WO2013105105A2 - Nouvelles compositions de vaccins viraux et méthodes de préparation de tels vaccins - Google Patents

Nouvelles compositions de vaccins viraux et méthodes de préparation de tels vaccins Download PDF

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Publication number
WO2013105105A2
WO2013105105A2 PCT/IN2012/000701 IN2012000701W WO2013105105A2 WO 2013105105 A2 WO2013105105 A2 WO 2013105105A2 IN 2012000701 W IN2012000701 W IN 2012000701W WO 2013105105 A2 WO2013105105 A2 WO 2013105105A2
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WIPO (PCT)
Prior art keywords
virus
rotavirus
aluminium
gel
group
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PCT/IN2012/000701
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English (en)
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WO2013105105A3 (fr
Inventor
Rajeev Mhalasakant DHERE
Jagdish Kamalaji ZADE
Rajendra Narayan SABALE
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Serum Institute Of India Ltd.
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Application filed by Serum Institute Of India Ltd. filed Critical Serum Institute Of India Ltd.
Publication of WO2013105105A2 publication Critical patent/WO2013105105A2/fr
Publication of WO2013105105A3 publication Critical patent/WO2013105105A3/fr

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A61K39/15—Reoviridae, e.g. calf diarrhea virus
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/525—Virus
    • A61K2039/5252—Virus inactivated (killed)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
    • A61K2039/541—Mucosal route
    • A61K2039/542—Mucosal route oral/gastrointestinal
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/545—Medicinal preparations containing antigens or antibodies characterised by the dose, timing or administration schedule
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55505—Inorganic adjuvants
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2720/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsRNA viruses
    • C12N2720/00011—Details
    • C12N2720/12011—Reoviridae
    • C12N2720/12311—Rotavirus, e.g. rotavirus A
    • C12N2720/12334—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • Group A rotaviruses are firmly established as the most important etiologic agents of dehydrating gastroenteritis in infants and young children worldwide. Severe rotavirus disease is preventable by vaccination. For Asia, Africa and Latin America, it has been estimated that there are between 3-4 billion cases of diarrhoea each year and of those cases about 5-10 million result in death (Walsh, J. A. et al.: N. Engl. J. Med., 301 :967-974 (1979)).
  • Rotaviruses are nonenveloped viruses whose genome comprises 11 segments of double-stranded RNA (dsRNA), contained in the core of the mature, triple-layered particle.
  • Rotavirus serotypes are determined by neutralizing antibody responses to each of the two outer capsid proteins, VP7 (termed G serotype) and VP4 (termed P serotype).
  • Ten G serotypes and 7 P serotypes have been identified in humans. Since serotypes Gl , G2, G3, and G4 together account for >80% of global human rotavirus strains, some vaccines aim to provide serotype-specific protection against these four serotypes. However, in certain geographic settings, other G types (e.g., G12, G8, and G9) may be epidemiologically important.
  • RotaShield (Wyeth-Lederle) was an oral, lyophilized, tetravalent (Gl-4) RRV rhesus/human reassortant vaccine supplied with a liquid diluent comprising sodium bicarbonate and citric acid having antacid property. The said vaccine was withdrawn later.
  • Rotarix (GSK) is an oral, lyophilized, G1P (8) vaccine supplied with a liquid diluent comprising calcium carbonate having antacid property.
  • Rotateq (Merck) is an oral, lyophilized, G1-4&P1 vaccine formulations.
  • lyophilized vaccines have a more sophisticated handling for administration as they may require more complex, hence relatively expensive devices such as multichamber/vial vaccines, with the active ingredient in one chamber and the reconstitution liquid in another chamber. Lyophilized vaccines are also associated with higher shipment and storage cost. These options may be inadequate for some countries in the developing world where the administration device has to be financially affordable and where the availability of production and storage infrastructure may be inadequate.
  • Rotavirus are conventionally administered orally to human infants, this route brings several challenges to immunizing rotavirus compositions.
  • Rotavirus is rapidly inactivated in an acidic environment, upon exposure to acid buffer or acidic gastric juice for example (C. Weiss and H.F.
  • rotavirus compositions are formulated in a way that they are stable during storage and after administration into the host recipient.
  • Rotavirus vaccines are primarily intended to be administered to babies, as early as at the age of 4 weeks.
  • a small vaccine dose volume such as lower than 2ml or even than 1.5 ml dose volume, will be advantageous for that population. Therefore, it is desirable that rotavirus compositions are formulated in a small volume dose.
  • J.Emerson et al discusses concentrating polio virus by aluminium hydroxide mediated sedimentation during water purification procedures, wherein the virus is not inactivated.
  • J.Emerson et al "Effect of Aluminum Hydroxide Sedimentation, Sand Filtration and Chlorination on the Virus of Poliomyelitis" Dec 1942, University of Michigan. However this method was utilized for poliovirus removal during water purification.
  • Samuel et al had reported removal of human rotavirus (SA-11) from stool suspensions or tap water by aluminum hydroxide floes, wherein SA-11 was reduced by 1 log or less. Further Samuel et also discussed comparison of rotavirus and poliovirus adsorption to aluminium hydroxide. Refer amuel et al "Comparison Between Adsorption of Poliovirus and Rotavirus by Aluminum Hydroxide and Activated Sludge Floes" University of Texas Applied And Environmental Microbiology, Feb. 1978, Vol. 35, No. 2 p. 360-363.However this method was utilized for rotavirus removal during water purification.
  • US7579008 discloses use of aluminium hydroxide as both, an adjuvant and an antacid in Gl rotavirus composition. This method however does not discuss concentrating rotavirus titre.
  • Andreas et al US 2008/0293123 discloses a novel method of concentrating viruses by utilizing a mixture of glycerol, magnesium salt, sodium salt in combination with tangential flow filtration. It also discusses that high virus concentration contribute significantly to virus instability and that there is a critical need to develop formulations that stabilize relatively high concentrations of virus.
  • said virus concentrating method does not involve alum adsorption and is only applicable to live recombinant adenovirus containing wild-type p53 gene.
  • the inventors have surprisingly found a novel method of concentrating live rotaviruses based on aluminium salt gel adsorption that could provide alternative liquid formulations possessing above mentioned advantageous features.
  • the present invention provides a stable gel adsorbed, liquid rotavirus vaccine, wherein the composition has following characteristics:
  • step (f) Mixing gel obtained in step (f) with an antacid combination in a ratio from about 60:40 to about 80:20.
  • An important aspect of the instant invention is that the process of concentrating rotavirus by aluminium salt having a concentration between 0.2 and 0.8 mg/ml, preferably between 0.5 and 0.8 mg/ml results in a final virus concentrate having atleast 5 times higher virus titre, more specifically 10 times with simultaneous removal of nucleic acid and protein contaminants.
  • aluminium salt is aluminium hydroxide, aluminium phosphate or a mixture of both.
  • a second aspect of the instant invention is that washing of sedimented gel is performed by utilizing a basal media selected from the group consisting of BME, MEM, DMEM, DMEM-F-12, IMDM, McCoy's 5 A, Media 199, Ham's F- 10, Ham's F-12, MS- 162, MS- 174, and RPMI, preferably MEM.
  • a basal media selected from the group consisting of BME, MEM, DMEM, DMEM-F-12, IMDM, McCoy's 5 A, Media 199, Ham's F- 10, Ham's F-12, MS- 162, MS- 174, and RPMI, preferably MEM.
  • the immunogenic composition of the present invention contains a mixture of atleast one aluminium hydroxide gel adsorbed rotavirus antigens, basal medium , antacid, stabilizers , buffers and has a pH of about 7.2 to 7.8, in a 1 ml dose volume.
  • said liquid immunogenic composition can be stable for at least 1 month at 36° C, atleast 3 months at 25° C and atleast 12 months at 2-8° C.
  • the vaccine formulations of the present invention preferably include one or more additional components, alone or in a biologically effective combination, which provides enhanced stability characteristics; including but not limited to amino acids, particularly glycine at a concentration between 2 and 5 % w/v.
  • Figure 1 Biological titer of virus preparation pre & post-aluminium hydroxide adsorption.
  • Figure 2 Effect of aluminium hydroxide gel adsorption & virus medium washing steps on concentration of nucleic acid & protein contaminants.
  • live or inactivated rotavirus antigen for inclusion in the claimed composition refers to a monovalent rotavirus strain, i.e. containing a single rotavirus strain, or be multivalent, i.e. containing at least two or more rotavirus strains.
  • the vaccine contains atleast one rotavirus serotype selected form a group of Gl G2, G3, G4, G5, G6, G7, G8, G9, G10, G i l, G 12, G13 and G14.
  • agentacid refers to MylantaTM, calcium carbonate (CaC0 3 ), magnesium carbonate, aluminium carbonate, aluminium phosphate, mix of aluminium hydroxide and magnesium carbonate, aluminium- magnesium- hydrycarbonate, aluminium hydroxide- magnesium carbonate- sorbitol- manitol, hydroxy- aluminium- sodium- carbonate, dihydroxy- aluminium- potassium- carbonate, magaldrate, hydrotalcite, almagcit, magnesium- aluminium- silicate- hydrate.
  • the immunogenic composition of the present invention contains a) atleast one aluminium hydroxide gel adsorbed rotavirus antigen, at a minimum concentration of 5.2 logi 0 FFU/ml/Serotype in combination with a citrate bicarbonate buffer in a suitable ratio,b) basal medium,c) phosphate and has a pH of about 7.2 to7.8, in a 1 ml dose volume.
  • the rotavirus antigen of the claimed composition may be produced according to routine production techniques.
  • rotavirus antigen preparations may be derived from tissue culture methods used to propagate the virus or express recombinant rotavirus antigens.
  • Suitable cell substrates for growing the virus include for example monkey kidney cells such as VERO or cells from a clone of VERO, VERO-like cells, monkey kidney cells such as AGMK cells MDCK, other cells lines of monkey kidney origin such as BSC-1, LLC-MK2 and MA 104, suitable pig cell lines, or any other mammalian cell type suitable for the production of rotavirus for vaccine purposes.
  • Suitable cell substrates also include human cells e.g. MRC-5 cells. Suitable cell substrates are not limited to cell lines; for example primary cells are also included.
  • mixture of sodium bicarbonate and citric acid is particularly suitable as antacid.
  • the concentrated gel containing virus is mixed with citrate bicarbonate buffer in a ratio selected from 60:40, 70:30 and 80:20, preferably 80:20.
  • the volume of a dose of vaccine according to the invention will normally be 2.5 ml or lower, typically between 0.5 ml and 2.5 ml.
  • a suitable vaccine dose will normally be 1.5 ml or suitably any volume smaller than 2.5 ml such as a volume of 2 ml or less, that is suitable for oral administration to babies or infants.
  • the dose volume will be such that the technical feasibility of the formulation is possible and there is no detrimental effect on the immunogenic potential of the formulation.
  • the rotavirus vaccine composition of the instant invention can be administered by oral or injectable route.
  • the immunogenic composition of the invention may also be formulated to contain other antigens, in particular antigens from other suitable live viruses for protection against other diseases, for example poliovirus.
  • Said additional active ingredients suitable for oral administration may be given either in admixture with the rotavirus composition, or alternatively may be co-administered (i.e. in a separate dose but on the same occasion) with the rotavirus composition claimed herein.
  • the sterilized stock solution (9mg ml) of aluminium hydroxide was kept at 4°C till the temperature of solution reached at 4°C. 6.66 ml of this solution was added to the 100ml of live rotavirus culture (titer 5.825 logio CCID 50 /ml) so that the final concentration of aluminium hydroxide was 0.6mg/ml.
  • This mixture was then incubated at 4°C for 15 to 20 hrs on rocker.
  • the container then was kept in vertical position to settle the gel at bottom for 1 to 2 hrs.
  • the supernatant was then removed and 10 to 15ml of gel was collected. The sample were removed before pre and post gel adsorption from gel and supernatant respectively.
  • This gel was then washed thrice with 50ml of minimum essential medium (MEM) followed by vortexing and samples after every wash were analyzed.
  • MEM minimum essential medium
  • Virus concentration was estimated by CCID50 assay. It was observed that post-gel adsorption, viral titers increased to about 10 fold (i.e. 6.625 logio CCID 50 /ml) from the pre-gel adsorption viral titer (i.e. 5.825 logio CCID 50 /ml. Also analysis of supernatant indicated that there was no detectable level of virus concentration in supernatant.
  • Residual DNA was estimated using Qubit and QUANT iT dsDNA BR assay. The adsorption alongwith subsequent washing with virus medium removed substantial amount of residual proteins and DNA.
  • aluminium hydroxide gel adsorbed rotavirus antigen at a minimum concentration of 5.2 logi 0 FFU/ml/Serotype in combination with a citrate bicarbonate buffer (1.92 mg/dose citric acid, 5mg/dose sodium bicarbonate) in 80:20 ratio, b) 8.48 gm/dose MEM, c) 0.7 mM phosphate, d) pH of about 7.2 to 7.8, in a 1 ml dose volume.
  • Stability of the inactivated gel adsorbed vaccine was studied for 1 month.
  • the inactivated rotaviruses were detected by ELISA and the total protein concentration was estimated using Bradford assay.
  • the vaccine was found to be stable at 2 to 8°C for at least 1 month, at 25°C for atleast 1 month and at 36 C for atleast 1 week.
  • Example 6
  • the gel adsorbed vaccine it is well known fact that aluminium gel act as an adsorbent, as well as adjuvant. Thus this preparation can be used for final vaccine preparation.
  • live rotavirus vaccines will be administered by oral route, the vaccine gel will pass through the stomach and exposed to acidic environment, thus resulting in dissolving of gel and subsequent release of virus.

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  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Virology (AREA)
  • Microbiology (AREA)
  • Immunology (AREA)
  • Medicinal Chemistry (AREA)
  • Chemical & Material Sciences (AREA)
  • Mycology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Abstract

L'invention concerne un procédé nouveau et économique de concentration de rotavirus pour obtenir des compositions liquides rotavirales adsorbées sur gel, le titre viral après adsorption étant au moins 10 fois supérieur au titre viral avant adsorption avec élimination partielle de contaminants d'acides nucléiques. La présente invention concerne également des formulations vaccinales stables d'un tel rotavirus inactivé et vivant.
PCT/IN2012/000701 2012-01-13 2012-10-25 Nouvelles compositions de vaccins viraux et méthodes de préparation de tels vaccins WO2013105105A2 (fr)

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IN135MU2012 2012-01-13
IN135/MUM/2012 2012-01-13

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WO2013105105A3 WO2013105105A3 (fr) 2013-09-06

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115011566A (zh) * 2022-05-25 2022-09-06 辽宁成大生物股份有限公司 一种人用狂犬病疫苗中残留dna的去除方法

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0294071A2 (fr) * 1987-06-03 1988-12-07 Merck & Co. Inc. Vaccins contre l'hépatite B comprenant des gels préfabriqués d'hydroxyde d'aluminium et leurs procédés de fabrication
WO2006087205A1 (fr) * 2005-02-17 2006-08-24 Glaxosmithkline Biologicals S.A. Vaccin a base de rotavirus vivant attenue pour administration orale
US20100068227A1 (en) * 2006-05-12 2010-03-18 Krishna Murthy Ella Composition Useful as a Vaccine

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0294071A2 (fr) * 1987-06-03 1988-12-07 Merck & Co. Inc. Vaccins contre l'hépatite B comprenant des gels préfabriqués d'hydroxyde d'aluminium et leurs procédés de fabrication
WO2006087205A1 (fr) * 2005-02-17 2006-08-24 Glaxosmithkline Biologicals S.A. Vaccin a base de rotavirus vivant attenue pour administration orale
US20100068227A1 (en) * 2006-05-12 2010-03-18 Krishna Murthy Ella Composition Useful as a Vaccine

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
SABIN: 'Experiments of the purification and concentration of the virus poliomyelitis' JOURNAL EXP. MED. vol. 56, no. 3, 31 August 1932, pages 307 - 317 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115011566A (zh) * 2022-05-25 2022-09-06 辽宁成大生物股份有限公司 一种人用狂犬病疫苗中残留dna的去除方法
CN115011566B (zh) * 2022-05-25 2024-01-23 辽宁成大生物股份有限公司 一种人用狂犬病疫苗中残留dna的去除方法

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