QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) All Versions of this Article: jpet.104.075895v1 312/2/517    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Jung, A. S. Articles by Margulies, K. B. Search for Related Content PubMed PubMed Citation Articles by Jung, A. S. Articles by Margulies, K. B.

CARDIOVASCULAR

Pharmacological Effects of ATI22-107 [2-(2-{2-[2-Chloro-4-(6-oxo-1,4,5,6-tetrahydro-pyridazin-3-yl)-phenoxy]-acetylamino}-ethoxymethyl)-4-(2-chloro-phenyl)-6-methyl-1,4-dihydro-pyridine-3,5-dicarboxylic Acid Dimethyl Ester)], a Novel Dual Pharmacophore, on Myocyte Calcium Cycling and Contractility

Departments of Physiology and Cardiology, Temple University, Philadelphia, Pennsylvania (A.S.J., M.P.Q., G.D.M., S.R.H., K.B.M.); and Department of Cardiovascular Biology, Artesian Therapeutics Inc., Gaithersburg, Maryland (D.P.B.)

Received for publication August 20, 2004
Accepted November 16, 2004.

	Abstract

Top Abstract Materials and Methods Results Discussion References

 
Historically, inhibitors of type III phosphodiesterases (PDE-III) have been effective inotropes in mammalian myocardium, but their clinical utility has been limited by adverse events, including arrhythmias that are considered to be due to Ca²⁺ overload. ATI22-107 [2-(2-{2-[2-chloro-4-(6-oxo-1,4,5,6-tetrahydro-pyridazin-3-yl)-phenoxy]-acetylamino}-ethoxymethyl)-4-(2-chlorophenyl)-6-methyl-1,4-dihydro-pyridine-3,5-dicarboxylic acid dimethyl ester)], a novel, dual pharmacophore compound, was designed to simultaneously inhibit the cardiac phosphodiesterase (PDE-III) and produce inotropic effects, whereas inhibiting the L-type calcium channel (LTCC) was designed to minimize increases in diastolic Ca²⁺. We compared the effects of ATI22-107 and enoximone, a pure PDE-III inhibitor, on the Fluo-3 calcium transient in normal feline ventricular myocytes and trabeculae. Enoximone-induced dose-dependent increases in peak [Ca²⁺]_i, diastolic [Ca²⁺]_i, T50, and T75. ATI22-107 demonstrated similar dose-dependent increases in peak [Ca²⁺]_i at 300 nM and 1.0 µM doses, with no further increases at higher doses. Throughout the dosing range, ATI22-107 induced much smaller, if any, increases in diastolic [Ca²⁺]_i, T₂₅, and T₇₅. Current measurement of LTCC via patch-clamp techniques revealed dose-dependent decreases in LTCC current with an increasing dose of ATI22-107, thereby validating the dual functionality of the drug that has been proposed in this study. Studies in isolated trabeculae demonstrated that enoximone-induced a dose-dependent augmentation of the entire force-frequency relation in normal myocardium, whereas augmentation of contractility was only observed at lower stimulation frequencies with ATI22-107. These results demonstrate the effects of the LTCC-antagonizing moiety of ATI22-107 and suggest that the novel simultaneous combination of PDE-III and LTCC inhibition by one molecule may produce a favorable profile of limited inotropy without detrimental effects of increased diastolic [Ca²⁺]_i.

Despite improvements in cardiac contractility, inotropes used to date have not been successful in decreasing mortality, largely because of increases in sudden death (Teerlink et al., 2000). In fact, recent studies suggest that excessive sarcoplasmic reticulum Ca²⁺ load may contribute to ventricular arrhythmias in failing hearts (Sipido et al., 2000) and could provide an explanation for increases in sudden death caused by existing inotropes, especially phosphodiesterase III (PDE-III) inhibitors (Naccarelli and Goldstein, 1989). Nevertheless, as recently reviewed, depressed cardiac contractility and reduced contractility reserve represent fundamental features of the pathophysiology of progressive heart failure (Houser and Margulies, 2003). Therefore, the development of novel new inotropes is still relevant.

Using proprietary techniques, a novel dual pharmacophore compound, 2-(2-{2-[2-chloro-4-(6-oxo-1,4,5,6-tetrahydro-pyridazin-3-yl)-phenoxy]-acetylamino}-ethoxymethyl)-4-(2-chlorophenyl)-6-methyl-1,4-dihydro-pyridine-3,5-dicarboxylic acid dimethyl ester, also called ATI22-107, has been developed with distinct chemical moieties that inhibit PDE-III and the L-type calcium channel (LTCC). We hypothesized that ATI22-107 would preserve the inotropic effects of a pure PDE-III inhibitor while minimizing the increased diastolic calcium levels by antagonizing the LTCC. To address this hypothesis, we compared in vitro pharmacologic actions of a pure PDE-III inhibitor, 1,3-dihydro-4-methyl-5-[4-(methylthio)-benzoyl]-2H-imidazole-2-one, also called enoximone, with ATI22-107 in isolated cardiac myocytes and thin trabeculae from normal feline hearts. Our findings demonstrate clear distinctions between the two compounds and indicate that ATI22-107 does indeed exhibit moderate inotropic activity while simultaneously enhancing rates of myocardial [Ca²⁺]_i decay and preventing increases in diastolic calcium [Ca²⁺]_i.

	Materials and Methods

Top Abstract Materials and Methods Results Discussion References

 
Pharmacology of ATI22-107 and PDE-III/LTCC Specificity. The title compound ATI22-107 possesses an inhibitory activity against PDE-III and LTCC and is a patented dual pharmacophore compound with a published molecular structure (Hamilton and Leighton, 2003). To ensure the PDE-III and LTCC inhibitory effects of ATI22-107, we performed separate PDE-III and LTCC inhibition assays as described previously (Lee et al., 1984; Weishaar et al., 1986).

Animal Preparation and Myocyte Cell Isolation. Normal adult felines were used for these studies. Animals were handled in accordance with the guidelines of the Guide for the Care and Use of Laboratory Animals published by the National Institutes of Health. Animals were anesthetized with pentobarbital, and feline left ventricular myocytes were isolated as described previously (duBell and Houser, 1989).

Intracellular Calcium ([Ca²⁺]_i) in Isolated Myocytes. Freshly isolated myocytes were loaded with Fluo-3 acetoxymethyl ester (Molecular Probes, Eugene, OR) at final concentration of 4 to 10 µM in the presence of 1 mM Ca²⁺. Myocytes were placed in a chamber on the stage of an inverted microscope and superfused at 1 to 2 ml/min at 37°C with Tyrode's solution (150 mM NaCl, 5.4 mM KCl, 1 mM CaCl, 1.2 mM MgCl, 10 mM glucose, 2 mM pyruvate, and 5 mM HEPES, pH 7.4). Calcium-tolerant, rod-shaped myocytes were chosen based on their appearance, including the absence of membrane irregularities and spontaneous contractions. Fluo-3 was excited at 480 nm with xenon light, and the emitted light was split with a 505-nm dichroic mirror. The emission at 530 nm was recorded to represent the cytosolic [Ca²⁺]_i transient. Myocytes were field-stimulated at 1.0 Hz and exposed to increasing doses (0, 0.3, 1.0, 3.0, and 10 µM) of either enoximone or ATI22-107. For every cell, 1.5 to 2 min of equilibration time was allowed upon each increase in dose until a steady state of the peak [Ca²⁺]_i was achieved. The [Ca²⁺]_i signal was recorded and saved on a computer for later analysis using pClamp software (Axon Instruments Inc., Union City, CA). Variables derived from raw [Ca²⁺]_i transients included peak [Ca²⁺]_i, diastolic [Ca²⁺]_i, time to 50% decay of the [Ca²⁺]_i transient (T₅₀), and time to 75% decay of the [Ca²⁺]_i transient (T₇₅).

Measurement of LTCC Currents (I_Ca,L). Myocytes were studied in a chamber mounted on an inverted microscope (Nikon, Tokyo, Japan) and initially perfused at 37°C with a normal physiological salt solution containing 150 mM/l NaCl, 5.4 mM/l KCl, 1 mM/l CaCl₂, 1.2 mM/l MgCl₂, 10 mM/l glucose, 2 mM/l sodium pyruvate, and 5 mM/l HEPES (pH 7.4). Low resistance (1–4 M) patch pipettes filled with a solution containing 130 mM/l cesium aspartate, 10 mM/l N-methyl-D-glucamine, 20 mM/l TEA-Cl, 2.5 mM/l Tris-ATP, 0.05 mM/l Tris-GTP, 1 mM/l MgCl₂, and 10 mM/l EGTA (pH 7.2) were used in whole-cell voltage clamp experiments. All myocytes were dialyzed with this solution and perfused with normal physiological salt solution for 10 min before experiments were initiated. After the initial dialysis period, myocytes were bathed with a sodium- and potassium-free bath solution containing 150 mM/l N-methyl-D-glucamine, 2 mM/l CaCl₂, 5.4 mM/l CsCl, 1.2 mM/l MgCl₂, 10 mM/l glucose, 5 mM/l HEPES, and 2 mM/l 4-aminopyridine (pH 7.4). All experiments were performed in sodium- and potassium-free (in and out) solutions so that Ca²⁺ currents were measured with minimal contamination from overlapping ionic currents. Precautions were made to ensure that leak current was never greater than 0.14 pA. Membrane potential and whole cell currents were measured with standard techniques as described in detail previously (Piacentino et al., 2002). Junction potentials were not corrected and were less than 10 mV. The cell capacitance was measured using small 10-mV hyperpolarizing test steps from -80 to 80 mV. Membrane potentials were controlled with an Axopatch 2B (Axon Instruments Inc.) voltage-clamp amplifier using pClamp8 (Axon Instruments Inc.) software and acquired with a Digidata 1200 analog to digital converter (Axon Instruments Inc.). The data were analyzed with Clampfit (Axon Instruments Inc.) and presented with Origin 6.0 (Microcal Inc., Studio City, CA). IV relationships were determined under control conditions, with 300 nM and 10 µM ATI22-107.

Force-Frequency Studies in Isolated Trabeculae. Right ventricular trabeculae were isolated from feline hearts, mounted in a custom-made force transducer apparatus, and allowed to equilibrate as described previously (Rossman et al., 2004). Once stabilized (at 0.5 Hz), trabeculae underwent control force-frequency experiments. Steady-state twitches were recorded at 0.5, 1.0, 1.5, 2.0, and 2.5 Hz. After control measurements, trabeculae were randomly chosen to be exposed to step doses of enoximone (300 nM and 1, 3, and 10 µM) or ATI22-107 (0.3, 1, 3, and 10 µM). A force-frequency experiment was performed at each dose of either enoximone or ATI22-107. After each dose change or change in stimulation frequency (0.5 to 2.5 Hz), equilibration was allowed until a steady state of force development was achieved.

Statistical Analysis. Results are presented as the mean ± S.E.M. unless otherwise stated. Statistical significance was determined by one-way, two-way, or repeated measures two-way analysis of variance and a Student-Newman-Keuls post hoc test [GraphPad Instat (GraphPad Software, Inc., San Diego, CA) and SPSS statistical software (SPSS Inc., Chicago, IL)]. Values of p < 0.05 were considered significant.

	Results

Top Abstract Materials and Methods Results Discussion References

 
PDE-III and LTCC Inhibition Assays. As seen in Table 1, ATI22-107 exhibits strong dose-dependent inhibition of both PDE-III and LTCC specific activity.

[Ca²⁺]_i Transients in Isolated Myocytes. Figure 1, A and B, shows typical Ca²⁺ transient traces of dose-response experiments for both enoximone and ATI22-107. Figure 1, C through F, shows that enoximone induced dose-dependent increases in peak [Ca²⁺]_i, diastolic [Ca²⁺]_i, T₅₀, and T₇₅. ATI22-107 demonstrated similar dose-dependent increases in peak [Ca²⁺]_i at 300 nM and 1.0 µM doses, with no further increases at higher doses. Moreover, throughout the dosing range, ATI22-107 induced little, if any, increase in diastolic [Ca²⁺]_i, T₅₀, and T₇₅ compared with enoximone (ATI22-107, n = 19; enoximone, n = 8).

View larger version (25K): [in this window] [in a new window]   Fig. 1. Dose-dependent Ca2+ transient relationship (normalized and raw traces) are shown for (A) enoximone and (B) ATI22-107. Enoximone exhibited a dose-dependent increase in inotropy throughout the dose range. ATI22-107 followed a similar trend consistently for only the first two doses. The higher doses caused no increase or diminished inotropy. Raw traces clearly show the much larger increases in diastolic calcium by enoximone compared with ATI22-107. Comparisons between enoximone and ATI22-107 for (C) peak Ca2+, (D) diastolic Ca2+, (E) T50, and (F) T75 (T50 and T75, time to 50 and 75% descent of the calcium transient, respectively) have been made. Enoximone induced dose-dependent increases in peak [Ca2+]i, diastolic [Ca2+]i, T50, and T75. ATI22-107 demonstrated similar dose-dependent increases at 300 nM and 1.0 µM doses with less or no further increases at higher doses. Throughout the dosing range, ATI22-107 induced much smaller, if any, increases in diastolic [Ca 2+]i, T50, and T75. This indicates an ability to increase inotropy while maintaining a relatively low [Ca2+]i. *, p < 0.05 versus enoximone at the equivalent dose.

I_Ca,L Measurements. As illustrated in Fig. 2, ATI22-107 induced dose-dependent decreases in I_Ca,L across a physiologic voltage range compared with baseline. These data demonstrate the activity of the LTCC moiety of the dual pharmacophore and indicate that the enhanced peak [Ca²⁺]_i, induced by ATI22-107, can only be attributed to the PDE inhibition moiety of the dual pharmacophore (ATI22-107, n = 3).

View larger version (16K): [in this window] [in a new window]   Fig. 2. Dose-dependent Ca2+ current-voltage relationship for 0, 300 nM, and 10 µM concentrations. A, representative currents for 0, 300 nM, and 10 µM concentrations of ATI22-107. Notice there is a noticeable decrease in calcium current with an increasing dose of ATI22-107, especially at 10- and 20-mV step voltages. B, current-voltage relationship at each dose found to be statistically different from one another (p < 0.001). There is a significant dose-dependent decrease in the current across a physiologic voltage range with increasing doses of ATI22-107. ATI22-107 clearly blocks the LTCC, which is most likely responsible for the unchanging diastolic Ca2+ levels across all doses and the diminished peak Ca2+ at higher doses.

Force-Frequency Studies. Figure 3, A through C, depicts force transients from isolated trabeculae during increments in stimulation frequency under control conditions and with a high dose of enoximone and ATI22-107. Data from all experiments are summarized in Table 2 and Fig. 3, D and E. Under control conditions, increasing stimulation frequency induces increased developed force (a positive-frequency response), faster rates of force development and force decline, and minimal changes in diastolic force. Enoximone induced dose-dependent increases in the developed tension and +dF/dt at each frequency. However, enoximone did not significantly enhance rate-dependent increases in the rate of force decline (-dF/dt). ATI22-107 induced similar dose-dependent increases in developed force and +dF/dt at lower stimulation frequencies (0.5, 1.0, and 1.5 Hz). However, in contrast to enoximone, ATI22-107 did not augment these parameters at higher stimulation frequencies (2.0 and 2.5 Hz). Another distinction between ATI22-107 and enoximone is that ATI22-107 produced dose-dependent increases in -dF/dt versus control at lower stimulation frequencies.

View larger version (25K): [in this window] [in a new window]   Fig. 3. Effects of ATI22-107 on feline trabeculae. Representative traces of raw data from a force-frequency experiment in normal feline cardiac trabeculae at (A) control, (B) a high dose (10 µM) of enoximone, and (C) a high dose (10 µM) of ATI22-107. Peak developed force is plotted against frequency separately for each dose of (D) ATI22-107 and (E) enoximone. The force-frequency relationship at each dose was found to be statistically different from one another (p < 0.05). There seemed to be a positive force-frequency relationship up to 2.0 Hz for all doses except the highest dose. The highest dose (10 µM) of ATI22-107 showed a decrease in developed tension at the two higher frequencies (2.0 and 2.5 Hz). #, p < 0.05 versus control at the same frequency; +, p < 0.05 versus previous dose at the same frequency; *, p < 0.05 versus the same dose of enoximone at the same frequency.

Figure 4 depicts the force-frequency responses as percentage change in developed tension versus 0.5 Hz of the same dose and drug. This depiction of the data clearly demonstrates that the normal positive force-frequency response is preserved by both high and low doses of enoximone with little change in the overall slope (gain) of frequency-dependent inotropic responses. In contrast, a moderate (1 µM) dose of ATI22-107 tends to reduce the gain of frequency-dependent inotropic responses, and a high (10 µM) dose of ATI22-107 prevents increases in developed force beyond 1.5 Hz (ATI22-107, n = 7; enoximone, n = 5).

View larger version (15K): [in this window] [in a new window]   Fig. 4. Percentage change of developed tension compared with 0.5 Hz of the same drug and dose. This depiction of the developed tension data clearly delineates the divergence of ATI22-107 and enoximone in generating force with increasing frequency and dose and, thus, the role of the LTCC blocking moiety of ATI22-107. #, p < 0.05 versus control at same frequency. *, p < 0.05 versus the same dose of enoximone at the same frequency.

	Discussion

Top Abstract Materials and Methods Results Discussion References

 
The primary new findings of these studies is that the dual pharmacophore ATI22-107 provides a positive inotropic effect that is limited by the increasing contribution of the LTCC-inhibiting moiety at the higher doses. The effect of the LTCC-inhibiting moiety is seen not only in the effects of the compound on the I_Ca,L in voltage-clamp experiments, but also in whole-cell [Ca²⁺]_i transients from isolated myocytes and the dynamic physiological responses of isolated cardiac trabeculae under physiological loading conditions. At low doses, the effects of ATI22-107 on peak [Ca²⁺]_i were equal to those observed with enoximone, a pure PDE-III inhibitor, but further increases in peak [Ca²⁺]_i were not observed at higher doses. Importantly, ATI22-107 induced dose-dependent increases in the decay rate of the [Ca²⁺]_i transients, as evidenced by the decreasing T₅₀ and T₇₅ seen in the myocyte experiments, and did not induce the increases in diastolic [Ca²⁺]_i observed with enoximone. In trabeculae, increases in -dF/dt with ATI22-107 are also consistent with improved diastolic function. Overall, the combination of PDE-III and LTCC-inhibiting moieties in a single compound has produced a novel pharmacological profile that demonstrates the effectiveness of the dual pharmacophore technology and may provide a new therapeutic modality for the treatment of heart failure.

The in vitro physiological responses to ATI22-107 clearly reflect the functional activities of each of the chemical moieties incorporated in this dual pharmacophore. At low doses of ATI22-107 (300 nM and 1 µM), the increases in peak cytosolic calcium in myocytes and increases in developed force in isolated trabeculae were virtually identical to those observed with enoximone. However, at both low and high doses of the two compounds, distinctions emerged between enoximone and ATI22-107 that are consistent with the additional activity of the LTCC inhibitor moiety. Specifically, differences in diastolic [Ca²⁺]_i levels, T₅₀, and T₇₅ are apparent even at 300 nM, and studies using voltage-clamp techniques confirm the modulation of I_Ca,L at this dose. Moreover, doses of ATI22-107 above 1 µM did not produce further increases in peak [Ca²⁺]_i in isolated myocytes. Finally, differences in modulation of the force-frequency responses by enoximone and ATI22-107 are also functionally important distinctions between the two compounds. Specifically, because a positive force-frequency response depends on progressive increases in Ca²⁺ entry via the LTCC, the divergence of frequency-dependent responses between ATI22-107 and enoximone attests to the activity of the LTCC-inhibiting moiety in the dual pharmacophore under dynamic physiological conditions.

Positive inotropes have long been considered an intuitive and tempting pharmacologic approach for treatment of heart failure. Although existing inotropic agents improve hemodynamic parameters and relieve symptoms among patients with advanced heart failure, these agents have typically been associated with increased mortality rates (Thackray et al., 2002; Klein et al., 2003; Rapezzi et al., 2003; Southworth, 2003). One putative mechanism contributing to increased mortality with existing inotropes is increased diastolic [Ca²⁺]_i levels that may impair diastolic relaxation and trigger malignant arrhythmias. In a previous study, ATI22-107 has been successfully shown to improve calcium homeostasis and provide antiarrhythmic effects in a pacing-induced heart failure model (Mazhari et al., 2004). Our results show the ability of ATI22-107 to provide limited inotropy, improve diastolic relaxation, and maintain stable diastolic calcium levels. This is a novel and favorable pharmacological profile that may provide a likely mechanism for improving function in heart failure models and might provide clinical utility.

Limitations. In our myocyte studies, we did not include simultaneous length change with the Ca²⁺ profile data. Although myocyte inotropy and lusitropy as evidenced by myocyte fractional shortening usually corresponds well with the [Ca²⁺]_i profile, we cannot entirely exclude the possibility that changes in myofilament Ca²⁺ sensitivity altered the relationship between [Ca²⁺]_i and myocyte shortening and contributed to increases in relaxation rates with ATI22-107. However, the general concordance between our myocyte [Ca²⁺]_i data and the force measurements in isolated trabeculae support the assertion that increases in peak [Ca²⁺]_i and rates of decline result in improved contractility and relaxation rates, respectively. We did not provide data that examine LTCC inhibitors alone or their comparison with our compound. However, the effects of LTCC inhibitors on normal myocytes and trabeculae are well established by previously published reports (Kass, 1982; Leonard and Talbert, 1982; Lindemann et al., 1982; Kanaya et al., 1983; Katz, 1986).

Conclusions. To our knowledge, this is the first demonstration of complementary effects of two distinct moieties in a dual pharmacophore designed for direct modulation of cardiac function. As such, our findings provide a proof of principle for the application of dual pharmacophore technology to produce specifically targeted therapeutics that exhibit the features of two or more well characterized compounds. At the same time, the ability of ATI22-107 to enhance inotropy and lusitropy without increasing diastolic [Ca²⁺]_i levels potentially allows enhancement of function without increasing the risks of arrhythmogenesis and/or ischemia. Due to the growing heart failure epidemic, including the increasing prevalence of patients with advanced symptoms despite application of optimized therapy with vasodilators and -blockers, the development of a safe and effective inotropic agent would provide a welcome addition to the heart failure therapeutic armamentarium. Further studies exploring the effects of this and other novel inotropes in failing hearts will be necessary to extend and complement the present investigations.

	Acknowledgements

 
We thank Dr. George Bratinov, who assisted with animal preparation and data analysis, and Drs. David Harris and Xioquen Chen, who assisted with cell isolation instruction.

	Footnotes

 
This research was supported by Grants HL33921 and HL61495 (to S.R.H.) and AG17022 (to K.B.M.) from the National Institutes of Health.

This work was previously published at the Biophysical Society 48th Annual Meeting; 2004 Feb 14–18; Baltimore, MD.

Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.

doi:10.1124/jpet.104.075895.

ABBREVIATIONS: ACE, angiotensin converting enzyme; NEP, neutral endopeptidase; PDE, phosphodiesterase; ATI22-107, 2-(2-{2-[2-chloro-4-(6-oxo-1,4,5,6-tetrahydro-pyridazin-3-yl)-phenoxy]-acetylamino}-ethoxymethyl)-4-(2-chloro-phenyl)-6-methyl-1,4-dihydro-pyridine-3,5-dicarboxylic acid dimethyl ester; LTCC, L-type calcium channel.

Address correspondence to: Dr. Kenneth B. Margulies, Cardiovascular Research Center, Temple University School of Medicine, 3420 N. Broad Street, Room 805 MRB, Philadelphia, PA 19140. E-mail: margul{at}temple.edu

	References

Top Abstract Materials and Methods Results Discussion References

 

Auvin S, Auguet M, Navet E, Harnett JJ, Viossat I, Schulz J, Bigg D, and Chabrier P-E (2003) Novel inhibitors of neuronal nitric oxide synthase with potent antioxidant properties. Bioorg Med Chem Lett 13: 209-212.[CrossRef][Medline]
Chodjania Y, Tharaux P-L, Ragueneau I, Dussaule J-C, Picker J-L, Funck-Brentano C, and Jaillon P (2002) Renal and vascular effects of S21402, a dual inhibitor of angiotensin-converting enzyme and neutral endopeptidase, in healthy subjects with hypovolemia. Clin Pharmacol Ther 71: 468-478.[CrossRef][Medline]
duBell WH and Houser SR (1989) Voltage and beat dependence of Ca2+ transient in feline ventricular myocytes. Am J Physiol 257: H746-H759.
Farina NKJC and Burrell LM (2000) Reversal of cardiac hypertrophy and fibrosis by S21402, a dual inhibitor of neutral endopeptidase and angiotensin converting enzyme in SHRs. J Hypertens 18: 749-755.[CrossRef][Medline]
Hamilton G and Leighton H (2003), inventors, Artesian Therapeutics Inc., assignee. Dihyrodpyridine compounds having simultaneous ability to block L-type calcium channels and to inhibit phosphodiesterase type 3 activity. WO2004033444. 2004 Apr 22.
Houser SR and Margulies KB (2003) Is depressed myocyte contractility centrally involved in heart failure? Circ Res 92: 350-358.[Abstract/Free Full Text]
Kanaya S, Arlock P, Katzung B, and Hondeghem L (1983) Diltiazem and verapamil preferentially block inactivated cardiac calcium channels. J Mol Cell Cardiol 14: 145-148.
Kass RS (1982) Nisoldipine: a new, more selective calcium current blocker in cardiac Purkinje fibers. J Pharmacol Exp Ther 223: 446-456.[Abstract/Free Full Text]
Katz AM (1986) Mechanisms of action and differences in calcium channel blockers. Am J Cardiol 58: 20D-22D.[CrossRef][Medline]
Klein L, O'Connor CM, Gattis WA, Zampino M, de Luca L, Vitarelli A, Fedele F, and Gheorghiade M (2003) Pharmacologic therapy for patients with chronic heart failure and reduced systolic function: review of trials and practical considerations. Am J Cardiol 91: 18-40.
Lee H, Roeske W, and Yamamura H (1984) High affinity specific [3H](+)PN 200-110 binding to dihydropyridine receptors associated with calcium channels in rat cerebral cortex and heart. Life Sci 35: 721-732.[CrossRef][Medline]
Leonard R and Talbert R (1982) Calcium-channel blocking agents. Clin Pharm 1: 17-33.[Medline]
Lindemann JP, Bailey JC, and Watanabe AM (1982) Potential biochemical mechanisms for regulation of the slow inward current: theoretical basis for drug action. Am Heart J 103: 746-756.[CrossRef][Medline]
Mazhari R, Derakhchan K, Hamilton G, Gillis MA, Bednarik D, Suzdak P, and Nattel S (2004) Improved calcium homeostasis and antiarrhythmic effects of a novel chimeric molecule that inhibits both type III phosphodiesterase and L-type calcium channel in failing hearts. J Am Coll Cardiol 43 (Suppl A): 189A.
Naccarelli GV and Goldstein RA (1989) Electrophysiology of phosphodiesterase inhibitors. Am J Cardiol 63: 35A-40A.[CrossRef][Medline]
Norton GR, Woodiwiss AJ, Hartford C, Trifunovic B, Middlemost S, Lee A, and Allen MJ (1999) Sustained antihypertensive actions of a dual angiotensin-converting enzyme neutral endopeptidase inhibitor, sampatrilat, in black hypertensive subjects. Am J Hypertension 12: 563-571.[CrossRef][Medline]
Piacentino V, III, Gaughan JP, and Houser SR (2002) L-type Ca2+ currents overlapping threshold Na+ currents: could they be responsible for the "slip-mode" phenomenon in cardiac myocytes? Circ Res 90: 435-442.[Abstract/Free Full Text]
Rapezzi C, Perugini E, Santi M, Bracchetti G, and Branzi A (2003) Inotropic therapy is unsuccessful: wrong conceptual target or wrong therapeutic tools? Ital Heart J 4 (Suppl 2): 22S-26S.
Rossman EI, Petre RE, Chaudhary KW, Piacentino I, Valentino, Janssen PML, Gaughan JP, Houser SR, and Margulies KB (2004) Abnormal frequency-dependent responses represent the pathophysiologic signature of contractile failure in human myocardium. J Mol Cell Cardiol 36: 33-42.[CrossRef][Medline]
Sipido KR, Volders PGA, de Groot SHM, Verdonck F, Van de Werf F, Wellens HJJ, and Vos MA (2000) Enhanced Ca2+ release and Na/Ca exchange activity in hypertrophied canine ventricular myocytes: potential link between contractile adaptation and arrhythmogenesis. Circulation 102: 2137-2144.[Abstract/Free Full Text]
Southworth MR (2003) Treatment options for acute decompensated heart failure. Am J Health Syst Pharm 60 (Suppl 4): S7-S15.
Teerlink JR, Jalaluddin M, Anderson S, Kukin ML, Eichhorn EJ, Francis G, Packer M, and Massie BM (2000) Ambulatory ventricular arrhythmias in patients with heart failure do not specifically predict an increased risk of sudden death. Circulation 101: 40-46.[Abstract/Free Full Text]
Thackray S, Easthaugh J, Freemantle N, and Cleland JG (2002) The effectiveness and relative effectiveness of intravenous inotropic drugs acting through the adrenergic pathway in patients with heart failure-a meta-regression analysis. Eur J Heart Fail 4: 515-529.[Abstract/Free Full Text]
Toda N, Tago K, Marumoto S, Takami K, Ori M, Yamada N, Koyama K, Naruto S, Abe K, and Yamazaki R (2003) Design, synthesis and structure-activity relationships of dual inhibitors of acetylcholinesterase and serotonin transporter as potential agents for Alzheimer's disease. Bioorg Med Chem 11: 1935-1955.[Medline]
Uddin MJ, Rao PNP, and Knaus EE (2003) Design and synthesis of novel celecoxib analogues as selective cyclooxygenase-2 (COX-2) inhibitors: replacement of the sulfonamide pharmacophore by a sulfonylazide bioisostere. Bioorg Med Chem 11: 5273-5280.[Medline]
Weishaar RE, Burrows SD, Kobylarz DC, Quade MM, and Evans DB (1986) Multiple molecular forms of cyclic nucleotide phosphodiesterase in cardiac and smooth muscle and in platelets. Isolation, characterization and effects of various reference phosphodiesterase inhibitors and cardiotonic agents. Biochem Pharmacol 35: 787-800.[CrossRef][Medline]
Yamamoto M, Ikeda S, Kondo H, and Inoue S (2002) Design and synthesis of dual inhibitors for matrix metalloproteinase and cathepsin. Bioorg Med Chem Lett 12: 375-378.[CrossRef][Medline]

This article has been cited by other articles:

				A. S. Jung, H. Kubo, R. Wilson, S. R. Houser, and K. B. Margulies Modulation of contractility by myocyte-derived arginase in normal and hypertrophied feline myocardium Am J Physiol Heart Circ Physiol, May 1, 2006; 290(5): H1756 - H1762. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) All Versions of this Article: jpet.104.075895v1 312/2/517    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Jung, A. S. Articles by Margulies, K. B. Search for Related Content PubMed PubMed Citation Articles by Jung, A. S. Articles by Margulies, K. B.