Resealed erythrocytes

Presentation byPresentation by SUNILBOREDDY SUNILBOREDDY

(M. Pharmacy)(M. Pharmacy)

Resealed ErythrocyteResealed ErythrocyteAs Drug CarrierAs Drug Carrier

Content Introduction Erythrocytes as drug carrier Basic concept of erythrocytesAdvantages & limitationSource & Isolation of ErythrocytesMethod of drug Loading CharacterizationDrug release, Storage Application Conclusion

INTRODUCTION

Current Research: Aimed at development of drug delivery system to get max. therapeutic effect.

Idea of drug carrier system: to get maximum therapeutic effects, Elimination of undesirable side effects and Increase patient compliance.

Magic Bullet Concept: Paul Ehrlich proposed. Decrease toxic effect and increase therapeutic effect to the targeted cell.

Resealed Erythrocytes

Properties as carrier :- Appropriate size, shape to permit passage

through the capillaries. Biocompatible & minimum toxic side effects Minimum leakage before target site is

achieved Should be able to carry broad spectrum of

drugs Appreciable stability during storage period Should have sufficient space & should carry

adequate amounts of drugs

Erythrocytes Erythro= redCytes = cell

Biconcave discs, anucleate.

Filled with hemoglobin (Hb), a protein that functions in gas transport

Contain the plasma membrane protein spectrin and other proteins that:Give erythrocytes their

flexibilityAllow them to change

shape as necessary

ErythrocytesCHEMICAL CONSTITUENTS:Water(63%)Lipid(0.5%)Glucose(0.8%)Minerals(0.7%)Non-hemoglobin protiens(0.9%)Methehemoglobin(0.5%) and Hemoglobin (33.67%)

Erythrocytes

Healthy adult male= 4.5 millions/µmlHealthy adult female= 4.8 millions/µml

Matured RBCs have no nucleus Ribosome & Mitochondria . Therefore all space is available for drug carrier.

Reticulocytes- immature erythrocytes

Advantages of erythrocyte as carrier:-Biodegradable Isolation is easy Non immunogenic

large volume of drug can be encapsulated in small volume of erythrocytesProlong systemic activity of drug

Protection from premature degration Prodrug concept (Bioreactor) Reduce Adverse Effect Peptide & Enzyme Delivery

Disadvantages :-Possibility of Leakage of cells & dose dumping

Molecule Alter Physiology Of cell

Source, fractionation & isolation of erythrocytes

Source:- mice, cattle, pig, dog, sheep, goat, monkey, chicken, rat, rabbit & human

Whole blood can be collected by venipuncture or from orbital sinus in heparinized tube

Red blood cells can be harvested & centrifuged(2500rpm/5min at 40c .

Different centrifugal force & different buffer composition(PBS,pH7.4) for different species is used

Fresh blood is used for loading of drugs.

Methods Of Drug Loading In Methods Of Drug Loading In ErythrocytesErythrocytes1. Hypo-osmotic lyses method a) Dilution method b) Dialysis method c) Pre-swell method d) Isotonic osmotic lyses

method2. Electric breakdown method3. Endocytosis method4. Membrane perturbation method5. Normal transport method6. Lipid fusion method

Dilutional Haemolysis

RBC Membrane ruptured RBC Loaded RBC

Resealed Loaded RBC

0.4% NaCl

Hypotonic

Drug

Loading buffer

Resealing buffer

Incubation at 250c

Efficiency 1-8%Enzymes delivery

Hypotonic med

Isotonic med.

Washed

Isotonic Osmotic Lysis

RBC Isotonically ruptured

RBC

Chemical – urea, polyethylene, polypropylene, and NH4Cl

Physical rupturing

Chemicalrupturing

Drug IsotonicBuffer

Loaded RBC

Resealed RBC

Incubation at 250 C

Preswell Dilutional Haemolysis

RBC0.6%w/v NaCl Swelled

RBC Drug + Loading buffer

5 min incubation at 0 0c

Loaded

RBC

Incubation at 25 0c

Resealing Buffer

Resealed

RBC

Efficiency 72%

Fig:- Preswell Method

Dialysis

RBC

Phosphate buffer

+

80 % Haematocri

t value

Placed in dialysis bag with air bubble

Dialysis bag placed in 200ml of lysis buffer with mechanical rotator 2hrs. 4c.

DrugLoading buffer

Loaded RBCDialysis bag placed in Resealing buffer with mechanical rotator 30 min 37c.

Resealed RBC

Efficiency 30-45%

Entrapment by Endocytosis

RBC

Drug Suspension

+Buffer containing

ATP, MgCl2, and CaCl2

At 250 CLoaded RBC Resealing Buffer Resealed RBC

Fig;- Entrapment By Endocytos Method

Membrane perturbation method

RBC

Amphotericin B

e.g. Chemical agents

Increased

permeability of RBCResealing Buffer

Drug

Resealed RBC

In vitro In vitro characterizationcharacterization

In vitro characterization

Mechanism of release of resealed erythrocytes

Shelf & storage stability of resealed erythrocytes

In vivo survival & immunological consequences

Some Important Aspects Of Resealed Erythrocytes

Applications of resealed erythrocytes

Erythrocytes as carrier for enzymes

Erythrocytes as carrier for drugs

Erythrocytes for drug targeting Drug targeting to reticuloendothelial system

Drug targeting to liver

-Treatment of liver tumors-Treatment of parasitic diseases-Removal of RES iron overload-Removal of toxic agents

Applications of resealed erythrocytes

Delivery of antiviral agentsOxygen deficiency therapyMicroinjection of macromolecules

Novel systemsNanoerythrosomesErythrosomes

References

Jain.S., Jain.N.K., resealed erythrocytes as drug carriers, Edited Jain N.K., Controlled And Novel Drug Delivery, New Delhi, CBS publishers, New Delhi, 2004, 256-281.

Vyas S.P., Khar R.K., Targeted And Controlled Drug Delivery: Novel Carrier Systems, New Delhi, CBS publisher, 2004, 387-413.