QUALITY CONTROL OF SNAKE ANTIVENOM

DR.M.V.KHADILKAR

Technical Director,

Premium Serums & Vaccines Pvt.Ltd.,Narayangaon,Dist.-Pune

Maharashtra

PRODUCTION PROCESS Equine(Horse/Mules/Ponies) procurement Quarantine/Complete Health screening Regular periodic Immunization with snake

venoms Periodic Titer Testing Plasma collection & storage Plasma Fractionation Batch pooling Sterile Filtration Containerization & Lyophilization Labeling /Inspection Finished Product Testing Marketing

ANTIVENOM QUALITY AND SAFETY DEPENDS ON

1.The control of source materials

2. Manufacturing procedures

BASIC RAW MATERIALS IN ANTIVENOM PRODUCTION

Snake Venoms

Hyperimmune Plasma

OBJECTIVES OF THE PURIFICATION PROCESS

High safety and efficacy

No impairment of the neutralizing activity during purification

Maintenance of high purityFree of microbial contaminants and

endotoxins WHO

Workshop,

PRECAUTIONS IN VENOM PRODUCTION

Venom is to be obtained from snakes in a controlled and documented manner.

It should be properly preserved and tested for quality

Venom being the prime raw material for the production of Antivenom ,it is a key factor in the production of effective antivenom.

PLASMA PRODUCTION

Animal health

Monitoring of immune response

Proper collection process

are the greatest contributors to good quality plasma production

COLLECTION OF PLASMA Maintenance of healthy & disease free herd of

animals Cleanliness of premises and complete asepsis

during procedure. Use of sterilized collection sets & containers

under controlled environmental conditions Properly designed clean room to separate plasma

and operations carried out by trained personnel. Storage in proper temperature controlled rooms. Plasma collection vessels must be labeled with

date, plasma volume, horse numbers and anti-coagulant use details.AVOIDS BIOBURDEN-HELPS IN CONTROLLING ENDOTOXINS

ENZYMATIC CLEAVAGE OF IMMUNOGLOBULIN MOLECULE

COMMON METHOD USED FOR PLASMA FRACTIONATION IN INDIA

Pepsin Digestion to get F(ab’)2 fragments from whole IgG.

Precipitation of unwanted proteins by either Ammonium Sulfate or Caprylic Acid

Concentration & Diafiltration of F(ab’)2

OTHER METHODS Use of whole IgG moleculePapain Digested Fab fragmentsChromatography (Ion-Exchange/Affinity)

PLASMA FRACTIONATION

Use of closed system with no exposure to environment.

Least product hold time during process Process validation-digestion, filtration,

ultrafiltration Strict supervision of critical process parameters

such as Temp,pH,duration etc Checks about cleaning & sterilization status IPQC checks -

QUALITY CONTROL TESTS

DESCRIPTION pH PROTEIN PRESERVATIVE CONTENT POTENCY ABNORMAL TOXICITY STERILITY ENDOTOXIN (SOLUBILITY & MOISTURE CONTENT)

ANTIVENOM SAFETYDepends on

1. Type of antivenom -Whole IgG/ F(ab)2 / FabEach fragment has different pharmacokineticsEfficacy reports not supported by clinical data

2. Dose3. Route & Speed of administration 4. Manufacturing practices

Hypersensitivity skin test has limited predictability value.

REASONS FOR REACTIONS

Contamination of plasma Enzymatic digestion & removal of unwanted

proteins Formulation Sterility

No data from Indian clinical trials is available, however , a lot has been reported by importing countries

Thought to vary from 3-80% with about 5-10% severe reactions

Reports are often subjective

EFFICACY

MONOVALENT / POLYVALENT –Has its own advantages/disadvantages

LEVEL OF REFINEMENT-Enzyme digestion,Ion exchange chromatography/Affinity chromatography. Effect on cost????

VENOM QUALITY GEOGRAPHICAL ANTIGENIC VARIATION SPECIFICITY/PARASPECIFICITY

CLINICAL MANAGEMENT-OUTCOME DEPENDS ON EARLY START OF TREATMENT ADEQUATE DOSES SUPPORT SYSTEMS

SNAKE ANTIVENOM I.P.

Name of Product Name of Product : : Snake Antivenom I.P.Snake Antivenom I.P. (Polyvalent, Lyophilized/liquid, Equine origin (Polyvalent, Lyophilized/liquid, Equine origin

Enzyme Refined Immunoglobulin)Enzyme Refined Immunoglobulin) Description of ProductDescription of Product :Lyophilized 20.0 ml Vial :Lyophilized 20.0 ml Vial Along with 10.0 ml W.F.I. I.P. OR Along with 10.0 ml W.F.I. I.P. OR

liquid 10 ml vialliquid 10 ml vial Potency Potency : When Reconstituted With 10.0 ml : When Reconstituted With 10.0 ml of Sterile Water For Injection I.P. of Sterile Water For Injection I.P. 1.0 ml of reconstituted mixture Neutralizes 1.0 ml of reconstituted mixture Neutralizes 0.60 mg. of Cobra Venom 0.60 mg. of Cobra Venom 0.45 mg. of Krait venom 0.45 mg. of Krait venom 0.60 mg. of Russell’s viper venom 0.60 mg. of Russell’s viper venom 0.45 mg. of Saw scaled viper venom 0.45 mg. of Saw scaled viper venom Nature and concentration Nature and concentration of preservative of preservative : Phenol /Cresol not more than < 0.25% : Phenol /Cresol not more than < 0.25% Storage conditionsStorage conditions :Store in cool dark place, avoid :Store in cool dark place, avoid exposure to sunlight for lyophilized exposure to sunlight for lyophilized

and to and to store between 2-8˚C for liquid store between 2-8˚C for liquid product.product.

HOW MUCH ANTIVENOM IS NEEDED??( TUMBARE & KHADILKAR,JOURNAL OF BOMBAY VETERINARY COLLEGE:2004,12(1&2):9-11)

Snake Average Venom Yield in ml (Liquid)

Average Venom Yield in mg (Lyophilized)

Theoretical dose of antivenom required for treatment(10 ml vial)

Indian Cobra

0.098-1.56 ml(0.435±0.01)

56.4-514.9mg(126.32±4.31)

21

Russell’s Viper

0.177 -1.356 ml

(0.412±0.02)

20 -277.5 mg(75.98±3.66)

13

Indian Common Krait

0.03-0.25 ml(0.101±0.01)

1.25-18.89 mg(8.35±0.64)

2

INDIAN SCENARIO

It is estimated that, India currently produces about 1.8-2.0 million Snake Antivenom vials per year

Almost 75-80% of total Snake Antivenom produced is supplied to different government institutions via tender procedure.

Current capacities can supply antivenom that meets about 60-70% of the requirements leading to unfortunate incidents of deaths

Many times the problem is compounded by improper distribution

This results in big demand-supply gap for this life saving drug

INDIAN SCENARIO Many Indian companies have been regularly

supplying this life saving medicine not only to its neighboring countries but also to countries in Africa and South Asia.

Due to the enforcement of various regulations after 2000, manufacturing of equine biological products was significantly reduced resulting in severe shortage of these life saving products.

Stringent regulatory requirements have also pushed up the production costs

INDIAN SCENARIO India still has very low price as compared to

many other countries It has been observed that 2010 prices for a 10 ml

vial of Indian polyvalent AVS range from about INR 300 to 500 (US$ 6.50–11.00), which is a fraction of the cost of a vial of CroFab antivenom in the USA (at over US$ 1900 per vial) or CSL antivenom in Australia (at US$ 1500 per vial)

AVS is supplied by Indian antivenom producers to government hospitals at Rs 115 per vial (US$ 2.50)

(Romulus Whitaker & Samir Whitaker in Current science, vol. 103, no. 6, 25 september 2012)

Most of the ASVS tenders were awarded at Rs. 190 -Rs. 210 per vial in 2011

MY VIEWS Dose finding studies can give guidelines for treatment

protocol based on syndromic approach. One protocol for big country like India is difficult considering

regional variation in venom composition. Use of polyvalent antivenom is more practical and

economical especially considering procurement and distribution system followed.

More legal sources of venoms should be established/promoted in different regions for its availability in production process.

The Indian antivenoms are reasonably priced. There is limit to purification considering its direct effect on

price. Collaboration between Industry & Academic institutions is

needed to improve yields & reducing costs.

THANK YOU