MEMORANDUM

**DIVISION OF CARDIO‑RENAL DRUG PRODUCTS INTERNAL CONSULTATION**

To: Douglas Throckmorton, M.D., Division Director

#  HFD 110, Division of Cardio‑Renal Drug Products

From: John Koerner, Ph.D., Pharmacologist

HFD 110, Division of Cardio‑Renal Drug Products

# Through: Charles Resnick, Ph.D., Supervisory Pharmacologist

HFD 110, Division of Cardio‑Renal Drug Products

CC: Elizabeth Hausner, D.V.M., Pharmacologist

HFD 110, Division of Cardio‑Renal Drug Products

#  Albert DeFelice, Ph.D., Supervisory Pharmacologist

HFD 110, Division of Cardio‑Renal Drug Products

[NDA]{.underline}: 21526

[Sponsor]{.underline}: CV Therapeutics, Inc.

[Drug]{.underline}: RANEXA, Ranolazine

[Proposed Clinical Indication]{.underline}: Angina

[Date Completed]{.underline}: 09/03/03

[Re]{.underline}: Nonclinical Electrophysiological and Proarrhythmic
Effects of Ranolazine

# **Executive Summary**

Ranolazine's nonclinical electrophysiological effects are generally
consistent with QT interval prolongation observed in clinical trials.
However, the sponsor provided information arguing against proarrhythmic
properties of ranolazine.

According to theory, torsade de pointes with QT prolonging drugs arises
from increased transmural dispersion of repolarization and triggered
activity (early afterdepolarizations) rather than from QT interval
prolongation, per se. The elegant models of Antzelevitch argue that
findings in isolated canine ventricular strips and wedge preparations,
in particular action potential lengthening and early
afterdepolarizations in M-cells, transmural dispersion of
repolarization, and induction of torsade-like arrhythmias in vitro can
predict proarrhythmic potential of a drug. Theory is supported in part
utilizing data from positive control drugs such as amiodarone,
cisapride, d-sotalol, erythromycin, quinidine and terfenadine.

While findings with ranolazine are primarily negative in the isolated
canine ventricular wedge preparation, ranolazine increases M-cell action
potential duration and transmural dispersion of repolarization under
conditions of hypokalemia, consistent with its ability to inhibit
repolarizing currents, I~Kr~ and I~Ks~, at concentrations similar to
those required for its proposed mechanism of action. Moreover,
proarrhythmic potential was not adequately evaluated since ranolazine
was not tested under conditions necessary to capture cisapride-induced
arrhythmias in this model. Additionally, in vivo evaluation for
proarrhythmia was not comprehensive.

Additional regulatory concerns reflect the complex nature of any
proarrhythmia model, lack of evaluation of metabolites in this in vitro
model, and adequacy of testing for proarrhythmia, utilizing known risk
factors for clinical arrhythmias, e.g. female gender, different pacing
modalities (pause, acceleration), adrenergic influences, and heart
failure.

For these reasons, study results provided by the sponsor cannot preclude
the risk of proarrhythmia with ranolazine.

# **Review and Evaluation**

## [Ionic Currents]{.underline}

Ranolazine inhibited both repolarizing currents (I~Kr~ and I~Ks~),
depolarizing currents (I~Na-Late~, and I~Ca-L~), and

I~NaCa~ in isolated canine ventricular myocytes.

Rank order for potency on native currents was I~Kr~ = I~Ks~ \>
I~Na-Late~ \> I~NaCa~ \> I~Ca-L~.

+------------------------------------------+---------------------------+
| Ionic Current                            | IC50 (μM)                 |
|                                          |                           |
| (Canine Ventricular Myocytes)            |                           |
+==========================================+===========================+
| I~Kr~                                    | 11.4                      |
+------------------------------------------+---------------------------+
| I~Ks~                                    | 13.4                      |
+------------------------------------------+---------------------------+
| I~K1~                                    | \> 100                    |
+------------------------------------------+---------------------------+
| I~Ca-L~                                  | 296                       |
+------------------------------------------+---------------------------+
| I~Na-Late~                               | 21                        |
+------------------------------------------+---------------------------+
| I~NaCa~                                  | 91                        |
+------------------------------------------+---------------------------+

Ranolazine was also shown to weakly inhibit cloned human potassium
channels (hERG and KvLQT1), with IC50s of 86 μM and 1.46 mM,
respectively. While this data demonstrates the ability of ranolazine to
block human repolarizing currents, it is not useful for potency
estimation since studies were performed using a *Xenopus* oocyte
expression system, which greatly underestimates potency. For example,
IC50s for concurrent positive controls dofetilide and verapamil were
approximately 20 -30 fold greater in oocytes than in mammalian
expression systems (1,2). Additionally, KvLQT1 channels were not
coexpressed with minK, which likely further underestimates ranolazine
potency on this repolarizing channel (3,4).

+--------------------------------+------------------+------------------+
| Ionic Current                  | IC50 (μM)        |                  |
|                                |                  |                  |
| (cloned human channel          |                  |                  |
| expressed in Xenopus oocytes)  |                  |                  |
+--------------------------------+------------------+------------------+
|                                | hERG             | KvLQT1           |
+--------------------------------+------------------+------------------+
| Ranolazine                     | 83               | 1460             |
+--------------------------------+------------------+------------------+
| Dofetilide\*                   | 0.3              | \-               |
+--------------------------------+------------------+------------------+
| Verapamil\*                    | 5.27             | \-               |
+--------------------------------+------------------+------------------+

> \* IC50s for inhibition of hERG expressed in mammalian expression
> system:
>
> dofetilide, 0.015 μM; verapamil 0.143 μM (literature reference values
> (1,2).

## The sponsor evaluated effects of several metabolites at a single concentration of 50 μM on I~Kr~ in canine ventricular myocytes. Several metabolites inhibited IKr by about 50% at this concentration. The sponsor did not evaluate effects of metabolites on other ionic currents, including I~Ks~. 

## [Action Potential Duration]{.underline}

Ranolazine was evaluated for effects on action potential duration in
several systems, including epicardium and M-cells from canine left
ventricular strips. Concentrations evaluated ranged from 1 to 100 μM. In
canine epicardial strips, ranolazine (≥ 5 μM) lengthened APD90 in a
concentration dependent way. Effects were potassium and rate dependent,
with lengthening slightly more pronounced with hypokalemia

(2 vs 4 mM potassium) and a more rapid stimulation rate (500 ms vs 2000
ms basic cycle lengths; i.e.,

120 vs 30 beats per minute). Ranolazine did not lengthen APD90 in M-cell
strips.

Ranolazine was also evaluated for effects on action potential duration
in isolated, buffer perfused canine ventricular wedge preparations.
Ranolazine was evaluated in epicardium and M-cells at concentrations of
1- 100 μM, under conditions of normokalemia and hypokalemia (4 and 2 mM
potassium) using basic cycle lengths of 500 and 2000 ms (stimulation
frequencies of 120 and 30 beats per min, respectively).

In the presence of normokalemia, ranolazine did not lengthen APD90 in
epicardium and M-cells. However, in the presence of hypokalemia,
ranolazine lengthened APD90 in a concentration dependent way in both of
these tissues.

![](media/image1.png){width="6.115277777777778in"
height="5.889583333333333in"}

Ranolazine also prolonged transmural QT interval, Tpeak to Tend, and
transmural dispersion of repolarization, and altered T-wave morphology
(wide, low, notched; see below). Ranolazine (10 and

100 μM) also lowered maximum upstroke velocity, indicating sodium
channel blockade at concentrations evaluated. Maximum upstroke velocity
was lowered by approximately 50% at 100 μM. Exposure duration and time
course of effects were not provided.

![](media/image2.png){width="7.415277777777778in"
height="4.409027777777778in"}

![](media/image3.png){width="5.635416666666667in"
height="6.500694444444444in"}

In comparison, most but not all positive control drugs showed pronounced
M-cell APD90 lengthening and increased transmural dispersion of
repolarization in canine ventricular strips or wedges (5-12).

  ----------------------------- -------------------- --------------------
  Drug                          Pronounced Effects   

                                M-cell               Transmural
                                                     Dispersion of
                                                     Repolarization

  Amiodarone (chronic)          no                   no

  Azimilide\*                   yes                  yes

  Cisapride\*                   yes                  yes

  Erythromycin                  yes                  yes

  Quinidine                     yes                  yes

  d-Sotalol                     yes                  yes

  Terfenadine\^                 yes                  yes
  ----------------------------- -------------------- --------------------

> \* Biphasic concentration -- response relationship, with prolongation
> at low concentrations, and shortening or attenuation of effect at high
> concentrations.

\^ Requires extended perfusion period (120 min) and hypokalemia (2 mM
potassium) for effects.

In canine Purkinje fibers, ranolazine (1 to 100 μM) shortened APD90 and
depressed maximal upstroke velocity. APD90 effects appeared independent
of extracellular potassium (2 vs 4 mM) and basic cycle length (500 vs
2000 ms). Similar to findings in M-cells, ranolazine (10 and 100 μM)
decreased maximum upstroke velocity, with approximately 50% inhibition
at 100 μM. Ranolazine (5 and 10 μM) shortened

d-sotalol induced APD lengthening and suppressed EADs in a concentration
related way. Lack of action potential duration lengthening in canine
Purkinje fibers with ranolazine is not impressive considering the poor
sensitivity of this model to torsadogenic drugs such as terfenadine and
terodiline (13, 14).

## 

## [Proarrhythmia]{.underline} 

Ranolazine (1- 100 μM) did not induce early afterdepolarizations (EADs)
in canine ventricular M-cell strips and Purkinje fibers. Ranolazine (5
and 10 μM) reversed d,l-sotalol induced APD90 lengthening and prevented
EADs in both canine M-cell strips and Purkinje fibers. Lower ranolazine
concentrations were not evaluated for this property.

In comparison, the historical positive control erythromycin induced EADs
in 2 of 10 M-cell strips at the highest concentration evaluated (8).
Quinidine (3.3 μM) induced EADs in 3 of 5 M-cell strips and EAD-induced
triggered activity in 2 of 5 M-cell strips with hypokalemia (2 mM
potassium) but not with normokalemia (4 mM potassium) (9). M-cell strips
from chronically amiodarone treated dogs did not show EADs or triggered
activity (5). Additionally, chronic amiodarone attenuated d-sotalol
induced APD90 lengthening and EADs.

The sponsor evaluated ranolazine for torsade-like polymorphic
arrhythmias (TdP, spontaneous and programmed electrical stimulation
(PES) inducible) in isolated canine ventricular wedge preparations.
Ranolazine was evaluated in the presence and absence of hypokalemia (2
mM potassium). PES consisted of a single extrastimulus (S2) applied to
the epicardial surface of the wedge preparation with the basic stimuli
(S1) applied to the endocardium at basic cycle lengths of 500 and 2000
ms.

Ranolazine did not induce torsade like arrhythmias in these studies.
However, the sample size was quite small (n = 4 for normokalemia, and n
= 3 for hypokalemia), and positive historical control data from the
Antzelevitch laboratory shows TdP incidences for d-sotalol, erythromycin
and cisapride to be fairly low in this in vitro model (7, 8, 10, 11).
Additionally, cisapride's proarrhythmic effects were limited to a single
concentration, with negative proarrhythmia findings at both lower and
higher concentrations, consistent with cisapride's biphasic effect on
APD90 and transmural dispersion of repolarization. Most importantly,
induction of TdP with cisapride required epicardial pacing (S1), which
doubled baseline transmural dispersion of repolarization. As only
endocardial pacing (S1) was utilized in the ranolazine experiments,
cisapride's positive torsadogenic effect in this model is not applicable
to the ranolazine experiments. Indeed, the inability of cisapride to
induce TdP in this model using endocardial pacing (S1), argues against
the model's sensitivity to torsadogenic drugs when evaluated in this
manner.

+------------------------------------+----------------+----------------+
| Drug                               | TdP Incidence  |                |
+------------------------------------+----------------+----------------+
|                                    | Spontaneous    | PES-Inducible  |
+------------------------------------+----------------+----------------+
| Ranolazine (1-100 μM)              | 0 of 4         | 0 of 3         |
+------------------------------------+----------------+----------------+
| d-Sotalol (100 μM)                 | 2 of 6         | 3 of 6         |
|                                    |                |                |
|                                    | 2 of 8         | 4 of 6         |
+------------------------------------+----------------+----------------+
| Erythromycin (100 μM)              | no data        | 3 of 4         |
+------------------------------------+----------------+----------------+
| Cisapride (0.1-5 μM)               | none           | 2 of 6\*       |
+------------------------------------+----------------+----------------+

\* TdP inducible at 0.2 μM cisapride (only); required epicardial pacing
(S1)

## [In Vivo Effects: QT Interval Prolongation and Proarrhythmia]{.underline}

Acute intravenous infusion of ranolazine was evaluated for effects on QT
interval and proarrhythmia in anesthetized dogs with acute AV block.
Ranolazine was administered at a dose of 0.5 mg/kg plus 1, 3 and 15
mg/kg/hr iv for 30 minutes/dose level (n=6). In one additional dog,
ranolazine was administered at a dose of 1.5 mg/kg plus 15 and 30
mg/kg/min iv for 30 minutes per dose level. Sotalol (8 mg/kg followed by
4 mg/kg/hr iv for 20 minutes served as a positive control (n=5). Left
ventricular effective refractory periods, QT interval, and QRS duration
were determined at basic cycle lengths of 300, 400, 600 and

1000 ms following each dose of ranolazine or sotalol. All dogs then
received bolus intravenous doses of phenylephrine (10, 20, 30, 40 and 50
μg/kg iv) in order to elicit arrhythmias.

In this study, ranolazine increased QT interval by approximately 10%
(not statistically significant). The QT effect was biphasic, with the
maximum increase observed at 3 mg/kg/hr. Ranolazine increased left
ventricular refractoriness but only at the fastest stimulation rate (300
ms basic cycle length). Ranolazine increased QRS interval by 10%
(statistically significant) at the next higher dose of 15 mg/kg/hr.
Proarrhythmia was not observed with ranolazine in the presence of
phenylephrine. The highest ranolazine dose of 30 mg/kg/hr induced acute
heart failure and death.

In contrast to findings with ranolazine, sotalol induced a large
increase in QT interval (33% increase at 1000 ms basic cycle length),
and increased left ventricular refractoriness without altering QRS
duration. Phenylephrine induced tachyarrhythmias, including TdP and
ventricular fibrillation, in sotalol treated dogs.

Limitations of this study design include its acute nature, since chronic
but not acute AV block has been shown to promote proarrhythmia by
electrical remodeling and downregulation of ion channels, including
I~Kr~ (15, 16). Additionally, ranolazine displaces radiolabeled ligand
from α-1 adrenergic receptors, suggestive of potential functional
antagonism (17). If ranolazine is indeed a functional α-1 adrenergic
antagonist at doses administered above, then the in vivo canine model
used for this study is inappropriate. Finally, positive control data is
quite limited for this model, lessening the regulatory importance of
negative findings.

# **References**

1.  Snyders DJ and Chaudhary A, High-affinity open-channel block by
    dofetilide of HERG expressed in a human cell line, *Mol Pharmacol*
    **49**: 949-955 (1996) \[PMID: 8649354\].

2.  Zhang S, Zhou ZF, Gong QM, Makielski C, and January CT, Mechanism of
    block and identification of the verapamil binding domain to HERG
    potassium channels, *Circ Res* **84**: 989-998 (1999) \[PMID:
    10325236\].

3.  Yang T, Hideaki K, Roden D, Phosphorylation of the IKs channel
    complex inhibits drug block: Novel mechanisms underlying variable
    antiarrhythmic drug actions, *Circulation* **108**: 132-134 (2003)
    \[PMID 12835205\].

4.  Busch A, Busch GL, Ford E, Suessbrich H, Lange HJ, etal, The role of
    IsK protein in the specific pharmacological properties of the IKs
    channel complex, *Br J Pharmacol* **122**: 187-189 (1997) \[PMID
    9313924\].

5.  Sicouri S, Moro S, Litovsky S, Elizari MV, Antzelevitch C, Chronic
    amiodarone reduces transmural dispersion of repolarization in the
    canine heart, *J Cardiovasc Electrophysiol* **8**:1269-79 (1997)
    \[PMID: 9395170\].

6.  Burashnikov A, Antzelevitch C, A combination of IKr, IKs, and ICa or
    INa block produced a relatively homogeneous prolongation of
    repolarization of cells spanning the canine left ventricular wall,
    *PACE* **20**: II-1216, (1997).

7.  DiDiego J, Belardinelli L, Antzelevitch C, Cisapride-induced
    transmural dispersion of repolarization and torsade de pointes in
    the canine left ventricular wedge preparation during epicardial
    stimulation, *Circulation*, **108**:1027-33 (2003) \[PMID:
    12912819\].

8.  Antzelevitch C, Sun Z, Zhang Z, Yan G, Cellular and ionic mechanisms
    underlying erythromycin-induced long QT intervals and torsade de
    pointes, *J Amer Coll Cardiol* **28**: 1836-1848 (1996) \[PMID
    8962574\].

9.  Sicouri S, Antzelevitch C, Drug-induced afterdepolarizations and
    triggered activity occur in a discrete subpopulation of ventricular
    muscle cells (M cells) in the canine heart: quinidine and digitalis,
    *J Cardiovasc Electrophysiol* **4**: 48-58, (1993) \[PMID:
    8287236\].

10. Shimizu W, Antzelevitch C, Effects of a K(+) channel opener to
    reduce transmural dispersion of repolarization and prevent torsade
    de pointes in LQT1, LQT2, and LQT3 models of the long-QT syndrome,
    *Circulation*, **102**:706-12 (2000) \[PMID: 10931813\].

11. Shimizu W, Antzelevitch C, Differential effects of beta-adrenergic
    agonists and antagonists in LQT1, LQT2 and LQT3 models of the long
    QT syndrome, *J Am Coll Cardiol*. **35**:778-86, (2000) PMID:
    10716483

12. Antzelevitch C, (personal communication).

13. Gintant GA, Limberis JT, McDermott JS, Wegner CD, Cox BF, The canine
    Purkinje fiber: an in vitro model system for acquired long QT
    syndrome and drug-induced arrhythmogenesis, *J Cardiovasc
    Pharmacol*. **37**:607-18 (2001) \[PMID: 11336111\].

14. Pressler ML, Warner MR, Rubart M, Rardon DP, Zipes DP, In vivo and
    in vitro electrophysiologic effects of terodiline on dog myocardium,
    *J Cardiovasc Electrophysiol* **6**: 443-54 (1995) \[PMID:
    7551314\].

15. Vos MA, de Groot SH, Verduyn SC, van der Zande J, Leunissen HD,
    Cleutjens JP, van Bilsen M, Daemen MJ, Schreuder JJ, Allessie MA,
    Wellens HJ, Enhanced susceptibility for acquired torsade de pointes
    arrhythmias in the dog with chronic, complete AV block is related to
    cardiac hypertrophy and electrical remodeling, *Circulation*:
    **98**:1125-35 (1998) \[PMID: 9736600\].

16. Volders PG, Sipido KR, Vos MA, Spatjens RL, Leunissen JD, Carmeliet
    E, Wellens HJ, Downregulation of delayed rectifier K(+) currents in
    dogs with chronic complete atrioventricular block and acquired
    torsades de pointes, *Circulation* **100**:2455-61 (1999) \[PMID:
    10595960\]

17. Hausner EA, Pharmacology Toxicology Review, NDA 21526.