12th USP Science & Standards Symposium - New Delhi

1. Track I, Session II: Chemical
Medicines and ExcipientsConsideration of Novel
Formulations
Wednesday, April 17, 2013 (11:30 a.m. to 1:30 p.m.)
IPC–USP Science & Standards Symposium
Partnering Globally for 21st Century Medicines

2. Moderator: Albinus D’Sa, Ph.D.
US Food and Drug Administration-India

3. Solid Oral Formulations Advances
Dr. Sukhjeet
Panacea Biotec

4. Solid Oral Formulation Advances
 During
the
past
biopharmaceutics,
three
decades,
pharmacokinetics
due
and
to
the
evolving
discipline
pharmacodynamics,
of
significant
advances have been made in the area of drug delivery.
 Advances in drug delivery Technologies
 Controlled drug delivery
 Oros
 matrix or reservoir system
 Site specific deliver systems
 Gastroretentive
 Colon Targeted
 Bioavailability enhancement
 Nanocrystals
 solid dispersion
 hot melt extrusion
4

5. BIOAVAILABILITY ENHANCEMENT OF POORLY
SOLUBLE DRUGS
5

6. Solid Oral Formulation Advances
Bioavailability Enhancement of Poorly Soluble Drugs

Oral bioavailability of drugs depends on its solubility and/or
dissolution rate.

40% of new chemical entities currently being discovered
are poorly water soluble. Therefore major problems
associated with these drugs was its very low solubility in
biological fluids, which results into poor bioavailability after
oral administration.

Many of these potential drugs are abandoned in the early
stages of development due to the solubility problems.

It is therefore important to realize the solubility problems of
these drugs and methods for overcoming the solubility
limitations are identified so that potential therapeutic
benefits of these active molecules can be realized.
6

7. Solid Oral Formulation Advances
Bioavailability Enhancement of Poorly Soluble Drugs
 Nanocrystals
 Solid Dispersion
7

8. NANOCRYSTALS

9. Nanocrystals
 They are nanoparticles with a crystalline character with a size in the
nanometer range
 Nanocrystals are composed of 100% drug; there is no carrier material
 Increased dissolution velocity
 Increased saturation solubility
 Increased cellular uptake – endocytosis , phagocytosis, Peyer’s patches??
9

10. Nanocrystals
Source : Nanotechnology in Ireland: A Snapshot
http://www.sciencecouncil.ie/media/icsti040714_nanotechnology_snapshot.pdf
10

11. Nanocrystals Advantages
 Enhanced oral bioavailability
 Improved dose proportionality
 Increased drug loading
 Reduced food effects
 Suitable for administration by all routes
 Possibility of sterile filtration due to decreased particle size range
 Simple composition - Does not require novel excipients, less regulatory issues
11

12. Case Study – Effect of particle size
Comparative PK Profile of Immunosuppressant in animals
Concentration
(ng/ml)
20
Pharmacokinetic Profile Comparison
15
Coarse API~10microns
Micronized API~1.5microns
Drug in Solution
Nanoparticulate API~less than 1microns
10
5
0
0
4
8
12
16
20 (hr)
Time 24
28
32
36
40
44
48
Animal PK study Results – Statistical Analysis
PK Parameter
AUC(0-t)
(hr*ng/ml)
Coarse
API~10microns
Micronized
API~1.5microns
Nanoparticulate
API~less than
1microns
Drug in Solution
194.66
494.08
1391.20
976.86
12

13. Nanocrystals- Marketed Preparations
Tradename
Drug
Indication
Company
Status
Rapamune®
Rapamycin
Immunesuppressive
Wyeth
marketed
Emend®
Aprepitant
Anti emetic
Merck
marketed
Tricor®
Fenofibrate
Hypercholesterolemia
Abbott
marketed
Megace ES®
Megestrol
Anti anorexic
Par Pharmaceutical
Companies
marketed
Triglide®
Fenofibrate
Hypercholesterolemia
Invega
Sustenna®
Paliperidone palimtate
Treatment of
schizophrenia
Semapimod®
Guanylhydrazone
TNF-α inhibitor
Cytokine
Pharmasciences
Phase II
Paxceed®
Paclitaxel
Anti inflammatory
Angiotech
Phase III
Theralux®
Thymectacin
Anti cancer
Celmed
Phase II
Nucryst®
Silver
Anti bacterial
Nucryst
Pharmaceuticals
Phase II
First Horizon
Pharmaceuticals
Johnson and Johnson
marketed
marketed
13

14. Nanocrystals Preparation Methods
 Precipitation (Bottom up)
 Milling (Top down)
 Homogenization (Top down)
 Top down and Bottom up
14

15. Nanocrystals Preparation Methods
 Milling (Top down):
 In this method, pearl, bead or ball mills can be utilized to prepare a nanocrystal
formulation.
 The drug substance and the stabilizer are dispersed in the dispersion medium, and
this mixture is then put into a grinder chamber. Balls are rotated at a very high
speed and particle size of the drug gets smaller until nanocrystals are obtained.
 Physicochemical characteristics of the nanocrystals depend on the number of
milling balls, the amount of drug and stabilizer, milling time and speed, type of
grinding chamber and temperature.
15

16. Nanocrystals Preparation Methods
 Homogenization (Top down)
 Ultasonification: Ultrasonic probes are used
to decrease the particle size in liquid or solid
dispersed phase.
 High Pressure Homogenization;
 Microfluidizers homogenizers: (Insoluble Drug Delivery – Particles, IDDP™ technology) is utilized to achieve production of submicron particles of
poorly soluble drugs.
 Piston gap Homogenizers: is performed in water (DissoCubes®), water
mixtures or nonaqueous media (Nanopure®)
 Top down and Bottom up :
Both Bottom up and Top down methods are used together eg. NanoEdge technology in
which precipitation is followed by high pressure homogenization.
16

17. Nanocrystals Preparation Methods
Advantages and disadvantages
Technology
Precipitation
Milling
Homogenization
Advantages
Disadvantages
•- finely dispersed drug
•- good control of desired size
•- needs to be stabilized
•- organic solvent residue
•- not universally applicable, only drugs
with certain properties are possible (e.g,
soluble in at least one solvent)
•- low energy technique
•- proven by 4 FDA approved drugs
•- residue from milling media
•- can be a slow process (several days)
•- needs to be stabilized
•- large batches difficult to produce due to
size of milling chamber
•- universally applicable
•- high energy technique
•- no problem with large batches
•- great experience needed
•- fast method (several minutes possibly)
17

18. Nanocrystals- Characterization
Characterization technique for drug Nanocrystals
Analytical purpose
Structural analysis
Analytical techniques
Conclusion from the results
Optical microscopy
Size distribution, flocculation tendency,
detection of large particles,
SEM, TEM, AFM
surface morphology of bulk and single
BET
particles,
Porosity, surface area
Solid state analysis
DSC, PXRD, Raman spectroscopy,
Amorphous content, polymorphism
IR spectroscopy,
Hot stage microscopy
Particle size analysis
Laser Diffraction , Dynamic light
Size and size distribution
scattering, coulter counter
Surface charge characteristics
laser Doppler anemometry (Zeta
Agglomeration tendency
potential), Capillary Zone
stability prediction
electrophoresis
Rheological assessment
Rheometer (Cone and plate,
Viscosity
rotational cylinder
18

19. Challenges in evaluation of nanocrystal
formulations
Dissolution Studies
1. What is a suitable dissolution method for drug nanocrystals?
2. How to separate undissolved nanocrystals from dissolution sample?
It is very important to separate dissolved particles before analysis, filtration using filters with pore sizes
of 0.1micron result in predictive dissolution profiles.
In situ analytical techniques, which avoid the need to separate dissolved API, are also promising
approach to assess nanocrystal dissolution
19

20. AMORPHOUS SOLID
DISPERSIONS

21. Solid Dispersion
 Group of solid products consisting of at least two different
components, generally a hydrophilic inert carrier or matrix and a
hydrophobic drug.
 The carrier can be either crystalline or amorphous in nature.
 The drug can be dispersed molecularly, in amorphous particles
(clusters) or in crystalline particles
21

22. Solid Dispersion
Conventional Dispersion
Pharmacokinetic profile of tacrolimus in animal model.
22

23. Solid Dispersion Classification
 Simple Eutectic Mixtures
 Solid Solutions
 Glass Solutions and Glass Suspensions
 Amorphous Precipitations in a Crystalline Carrier
 Compound or Complex Formation;
 Combinations of the previous five types.
23

24. Solid Dispersion Classification
 Simple Eutectic Mixtures : These are prepared by rapid solidification of the
fused melt of two components that show complete liquid miscibility but
negligible solid–solid solubility. In a simple eutectic mixture, the drug is
precipitated out in a crystalline form.
 Solid Solutions : The two components crystallize together in a homogeneous
one-phase system.
 Glass Solutions and Glass Suspensions: A glass solution is a
homogeneous glassy (amorphous) system in which a solute dissolves in the
glassy carrier. A glass suspension refers to a mixture in which precipitated
particles are suspended in a glassy solvent.
24

25. Solid Dispersion Classification
 Amorphous Precipitations in a Crystalline Carrier: the API is at the
molecular level dispersed in a polymer matrix
 Compound or Complex Formation;
 Combinations of the previous five types.
25

26. Solid Dispersion Advantages
 Reduced particle size
 Improved wettability
 Higher porosity
 Drugs in amorphous state
26

27. Solid Dispersion Preparation Method
 Fusion method
 Hot melt extrusion
 Solvent method
 Supercritical fluid method
27

28. Solid Dispersion Preparation Method
 Fusion method: The drug was melted in a carrier and after cooling the dry mass
obtained was pulverized and sieved to obtain powder
 Hot melt extrusion: The drug/carrier mix is typically processed with a twin-screw
extruder. The drug/carrier mix is simultaneously melted, homogenized and then
extruded
 Solvent method: The physical mixture of the drug and carrier is dissolved in a
common solvent, which is evaporated and resulted in formation of solid dispersion.
 Supercritical fluid method: It is mostly applied with CO2 , which is used as either
a solvent for drug and matrix or as an anti solvent. In this method the drug and matrix
are dissolved in CO2 and sprayed through a nozzle into an expansion vessel with lower
pressure resulting in immediate formation of particles.
28

29. Solid Dispersion Preparation Method
Advantages and Disadvantages
Method
Advantages
Disadvantages
Hot melt
extrusion
Solvent Method
Supercritical
fluid drying





Short time process
Solvent free
Solvent free
Good controlled temperature system
Large scale production available

Not suitable for thermally labile drugs


Not suitable for thermally labile drugs
Carriers without proper thermoplastic
properties can not be used

Short time process Micro to nanoparticulates obtained
Robust process
Large scale production available



Fusion
Possible solvents residue in the
product

Mild production condition

Possible solvent residue in the
product
Solubilizing power of supercritical
fluid (CO2) limited

29

30. Solid Dispersion- Characterization
 DETECTION OF MOLECULAR STRUCTURE IN AMORPHOUS SOLID
DISPERSION.
The properties of a solid dispersion are highly affected by the uniformity of
distribution of the drug in the matrix.
1. Confocal Raman Spectroscopy was used to measure the homogeneity of
the solid mixture.
2. IR or FTIR, can be used to measure the extent of interactions between drug
and matrix.
3. Temperature Modulated Differential Scanning Calorimetry can be used to
assess the degree of mixing of an incorporated drug.
30

31. QUANTIFICATION OF CRYSTALLINITY IN AMORPHOUS SOLID
DISPERSION
 Need : To quantify conversion of amorphous form to crystalline during processing/
ageing.
 Acceptance Criteria: The method should be able to quantify at least 5% change w.r.t.
API in the formulation.
 Challenge : Quantification is difficult due to
a) Dilution effect of excipients
b) Interference of crystalline excipients.
31

32. Solid Dispersion- Characterization
 QUANTIFICATION OF CRYSTALLINITY IN AMORPHOUS SOLID DISPERSION:
Following techniques are available to quantify crystallinity:
1. Powder X-ray Diffraction
2. Terahertz Pulsed Spectroscopy
3. Raman Spectroscopy
32

33. Solid Dispersion- Marketed Preparations
Product/Substance
Dispersion Polymer or
Technology used
Company
Carrier
Gris-PEG ®
Polyethylene glycol
(Griseofulvin)
Melt process, exact
Novartis
process unknown
Sproramax capsules
Hydroxypropyl
Spray layering
Janseen
(Itraconazole)
methylcellulose (HPMC)
Cesamet® (Nabilone)
Providone
Process unknown
Lilly
Kaletra (Lopinavir and
Polyvinylpyrolidone
Melt - extrusion
Abbot
ritonavir)
(PVP)/polyvinyl acetate
Ibuprofen
Various
Melt - extrusion
Soliqs
Isoptin SRE-240
Various
Melt-extrusion
Soliqs
LCP-Tacro (Tracrolimus) HPMC
Melt-granulation
Life Cycle Pharma
Intelence (Etravirine)
HPMC
Spray drying
Tibotec
Certican (Everolimus)
HPMC
Melt or spray drying
Novartis
Afeditab (Nifedipine)
Poloxomer or PVP
Melt/absorb on carrier
Elan Corp.
Pharmaceutica
Laboratories
(Verapamil)
33

34. REGULATORY CHALLENGES

35. Regulatory Challenges
 The emergence of products based on new technologies, posed an urgent
need for the regulatory agencies to develop a comprehensive list of tests and
a streamlined approval process.
 Currently, the regulatory agencies examine such drug products on a productby-product basis.
 There is generally a lack of standards in the examination of products based
on new technologies (Nanocrystals/solid dispersions) as a unique category of
therapeutic agents.
35

36. Regulatory Challenges
 A few fundamental and logical questions may help simplify the discussion
around potential regulatory complexity of new technologies products
 Physico-chemical characteristics: what are the key characteristics of
the product that are essential for its activity and safety, and are those
critical characteristics of the product reproduced within acceptable
pharmaceutical tolerances in manufacturing?
 Definition: does the product meet the criteria of an acceptable,
scientifically sound description or definition (these are still evolving) for
eg. what can be considered a nanomedicine (certain size constraints as
well as unique function)?
36

37. Regulatory Challenges
 Biodistribution: are there any particular properties of the product that one
would expect unusual biodistribution or more importantly cause persistence of
the product in particular tissues over extended periods of time, intentionally or
otherwise? If so, what are their effects?
 Clinical: what human clinical data should be collected to evaluate potential
safety risks specialy to the nanomedicine, whether acute or on a longer term
basis upon repeated administration?
37

38. Regulatory Challenges
 Thus in order to proactively address rapid advances in drug development,
appropriate processes to develop definitions, quality standards, and
requirements for development studies including clinical trial must be in place.
38

42. Advances in Topical Drug Delivery
Vinod P. Shah, Ph. D.
Consultant, USP

43. Topical Drug Products

Challenges in BE Evaluation
– Pros and cons of different methods for BE

Standards for topical drug products
– USP <3>, <724> and <1724>

Future steps
– Standardization of DPK methodology
– Explore potential use of In vitro release (IVR)
– Combination techniques for BE – e.g., DPK with
DMD; DPK with IVR; DMD with IVR

44. Topical Dosage Forms

Transdermals - For systemic effect

Topical drug delivery - For local action (in
skin)

Topical dosage forms - Generic drugs
– Generic Product: PE + BE = TE = TI
– Topical: Q1 and Q2
– Bioequivalence testing - Challenge
Consider site and mechanism of action
Sensitivity and feasibility of approach
Complexity of the formulation
Case-by-case approach
– In Vitro testing

45. Locally Acting Drug Products
• Methods for BE (identified in 21 CFR 320.24)
–
–
–
–
Pharmacokinetic study
Pharmacodynamic study
Clinical study (comparative clinical trials) and
In vitro dissolution / release
• A 2003 addition to the Federal FD & C Act at Section
505 (j)(8)(A)(ii) indicates that “For a drug that is not
intended to be absorbed into the bloodstream, the
Secretary may assess bioavailability by scientifically
valid measurements to reflect the rate and extent to
which the active ingredient or therapeutic ingredient
becomes available at the site of drug action”.

46. Methods of BE of Topical Dermatological Drug Products
Experimental Procedures
Acceptable
Promising
Clinical
DPK
Pharmacodynamic
Microdialyss
Unacceptable
Spectroscopy
Suction Blister
PK
In Vitro
Skin Biopsy
Grafted Skin
Surface Recovery

47. Dermatopharmacokinetics (DPK)
Lessons Learned from US DPK studies
The methodology must be standardized and
validated
- drug application area
- drug / stratum corneum removal area
• In Japan DPK is accepted
• DPK is suitable for superficial skin infection
• How to resurrect DPK?

48. Promising Methodology
In Vitro Methods
• Synthetic Membrane
- QC measure
- Can provide supportive data with other
promising methods
- With Q1 and Q2, can provide information on Q3,
and can be used for drug approval*
* Draft Guidance on Acyclovir – March 2012

49. BE of Topical Drugs - Case-by-Case
• PK approach: Topical patch – Lidocaine 5%
- Lidocaine concentration in plasma – it is proportional
to the concentration at site of action
• PD approach: Flucocinolone acetonide topical oil
-Vasoconstriction . If Q1 and Q2 then biowaiver
• Clinical approach: 5-Flourouracil cream 5%
- Clinical endpoint BE study using actinic keratoses
lesions (100% clearance)
• PK & Clinical approach: Diclofenac sodium gel 1%
• In Vitro approach: Acyclovir Ointment 5%
- If generic and RLD are Q1 and Q2  Q3 (IVR)
- If not Q1 and Q2  clinical end point study

50. In vitro Release (IVR) Test

Reasonable test

Batch-to-batch uniformity

QbD emphasizes development of a meaningful
drug development specification based on
clinical performance. IVR is the first step
towards this goal.

To be implemented as a required drug
product release and stability test.
Ref: AAPS Journal, 15 (1), 41-52, 2013.

51. Q1, Q2 and Q3. In vitro Release

Q1 – Same ingredients/components as RLD

Q2 – Same ingredients/components in the same
concentration as RLD

Q3 – Same ingredients/components/in the same
concentration with same arrangement of matter
(microstructure) as RLD

Acceptable comparative physicochemical characterization
and equivalent in vitro release (Q3) to RLD

Biowaiver may be granted with supportive data to
demonstrate Q1 and Q2 same and similar physicochemical
characteristics (Q3 – IVR)
Ref: AAPS Journal, 15 (1), 41-52, 2013.

52. Product Quality & Product Performance Tests
• Product Quality Test
Intended to assess attributes such as identity, strength, purity,
content uniformity, pH, minimum fill, microbial limits.
• Product Performance Test
Designed to assess product performance and in many cases
relates to drug release from the dosage form.
• Quality tests assess the integrity of the dosage form, whereas
performance tests assess drug release and other attributes that
relate to in vivo drug performance.
• Taken together, quality and performance tests assure the
identity, strength, quality, and purity of the pharmaceutical
dosage form.

53. Product Quality & Product Performance Test
Chapters in USP:

<3> Topical and transdermal drug products –
product quality tests. Official in USP

<724> Drug release (for TDS)

<1724> Semisolid drug products – Performance
test Official in USP36/NF 31, Supplement 1;
Official in August 1, 2013.

54. Drug Product Quality Tests <3>

Strength, efficacy, purity and safety characterization

Qualitative description organoleptic qualities and product
consistency

Visual test of homogenity

Identification

pH potential effects

Variation is specific gravity

Monitoring water content and alcohol content (where applicable)

Container closure system

Preservative

Antioxidants

impurity

55. In Vitro Release <1724>

Vertical Diffusion Cell System – based on passive
diffusion of the active into the receptor fluid - most
experience, widely used, well studied

Sensitive, reproducible, rugged, robust

Synthetic membrane – support membrane

Medium – aqueous or hydro-alcoholic mixture

Degas the medium, 320

Release rate dependent on formulation composition,
particle size and strength

Release rate is formulation specific (similar to MR
dosage forms)

56. Semisolid Dosage Forms: Creams, Ointments, Gels
• Method of choice:
Vertical diffusion cell with a synthetic membrane
VDC method is described in USP <1724>
(USP36/NF31, 1st Supplement,
Official August 1, 2013)
• It is a measure of product quality and sameness with
SUPAC related changes.
• With Q1, Q2 and Q3 (in vitro release) the method can be
used for biowaiver of acyclovir ointment
• FDA is advocating use of IVR as a specification for
product release – similar to dissolution

57. In Vitro Release

QC tool – batch release test !

Formulation development

Selection of formulation for clinical testing

Product performance assessment

Post approval changes (SUPAC)

Compare with RLD

IVR is a useful tool

Possible application for biowaiver of lower
strength(s)

59. Conclusions

Bioequivalence evaluation – case-by-case

DPK method needs to be standardized and
validated

IVR can be used as a product performance test

Potential application of IVR should be explored

Application of combination techniques should
be explored for generic drug approval

61. Target Delivery of Injectables
N. Subramanian, M.Pharm., Ph.D.
Agila Specialities Ltd., (Division of Strides Arcolabs Ltd.)

62. Topics Covered

Market Trend of Target delivery Systems

Classification

Design requirements

Liposomal Drug Delivery System
– Manufacturing & Characterization aspects.

Regulatory Viewpoint on Liposomal drug product

Conclusion

63. Drug Discovery Timelines
Preclinical
Phase
I/II/III
Phase IV
• 1000 Molecules
• 3-6 years
• 25 Molecules
• 6-7 years
• 1 Molecule
New Drug to Market

64. Drug Delivery System – Global market View

The global market for drug delivery systems in 2010 was $131.6 billion. The market is expected to rise
at a compound annual growth rate (CAGR) of 5% and reach nearly $175.6 billion by 2016.

The U.S. constituted approximately 59% of the total drug delivery market in 2010. It is forecast to grow
from $78 billion in 2010 to $103 billion in 2016.
*Source – BCC Research

65. Drug Delivery System – Global market View
 The largest segment of the market is targeted drug delivery, which reached $50.9 billion in 2009 and
is expected to increase to $80.2 billion in 2014, for a CAGR of 9.5%.
 Sustained-release products have the second-largest market share, with estimated sales of $36.1
billion in 2009 and $45.8 billion in 2014, for a CAGR of 4.9%.

66. Global Cancer therapy Market
The global cancer therapy market will increase from $40.0 billion in 2007 to an estimated $110.6 billion in
2013, a compound annual growth rate (CAGR) of 18.5%.
 Target therapy dominated in 2007 with a 45% share of the total cancer therapy market. This is expected to
increase to 62.5% in 2013.

67. Nanocarrier Market Share by Technology in 2021
Nanoshells
Polymeric nanocarriers
Liposomes
Micelles
Dendrimers
AU nanocarriers

68. Classification of Current Targeted Drug Delivery Processes
1. Systemic targeting based on blood circulation and extravasation
a) Ligand–receptor interaction mediated
b) Locally-activated delivery
i.
Self-triggered release of the drug at the target cells
ii.
Externally-activated release of the drug at the target cells
2. Intracellular targeting
a) Low-pH activation technologies that use default pathway delivery
to lysosomes
b) Mechanisms that avoid (default) lysosomal delivery

69. Therapeutic Monoclonal Antibodies in Cancer Therapy

70. Antibody Drug Conjugates in Cancer Therapy

71. Target Drug Delivery Systems
Brand Name
Structure
Type of Drug association
Tocosol
Drug solubilized in emulsion
droplets
Abraxane
Drug in Albumin nanoparticles
Genexol
Drug in Polymer micelles
Taxol
Drug solubilized in cremophor
micelles
Xyotax
Micelle/aggregate of
Drug/Glutamic acid derivative
Ambisome
Drug solubilized in lipid bilayer

72. Drug Targeting Concepts of I.V. Administered Systems
 EPR effect
 Nanoparticle properties and design
 Increased retention in the circulation due to PEGylation
 Ligand–receptor type interactions

73. Design Requirement of Drug Delivery Systems
Route of
Administration
Duration of
delivery
Drug
Properties
GOAL
Mechanism
of drug
release
Biocompatibility
Nature of
delivery
vehicle
Ability to
Targeting

74. Liposomes (Vesicles)
The Technology
• Self assembled, closed systems made of lipids in the form of one or
multiple concentric bilayers capable
hydrophobic/ amphiphilic drugs
of
delivering
hydrophilic/
74

75. Liposomes (Vesicles)
Key Advantages
•
Suitable for delivery of hydrophobic, hydrophilic and amphipatic
drugs and agents
•
Chemically and physically well characterized entities
•
Biocompatible
•
Suitable for controlled release
•
Suitable to give localized action in particular tissues.
•
Suitable to administer via various routes
•
Protect drug from degradation in body
•
Reduces side effects
•
Improves patient quality of life

76. Liposome Forming Material - Phospholipid
Key Features:
- Biodegradable and biocompatible
- Non-immunogenic and component of cell membrane
- Approved by regulatory authorities for pharmaceutical use
76

77. Classification Based on Size
• Small unilamellar vesicles
• Medium sized unilamellar vesicles
• Large unilamellar vesicles
• Oligolamellar vesicles
• Multilamellar large vesicles
• Multivesicular vesicles

78. Classification Based on Specific Properties
Conventional and Stealth liposomes
• Conventional liposomes are encaptured by
macrophages
• Stealth liposomes evades this process and
remain long-circulating in the blood.
78

79. Liposome Targeting
Passive Targeting

Accumulates in tumor through leaky
micro-vasculature of tumor tissue

Retained at site due to insufficient
lymphatic drainage from the tumor

Maintain higher concentration of
liposomal drug in the tumor.

Intact vasculature of heart and healthy
organs prevents drug exposure lowering
the toxicity to cardiac muscle and
healthy organs
Lower toxicity and Higher tumor accumulation (EPR effect)
79

80. Drug Release from Liposome into Tumor
The tumor microenvironment contributes to destabilizing the lipid carrier through
the action of the slightly acidic pH of interstitial fluids, the release of lipases from
dying tumor cells, and the release of enzymes and oxidizing agents by tumor
infiltrating inflammatory cells. In addition, phagocytic cells residing in tumors could
metabolize liposomes and release free drug, killing neighboring cells via the
bystander effect.
Higher concentration of the drug is
therefore maintained inside the tumor by
pegylated liposomes for prolonged period
of time.
80

81. Marketed Liposomal Products
Approved Liposome Products in US
Doxil
Doxorubicin
1995
Daunoxome
Daunorubicin
1996
Ambisome
Amphotericin B
1997
Depocyt
Cytarabine
1999
First Generic Liposomal Injection approval
Doxorubicin
Sun Pharma
2013
liposome inj
81

82. Steps Involved in Manufacture of Liposomes
•
Selection and Analysis of raw materials – Drug, Phospholipids,
cholesterol, buffer etc.
•
Optimization of composition of formulation (Drug: lipid).
•
Preparation of multilamellar vesicles by Spray drying, ethanol injection,
thin film hydration etc.
•
Preparation of unilamellar vesicles by size reduction.
•
Drug loading – Active or Passive methods.
•
Removal of free drug.
•
Sterilization process.
82

83. Analytical Challenges
General Tests
Description
Assay
Related substances
pH
Osmolality
Particulate matter
Sterility
Bacterial endotoxin
Absorbance
Transmittance
Fill range
Volume variation
Reconstitution time (lyo products)
Content Uniformity (lyo products)
83

84. Analytical Challenges
Liposome specific tests
Instrument Used
Lipid content
HPLC
Phosphorus content
ICP
Lyso lipids
HPLC
Physical form of drug inside liposome
Fluorescence spectroscopy, TEM, SAXS
In vitro drug release study
HPLC
In vitro Plasma stability
HPLC, LCMS
Internal volume
NMR
Phase transition temperature
DSC
Particle size and PDI
Particle Size Analyzer
Particle Shape and Lamellarity
TEM
% Free drug & % encapsulated drug
HPLC
Zeta potential
Zeta Sizer

85. Assay of Drug in Liposomal Products

Total Drug present in the formulation by HPLC, LCMS methods.

Entrapped drug in liposomes by separation process like centrifugation
or by column separation (example – Sephadex) and then quantification.

Free drug (Unentrapped drug) in formulation.
85

86. Lipid Analysis in Liposomal Products

Analysis of Individual lipids present in the formulation like HSPC,
Cholesterol, DSPG, mPEG-DSPE depending on the formulation.

Analysis of the degradation products of the lipids in formulation like LysoPC, which will influence the stability of the formulation.

Cholesterol will determine the rigidity of the bilayer and thereby the drug
release from liposomes

Analysis of the lipids is very critical as the change in composition will alter
the product performance
86

87. Phase Transition Temperature of Liposomal Products
 Phase transition temperature of the liposomes depends on the
type of lipids used.
 This determines the conditions for the formation of the
liposomes, drug loading, in-vitro and in-vivo drug release.
 This also determines the stability and storage condition of the
liposomal formulations.
 Examples of Phase transition temperatures of lipids:
HSPC – 52° - 55°C
DPPC – 38° - 42°C
DMPC – 18° - 23.2°C
87

88. Particle Size Analysis of Liposomal Drug Products

Particle size plays a major role in the success of the performance of the
product.
88

89. Cryo TEM image of Multilamellar Vesicles Before Size Reduction
89

90. Cryo TEM image of Multilamellar Vesicles After Size Reduction
90

91. Cryo TEM Image of Hydrophilic Drug Containing Liposomes
91

92. Cryo TEM Image of Hydrophobic Drug Containing Liposomes
92

93. Zeta Potential Analysis of Liposomal Drug Products

The magnitude of the zeta potential gives an indication of the potential stability of
the colloidal system.

If all the particles in suspension have a large negative or positive zeta potential then
they will tend to repel each other and there is no tendency to flocculate. However, if
the particles have low zeta potential values then there is no force to prevent the
particles coming together and flocculate. The general dividing line between stable
and unstable suspensions is generally taken at either +30mV or -30mV.

Generally Particles with zeta potentials more positive than +30mV or more negative
than -30mV are normally considered stable.

In case of pegylated liposomes, the zeta potential is around -10mV. The liposomes
is stable in spite of low zeta potential due to steric hindrance provided by the PEG
layer on the surface of the liposomes.
93

94. Invitro Drug Release of Liposomal Drug Products

This study is performed to understand the release pattern of drug
from the liposomes when administered in-vivo.

This is performed using dialysis membranes / HLB cartridges
having specific affinity for the free drug.

Selection of media, experimental conditions like pH, temperature,
ionic concentrations play important role in the drug release.

Free drug and entrapped drug are separated after specific time
points and Drug release curve is plotted with respect to time.
94

95. Preclinical Studies of Liposomal Drug Products
At the preclinical stage, Sponsors should perform minimum of the following studies:
Toxicokinetic and Pharmacokinetic Studies:
Single and multiple dose pharmacokinetics, toxicokinetics, and tissue distribution
studies in relevant species.
a)
Pharmacokinetic study in tumor bearing mice (in case of anticancer drug) /
normal mice (in case of other drugs) and also in rats.
b)
Organ Distribution study in mice and rats.
c)
Pharmacokinetic / Toxicokinetic study in higher animal like dog/monkey.
Efficacy Studies:
a) Tumor regression study in case of anti cancer drugs (Ex. Doxorubicin)
b) Antifungal activity in case of antifungal agents (Ex. Amphotericin B)
95

96. FDA’s Draft Guidance on Liposome Drug Products
Draft Guidance
since August 2002

97. Physicochemical Characterization Requirement
•
Description
•
•
morphology of the liposome, including
unencapsulated (i.e., free) drug
lamellarity determination, if applicable
substance
assay for encapsulated and
•
net charge
•
degradation products related to the lipids
•
volume of entrapment in liposomal
•
assay of lipid components
vesicles
•
in vitro test for release of drug substance
•
particle size (mean and distribution
profile)
•
phase transition temperature
•
spectroscopic data, as applicable
•
in vitro release of the drug substance
from the liposome drug product
•
osmotic properties
•
light scattering index
from the liposome

98. Pharmacokinetic and Bioavailability Requirement
• a single-dose pharmacokinetic study; this should be a
comparative study between the liposome and nonliposome
drug product, when appropriate
• a multiple-dose study evaluating the pharmacokinetics of the
drug substance after administration of the liposome drug
product
• a dose-proportionality study over the expected therapeutic
dose range after administration of the liposome drug product

99. Pharmacopoeial Status
• Indian Pharmacopoeia has published a
chapter on Liposomal products in its 2010
edition
• Also a monograph on Liposomal
Amphotericin B for injection is included in IP.

100. Guidances for Generic Liposomal Products

101. Pharmaceutical Equivalence Tests
Draft Guidance on Doxorubicin Hydrochloride - USFDA
•
•
•
•
•
•
•
•
•
•
•
•
•
•
Lipid content
Free and encapsulated drug
Internal and total sulfate
Ammonium concentration
Histidine concentration
Sucrose concentration
State of encapsulated drug
Form of a doxorubicin sulfate precipitate inside the liposome
Internal environment (volume, pH, sulfate and ammonium ion concentration)
Liposome morphology and number of lamellae
Lipid bilayer phase transitions
Liposome size distribution
Grafted PEG at the liposome surface
Electrical surface potential or charge

102. Bioequivalence Requirement
1. Bioequivalence Study

103. In vitro Leakage Under Multiple Conditions

104. Pharmaceutical Equivalence Tests
EMEA Reflection Paper
•
•
•
•
•
•
•
•
•
•
•
•
Lipidic components (description, source and characterisation, manufacture, specification
and stability);
Quality, purity and stability of other nonlipidic starting materials and critical excipients;
Identification and control of key intermediates in the manufacturing process;
Active substance/ lipidic moiety ratio at relevant manufacturing steps to ensure consistent
formulation;
Liposome morphology, size and size distribution,
Fraction of encapsulated active substance (amount of free/entrapped)
Assay of lipidic components;
Osmolarity;
Stability of the active substance, lipids and functional excipients in the finished product,
including quantification of critical degradation products (e.g. Lyso phosphatidylcholine,
oxidated/ hydrolytic moieties)
Stability studies under proposed in-use conditions;
In vitro drug substance release rate from the liposome in relevant media and stress
conditions;
Validated process for reconstitution and/or pharmacy preparation

105. Pharmaceutical Equivalence Tests
 Maintenance of liposomal formulation integrity in plasma;
 Characterization/ specification testing for lipid bilayer phase transition; temperature and/or
liposomal ‘surface’ charge;
 Confirmation of physical state of the active substance inside the liposome
 for pegylated liposomal formulations:
 details of linkage chemistry (PEG-lipid),
 molecular weight of pegylated lipid and size distribution,
 disposition of PEG at surface,
 stability of pegylation;
 Discriminating validated in-vitro release methods should be developed to:
 monitor the simulated release of the active substance from the liposomes when in
circulation and if possible around the targeted site of action (e.g. different pH
environments at site of action).

106. Non-Clinical and Clinical Studies
 Non clinical Studies
 Pharmacokinetic studies
o accumulation in target organs, pharmacokinetics and distribution
 Pharmacodynamic
o demonstration of similarity in pharmacodynamic response at different dose
levels using adequate models
o in-vitro tests which characterize the interaction between liposomes and
target cells or with other cells where the interaction is toxicologically relevant
and important
 Toxicological studies
 Clinical Studies
 Comparative pharmacokinetic studies

107. Conclusions
• Target Delivery system expected to grow at a faster phase in the next
decade.
• Better understanding in the manufacturing and characterization of TDS
in the last 2 decades will boost the development of the injectable target
delivery system.
• Availability of various guidance documents and monographs will provide
better clarity of the requirements for development of generic liposomal
dosage form.
• This will help generic manufacturers in bringing generic liposomal
formulation to the market faster thereby enable affordable therapy to
needy patients.

109. Performance Test for Novel Dosage
Forms
Vinod P. Shah, Ph. D.
Consultant, USP

110. Outline

Taxonomy of Dosage Forms

Novel Dosage Forms

Product Quality and
Product Performance Test

Performance Test for Novel Dosage Forms
– Transdermal Drug Delivery System
– Semisolids: creams, ointments and gels
– Liposomes

111. Dosage Form Taxonomy (USP)
Route of
Intended site
Administration of release
Dosage Form
Examples
Dosage Form Dosage Form
Quality Tests Performance
Tests*
Parenteral
Body tissues
and fluids
Injectables,
Liposomes, micro and
nano particles,
implants, stents
<1>
<1001>**
Oral
Gastro intestinal Tablets and capsules,
tract
liquids
<2>
<701>, <711>
Topical /
Skin
Transdermal
Mucosal
Mouth, eye, ear,
(Local or Systemic) rectum, vagina,
intra-uterine
Semisolids, TDS
<3>
<724>, <1724>
Films, tablets, liquids,
suspensions,
suppositories
<4>
<1004>**
Inhalation
Liquids, aerosols,
powders
<5>
<601>, <602>,
<603>, <604>,
<1601>
Nasal cavity,
lung
* CK Brown et. al., FIP/AAPS Workshop Report: Dissolution/in vitro release testing of
novel/special dosage forms. AAPS PharmSci Tech. 12(2): 782-794, 2011
** Under Development
111

112. Pharmaceutical Dosage Forms

Traditional solid oral dosage forms 
dissolution test e.g., tablets, capsules,
suspensions

Novel (non-oral) dosage forms 
In vitro release test e.g., transdermal patches,
liposomes, stents, implants

Drug-device eluting dosage forms 
Drug elution test

113. Novel Dosage Forms

Orally disintegrating tablets

Chewable tablets

Medicated chewing gum

Suspensions

Suppositories

Transdermal patches

Topical semisolids – cream, ointment, gel

Subcutaneous implants

Injectable microparticulate formulations, Microspheres

Liposomes

Drug eluting stents

114. Product Quality & Product Performance Tests

Product Quality Test
Intended to assess attributes such as identity, strength,
purity, content uniformity, pH, minimum fill, microbial limits.

Product Performance Test
Designed to assess product performance and in many cases
relates to drug release from the dosage form.

Quality tests assess the integrity of the dosage form,
whereas performance tests assess drug release and other
attributes that relate to in vivo drug performance.

Taken together, quality and performance tests assure the
identity, strength, quality, and purity of the pharmaceutical
dosage form.

115. Product Quality Tests

USP General Chapters <1> through <5> provide
– information about the product quality tests
– a framework to support new individual monographs that are
“moving forward” documents and are not intended to replace the
need for individual monographs.
– a pick list of consolidated common product quality test
requirements in a concise and a coherent fashion.

If a validated performance test procedure is available for the
specific drug product, it is identified in general chapter below
<1000>. Additional information, or information on promising
technologies that have not yet been fully validated, may be
presented in informational chapters above <1000>.

116. Drug Product Performance Test

Product quality and performance tests link with
establishment of BA or BE  Assessment of
Drug Product Performance – Bioavailability,
Bioequivalence and Dissolution <1090>.

When documentation of BA or BE is less
certain  USP Medicines Compendium general
chapter Drug Product Performance <12>
provides information on optimum drug product
performance

117. In Vitro Release: Novel Dosage Forms

Why in vitro testing?

It is a product quality test
– As a QC measure
– Drug release as a means of product sameness under
SUPAC related changes

It is a product performance test

It is a tool to biopharmaceutics characterization
of the product

118. Concept of In Vitro Testing

Dissolution tests for solid dosage forms are well established

General principles of dissolution test should be applicable to
in vitro release of novel dosage forms has been expanded to a variety of novel / special
dosage forms such as TDS (patches), gel, creams, lotions,
ointments, suppositories, injectable microparticulate system,
liposomes, drug eluting stents, implants, aerosols

Due to complexity and drug delivery of novel / special
dosage forms, different apparatus and procedures need to
be employed – on a case-by-case basis.

119. Transdermal Patches
•
Current compendial apparatus
–
- Paddle over disk – USP Apparatus 5
–
- Rotating cylinder – USP Apparatus 6
–
- Reciprocating disk – USP Apparatus 7
•
•
Ensures patch is prevented from floating during test
•
pH of the medium ideally 5-6
•

Method of Choice: Paddle over disk with watch
glass-patch-screen sandwich assembly (Apparatus 5)
Test temperature - 32oC
Unnecessary proliferation of
dissolution equipment should be avoided

121. Semisolid Dosage Forms: creams, Ointments, Gels
• Method of choice:
Vertical diffusion cell with a synthetic membrane
VDC method is described in USP <1724>
(USP36/NF31, 1st Supplement, Official August 1, 2013)
• It is a measure of product quality and sameness with SUPAC
related changes.
• With Q1, Q2 and Q3 (in vitro release) the method can be
used for biowaiver of acyclovir ointment

123. Drug Release from Microspheres
Risperdal® Consta® 25 mg long acting injection
Manufacturer:
McNEIL JANSSEN
API:
Risperidone
Route of Administration:
Intra-muscular
Indication:
Long term treatment of
Schizophrenia
Archana Rawat Ph. D. Thesis 2011. University of Connecticut

124. Accelerated Release Testing: Risperdal Consta
Temperature:
Flow rate:
Release medium:
45, 50 and 54.5C
8ml/min
Phosphate buffer saline (PBS pH 7.4)
Fraction released
1
0.8
0.6
45°C
0.4
50°C
0.2
54.5°C
0
0
1
2
3
4
5
6
7
8
Time (Days)
 Microspheres release for:
• ~ 2 days at 54.5°C
• ~ 3 days at 50°C
• ~ 7 days at 45°C
Archana Rawat Ph. D. Thesis 2011. University of Connecticut

125. Plasma Profile Deconvolution: Risperdal® Consta®
Plasma profile
Deconvoluted plasma profile
Fraction Absorbed
1
0.8
In vivo profile
0.6
0.4
0.2
0
0
10
20
30
40
50
Time (Days)
 Plasma profile deconvolution using Loo-Riegelman method
Schizophrenia Research 70 (2004) 91– 100
60
70

126. In Vitro-In Vivo Comparison: Risperdal® Consta®
In vivo profile (scaling factor: 7)
1
In vivo profile (scaling factor: 19)
In vitro profile (45°C)
0.8
Fraction Release/Absorbed
Fraction Released/Absorbed
1
0.6
0.4
0.2
0
0.8
In vitro profile (50°C)
0.6
0.4
0.2
0
0
2
4
6
8
10
0
1
2
Time (Days)
4
1
0.8
0.6
0.4
In vivo profile (scaling factor: 39)
In vitro profile (54.5°C)
0.2
Fraction Released (In vitro)
1
Fraction released/absorbed
3
Tim e (Days)
0.8
y = 0.9687x + 0.017
R2 = 0.995
0.6
0.4
0.2
0
0
0
0.5
1
Time (Days)
1.5
2
0
0.2
0.4
0.6
Fraction Absorbed (In vivo)
Archana Rawat Ph. D. Thesis 2011. University of Connecticut
0.8
1

127. Aerosol Products – MDI and DPI
• Oral inhalation aerosols
• Nasal inhalation aerosols
• Aerodynamic particle size distribution
measured by multistage cascade impactor
• Delivered dose uniformity

128. Summary
Preferred / Recommended Apparatus
Type of Dosage Form
• Solid oral dosage forms
• Oral suspensions
• Orally disintegrating tablets
• Chewable tablets
• Transdermal patches
• Topicals – Semisolids
• Suppositories
• Medicated gums
• Microparticle formulation
• Implants
Release Method
• Basket or Paddle
• Paddle
• Paddle
• Basket, Paddle,
• Paddle over disk
• Vertical diffusion cell
• Paddle, Modified basket
• Special apparatus
• Modified flow through cell
• Modified flow through cell

129. Conclusions
• An appropriate drug release test is required to characterize
the drug product and to assure batch-to-batch reproducibility
for consistent in vivo performance
• The in vitro drug release test for some ‘special’ dosage forms
such as semi-solid dosage forms and transdermal drug
delivery systems have proven to be equally valuable as the
dissolution test for solid dosage forms
• The in vitro drug release test shows promise for other
dosage forms such as chewable tablets, suspensions and
suppositories
•For other dosage forms such as chewing gums, powders,
parenterals, further method development and refinement is
needed to make the drug release test a valuable tool

130. Novel / Special Dosage Forms - Report
FIP/AAPS Joint Workshop Report: Dissolution / In vitro
Release Testing of Novel / Special Dosage Forms:
CK Brown, HD Friedel, AR Barker, LF Buhse, S Keitel, TL Cecil, J
Kraemer, JM Morris, C Reppas, MP Sticklemeyer, C Yomota, VP
Shah.
- AAPS PharmSciTech: Vol 12, Issue 2,
782-794, 2011
- Dissolution Technologies: Vol 18 (4), 51-64, 2011.
- Die Pharmazeutische Industrie:
- Indian J of Pharm Sci: 73(3), 338-353, 2011.
FIP/RPSGB Workshop in London – October 20-21, 2008
AAPS/FIP Workshop in Los Angeles – November 7-8, 2009