A Small Molecule Inhibitor of Scavenger Receptor BI-mediated Lipid Uptake - Probe 3

Faloon PW, Dockendorff C, Yu M, et al.

Publication Details

Scavenger receptor class B, type I (SR-BI) mediates selective uptake of cholesterol from high-density lipoprotein (HDL) particles, a poorly understood process that is distinct from endocytic uptake of lipoproteins, such as low-density lipoprotein (LDL). We set out to find small molecules that inhibit SR-BI function and could be used to characterize the mechanisms involved in HDL uptake. Using a cell-based DiI-HDL uptake assay, we performed a high-throughput screen (HTS) of the NIH MLPCN compound library. Of the 319,533 compounds tested, 3,046 compounds (0.96%) were classified as inhibitors of DiI-HDL uptake. MLS002474293 (SID 85791767, CID 44202943) was identified in the primary HTS as an inhibitor. It had potent activity in the primary assay, a low hit rate in other MLPCN screens, and possessed structural properties suitable for analog synthesis. Structure/activity relationship (SAR) studies led to the probe (CID 44487061/ML312) that showed nanomolar inhibition of HDL uptake through SR-BI and no measurable activity in the cytotoxicity assay. ML312 was tested for lipid efflux inhibition, modulation of HDL binding to SR-BI, and inhibition of endocytosis. ML312 functions by inhibiting both SR-BI-mediated lipid uptake and efflux of free cholesterol to HDL particles. As a tool compound, ML312 is distinct from prior small-molecule inhibitors of SR-BI (e.g., BLT-1 and ITX-5061), and it has better solubility than the existing probes identified concurrently in this project, ML278 and ML279. ML312 will be useful in elucidating how SR-BI mediates lipid transport. In addition, it could clarify the role of SR-BI in a number of biological processes where it plays a crucial role.

Assigned Assay Grant No.: 2 R01 HL052212-11

Screening Center Name & PI: Broad Institute Probe Development Center, Stuart Schreiber

Chemistry Center Name & PI: Broad Institute Probe Development Center, Stuart Schreiber

Assay Submitter & Institution: Monty Krieger, Biology Department, Massachusetts Institute of Technology

PubChem Summary Bioassay Identifier (AID): 488952

Probe Structure and Characteristics

ML312.

ML312

1. Recommendations for Scientific Use of the Probe

We have developed a small-molecule probe (ML312) that inhibits the transfer of lipids between high-density lipoprotein (HDL) and cells mediated by the HDL Scavenger receptor class B, type I (SR-BI). ML312 inhibits both cellular selective lipid uptake of HDL cholesteryl ester and efflux of cellular cholesterol to HDL. This probe will enable deeper understanding of the molecular and cellular functions of SR-BI, across diverse areas of physiology and medicine, and represents a novel lead for further optimization prior to use in in vivo studies.

SR-BI influences multiple facets of lipoprotein/lipid metabolism, and in vitro and in vivo studies (e.g., transgenic and knockout mice) have established a role for SR-BI in many mammalian physiologic and pathophysiologic systems (1–2). SR-BI knockout (KO) mice display increased total plasma cholesterol levels and reduced adrenal cholesterol levels. Female KO mice are infertile due to the importance of lipoprotein metabolism in ovarian function and oocyte maturation (2). Lipoprotein metabolism also impacts endothelial biology, platelet function, bile secretion, steroidogenesis, and cholesterol homeostasis (2). SR-BI is considered to be a pattern-recognition receptor (PRR), a type of immune recognition receptor for microbial substances, such as lipopolysaccharide (LPS; 3), and has the ability to clear LPS and to suppress stimulation of NF-kB and cytokine stimulation via Toll-like receptors (TLRs; 4,5). Further, SR-BI serves as a co-receptor for Hepatitis C (HCV) viral entry, and interference with compound (e.g., ITX-5061) or blocking antibodies can reduce cellular infection (6–9). Additionally, the presence of SR-BI enhances sporozoite invasion efficiency of hepatocytes by the malaria parasite, Plasmodium falciparum (10,11). Thus, ML312 will be a useful tool with which to further explore the contributions of SR-BI in each of these diverse areas in vitro and in vivo.

At the molecular level, SR-BI controls the structure and composition of plasma HDL, and the level and fate of HDL cholesterol, including delivery to the liver and steroidogenic tissues. SR-BI binds HDL and functions as a cell surface transporter to move cholesterol or its esters into or out of cells and as a signaling receptor to control cell function. SR-BI can also interact with and transport a wide variety of other ligands. A new probe could be used to map important sites of interaction on SR-BI for these processes, to help identify possible intracellular binding partners, to verify whether SR-BI oligomerizes to mediate HDL interactions, or to improve our knowledge of other aspects of SR-BI biology.

2. Materials and Methods

See subsections for a detailed description of the materials and methods used for each assay.

Materials and Reagents

  • DiI-HDL, custom purified HDL particles derived from human blood were prepared by the Assay Provider and labeled with 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiI; Catalog No. D-282; Invitrogen, Carlsbad, CA).
  • Alexa 488 HDL, human HDL particles labeled with the Alexa Fluor® 488 Protein Labeling Kit (Catalog No. A-10235, Invitrogen; Carlsbad, CA) were purified and labeled by the Assay Provider.
  • CellTiter-Glo® Luminescent Cell Viability Assay was purchased from Promega (Catalog No. G7573; Madison, WI).
  • Radiolabeled cholesterol [1,2-3H(N)]-, 1 mCi (37 MBq) was obtained from PerkinElmer – NEN (Catalog No. NET139001MC; Waltham, MA).
  • Alexa Fluor-594 conjugated human transferrin (Catalog No. T-13343) was obtained from Invitrogen.

Cell Lines

The following cell lines were used in this study:

  • ldlA[mSR-BI] is a Chinese hamster ovary (CHO) cell line that overexpresses murine SR-BI, isoform 1 (NP_058021) and lacks the LDL receptor was obtained from the Assay Provider (Krieger Laboratory). This cell line was used for the primary assay and several secondary assays. A variant of this cell line expressing mutant SR-BI, where a cysteine required for interaction with BLT-1 is mutated to serine (C384S SR-BI), was used in several secondary assays.
  • [ldlA7] is the parental cell line to ldlA[mSR-BI] cells, and does not overexpresses SR-BI, and can be used to rule out compound activity independent of SR-BI. This cell line was obtained from the Assay Provider.

2.1. Assays

DiI-HDL Uptake Assay (Primary Assay)

ldlA[mSR-BI] cells were plated into 384-well plates at 30 uL per well and incubated overnight. As a measurable surrogate for cholesterol uptake, human HDL particles were treated with the lipophilic fluorescent dye DiI and exposed to ldlA[mSR-BI] cells in lipoprotein-free media (Ham’s F12/0.5% fatty acid-free Bovine Serum Albumin (BSA)/25 mM HEPES pH 7.4 plus 10 μg protein/ml DiI-HDL). Cells took up the Dil via SR-BI over 3 hours in the presence of compound. After significant uptake of the DiI, the cells became fluorescent. The level of fluorescence correlates with the amount of Dil uptake and can be measured with a standard plate reader (here a PerkinElmer EnVision plate reader was used). The uptake of lipid (represented by DiI) was inhibited by the positive control compound BLT-1 or with an excess of HDL untreated with DiI. Primary HTS data were analyzed in GeneData Screener Assay Analyzer, and were normalized against DMSO and the positive control (1 μM BLT-1). For the HTS, the average of two replicates was used to rank order activity and to choose compounds for retests. For dose studies, percent (%) activity was determined for each concentration and the concentration response curves (CRCs) were generated with GeneData Condeseo.

Fluorescence Quencher Counter screen

The primary assay for this project measures a reduction in fluorescence by potential inhibitors. One possible explanation for the loss of signal in the primary assay is the compounds have inherent fluorescence-quenching properties and reduce signal in a dose-dependent manner without actually altering the uptake of the fluorescent substrate. We developed an assay where compounds were pinned into assay buffer containing DiI-HDL without any cells. The compounds were incubated with DiI-HDL at 5 μg protein/mL in Ham’s/25 mM HEPES/0.5% fatty acid-free BSA media for 30 minutes. After the incubation, the assay plates were measured with the identical settings for the primary assay. Compounds that quench DiI fluorescence led to a dose-dependent loss of signal in this assay. Any compound that altered fluorescence or had an IC50 value of <30 μM was not considered for further studies.

Cell Cytotoxicity Assay

It is possible that a compound will cause cells to decrease HDL-mediated Dil uptake due to nonspecific consequences of cellular toxicity, representing a second class of false positive hits. The cells were treated with compounds for 3 hours or 24 hours, and then cell viability was measured using the CellTiter-Glo Assay (Promega), a luciferase-based reagent that measures cellular ATP levels. The compounds were tested at different concentrations to determine IC50 values. Compounds that were toxic after 3 hours with an IC50 value of less than 30 μM were excluded from additional studies. Compounds that were active in the primary assay but toxic below 30 μM at 24 hours (but nontoxic at 3 hours) were still considered. None of our preferred scaffolds showed cytotoxicity at either time point. Data were normalized against DMSO and the positive control (1 μM BLT-1) in GeneData Assay Analyzer. Curves were generated with GeneData Condeseo and showed percent (%) activity for the individual doses.

HDL Binding Assay

HDL binding was assessed using Alexa Fluor 488-labeled HDL particles. For this assay, the Alexa 488 dye was covalently bound to apolipoprotein components of the HDL particle via primary amines; thus, no transfer of the fluorophore to cell membranes occurs. In this manner, direct binding of the HDL particles to SR-BI can be measured. As a positive control, BLT-1, which is known to increase binding of HDL to SR-BI, was used at 1 μM (12). It is possible that a compound can reduce binding of HDL to the receptor, and this would lead to a decrease in signal. This assay is used to characterize the mechanism of action of a particular compound; therefore, any outcome in the assay is acceptable. Data were normalized against DMSO and the positive control (1 μM BLT-1) in Genedata Assay Analyzer. Curves were generated with Genedata Condeseo and showed percent (%) activity for the individual doses.

Cholesterol Efflux Assay

SR-BI also mediates bidirectional flux (e.g. efflux) of unesterified or ‘free’ cholesterol (FC) between cells and HDL or other acceptors. In vivo, the greatest SR-BI-mediated selective uptake occurs in the liver and steroidogenic organs. This assay is used to determine if the compounds alter the efflux of FC to HDL. Compounds that do and do not inhibit efflux in a dose-dependent manner will be of value.

On Day 0, 50,000 cells per well were plated in 24-well plates. On Day 1, the medium was replaced with Ham’s F12 medium supplemented with 10% bovine lipoprotein deficient serum with 1 μCi/mL[1,2-3H]cholesterol (40–60 Ci/mmol). On Day 3, the cells were washed to remove serum and incubated in Ham’s F12 plus 1% fatty acid-free BSA. On Day 4, the cells were washed and pretreated with compounds for 1 hour. Subsequently, the cells were incubated for an additional 2 hours with the same concentrations of small molecules and with unlabeled HDL (final HDL concentration of 100 μg protein/mL). The medium was collected to determine released cholesterol, and the cells were lysed. The amount of [3H]cholesterol in the medium and cells was determined using liquid scintillation counting. Total cellular [3H]cholesterol was calculated as the sum of the radioactivity in the efflux medium plus the radioactivity in the cells and was used to calculate the [3H]cholesterol efflux (percent of total [3H]cholesterol released into the medium).

Mutant SR-BI Cholesterol Efflux Assay

BLT-1 is known to interact at cysteine 384 of SR-BI (16). If the cysteine is mutated to serine, BLT-1 can no longer bind and shows reduced ability to inhibit cholesterol uptake and efflux. This assay is used to determine if the compounds work in a similar fashion to BLT-1 and require Cys384 to alter the efflux of free cholesterol to HDL. Compounds that do and do not inhibit efflux in a dose-dependent manner will be of value.

On Day 0, 50,000 C384S mutant cells per well were plated in 24-well plates. On Day 1, the medium was replaced with Ham’s F12 medium supplemented with 10% bovine lipoprotein deficient serum with 1 uCi/mL [1,2-3H]cholesterol (40–60 Ci/mmol). On Day 3, the cells were washed to remove serum and incubated in Ham’s F12 plus 1% fatty acid-free BSA. On Day 4, the cells were washed and pretreated with compounds for 1 hour. Subsequently, the cells were incubated for an additional 2 hours with the same concentrations of small molecules and with unlabeled HDL (final HDL concentration of 100 μg protein/mL). The medium was collected to determine released cholesterol, and the cells were lysed. The amount of [3H]cholesterol in the medium and within cells was calculated using liquid scintillation counting. Total cellular [3H]cholesterol was determined as the sum of the radioactivity in the efflux medium plus the radioactivity in the cells and was used to calculate the [3H]cholesterol efflux (percent of total [3H]cholesterol released into the medium).

DiI-HDL Uptake Assay in the Absence of SR-BI

In the primary assay, the cell line used to measure DiI-HDL uptake lacks the LDL receptor and overexpresses SR-BI. In this assay, the parental CHO cell line [ldlA7], which lacks the LDL receptor but does not overexpress SR-BI, was used to determine if inhibitors work via non-SR-BI-mediated mechanisms. As in the primary assay, 10 μg protein/mL of DiI-HDL was used to measure Dil uptake into the cells after 3 hours of incubation with different concentrations of compound. Little to no uptake of DiI-HDL was observed in these cells, and the only signal observed was minimal background staining.

Transferrin Endocytosis Assay

This assay measures the endocytosis of an independent ligand, transferrin, which is not taken up via SR-BI but by clathrin-mediated endocytosis. This assay provides a measure of the selectivity of the inhibitors. Alexa-594-labeled transferrin is taken into the cell via endocytosis and localization of labeled transferrin was quantitated in the various intracellular compartments. If a compound inhibits endocytosis, the labeled transferrin will not enter the cell leading to a decrease of fluorescent signal. Inhibitors of interest should act selectively at SR-BI and should have no activity in this assay.

Cells were pre-treated with compound for 3 hours and then treated with the Alexa-594 transferrin reagent for 30 minutes (Catalog No.T-13343; Invitrogen) in serum-free media. Plates of cells were then placed onto ice, washed with ice cold PBS, and then fixed with 4% paraformaldehyde. In addition, the nuclei were stained with 300 nM 4′,6-diamidino-2-phenylindole (DAPI) (Catalog No. 21490; Invitrogen). The cells were imaged with the Molecular Devices IXM microscope. Translocation measurements were performed using MetaExpress software and normalized for cell number.

Radioactive HDL Uptake Assay (wild type SR-BI)

The goal of this assay is to verify compounds that disrupt the binding of HDL particles to SR-BI scavenger receptor using an alternative means of labeling HDL-cholesterol particles and avoiding any type of fluorescence measurement. To measure this binding event, HDL particles are labeled with [3H]cholesterol and added to mSR-BI cells. Cells take up HDL via the SR-BI scavenger receptor in 2 to 3 hours. After significant uptake of the HDL, the radiolabel can be detected by liquid scintillation counting. The level of radioactivity correlates with the amount of HDL uptake. The uptake of lipid particles can be inhibited by the compound BLT-1 or when co-treated with an excess of unlabeled HDL. The ldlA[mSR-BI] cells utilized in the assay are a CHO cell line lacking expression of the LDL receptors and overexpressing the scavenger receptor, SR-BI. Inhibitors of SR-BI and HDL uptake will have a reduction in liquid scintillation counts.

Radioactive HDL Uptake Assay (mutant SR-BI)

BLT-1 is known to interact at cysteine 384 of SR-BI (16). If the cysteine is mutated to serine, BLT-1 can no longer bind and shows no ability to inhibit cholesterol uptake. This assay is used to determine if the compounds work in a similar fashion to BLT-1 and require Cys384 to alter the binding of HDL particles to mutant SR-BI scavenger receptor, using an alternative means of labeling HDL-cholesterol particles and avoiding any type of fluorescence measurement. To measure this binding event, HDL particles are labeled with [3H]cholesterol and added to C384S SR-BI cells. Cells take up HDL via the SR-BI scavenger receptor in 2 to 3 hours. After significant uptake of the HDL, the radiolabel is detected by liquid scintillation counting. The level of radioactivity correlates with the amount of HDL uptake. The uptake of lipid particles can be inhibited by the treatment with an excess of unlabeled HDL. The mSR-BI cells utilized in the assay are a CHO cell line lacking expression of the LDL receptors and overexpressing the mutant scavenger receptor, C384S SR-BI.

2.2. Probe Chemical Characterization

The probe (ML312) was prepared as described in Section 2.3, and was analyzed by UPLC, 1H and 13C NMR spectroscopy, and high-resolution mass spectrometry. The data obtained from NMR and mass spectrometry were consistent with the structure of the probe, and UPLC analysis showed purity of >95%.

The solubility of the probe was determined to be 79 μM in phosphate-buffered saline (PBS; pH 7.4, 23°C) solution with 1% DMSO. The probe is stable in PBS (1% DMSO), with 94% remaining after 48 hours (Figure 1A). The probe is also stable in the presence of GSH, with 91% of the compound remaining after 6 hours (Figure 1B).

Figure 1A. Stability of Probe (ML312) in PBS Buffer (with 1% DMSO, pH 7.4, 23°C).

Figure 1A

Stability of Probe (ML312) in PBS Buffer (with 1% DMSO, pH 7.4, 23°C).

Figure 1B. Stability of Probe (ML312) to GSH (with 1% DMSO).

Figure 1B

Stability of Probe (ML312) to GSH (with 1% DMSO).

The compound is also very stable in human and mouse plasma, with >99% remaining after a 5-hour incubation period. Plasma protein binding (PPB) studies showed that it was 94.5% bound in human plasma, and 94.7% bound in mouse plasma.

Table 1 summarizes known properties of the probe.

Table 1. Summary of Known Probe (ML312) Properties Computed from Structure.

Table 1

Summary of Known Probe (ML312) Properties Computed from Structure.

2.3. Probe Preparation

The probe and several related analogs were prepared in a straightforward fashion as described in Scheme 1. Other analogs were previously prepared using solid-phase synthesis as described by Marcaurelle at al. (19). Lactam 1 was previously prepared via a “build-couple-pair” strategy using a SNAr reaction as the key ring-forming (pairing) step (19). This compound was elaborated by protecting the hydroxyl group as a t-butyldimethylsilyl (TBS) ether, then removing the aniline protecting group (Fmoc) with piperidine. The free aniline was then reacted with isopropylisocyanate to provide the urea 4 in good yield. Catalytic Pd(PPh3)4 with N,N-dimethylbarbituric acid was then used to liberate the secondary amine 5, which was subsequently reacted with 4-chlorophenylsulfonyl chloride to give sulfonamide 6. The TBS protecting group was finally removed with a fluoride source (TBAF) to provide the probe (ML312) in good yield and excellent purity after column chromatography.

Scheme 1. Synthesis of Probe 3 (ML312).

Scheme 1

Synthesis of Probe 3 (ML312).

3. Results

3.1. Dose Response Curves for Probe

Figure 2 shows the dose response curves for the probe (ML312).

Figure 2. Dose Response Curves for the Probe (ML312).

Figure 2

Dose Response Curves for the Probe (ML312). ML312 was used over a range of concentrations up to 35 μM in the primary DiI-HDL uptake assay, IC50=0.06 μM (AID 623880) (A), Alexa-488 HDL binding assay, AC50 = 0.44 μM (AID 624024) (more...)

3.2. Cellular Activity

The primary assay and several of the secondary assays are cell-based experiments. The compound is active in cells and the effective IC50 value for the probe in the primary assay (averaging 0.10 μM over multiple assays) is well below the measured PBS solubility (79 μM). This inhibition of uptake was confirmed with a non-fluorescent, radiolabeled version of the assay with an average IC50 value of 1.0 μM (AID 624022). Since SR-BI is a cell-surface receptor (and the compound is presumed to act on the extracellular surface), cell permeability is not an issue, though the probe is expected to have reasonable permeability. ML312 showed no cytotoxicity at 24 hours (Figure 2C, AID 623895). The probe also shows no effect on the endocytosis of transferrin, at concentrations up to 35 μM (AID 602134).

3.3. Profiling Assays

The probe will be sent to Ricerca for evaluation of off-target binding to a broad panel of receptors, ion channels and enzymes. Metabolic stability will also be evaluated.

4. Discussion

4.1. Comparison to Existing Art and How the New Probe is an Improvement

We decided to compare the performance of ML312 with BLT-1, the prior art compounds ITX-5061 and R-154716, plus our probes ML278 and ML279. These results are presented in Table 2 and Figure 3 (AID 624029). BLT-1 is a potent inhibitor of SR-BI-mediated lipid uptake and of free cholesterol efflux, however it is a non-reversible covalent modifier of SR-BI and is highly toxic to cells. In contrast, ML312 is presumably a reversible inhibitor of HDL uptake (it possesses no functionality that would make it susceptible to covalent reaction), and shows no cytotoxicity in [ldlA]mSR-BI cells (Figure 2C, AID 588829). As discussed earlier, ITX-5061 is an SR-BI inhibitor that is currently (as of April, 2012) in a Phase 1b clinical trial for HCV infection. We synthesized both ITX-5061 and R-154716 for direct comparison to ML312 in multiple assays. Although ML312 is not as potent as R-154716 or our other probes ML278 and ML279, it is has significantly better solubility than any of the probes or prior art thus identified. We also expect it to have better selectivity over secondary targets, in part due to its multiple stereocenters and increased fraction of sp3 carbons (20). The testing of this hypothesis is anticipated by screening our probes and the prior art against an extended panel of pertinent receptors and other off-target proteins.

Table 2. Comparison of Probes to Select Prior Art Compounds.

Table 2

Comparison of Probes to Select Prior Art Compounds.

Figure 3. Comparison of ML312 to Prior Art Compounds with DiI-HDL Uptake.

Figure 3

Comparison of ML312 to Prior Art Compounds with DiI-HDL Uptake. ML312 was tested against prior art compounds (ML278, ML279, R-154716, BLT-1 and ITX-5061) in the DiI-HDL uptake assay. Each compound was tested over a range of concentrations up to 35 μM. (more...)

Investigation into relevant prior art entailed searching the following databases: SciFinder, Web of Science, PubMed, and Patent Lens. Abstracts were obtained and were analyzed for relevance to the current project. The searches are current as of February 24, 2012.

5. References

1.
Acton S, Rigotti A, Landschulz KT, Xu S, Hobbs HH, Krieger M. Identification of scavenger receptor SR-BI as a high density lipoprotein receptor. Science. 1996 Jan 26;271(5248):518–20. [PubMed: 8560269]
2.
Rigotti A, Miettinen HE, Krieger M. The role of the high-density lipoprotein receptor SR-BI in the lipid metabolism of endocrine and other tissues. Endocr Rev. 2003 Jun;24(3):357–87. [PubMed: 12788804]
3.
Fioravanti J, Medina-Echeverz J, Berraondo P. Scavenger receptor type B, class I: A promising immunotherapy target. Immunotherapy. 2011 Mar;3(3):395–406. [PubMed: 21395381]
4.
Guo L, Song Z, Li M, Wu Q, Wang D, Feng H, Bernard P, Daugherty A, Huang B, Li XA. Scavenger receptor BI protects against septic death through its role in modulating inflammatory response. J Biol Chem. 2009 Jul 24;284(30):19826–34. Epub 2009 Jun 2. [PMC free article: PMC2740408] [PubMed: 19491399]
5.
Zhu P, Liu X, Treml LS, Cancro MP, Freedman BD. Mechanism and regulatory function of CpG signaling via scavenger receptor B1 in primary B cells. J Biol Chem. 2009 Aug 21;284(34):22878–87. Epub 2009 Jun 19. [PMC free article: PMC2755695] [PubMed: 19542230]
6.
Catanese MT, Graziani R, von Hahn T, Moreau M, Huby T, Paonessa G, Santini C, Luzzago A, Rice CM, Cortese R, Vitelli A, Nicosia A. High-avidity monoclonal antibodies against the human scavenger class B type I receptor efficiently block hepatitis C virus infection in the presence of high-density lipoprotein. J Virol. 2007 Aug;81(15):8063–71. Epub 2007 May 16. [PMC free article: PMC1951280] [PubMed: 17507483]
7.
Catanese MT, Ansuini H, Graziani R, Huby T, Moreau M, Ball JK, Paonessa G, Rice CM, Cortese R, Vitelli A, Nicosia A. Role of scavenger receptor class B type I in hepatitis C virus entry: kinetics and molecular determinants. J Virol. 2010 Jan;84(1):34–43. [PMC free article: PMC2798406] [PubMed: 19828610]
8.
Voisset C, Callens N, Blanchard E, Op De, Beeck A, Dubuisson J, Vu-Dac N. High density lipoproteins facilitate hepatitis C virus entry through the scavenger receptor class B type I. J Biol Chem. 2005 Mar 4;280(9):7793–9. Epub 2005 Jan. [PubMed: 15632171]
9.
Syder AJ, Lee H, Zeisel MB, Grove J, Soulier E, Macdonald J, Chow S, Chang J, Baumert TF, McKeating JA, McKelvy J, Wong-Staal F. Small molecule scavenger receptor BI antagonists are potent HCV entry inhibitors. J Hepatol. 2011 Jan;54(1):48–55. Epub 2010 Aug 21. [PubMed: 20932595]
10.
Rodrigues CD, Hannus M, Prudêncio M, Martin C, Gonçalves LA, Portugal S, Epiphanio S, Akinc A, Hadwiger P, Jahn-Hofmann K, Röhl I, van Gemert GJ, Franetich JF, Luty AJ, Sauerwein R, Mazier D, Koteliansky V, Vornlocher HP, Echeverri CJ, Mota MM. Host scavenger receptor SR-BI plays a dual role in the establishment of malaria parasite liver infection. Cell Host Microbe. 2008 Sep 11;4(3):271–82. [PubMed: 18779053]
11.
Yalaoui S, Huby T, Franetich JF, Gego A, Rametti A, Moreau M, Collet X, Siau A, van Gemert GJ, Sauerwein RW, Luty AJ, Vaillant JC, Hannoun L, Chapman J, Mazier D, Froissard P. Scavenger receptor BI boosts hepatocyte permissiveness to Plasmodium infection. Cell Host Microbe. 2008 Sep 11;4(3):283–92. [PubMed: 18779054]
12.
Nieland TJ, Penman M, Dori L, Krieger M, Kirchhausen T. Discovery of chemical inhibitors of the selective transfer of lipids mediated by the HDL receptor SR-BI. Proc Natl Acad Sci U S A. 2002 Nov 26;99(24):15422–7. Epub 2002 Nov 18. [PMC free article: PMC137732] [PubMed: 12438696]
13.
Masson D, Koseki M, Ishibashi M, Larson CJ, Miller SG, King BD, Tall AR. Increased HDL cholesterol and apoA-I in humans and mice treated with a novel SR-BI inhibitor. Arterioscler Thromb Vasc Biol. 2009 Dec;29(12):2054–60. Epub 2009 Oct 8. [PMC free article: PMC2783626] [PubMed: 19815817]
14.
Nieland TJ, Chroni A, Fitzgerald ML, Maliga Z, Zannis VI, Kirchhausen T, Krieger M. Cross-inhibition of SR-BI- and ABCA1-mediated cholesterol transport by the small molecules BLT-4 and glyburide. J Lipid Res. 2004 Jul;45(7):1256–65. Epub 2004 Apr 21. [PubMed: 15102890]
15.
Nieland TJ, Shaw JT, Jaipuri FA, Maliga Z, Duffner JL, Koehler AN, Krieger M. Influence of HDL-cholesterol-elevating drugs on the in vitro activity of the HDL receptor SR-BI. J Lipid Res. 2007 Aug;48(8):1832–45. Epub 2007 May 28. [PubMed: 17533223]
16.
Yu M, Romer KA, Nieland TJ, Xu S, Saenz-Vash V, Penman M, Yesilaltay A, Carr SA, Krieger M. Exoplasmic cysteine Cys384 of the HDL receptor SR-BI is critical for its sensitivity to a small-molecule inhibitor and normal lipid transport activity. Proc Natl Acad Sci U S A. 2011 Jul 26;108(30):12243–8. Epub 2011 Jul 11. [PMC free article: PMC3145699] [PubMed: 21746906]
17.
Nieland TJ, Shaw JT, Jaipuri FA, Duffner JL, Koehler AN, Banakos S, Zannis VI, Kirchhausen T, Krieger M. Identification of the molecular target of small molecule inhibitors of HDL receptor SR-BI activity. Biochemistry. 2008 Jan 8;47(1):460–72. Epub 2007 Dec 8. [PMC free article: PMC2736594] [PubMed: 18067275]
18.
Nishizawa T, Kitayama K, Wakabayashi K, Yamada M, Uchiyama M, Abe K, Ubukata N, Inaba T, Oda T, Amemiya Y. A novel compound, R-138329, increases plasma HDL cholesterol via inhibition of scavenger receptor BI- mediated selective lipid uptake. Atherosclerosis. 2007 Oct;194(2):300–8. Epub 2006 Dec 12. [PubMed: 17166497]
19.
Marcaurelle L, et al. An Aldol-based build/couple/pair strategy for the synthesis of medium- and large-sized rings: Discovery of macrocyclic histone deacetylase inhibitors. J Am Chem Soc. 2010 Dec 1;132(47):16962–76. Epub 2010 Nov 10. [PMC free article: PMC3004530] [PubMed: 21067169]
20.
Lovering F, Bikker J, Humblet C. Escape from flatland: Increasing saturation as an approach to improving clinical success. J Med Chem. 2009 Nov 12;52(21):6752–6. Epub 2009 Oct 14. [PubMed: 19827778]
21.
Gaidukov L, Nager AR, Xu S, Penman M, Krieger M. Glycine dimerization motif in the N-terminal transmembrane domain of the high density lipoprotein receptor SR-BI required for normal receptor oligomerization and lipid transport. J Biol Chem. 2011 May 27;286(21):18452–64. Epub 2011 Mar 25. [PMC free article: PMC3099662] [PubMed: 21454587]
22.
Kocher O, Krieger M. Role of the adaptor protein PDZK1 in controlling the HDL dSR-BI. Curr Opin Lipidol. 2009 Jun;20(3):236–41. [PMC free article: PMC2849150] [PubMed: 19421056]
23.
Dorner M, Horwitz J, Robbins J, Barry W, Feng Q, Mu K, Jones C, Schoggins J, Catanese M, Burton D, Law M, Rice C, Ploss A. A genetically humanized mouse model of hepatitis C virus infection. Nature. 2011 Jun;474(7350):208–13. [PMC free article: PMC3159410] [PubMed: 21654804]