using-multi-fluorinated-bile-acids-vivo-magnetic-resonance-imaging-to

Using Multi-fluorinated Bile Acids and <em>In Vivo</em> Magnetic Resonance Imaging to Measure Bile Acid Transport

Along with their traditional role as detergents that facilitate fat absorption, emerging literature indicates that bile acids are potent signaling ...

Food and Drug Administration

To synthesize a trifluorinated bile acid that can be used for (19)F magnetic resonance imaging (MRI) of bile acid enterohepatic circulation, characterize its in vitro transporter affinity, stability, and (19)F-MRI signal, and assess its ability to concentrate in the gallbladder of C57BL/6 mice.

Design and characterization of a novel fluorinated magnetic resonance imaging agent for functional analysis of bile Acid transporter activity.

We used live-animal magnetic resonance imaging (MRI) to examine the gallbladders of male mice. These healthy mice were fasted overnight before the study and anesthetized in an animal chamber, with a gas mixture of oxygen and isoflurane for small animal MRI. In the course of these live-animal MRI studies, we observed a Phrygian cap appearance to the gallbladder of one healthy-appearing 6-week-old male mouse, similar to that of the human gallbladder described in many reports. After euthanasia for measurement of bile content, this mouse's gallbladder appeared anatomically normal. To our knowledge, this is the first report of a Phrygian cap appearance of the murine gallbladder.

Phrygian Cap Appearance of a Mouse Gallbladder on Magnetic Resonance Imaging.

A novel trifluorinated cholic acid derivative, CA-lys-TFA, was designed and synthesized for use as a tool to measure bile acid transport noninvasively using magnetic resonance imaging (MRI). In the present study, the in vivo performance of CA-lys-TFA for measuring bile acid transport by MRI was investigated in mice. Gallbladder CA-lys-TFA content was quantified using MRI and liquid chromatography/tandem mass spectrometry. Results in wild-type (WT) C57BL/6J mice were compared to those in mice lacking expression of Asbt, the ileal bile acid transporter. (19)F signals emanating from the gallbladders of WT mice 7 h after oral gavage with 150 mg/kg CA-lys-TFA were reproducibly detected by MRI. Asbt-deficient mice administered the same dose had undetectable (19)F signals by MRI, and gallbladder bile CA-lys-TFA levels were 30-fold lower compared to WT animals. To our knowledge, this represents the first report of in vivo imaging of an orally absorbed drug using (19)F MRI. Fluorinated bile acid analogues have potential as tools to measure and detect abnormal bile acid transport by MRI.

In vivo performance of a novel fluorinated magnetic resonance imaging agent for functional analysis of bile acid transport.

The aim was to elucidate how steps in drug translocation by a solute carrier transporter impact Michaelis-Menten parameters Km, Ki, and Vmax. The first objective was to derive a model for carrier-mediated substrate translocation and perform sensitivity analysis with regard to the impact of individual microrate constants on Km, Ki, and Vmax. The second objective was to compare underpinning microrate constants between compounds translocated by the same transporter. Equations for Km, Ki, and Vmax were derived from a six-state model involving unidirectional transporter flipping and reconfiguration. This unidirectional model is applicable to co-transporter type solute carriers, like the apical sodium-dependent bile acid transporter (ASBT) and the proton-coupled peptide cotransporter (PEPT1). Sensitivity analysis identified the microrate constants that impacted Km, Ki, and Vmax. Compound comparison using the six-state model employed regression to identify microrate constant values that can explain observed Km and Vmax values. Results yielded some expected findings, as well as some unanticipated effects of microrate constants on Km, Ki, and Vmax. Km and Ki were found to be equal for inhibitors that are also substrates. Additionally, microrate constant values for certain steps in transporter functioning influenced Km and Vmax to be low or high.

Mechanistic interpretation of conventional Michaelis-Menten parameters in a transporter system.

Previously, we developed a trifluorinated bile acid, CA-lys-TFA, with the objective of noninvasively assessing bile acid transport in vivo using (19) F magnetic resonance imaging (MRI). CA-lys-TFA was successfully imaged in the mouse gallbladder, but was susceptible to deconjugation in vitro by choloylglycine hydrolase (CGH), a bacterial bile acid deconjugating enzyme found in the terminal ileum and colon. The objective of the present study was to develop a novel trifluorinated bile acid resistant to deconjugation by CGH. CA-sar-TFMA was designed, synthesized, and tested for in vitro transport properties, stability, imaging properties, and its ability to differentially accumulate in the gallbladders of normal mice, compared with mice with known impaired bile acid transport (deficient in the apical sodium-dependent bile acid transporter, ASBT). CA-sar-TFMA was a potent inhibitor and substrate of ASBT and the Na(+) /taurocholate cotransporting polypeptide. Stability was favorable in all conditions tested, including the presence of CGH. CA-sar-TFMA was successfully imaged and accumulated at 16.1-fold higher concentrations in gallbladders from wild-type mice compared with those from Asbt-deficient mice. Our results support the potential of using MRI with CA-sar-TFMA as a noninvasive method to assess bile acid transport in vivo.

Design and evaluation of a novel trifluorinated imaging agent for assessment of bile acid transport using fluorine magnetic resonance imaging.

Floxuridine is often used to treat metastatic liver disease and is given as an infusion directly into the hepatic artery to increase the amount of intact drug that reaches the liver. The objective of this work was to design and synthesize prodrugs of floxuridine through conjugation to chenodeoxycholic acid (CDCA) to target the liver via the bile acid liver uptake transporter Na(+)/taurocholate cotransporting polypeptide (NTCP, SLC10A1). Two isomeric prodrugs of floxuridine were synthesized: floxuridine 3'glutamic acid-CDCA and floxuridine 5'-glutamic acid-CDCA. Both were potent inhibitors and substrates of NTCP. Floxuridine 3'glutamic acid-CDCA showed Ki=6.86±1.37 μM, Km=10.7±2.1 μM, and passive permeability=0.663(±0.121)×10(-7) cm/s while floxuridine 5'-glutamic acid-CDCA showed Ki=0.397±0.038 μM, Km=40.4±15.2 μM, and passive permeability=1.72(±0.18)×10(-7) cm/s. Floxuridine itself had a higher passively permeability of 7.54(±0.45)×10(-7) cm/s in the same cell line, indicating that both prodrugs have the potential for lower non-specific effects than the drug alone. Prodrugs were stable in rat plasma (t=3 h), but quickly released in rat liver s9 fraction, suggesting future in vivo evaluation.

Article	Year
Design and characterization of a novel fluorinated magnetic resonance imaging agent for functional analysis of bile Acid transporter activity. Pharmaceutical research\| PubMed ID: 23319170	2013
Phrygian Cap Appearance of a Mouse Gallbladder on Magnetic Resonance Imaging. Journal of veterinary science & medical diagnosis\| PubMed ID: 24371838	2013
In vivo performance of a novel fluorinated magnetic resonance imaging agent for functional analysis of bile acid transport. Molecular pharmaceutics\| PubMed ID: 24708306	2014
Mechanistic interpretation of conventional Michaelis-Menten parameters in a transporter system. European journal of pharmaceutical sciences : official journal of the European Federation for Pharmaceutical Sciences\| PubMed ID: 25169756	2014
Design and evaluation of a novel trifluorinated imaging agent for assessment of bile acid transport using fluorine magnetic resonance imaging. Journal of pharmaceutical sciences\| PubMed ID: 25196788	2014
Synthesis and in vitro evaluation of bile acid prodrugs of floxuridine to target the liver. International journal of pharmaceutics\| PubMed ID: 25219859	2014

Diana Vivian

Biography

JoVE Journal Publications

Other Publications