The deep pocket in the Switch II region is also explored by modifying the aromatic ring of the fragment that is ensconced into the pocket. Compounds 19 (SOF-658) and 26 (SOF-648) formed a single robust adduct specifically at Tyr-82, inhibited Ral GTPase exchange in buffer and in mammalian cells, and blocked invasion of pancreatic ductal adenocarcinoma cancer cells. Compound 19 (SOF-658) was stable in buffer, mouse, and human microsomes suggesting that further optimization could lead to small molecules to probe Ral activity in tumor models.

Small-Molecule Cyanamide Pan-TEAD·YAP1 Covalent Antagonists.

Transcriptional enhanced associate domains (TEADs) are transcription factors that bind to cotranscriptional activators like the yes-associated protein (YAP) or its paralog transcriptional coactivator with a PDZ-binding motif (TAZ). TEAD·YAP/TAZ target genes are involved in tissue and immune homeostasis, organ size control, tumor growth, and metastasis. Here, we report isoindoline and octahydroisoindole small molecules with a cyanamide electrophile that forms a covalent bond with a conserved cysteine in the TEAD palmitate-binding cavity. Time- and concentration-dependent studies against TEAD1-4 yielded second-order rate constants kinact/KI greater than 100 M-1 s-1. Compounds inhibited YAP1 binding to TEADs with submicromolar IC50 values. Cocrystal structures with TEAD2 enabled structure-activity relationship studies. In mammalian cells, compounds suppressed CTGF mRNA levels and inhibited TEAD1-4 transcriptional activity with submicromolar IC50 values. Inhibition of TEAD binding to YAP1 in mammalian cells was also observed. Several compounds inhibited the cell viability of sarcoma, hepatocellular carcinoma, glioblastoma, and breast cancer cells with single-digit micromolar IC50 values.

Novel Selective Galectin-3 Antagonists Are Cytotoxic to Acute Lymphoblastic Leukemia.

Galectin-3 is a ß-galactoside-specific, carbohydrate-recognizing protein (lectin) that is strongly implicated in cancer development, metastasis, and drug resistance. Galectin-3 promotes migration and ability to withstand drug treatment of B-cell precursor acute lymphoblastic leukemia (BCP-ALL) cells. Due to high amino acid conservation among galectins and the shallow nature of their glycan-binding site, the design of selective potent antagonists targeting galectin-3 is challenging. Herein, we report the design and synthesis of novel taloside-based antagonists of galectin-3 with enhanced affinity and selectivity. The molecules were optimized by in silico docking, selectivity was established against four galectins, and the binding modes were confirmed by elucidation of X-ray crystal structures. Critically, the specific inhibition of galectin-3-induced BCP-ALL cell agglutination was demonstrated. The compounds decreased the viability of ALL cells even when grown in the presence of protective stromal cells. We conclude that these compounds are promising leads for therapeutics, targeting the tumor-supportive activities of galectin-3 in cancer.

Covalent Fragment Screening Identifies Rgl2 RalGEF Cysteine for Targeted Covalent Inhibition of Ral GTPase Activation.

Ral GTPases belong to the RAS superfamily, and they are directly activated by K-RAS. The RalGEF pathway is one of the three major K-RAS signaling pathways. Ral GTPases do not possess a cysteine nucleophile to develop a covalent inhibitor following the strategy that led to a K-RAS G12C therapeutic agent. However, several cysteine amino acids exist on the surface of guanine exchange factors that activate Ral GTPases, such as Rgl2. Here, we screen a library of cysteine electrophile fragments to determine if covalent bond formation at one of the Rgl2 surface cysteines could inhibit Ral GTPase activation. We found several chloroacetamide and acrylamide fragments that inhibited Ral GTPase exchange by Rgl2. Site-directed mutagenesis showed that covalent bond formation at Cys-284, but not other cysteines, leads to inhibition of Ral activation by Rgl2. Follow-up time- and concentration-dependent studies of derivatives identified by substructure search of commercial libraries further confirmed Cys-284 as the reaction site and identified the indoline fragments as the most promising series for further development. Cys-284 is located outside of the Ral ⋅ Rgl2 interface on a loop that has several residues that come in direct contact with Ral GTPases. Our allosteric covalent fragment inhibitors provide a starting point for the development of small-molecule covalent inhibitors to probe Ral GTPases in animal models.

Overcoming Microenvironment-Mediated Chemoprotection through Stromal Galectin-3 Inhibition in Acute Lymphoblastic Leukemia.

Environmentally-mediated drug resistance in B-cell precursor acute lymphoblastic leukemia (BCP-ALL) significantly contributes to relapse. Stromal cells in the bone marrow environment protect leukemia cells by secretion of chemokines as cues for BCP-ALL migration towards, and adhesion to, stroma. Stromal cells and BCP-ALL cells communicate through stromal galectin-3. Here, we investigated the significance of stromal galectin-3 to BCP-ALL cells. We used CRISPR/Cas9 genome editing to ablate galectin-3 in stromal cells and found that galectin-3 is dispensable for steady-state BCP-ALL proliferation and viability. However, efficient leukemia migration and adhesion to stromal cells are significantly dependent on stromal galectin-3. Importantly, the loss of stromal galectin-3 production sensitized BCP-ALL cells to conventional chemotherapy. We therefore tested novel carbohydrate-based small molecule compounds (Cpd14 and Cpd17) with high specificity for galectin-3. Consistent with results obtained using galectin-3-knockout stromal cells, treatment of stromal-BCP-ALL co-cultures inhibited BCP-ALL migration and adhesion. Moreover, these compounds induced anti-leukemic responses in BCP-ALL cells, including a dose-dependent reduction of viability and proliferation, the induction of apoptosis and, importantly, the inhibition of drug resistance. Collectively, these findings indicate galectin-3 regulates BCP-ALL cell responses to chemotherapy through the interactions between leukemia cells and the stroma, and show that a combination of galectin-3 inhibition with conventional drugs can sensitize the leukemia cells to chemotherapy.

Design and Synthesis of Fragment Derivatives with a Unique Inhibition Mechanism of the uPAR·uPA Interaction.

There is substantial interest in the development of small molecules that inhibit the tight and highly challenging protein-protein interaction between the glycophosphatidylinositol (GPI)-anchored cell surface receptor uPAR and the serine protease uPA. While preparing derivatives of a fragment-like compound that previously emerged from a computational screen, we identified compound 5 (IPR-3242), which inhibited binding of uPA to uPAR with submicromolar IC50s. The high inhibition potency prompted us to carry out studies to rule out potential aggregation, lack of stability, reactivity, and nonspecific inhibition. We designed and prepared 16 derivatives to further explore the role of each substituent. Interestingly, the compounds only partially inhibited binding of a fluorescently labeled α-helical peptide that binds to uPAR at the uPAR·uPA interface. Collectively, the results suggest that the compounds bind to uPAR outside of the uPAR·uPA interface, trapping the receptor into a conformation that is not able to bind to uPA. Additional studies will have to be carried out to determine whether this unique inhibition mechanism can occur at the cell surface.

Small-Molecule Inhibition of the uPAR ⋠uPA Interaction by Conformational Selection.

The urokinase receptor (uPAR) is a cell surface receptor that binds to the serine protease urokinase-type plasminogen activator (uPA) with high affinity. This interaction is beneficial for extravascular fibrin clearance, but it has also been associated with a broad range of pathological conditions including cancer, atherosclerosis, and kidney disease. Here, starting with a small molecule that we previously discovered by virtual screening and cheminformatics analysis, we design and synthesize several derivatives that were tested for binding and inhibition of the uPAR ⋅ uPA interaction. To confirm the binding site and establish a binding mode of the compounds, we carried out biophysical studies using uPAR mutants, among them uPARH47C-N259C , a mutant previously developed to mimic the structure of uPA-bound uPAR. Remarkably, a substantial increase in potency is observed for inhibition of uPARH47C-N259C binding to uPA compared to wild-type uPAR, consistent with our use of the structure of uPAR in its uPA-bound state to design small-molecule uPAR ⋅ uPA antagonists. Combined with the biophysical studies, molecular docking followed by extensive explicit-solvent molecular dynamics simulations and MM-GBSA free energy calculations yielded the most favorable binding pose of the compound. Collectively, these results suggest that potent inhibition of uPAR binding to uPA with small molecules will likely only be achieved by developing small molecules that exhibit high-affinity to solution apo structures of uPAR, rather than uPA-bound structures of the receptor.

Phenotypic Screening of Chemical Libraries Enriched by Molecular Docking to Multiple Targets Selected from Glioblastoma Genomic Data.

Like most solid tumors, glioblastoma multiforme (GBM) harbors multiple overexpressed and mutated genes that affect several signaling pathways. Suppressing tumor growth of solid tumors like GBM without toxicity may be achieved by small molecules that selectively modulate a collection of targets across different signaling pathways, also known as selective polypharmacology. Phenotypic screening can be an effective method to uncover such compounds, but the lack of approaches to create focused libraries tailored to tumor targets has limited its impact. Here, we create rational libraries for phenotypic screening by structure-based molecular docking chemical libraries to GBM-specific targets identified using the tumor's RNA sequence and mutation data along with cellular protein-protein interaction data. Screening this enriched library of 47 candidates led to several active compounds, including 1 (IPR-2025), which (i) inhibited cell viability of low-passage patient-derived GBM spheroids with single-digit micromolar IC50 values that are substantially better than standard-of-care temozolomide, (ii) blocked tube-formation of endothelial cells in Matrigel with submicromolar IC50 values, and (iii) had no effect on primary hematopoietic CD34+ progenitor spheroids or astrocyte cell viability. RNA sequencing provided the potential mechanism of action for 1, and mass spectrometry-based thermal proteome profiling confirmed that the compound engages multiple targets. The ability of 1 to inhibit GBM phenotypes without affecting normal cell viability suggests that our screening approach may hold promise for generating lead compounds with selective polypharmacology for the development of treatments of incurable diseases like GBM.

Small-molecule covalent bond formation at tyrosine creates a binding site and inhibits activation of Ral GTPases.

Ral (Ras-like) GTPases are directly activated by oncogenic Ras GTPases. Mutant K-Ras (G12C) has enabled the development of covalent K-Ras inhibitors currently in clinical trials. However, Ral, and the overwhelming majority of mutant oncogenic K-Ras, are devoid of a druggable pocket and lack an accessible cysteine for the development of a covalent inhibitor. Here, we report that covalent bond formation by an aryl sulfonyl fluoride electrophile at a tyrosine residue (Tyr-82) inhibits guanine exchange factor Rgl2-mediated nucleotide exchange of Ral GTPase. A high-resolution 1.18-Å X-ray cocrystal structure shows that the compound binds to a well-defined binding site in RalA as a result of a switch II loop conformational change. The structure, along with additional high-resolution crystal structures of several analogs in complex with RalA, confirm the importance of key hydrogen bond anchors between compound sulfone oxygen atoms and Ral backbone nitrogen atoms. Our discovery of a pocket with features found on known druggable sites and covalent modification of a bystander tyrosine residue present in Ral and Ras GTPases provide a strategy that could lead to therapeutic agent targeting oncogenic Ras mutants that are devoid of a cysteine nucleophile.

Chemical Space Overlap with Critical Protein-Protein Interface Residues in Commercial and Specialized Small-Molecule Libraries.

There is growing interest in the use of structure-based virtual screening to identify small molecules that inhibit challenging protein-protein interactions (PPIs). In this study, we investigated how effectively chemical library members docked at the PPI interface mimic the position of critical side-chain residues known as "hot spots". Three compound collections were considered, a commercially available screening collection (ChemDiv), a collection of diversity-oriented synthesis (DOS) compounds that contains natural-product-like small molecules, and a library constructed using established reactions (the "screenable chemical universe based on intuitive data organization", SCUBIDOO). Three different tight PPIs for which hot-spot residues have been identified were selected for analysis: uPAR⋅uPA, TEAD4⋅Yap1, and CaV α⋅CaV ß. Analysis of library physicochemical properties was followed by docking to the PPI receptors. A pharmacophore method was used to measure overlap between small-molecule substituents and hot-spot side chains. Fragment-like conformationally restricted small molecules showed better hot-spot overlap for interfaces with well-defined pockets such as uPAR⋅uPA, whereas better overlap was observed for more complex DOS compounds in interfaces lacking a well-defined binding site such as TEAD4⋅Yap1. Virtual screening of conformationally restricted compounds targeting uPAR⋅uPA and TEAD4⋅Yap1 followed by experimental validation reinforce these findings, as the best hits were fragment-like and had few rotatable bonds for the former, while no hits were identified for the latter. Overall, such studies provide a framework for understanding PPIs in the context of additional chemical matter and new PPI definitions.

Small-Molecule Covalent Modification of Conserved Cysteine Leads to Allosteric Inhibition of the TEAD⋠Yap Protein-Protein Interaction.

The Hippo pathway coordinates extracellular signals onto the control of tissue homeostasis and organ size. Hippo signaling primarily regulates the ability of Yap1 to bind and co-activate TEA domain (TEAD) transcription factors. Yap1 tightly binds to TEAD4 via a large flat interface, making the development of small-molecule orthosteric inhibitors highly challenging. Here, we report small-molecule TEAD⋅Yap inhibitors that rapidly and selectively form a covalent bond with a conserved cysteine located within the unique deep hydrophobic palmitate-binding pocket of TEADs. Inhibition of TEAD4 binding to Yap1 by these compounds was irreversible and occurred on a longer time scale. In mammalian cells, the compounds formed a covalent complex with TEAD4, inhibited its binding to Yap1, blocked its transcriptional activity, and suppressed expression of connective tissue growth factor. The compounds inhibited cell viability of patient-derived glioblastoma spheroids, making them suitable as chemical probes to explore Hippo signaling in cancer.

Small molecules inhibit ex vivo tumor growth in bone.

Bone is a common site of metastasis for breast, prostate, lung, kidney and other cancers. Bone metastases are incurable, and substantially reduce patient quality of life. To date, there exists no small-molecule therapeutic agent that can reduce tumor burden in bone. This is partly attributed to the lack of suitable in vitro assays that are good models of tumor growth in bone. Here, we take advantage of a novel ex vivo model of bone colonization to report a series of pyrrolopyrazolone small molecules that inhibit cancer cell invasion and ex vivo tumor growth in bone at single-digit micromolar concentration. We find that the compounds modulated the expression levels of genes associated with bone-forming osteoblasts, bone-destroying osteoclasts, cancer cell viability and metastasis. Our compounds provide chemical tools to uncover novel targets and pathways associated with bone metastasis, as well as for the development of compounds to prevent and reverse bone tumor growth in vivo.

Mimicking Intermolecular Interactions of Tight Protein-Protein Complexes for Small-Molecule Antagonists.

Tight protein-protein interactions (Kd <100 nm) that occur over a large binding interface (>1000 Å2 ) are highly challenging to disrupt with small molecules. Historically, the design of small molecules to inhibit protein-protein interactions has focused on mimicking the position of interface protein ligand side chains. Here, we explore mimicry of the pairwise intermolecular interactions of the native protein ligand with residues of the protein receptor to enrich commercial libraries for small-molecule inhibitors of tight protein-protein interactions. We use the high-affinity interaction (Kd =1 nm) between the urokinase receptor (uPAR) and its ligand urokinase (uPA) to test our methods. We introduce three methods for rank-ordering small molecules docked to uPAR: 1) a new fingerprint approach that represents uPA's pairwise interaction energies with uPAR residues; 2) a pharmacophore approach to identify small molecules that mimic the position of uPA interface residues; and 3) a combined fingerprint and pharmacophore approach. Our work led to small molecules with novel chemotypes that inhibited a tight uPAR⋅uPA protein-protein interaction with single-digit micromolar IC50 values. We also report the extensive work that identified several of the hits as either lacking stability, thiol reactive, or redox active. This work suggests that mimicking the binding profile of the native ligand and the position of interface residues can be an effective strategy to enrich commercial libraries for small-molecule inhibitors of tight protein-protein interactions.

Galectin-1 inhibitors and their potential therapeutic applications: a patent review.

INTRODUCTION: Galectins have affinity for ß-galactosides. Human galectin-1 is ubiquitously expressed in the body and its expression level can be a marker in disease. Targeted inhibition of galectin-1 gives potential for treatment of inflammatory disorders and anti-cancer therapeutics. AREAS COVERED: This review discusses progress in galectin-1 inhibitor discovery and development. Patent applications pertaining to galectin-1 inhibitors are categorised as monovalent- and multivalent-carbohydrate-based inhibitors, peptides- and peptidomimetics. Furthermore, the potential of galectin-1 protein as a therapeutic is discussed along with consideration of the unique challenges that galectin-1 presents, including its monomer-dimer equilibrium and oxidized and reduced forms, with regard to delivering an intact protein to a pathologically relevant site. EXPERT OPINION: Significant evidence implicates galectin-1's involvement in cancer progression, inflammation, and host-pathogen interactions. Conserved sequence similarity of the carbohydrate-binding sites of different galectins makes design of specific antagonists (blocking agents/inhibitors of function) difficult. Key challenges pertaining to the therapeutic use of galectin-1 are its monomer-dimer equilibrium, its redox state, and delivery of intact galectin-1 to the desired site. Developing modified forms of galectin-1 has resulted in increased stability and functional potency. Gene and protein therapy approaches that deliver the protein toward the target are under exploration as is exploitation of different inhibitor scaffolds.

Structural characterisation of human galectin-4 N-terminal carbohydrate recognition domain in complex with glycerol, lactose, 3'-sulfo-lactose, and 2'-fucosyllactose.

Galectin-4 is a tandem-repeat galectin with two distinct carbohydrate recognition domains (CRD). Galectin-4 is expressed mainly in the alimentary tract and is proposed to function as a lipid raft and adherens junction stabilizer by its glycan cross-linking capacity. Galectin-4 plays divergent roles in cancer and inflammatory conditions, either promoting or inhibiting each disease progression, depending on the specific pathological condition. The study of galectin-4's ligand-binding profile may help decipher its roles under specific conditions. Here we present the X-ray structures of human galectin-4 N-terminal CRD (galectin-4N) bound to different saccharide ligands. Galectin-4's overall fold and its core interactions to lactose are similar to other galectin CRDs. Galectin-4N recognises the sulfate cap of 3'-sulfated glycans by a weak interaction through Arg45 and two water-mediated hydrogen bonds via Trp84 and Asn49. When galectin-4N interacts with the H-antigen mimic, 2'-fucosyllactose, an interaction is formed between the ring oxygen of fucose and Arg45. The extended binding site of galectin-4N may not be well suited to the A/B-antigen determinants, α-GalNAc/α-Gal, specifically due to clashes with residue Phe47. Overall, galectin-4N favours sulfated glycans whilst galectin-4C prefers blood group determinants. However, the two CRDs of galectin-4 can, to a less extent, recognise each other's ligands.

Structural characterization of human galectin-4 C-terminal domain: elucidating the molecular basis for recognition of glycosphingolipids, sulfated saccharides and blood group antigens.

Human galectin-4 is a lectin that is expressed mainly in the gastrointestinal tract and exhibits metastasis-promoting roles in some cancers. Its tandem-repeat nature exhibits two distinct carbohydrate recognition domains allowing crosslinking by simultaneous binding to sulfated and non-sulfated (but not sialylated) glycosphingolipids and glycoproteins, facilitating stabilization of lipid rafts. Critically, galectin-4 exerts favourable or unfavourable effects depending upon the cancer. Here we report the first X-ray crystallographic structural information on human galectin-4, specifically the C-terminal carbohydrate recognition domain of human (galectin-4C) in complex with lactose, lactose-3'-sulfate, 2'-fucosyllactose, lacto-N-tetraose and lacto-N-neotetraose. These structures enable elucidation of galectin-4C binding fine-specificity towards sulfated and non-sulfated lacto- and neolacto-series sphingolipids as well as to human blood group antigens. Analysis of the lactose-3'-sulfate complex structure shows that galectin-4C does not recognize the sulfate group using any specific amino acid, but binds the ligand nonetheless. Complex structures with lacto-N-tetraose and lacto-N-neotetraose displayed differences in binding interactions exhibited by the non-reducing-end galactose. That of lacto-N-tetraose points outward from the protein surface whereas that of lacto-N-neotetraose interacts directly with the protein. Recognition patterns of human galectin-4C towards lacto- and neolacto-series glycosphingolipids are similar to those of human galectin-3; however, detailed scrutiny revealed differences stemming from the extended binding site that offer distinction in ligand profiles of these two galectins. Structural characterization of the complex with 2'-fucosyllactose, a carbohydrate with similarity to the H antigen, and molecular dynamics studies highlight structural features that allow specific recognition of A and B antigens, whilst a lack of interaction with the 2'-fucose of blood group antigens was revealed. DATABASE ACCESSION CODES: 4YLZ, 4YM0, 4YM1, 4YM2, 4YM3.

Galectin-3 inhibitors: a patent review (2008-present).

INTRODUCTION: Galectin-3 (Gal-3), a lectin with preference for ß-galactoside-containing carbohydrates, is a structurally unique member of the galectin family. It is ubiquitously expressed in various mammalian tissues with a wide distribution from the intracellular environment to the extracellular space. Gal-3 is a well-established player in numerous diseases, from infections to heart failure. Notably, as Gal-3 overexpression is associated with cancer drug resistance, it has been identified as a valuable therapeutic target in the fight against cancers. AREAS COVERED: This review discusses the recent progress of patent applications (2008-present) and the current knowledge of pertinent Gal-3-inhibitor interactions in an effort to progress the development of selective and high affinity carbohydrate-based inhibitors targeting Gal-3, with an emphasis on engaging a structure-based drug design rationale. EXPERT OPINION: The lack of commercially available anti-Gal-3 therapeutic reagents and its clear involvement in serious disease, notably cancer, leads to an urgent need for development of inhibitors that specifically target Gal-3. Design of potent and selective carbohydrate inhibitors targeting Gal-3 is challenging due to relative weak protein-carbohydrate interactions along with the high sequence homology in the carbohydrate binding site region among galectins. To date, some chemical scaffolds have been exploited for design of promising effective Gal-3 inhibitors for cancer therapy.

Exploring a structural protein-drug interactome for new therapeutics in lung cancer.

The pharmacology of drugs is often defined by more than one protein target. This property can be exploited to use approved drugs to uncover new targets and signaling pathways in cancer. Towards enabling a rational approach to uncover new targets, we expand a structural protein-ligand interactome () by scoring the interaction among 1000 FDA-approved drugs docked to 2500 pockets on protein structures of the human genome. This afforded a drug-target network whose properties compared favorably with previous networks constructed using experimental data. Among drugs with the highest degree and betweenness two are cancer drugs and one is currently used for treatment of lung cancer. Comparison of predicted cancer and non-cancer targets reveals that the most cancer-specific compounds were also the most selective compounds. Analysis of compound flexibility, hydrophobicity, and size showed that the most selective compounds were low molecular weight fragment-like heterocycles. We use a previously-developed screening approach using the cancer drug erlotinib as a template to screen other approved drugs that mimic its properties. Among the top 12 ranking candidates, four are cancer drugs, two of them kinase inhibitors (like erlotinib). Cellular studies using non-small cell lung cancer (NSCLC) cells revealed that several drugs inhibited lung cancer cell proliferation. We mined patient records at the Regenstrief Medical Record System to explore the possible association of exposure to three of these drugs with occurrence of lung cancer. Preliminary in vivo studies using the non-small cell lung cancer (NCLSC) xenograft model showed that losartan- and astemizole-treated mice had tumors that weighed 50 (p < 0.01) and 15 (p < 0.01) percent less than the treated controls. These results set the stage for further exploration of these drugs and to uncover new drugs for lung cancer therapy.