In Vitro Integration of Permeability with the Dissolution Profile of Deferasirox

Dispersible Tablets and its Potential for Establishing In Vitro—In Vivo Correlation

Amal Al-Durdunji, Hatim Al-khatib and Mutasim Al-Ghazawi*

Faculty of Pharmacy-The University of Jordan

INTRODUCTION (IVIVC) Over years, the in vitro dissolution (release) test has emerged as the single most important and useful quality control procedure to assure product quality and performance. The value of an in vitro dissolution test to assess drug quality is increased when differences in the dissolution rate can be correlated with known differences in in vivo product performance

Dissolution test Biowaiver: permission to use dissolution

test as a surrogate of pharmacokinetic data:

• For accepting product sameness under

SUPAC related changes. • To waive bioequivalence requirements

for lower strengths of a dosage form. • To support waivers for other

bioequivalence requirements.

IVIVC Definition

An In-vitro in-vivo correlation (IVIVC) has been defined by the Food and Drug Administration (FDA) as

“a predictive mathematical model describing the relationship between an in-vitro property of a dosage form and an in-vivo response”.

IVIVC In-vitro In-vivo correlation can be used in the development of new pharmaceuticals to reduce the number of human studies during the formulation development; the main objective of an IVIVC is to serve as a surrogate for in-vivo bioavailability and to support biowaivers. It supports and/or validates the use of dissolution methods and specification setting.

IVIVC Levels Level A correlation is a point to point relationship between in vitro dissolution and the in vivo absorption rate of a drug from the dosage form. Level B compares the mean in vitro dissolution time (MDTvitro) to the mean in vivo dissolution time (MDTvivo). Level C is a single point comparison of the amount of drug dissolved at one dissolution time point to one pharmacokinetic parameter. Multiple Level C is a correlation involving one or several pharmacokinetic parameters to the amount of drug dissolved at various time points.

IVIVC can be established when drug absorption is controlled by drug dissolution, so extended release dosage forms are good candidates for IVIVC. IVIVCs can be also developed for immediate release dosage forms provided that dissolution is the rate limiting step for drug absorption, in such case, biopharmaceutics classification system (BCS) type II compounds which have high intestinal permeability and low solubility are most suitable (Amidon et al., 1995).

Introduction (Biorelevant Dissolution)

Bio-relevant conditions are needed to establish a relationship between in vitro dissolution profiles and in vivo pharmacokinetics.

Drug absorption from oral solid dosage forms

requires disintegration, dissolution, transit in the gastro-intestinal tract and permeation through intestinal membranes (Takano et al., 2012)*.

* Takano, R. Kataoka, M. and Yamashita, S. (2012), Integrating Drug Permeability with Dissolution Profile to Develop IVIVC. Biopharmaceutics and Drug Disposition, 33, 354-365.

Introduction

So evaluation of not only solubility properties but also permeability is needed to establish bio-relevant dissolution conditions.

Permeability can be simulated by the use of

water immiscible organic solvent, this simulates drug partitioning in the gastro-intestinal membranes.

Introduction Biphasic dissolution system involves the use of

two immiscible liquid layers, one is an aqueous phase and the other is an organic one. A drug is released into the aqueous phase then partitions to the organic layer depending on its distribution coefficient. Further aqueous dissolution can occur and the cycle of dissolution-partitioning can continue preventing the drug from accumulating in the aqueous phase.

Biphasic Dissolution Testing

Introduction

The biphasic dissolution system is expected to be suitable for BCS class II compounds, because it overcomes the non-sink problems associated with the dissolution of such compounds.

Research Objectives • Deferasirox dispersible tablets contain 500 mg

of drug. • Deferasirox is a BCS class II drug (FDA,

2005)* with a reported aqueous solubility of 0.04 mg/mL (Yalkowsky et al., 1992)**.

• The absolute bioavailability of Deferasirox from Exjade is 73% (FDA, 2005)*.

*Food and Drug Administration (2005), Clinical Pharmacology and Biopharmaceutics Review NDA 21-882. Retrieved October 27, 2012, from http://www.accessdata.fda.gov/drugsatfda_docs/nda/2005/021882_s000_Exjade_BioPharmr.pdf

**Yalkowsky, S. and Dannenfelser, R. (1992), DFX. Retrieved October 27, 2012, from http://www.drugbank.ca/drugs/DB01609.

Research Objectives

(1) To examine the discriminatory power of a dissolution system utilizing two-phase dissolution medium using different Deferasirox formulations.

(2) To compare the discriminatory power of a two-phase dissolution with a mono- phasic dissolution system.

(3) To establish in vitro-in vivo correlation for Deferasirox tablets.

Tested Batches Component Name DFSHST/017A DFSHST/028 126560

(mg/tab) (mg/tab) (mg/tab) Deferasirox 500 (d0.9 41.5 μm) 500 (d0.9 ˂10 μm) 500 (d0.9 ˂10 μm) Lactose mesh 200 290.4 186.9 186.9 Crospovidone XL 10 254.8 164.9 164.9 Povidone K30 52 33.65 33.65 Sodium Lauryl Sulfate 8.4 11 11 Water QS QS QS Crospovidone XL 10 85.2 51.7 180 Avicel 102 - 68.05 603.55 Prosolve 50 SMCC 244.4 - - Lactose Fast Flow 243.2 - - Lactose DCL11 - 70.9 - Aerosil 3.6 3.2 5 Magnesium Stearate 18 9.7 15 Total 1700 1100 1700

Component Name Exjade Deferasirox Lactose mesh 200 Crospovidone XL 10 Povidone K30 Sodium Lauryl Sulfate Water Crospovidone XL 10 Avicel 102 Prosolve 50 SMCC Lactose Fast Flow Lactose DCL11 Aerosil Magnesium Stearate Total

Biphasic Dissolution Testing Variable Chosen

Parameter Comments

Organic phase Octanol Deferasirox has the highest solubility in this solvent

Volume of organic phase

500 mL Sink conditions

Aqueous phase Sodium phosphate

buffer pH 6.8

Physiologically-relevant, Deferasirox has the highest solubility in this buffer

Volume of aqueous phase

300 mL The lower the volume the better the partitioning into the organic phase. It is the lowest volume that enables practical sampling

Biphasic Dissolution Testing Variable Chosen

Parameter Comments

Apparatus 2 RPM

120 At 150 rpm, mixing of the two phases occurred. 100 rpm didn’t discriminate between Deferasirox formulations

Flow rate in apparatus 4

30 mL/min Since Deferasirox is poorly soluble in aqueous solutions, the choice of the flow rate was limited to high speeds

Apparatus 4 cell 22.6 mm in diameter

The cell is suitable for large tablets

Glass beads in apparatus 4 cell

Present The presence or absence of glass beads gave similar results

Biphasic Dissolution Testing

Biphasic Dissolution Testing

Single Phase Dissolution Testing (FDA dissolution database 06/21/2006)

• USP apparatus II. • 50 rpm • Phosphate buffer pH 6.8+0.5% tween 20 • 900 mL

Dissolution Results Release in the Organic Phase

Dissolution Results Release in the Aqueous Phase

Dissolution Results Single Phase Dissolution Medium

Human in vivo data were generated in three single-dose, fasted-state, bioequivalence studies of crossover design for the three test formulations against the originator Exjade®. Data was generously provided by Hikma Pharmaceuticals (Amman, Jordan).

Bioequivalence Studies

Bioequivalence Studies The studies were all sponsored by Hikma Pharmaceuticals and performed in the International Pharmaceutical Research Center (IPRC, Amman, Jordan) and Pharma Medica (Toronto, Canada) in compliance with the Declaration of Helsinki after obtaining the respective Institutional Review Board’s approval of the protocol in each case.

Bioequivalence Studies

The pharmacokinetic parameters were calculated using non-compartmental analysis of average plasma concentration-time profiles using WinNonlin (version 5.3; Pharsight Corporation, Mountain View, CA), AUC was calculated by trapezoidal rule, maximum plasma concentration Cmax and time to peak concentration (Tmax) were directly obtained from the average plasma concentration-time profiles.

Pharmacokinetic parameters

Bioequivalence Studies

Formula AUClast

(μg.hr/mL) Cmax (μg/mL) Tmax (hr)

DFSHST/017A 87.70 5.68 4.00

DFSHST/028 153.53 10.68 4.50

126560 128.58 10.93 3.67

Exjade® 128.39 10.29 3.33

Study 90% confidence interval for the ratio

(point estimate)

Cmax AUC0-t AUCinf

DSFHST/017A against

Exjade® (n=8)

53.16%-70.18%

(61.08%)

61.81%-76.69%

(68.85%)

60.78%-90.91%

(76.84%)

DFSHST/028 against

Exjade® (n=6)

61.74%-131.78% (90.20%)

69.77%-159.38%

(105.45%)

75.38%-150.89%

(106.65%)

126560 against Exjade®

(n=40)

93.56%-106.57% (99.86%)

95.11%-104.40% (99.65%)

95.93%-104.96%

(100.34%)

Study f2 DSFHST/017A against

Exjade® 42.7

DFSHST/028 against Exjade® 73

126560 against Exjade® 80

Similarity Factor (f2) Release in the Organic Phase

In vitro dissolved fractions in the organic phase of a biphasic dissolution medium fitted to double Weibull model.

In Vitro model

Exploring IVIVC: Deconvolution

0

2

4

6

8

10

12

0 20 40 60 80

Con

cent

ratio

n(m

cg/m

L)

Time (Hours)

126560

Fractions absorbed over time of DFX formulations

In Vivo model

Exploring IVIVC: Correlation

Exploring IVIVC: Correlation

Exploring IVIVC: Predictability

Formulation Parameter Predicted Observed Prediction

error %

DFSHST/017A

Cmax 5.3 5.7 -6.9

AUCinf 89.4 87.7 2.0

DFSHST/028 Cmax 9.6 10.7 -10.4

AUCinf 154.5 153.5 0.6

126560 Cmax 10.5 10.9 -4.0

AUCinf 138.5 128.6 7.7

Exjade Cmax 9.5 10.3 -7.3

AUCinf 134.1 128.4 4.5

Level A correlation, which is the most preferred level by the regulatory authorities, was successfully achieved employing the two-phase dissolution. The internal validation results were conclusive, the prediction error values for Cmax and AUCinf for each formulation used in building the model were below 15%. The average absolute prediction error was below 10% for both pharmacokinetic parameters, which is the acceptance criterion by the FDA for an IVIVC model.

Exploring IVIVC: Predictability

Predictability after Employing Single Phase Dissolution

Formulation Parameter Predicted Observed Prediction

error %

DFSHST/017A

Cmax 5.8 5.7 2.6

AUCinf 78.8 87.7 -10.1

DFSHST/028 Cmax 15.6 10.7 46.0

AUCinf 158.2 153.5 3.1

126560 Cmax 15.5 10.9 41.6

AUCinf 123.2 128.6 -4.2

Exjade Cmax 14.7 10.3 43.0

AUCinf 124.8 128.4 -2.8

Predictability after Employing Single Phase Dissolution

Conclusion A biphasic dissolution method employing flow-

through apparatus and USP paddles apparatus for discriminating the performance among different formulations of Deferasirox dispersible tablets was developed.

It simulates two in vivo processes that contribute to drug absorption; dissolution and permeation, it also provides sink condition which is critical for the dissolution of drugs of low solubility and high permeability.

A valid IVIVC was successfully established.

Acknowledgement Hikma Pharmaceuticals for providing the results for the bioequivalence studies. Deanship of Academic Research at the University of Jordan

Pharsight (PAL for WinNonlin and IVIVC toolkit)

Development of Biorelevant Dissolution Testing Conditions for Fat-based Progesterone Pessaries and their Potential Application for in vitro-in vivo

correlation (IVIVC)

By Jozef Jawad Adel Al-Gousous

Supervisor Dr. Mutasim Al-Ghazawi

Co-Supervisor Dr. Hatim Alkhatib

Objective

• The aim of this work was to develop a reliable dissolution testing procedure for fat-based progesterone pessaries to provide a reliable quality control test for them, and to evaluate the potential of this method to develop an in vitro-in vivo correlation (IVIVC).

Intro (cont’d) • Overwhelming majority of biorelevant dissolution tests

reported in literature are confined to oral dosage forms.

• In this work we attempted to develop one for a vaginal dosage form.

• The vaginal wall is, indeed, suitable for the systemic absorption of drugs due to its rich blood supply and avoidance of first-pass effect.

• Lipophilic suppositories (pessaries) can be used as in the case of progesterone.

• Relaese by melting, sedimentation and dissolution or melting and partitioning.

Introduction (cont’d) • Main progestational hormone. • For secondary amenorrhea,

prevention of endometrial hyperplasia in HRT and part of ART.

• Very poorly water soluble (3.4 mg/L) and high dose (400 mg).

• No ionisable groups. • Challenging to develop sink

conditions. • Difficult to recommend a

dissolution method of choice for lipophilic suppositories. O

O

Experimental part • Three registered progesterone pessary products (A, B and C),

each manufactured by a different manufacturer were used. • Reference (A) pessaries were obtained from the market. • B and C pessaries were obtained from the Pharmaceutical

Research Unit (PRU, Amman, Jordan). • In addition to progesterone, product A contained vegetable

fat the exact identity of which was not disclosed by the manufacturer, while products B and C contained Suppocire BS2.

Experimental part (HPLC) • HPLC analysis using a a C18 analytical column • Shimadzu with PDA detector and autosampler. • Also validated on Knauer chromatograph equipped with a UV

detector 2500 and a 20 µL injection loop. • Spironolactone IS • Water:methanol: acetonitrile 30:35:35 at 1 ml/min. • 20 microliter injection volume. • 240 nm detection wv. • Validation of accuracy precision and selectivity. • 5-point calibration curve with each conc. in triplicate

Results of HPLC method development

Progesterone conc. (µg/ml)

Progesterone conc. determined (µg/ml) %RSD %Accuracy

3 3.09 1.58 3.00

15 14.66 0.29 2.27

30 30.15 0.75 0.50

48 47.73 0.091 0.56

60 60.21 0.15 0.35

Progesterone conc. (µg/ml)

Progesterone conc. determined (µg/ml) %RSD

% Accuracy

4 3.93 0.57 1.75

10 9.84 1.99 1.6

30 30.51 1.15 1.7

50 49.96 0.73 0.08

60 59.79 1.43 0.35

Calibration curve for progesterone

y = 0.0564x + 0.0099R2 = 0.9999

00.5

11.5

22.5

33.5

4

0 20 40 60 80

Progesterone conc. (µg/ml)

AU

C r

atio

Results of HPLC method development

Experimental part ( solubility)

• By incubating excess progesterone with solutions having different conc.’s of either HPBCD or SLS.

• 37oC • LabTech shaking incubator. • Filter with 0.45 micron Nylon filters and

analyze using HPLC.

Solubility results SLS concentration (%w/v) Progesterone solubility (mg/ml) Volume saturated by 400 mg of

progesterone (ml)

0.50 0.726 551

1.00 1.55 258

1.50 2.72 147

2.00 3.57 112

2.50 4.55 88

3.00 5.34 75

HPBCD concentration (%w/w) Progesterone solubility (mg/ml) Volume saturated by 400 mg of progesterone (ml)

2.00 0.956 418

3.85 2.066 194

7.41 5.064 79

16.67 10.706 37

28.57 20.955 19

Experimental part (Dissolution) • Triplicate, in USP type II apparatus dissolution vessel containing 900 ml of 10%

w/w HPBCD at 370C, and the paddle was rotated at a speed of 100 rpm. • The samples were appropriately filtered through 0.45 micron cellulose acetate

filters, then, to a portion of each sample • The sampling times were 15, 20, 25, 30, 35, 40, 60 and 90 minutes. • Performing the test at 50 and 75 rpm was also attempted. • USP type I (basket) method was also attempted using the same medium, 100 rpm

and a Vankel VK700 dissolution testing apparatus at 370C with an autosampler. In addition, a release test using the Vankel VK700.

• Also a dialysis cell was constructed from a dissolution basket enclosed by a cellulose acetate dialysis membrane with a molecular weight cut-off of 14 kDa. This dialysis cell was immersed into the dissolution medium.

Dissolution results

• Very fine dispersion that cannot be filtered with SLS.

• Acceptable results with HPBCD using type II USP apparatus at 100 rpm.

• Too high %RSD (>20% at >30% release) with 10% HPBCD at 50 and 75 rpm.

• Too slow using type I apparatus and even slower when using dialysis membranes.

Dissolution results (cont’d)

time (min) Product A B C

%release SD %RSD %release SD %RSD %release SD %RSD

15 19.5 3.8 19.5 9.2 1.5 16.3 9.1 6.8 74.7

20 29.4 4 13.6 18.2 4.8 26.4 28.0 7.4 26.4

25 37.6 4.6 12.2 33.5 4.8 14.3 40.8 6.9 16.9

30 43.9 2.7 6.2 49.7 5 10.1 52.8 8.2 15.5

35 52.6 2.4 4.6 62.4 4.7 7.5 62.6 8.8 14.1

40 60.0 1.7 2.8 71.7 4.8 6.7 72.3 8.0 11.1

60 79.7 2.8 3.5 84.5 4.6 5.4 86.6 2.9 3.3

90 87.9 2.8 3.2 91.0 0.9 1.0 91.7 0.8 0.9

Dissolution results (cont’d)

Dissolution results (cont’d)

Reference Test f1 f2

A B 14.9 54

A C 13.9 54.7

B C 5.7 66.7

C B 5.3 66.7

In vivo PK studies

• These were performed at the PRU as two bioequivalence studies, one comparing product B (test) to product A (reference), and the other comparing product C (test) to product A (reference). The data were analyzed using WinNonlin 5.2 software.

In vivo PK studies results

In vivo PK studies results A I A II B C

Tmax (h) Mean 7.27 6.56 6.71 5.88

SD 7.94 5.58 4.47 2.86

Cmax

(µg/ml)*

Mean 12.70 19.40 13.87 19.53

SD 1.60 1.49 1.72 1.40

AUClast

(µg.h/ml)*

Mean 213.1 264.6 206.1 252.5

SD 2.06 1.56 2.03 1.57

AUC0∞

(µg.h/ml)*

Mean 209.9 296.2 211.7 286.7

SD 1.84 1.62 2.01 1.53

MRT (h)# Mean 18.97 16.40 17.15 15.85

SD 59.7 41.8 63.3 36.6

A I A II B C

ka (h-1) 0.0915±0.0676 0.0731±0.0412 0.109±0.0790 0.0832±0.0506

ke (h-1) 0.495±0.345 0.672±0.975 0.651±0.996 0.751±0.647

In vivo PK studies results

In vivo PK studies results

Conclusion • Despite their apparent simplicity, progesterone vaginal

suppositories are a challenging dosage form in terms of developing a suitable in vitro release testing procedure due to the variability in their behavior. HPBCD can be used to provide sink conditions for the release of progesterone from the pessaries as it is capable of sufficiently solubilizing the drug without dispersing the base and so providing gentle conditions unlike the use of extremely high concentrations of surfactants. Our method showed good potential of being biorelevant, but in vitro and in vivo evaluation of another product with significantly different release rate is required to validate the IVIVC and check its discriminating power.

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