GE-511

PRESENTED BY

GANDRATHI KULDEEP KUMAR

M.S.(PHARM), I SEMESTER

DEPARTMENT OF PHARMACEUTICS

NATIONAL INSTITUTE OF PHARMACEUTICAL EDUCATION AND RESEARCH

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Introduction

Antibody-Directed Enzyme-Prodrug Therapy

Advances in ADEPT

Conclusion

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Many therapies are used for the treatment of cancer

These therapies suffer from many limitations

Limitation to the on-going treatments is due to

Drug resistance

Lack of selectivity

Pathway redundancy

Many chemotherapeutic agents-narrow therapeutic index

A more efficient approach is needed

3

It was proposed by Dr. Bagshawe et al

Advantageous over conventional therapies

Involves 2 steps:

Administration and localization of antibody-enzyme complex in tumor cell

Selective conversion of prodrug by antibody-enzyme fusion protein

4Afshar, S et al., Molecular cancer therapeutics 8, 185.

TUMOR CELL

Mechanism of ADEPT

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6

Bagshawe, K.D et al., 1999. Current opinion in immunology 11, 579-583.

Better understanding of the therapy

Identification of antigen

Penetration of antibody-enzyme fusion protein

Elimination of biological molecules might be

non-linear

Enzyme specific prodrug activation

7Fang et al., 2008. Drug Metabolism and Disposition 36, 1153-1165.

Use of different

enzymes in

ADEPT

Modifications in

antibody-enzyme

complexes

Phase-I clinical

studies

Optimization of

ADEPT- mathematical

models

ADVANCEMENTS IN ADEPT

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9

Enzymes

employed

in ADEPT

Non-human

enzymes

Intracellular

human

enzymes

Engineered

human

enzymes

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Vast number of prodrugs can be designed

Bacterial enzymes are generally employed

Many studies were reported on bacterial carboxypeptidase G2

Limited by their immunogenecity

Modified, in the recent times, to reduce immunogenecity

Example: Reduction in immunogenecity of β-lactamase

11

Contd...

Non-human enzymes are more efficient compared to human enzymes

Example:

Activation of prodrug CPT-11 to the active drug SN-38 is very fast using rabbit carboxylesterasescompared to the human enzymes

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These are produced by inducing mutations in human enzymes

Less immunogenic compared to non human enzymes

Substrate specificity of wild type enzymes is altered

Example:

Double mutant (hDM) of human Poly Nucleoside Phosphorylase (hPNP)

Afshar, S et al., Molecular cancer therapeutics 8, 185.

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They are present only within the cell

No systemic activation of prodrugs will be observed

Recent strategy employed in ADEPT

Less immunogenic compared to non human enzymes and engineered human enzymes

Example: Post Proline cleaving endopeptidase

N-Protected glycine-proline

dipeptidedoxorubicin

Doxorubicin

Heinis, C et al., 2004. Biochemistry 43, 6293-6303.

S.No Enzyme Prodrug Drug

1 DT diaphorase 5-(Aziridin-1-yl)-2,4-

nitrobenzamide (CB 1954)

5-(Aziridin-1-yl)-4-

hydroxyl amino-2-

nitrobenzamide.

2 Plasmin Peptidyl-p-phenylene diamine-

mustard

Phenylenediamine-

mustard

3 Carboxypeptidase G2 Benzoic acid mustard

glutamates

Benzoic acid

mustards (various)

4 Thymidine kinase (viral) Ganciclovir* Ganciclovir

triphosphate

5 Cytosine deaminase 5-Fluorocytosine* 5-Fluorouracil

6 β –Glucosidase Amygdalin Cyanide

7 β –Lactamase Nitrogen-mustard-

cephalosporin

Nitrogen mustards

(various)

8 Alkaline phosphatase Phenol mustard phosphatase*

Doxorubicin phosphatase*

Phenol mustard

Doxorubicin

9 Cytochrome P-450 Cyclophosphamide

Ifosfamide

Phosphoamide

mustard

(+ acrolein)

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Enzymes and prodrugs proposed for cancer therapy

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Use of humanized antibodies and enzymes

Example: humanized disulfide-stabilized anti

p185HER2 Fv-β-lactamase fusion protein

Accelerated clearance of Ab-E fusion protein

Usage of clearance antibody

Hypermannosylation of recombinant antibody-enzyme fusion protein

Rodrigues et al., 1995. Cancer Res 55, 63-70.

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Produced via post translational modifications in

Pichia pastoris

Elimination is well understood

Complex eliminates via mannose receptors

Macrophages (spleen) and endothelial cells (liver) are

mainly responsible for the elimination of complex

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With A5CP conjugate and CMDA prodrug

It required additional clearance antibody

100% patients developed HAMA and 97% HACA

With recombinant MFECP1 and Bis-iodo phenol

prodrug

Only 31 patients were taken

36% patients developed HACA and none of them developed HAMA

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Treatment with MFECP1

(units/m2) and BIP prodrug

(mg/m2 X3)

No. of patients Toxicity

5000 MFECP1 + 12.42 BIP

prodrug

Plasma CPG2

0.0111units/mL (median)

3 G3 thrombocytopenia (1), G3

neutropenia (1), G3 leukopenia (1)

5000 MFECP1 + 1075 BIP

prodrug

Plasma CPG2

<0.002units/mL

1 G4 ALT/AST, G3 GGT, G4 Cr/urea,

G3 anemia, G3 leukopenia, G3

thrombocytopenia

3000 MFECP1 + 537.6 BIP

prodrug

Plasma CPG2

<0.002units/mL

2 G4 ALT/AST (1), G3 anemia (1), G3

thrombocytopenia (1), G3

leukopenia (1), G3 fatigue

1500 MFECP1 +200 BIP

prodrug

1 G3 thrombocytopenia (1)

Abbrevations: G3 :Grade 3, G4: Grade 4, ALT: Alanine aminotransferase, AST: Aspartate aminotransferase

GGT: ϒ-glutamyl transferase

Dose escalation and toxicity of BIP prodrug

Mayer, A. et al., 2006. Clinical cancer research 12, 6509-6516.

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To predict therapeutic outcome before preclinical

and clinical studies

Two models were applied:

Compartmental model: Failed to describe pharmacokinetic

properties of biological molecules

Physiology based Pharmacokinetic (PBPK) model: Applied to

analyse each compartment or organ.

Galluppi et al., 2001. Clinical pharmacology and therapeutics 69, 387.

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For an effective ADEPT

Optimal clearance of Ab-E is 1.5X10-3 ml/min

Optimal Emax of Ab-E for prodrug conversion is 600 min-1

Optimal Permeability of prodrug is 1.4 X 10-4 cm/sec

Optimal dosing interval of Ab-E and prodrug is 5 days

Galluppi et al., 2001. Clinical pharmacology and therapeutics 69, 387.

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ADEPT was proved to be a potential therapy for the

treatment of cancer. Clinical trials of ADEPT proved

this fact but the therapy is limited by toxic effects,

many of which were being addressed in the recent

years

Further research has to be encouraged in the future

as ADEPT has the potential of being the successful

therapy in the treatment of cancer in the future

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