WO2000071156A2 - Nouvelles formulations vaccinales-3 - Google Patents

Nouvelles formulations vaccinales-3 Download PDF

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Publication number
WO2000071156A2
WO2000071156A2 PCT/GB2000/001921 GB0001921W WO0071156A2 WO 2000071156 A2 WO2000071156 A2 WO 2000071156A2 GB 0001921 W GB0001921 W GB 0001921W WO 0071156 A2 WO0071156 A2 WO 0071156A2
Authority
WO
WIPO (PCT)
Prior art keywords
cells
cultured
cancer vaccine
peptides
malignant
Prior art date
Application number
PCT/GB2000/001921
Other languages
English (en)
Other versions
WO2000071156A3 (fr
Inventor
Andrew Sutton
Peter Smith
Darren Stevenson
Haj Chana
Peter Thraves
Original Assignee
Onyvax Limited
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from GBGB9911824.2A external-priority patent/GB9911824D0/en
Priority claimed from GBGB9911825.9A external-priority patent/GB9911825D0/en
Priority claimed from GBGB9911823.4A external-priority patent/GB9911823D0/en
Priority claimed from GB0008032A external-priority patent/GB0008032D0/en
Priority claimed from GB0008029A external-priority patent/GB0008029D0/en
Application filed by Onyvax Limited filed Critical Onyvax Limited
Priority to AU49357/00A priority Critical patent/AU4935700A/en
Publication of WO2000071156A2 publication Critical patent/WO2000071156A2/fr
Publication of WO2000071156A3 publication Critical patent/WO2000071156A3/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
    • A61K39/0005—Vertebrate antigens
    • A61K39/0011—Cancer antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • 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
    • A61K2039/515—Animal cells
    • A61K2039/5152—Tumor cells
    • 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
    • A61K2039/80—Vaccine for a specifically defined cancer

Definitions

  • Cell-based vaccines fall into two categories. The first, based on autologous cells, involves the removal of a biopsy from a patient, cultivating tumour cells in vitro, modifying the cells through transfection and/or other means, irradiating the cells to render them replication-incompetent and then injecting the cells back into the same patient as a vaccine. Although this approach enjoyed considerable attention over the past decade, it has been increasingly apparent that this individually-tailored therapy is inherently impractical for several reasons. The approach is time consuming (often the lead-time for producing clinical doses of vaccine exceeds the patients life expectancy), expensive and, as a 'bespoke' product, it is not possible to specify a standardised product (only the procedure, not the product, can be standardised and hence optimised and quality controlled).
  • This invention describes a methodology for extracting immunogenic processed and semi-processed peptides from tumour cell lines, combining the hallmarks of polyvalency/heterogeneity of the whole cell approach with the more defined nature of a sub-unit vaccine
  • the disclosure describes ways of exposing cell lines to various types of phenotype shift (e g anoxia, nutrient deprivation, novel culture techniques or co-culture) and then extracting immunogenic peptides in a manner that reflects the state of the cell at the moment of extraction
  • phenotype shift e g anoxia, nutrient deprivation, novel culture techniques or co-culture
  • the invention allows for complex phenotype shifts in cells to be captured in an antigenic profile in the form of a 'peptide snapshot', providing significant advantages over whole-cell approaches where it is not possible to maintain the antigenic profile during the time taken to harvest and irradiate the cells and formulate a vaccine
  • Mouse melanoma cell line K1735, immortalised human non-malignant prostate epithelial line PNT2-C2, immortalised human prostate tumour line NIH1542, and a human metastatic tumour line DU145 were cultured in 2D culture to near confluencey Analysis of small immunogenic peptides extracted by the method of Nair et al (1997 Eur J Immunol 27, p589-597) was performed using Surface Enhanced Laser Desorption lonisation (SELDI) mass spectrometry Each cell line was shown to posses a unique fingerprint of processed or semiprocessed peptides (Figure 2) Cell Lines with Differing Apoptotic Indices Show Differing Peptide Profiles Following Irradiation
  • a double extraction was performed using TRI REAGENT (Sigma #T9424).
  • the reagent was added directly to the washed cell pellets and samples were allowed to stand for 5 minutes before the addition of chloroform. Samples were vortexed and allowed to stand for a further 10 minutes at room temperature then centrifuged at 12,000xg for 15 minutes.
  • Reverse transcription was performed using the 1st strand cDNA synthesis kit for RT-PCR (AMV) from Boehringer Mannheim (#1 483 188). Reactions were incubated at 25°C for 10 minutes then at 42°C for 1 hour. The AMV enzyme was denatured by heating to 99°C for 5 minutes then the reaction was cooled to 4°C.
  • AMV 1st strand cDNA synthesis kit for RT-PCR
  • PCR primers were designed from published sequences. Verification of cDNA integrity was achieved by PCR amplification using GapdH housekeeping primers.
  • thermocycler program (1 ) The internal nested PCR set was run on program (2) below.
  • (1 ) 95°C 3 min
  • (2) 95°C 3 min
  • the PCR samples were run in pairs.
  • the prostate cells alone then the same cells grown in co-culture with NHBMSC followed by NHBMSC alone then a positive and negative control Figure 6.
  • PSA peptides and oligoepitope peptides of PSA to elicit an T cell response to cells containing PSA (Correale P et al J Immunol 1998 Sep 15,161 (6) 3186-)
  • PSA prostate tumour cells
  • bone marrow stromal cells it may be possible to enhance the underlying immunogenicity of the cell lines and allow for extraction of peptides from the co-cultured cells which will contain both known (PSA) and unknown TAA's or TSA's
  • P4E6 and ShMac-5 cells were seeded at 1 x 10 6 in T175 culture flasks The cultures were maintained for 2-3 days using keratinocyte growth medium supplemented with 5% foetal calf serum until near confluent cultures were obtained Synthetic androgen R1881 was added to the near confluent cultures at a concentration of 1 x10 8 M and the culture grown on for a further 24 hours
  • RNA pellets were washed with 75% ethanol, dried and re-suspended in TE buffer.
  • RNA sample was treated with Deoxyribonuclease I (Life Technologies #18068-015) to ensure there was no contamination with genomic DNA. Reactions were incubated for 15 minutes at room temperature then the DNase inactivated by the addition of 25mM EDTA and heating to 65°C for 10 minutes.
  • Reverse transcription was performed using the 1st strand cDNA synthesis kit for RT-PCR (AMV) from Boehringer Mannheim (#1 483 188). Reactions were incubated at 25°C for 10 minutes then at 42°C for 1 hour. The AMV enzyme was denatured by heating to 99°C for 5 minutes then the reaction was cooled to 4°C. 32P or 33P deoxynucleotides were used to label the cDNA for subsequent probing of the arrays. RNA from DU145 and NHBSF cultures were mixed in equal proportions and the same quantity of RNA used in the reverse transcription as was used for the co-culture RNA.
  • AMV 1st strand cDNA synthesis kit for RT-PCR
  • Labeled cDNA was used to probe a Research Genetics Human Named Array (Named Array) following the manufacturers protocol for hybridisation and washing. Images were recorded utilising a phosphor imager and recorded files analysed with the Research Genetics PathwaysTM software. Results
  • Bone morphogenetic protein 8 (osteogenic protein 2)
  • Figure 1 a. B16-F10 Roller bottle 5 day growth b. B16-F10 5 xT175 5 day growth c. K1735 5 x T175 5 day growth d. K1735 Roller bottle 5 day growth
  • Figure 2 a. PNT2 Not irradiated b. 1542 Not irradiated c. Du-145 Not irradiated d. K1735 Not irradiated
  • FIG. 3 a. PNT2 Not irradiated b. PNT2 irradiated c. 1542 Not irradiated d. 1542 irradiated

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Chemical & Material Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Medicinal Chemistry (AREA)
  • Public Health (AREA)
  • General Health & Medical Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Microbiology (AREA)
  • Oncology (AREA)
  • Immunology (AREA)
  • Mycology (AREA)
  • Epidemiology (AREA)
  • Organic Chemistry (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Peptides Or Proteins (AREA)

Abstract

Il est désormais établi que les cellules cancéreuses contiennent des antigènes spécifiques de la tumeur et associés à la tumeur, lesquels antigènes sont susceptibles de reconnaissance par le système immunitaire. La présente invention concerne ainsi un vaccin du cancer, essentiellement, mais de façon non limitative, dirigé contre le cancer de la prostate. Ce vaccin contient un composant immunogène qui inclut un mélange peptidique extrait de cellules malignes ou bénignes dont l'immunogénicité est altérée. En outre, les cellules desquelles le peptide est extrait peuvent être des cellules allogènes ou des cellules autologues. L'invention concerne également des procédés de production de vaccins anticancéreux.
PCT/GB2000/001921 1999-05-21 2000-05-19 Nouvelles formulations vaccinales-3 WO2000071156A2 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU49357/00A AU4935700A (en) 1999-05-21 2000-05-19 New vaccine formulations-3

Applications Claiming Priority (10)

Application Number Priority Date Filing Date Title
GBGB9911824.2A GB9911824D0 (en) 1999-05-21 1999-05-21 New vaccine formulations
GBGB9911825.9A GB9911825D0 (en) 1999-05-21 1999-05-21 New vaccine formulations
GBGB9911823.4A GB9911823D0 (en) 1999-05-21 1999-05-21 New vaccine formulations
GB9911823.4 1999-05-21
GB9911825.9 1999-05-21
GB9911824.2 1999-05-21
GB0008032A GB0008032D0 (en) 2000-04-01 2000-04-01 New prostate cell lines
GB0008029.1 2000-04-01
GB0008029A GB0008029D0 (en) 2000-04-01 2000-04-01 New vaccine formulations
GB0008032.5 2000-04-01

Publications (2)

Publication Number Publication Date
WO2000071156A2 true WO2000071156A2 (fr) 2000-11-30
WO2000071156A3 WO2000071156A3 (fr) 2001-02-22

Family

ID=27515936

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/GB2000/001921 WO2000071156A2 (fr) 1999-05-21 2000-05-19 Nouvelles formulations vaccinales-3

Country Status (2)

Country Link
AU (1) AU4935700A (fr)
WO (1) WO2000071156A2 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2005011729A1 (fr) * 2003-07-23 2005-02-10 Onyvax Limited Lignees cellulaires provenant de la prostate humaine, utilisees dans le traitement du cancer

Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1990003183A1 (fr) * 1988-09-23 1990-04-05 University Of Southern California Vaccin immunotherapeutique pour tumeurs melaniques
WO1997024132A1 (fr) * 1995-12-28 1997-07-10 The Johns Hopkins University School Of Medicine Vaccins a base de tumeurs de cytokine paracrine allogenique
WO1997028255A1 (fr) * 1996-02-02 1997-08-07 The Government Of The United States Of America, Represented By The Secretary, Department Of Health And Human Services Lignees de cellules epitheliales immortalisees provenant de prostate d'origine humaine, clones correspondants et leurs applications s'agissant de recherche et de therapie du cancer de la prostate
WO1998054343A1 (fr) * 1997-05-29 1998-12-03 Bystryn Jean Claude Vaccin anticancereux
WO1999019462A1 (fr) * 1997-10-10 1999-04-22 The Regents Of The University Of California Populations renforcees de cellules immunogenes preparees avec utilisation d'antagonistes du recepteur h2
WO2000033869A2 (fr) * 1998-12-10 2000-06-15 Onyvax Limited Nouveaux traitements contre le cancer

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1990003183A1 (fr) * 1988-09-23 1990-04-05 University Of Southern California Vaccin immunotherapeutique pour tumeurs melaniques
WO1997024132A1 (fr) * 1995-12-28 1997-07-10 The Johns Hopkins University School Of Medicine Vaccins a base de tumeurs de cytokine paracrine allogenique
WO1997028255A1 (fr) * 1996-02-02 1997-08-07 The Government Of The United States Of America, Represented By The Secretary, Department Of Health And Human Services Lignees de cellules epitheliales immortalisees provenant de prostate d'origine humaine, clones correspondants et leurs applications s'agissant de recherche et de therapie du cancer de la prostate
WO1998054343A1 (fr) * 1997-05-29 1998-12-03 Bystryn Jean Claude Vaccin anticancereux
WO1999019462A1 (fr) * 1997-10-10 1999-04-22 The Regents Of The University Of California Populations renforcees de cellules immunogenes preparees avec utilisation d'antagonistes du recepteur h2
WO2000033869A2 (fr) * 1998-12-10 2000-06-15 Onyvax Limited Nouveaux traitements contre le cancer

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
FRESHNEY R. I.: "Culture of animal cells" 1987 , ALAN R. LISS , US, NEW YORK XP002152704 154920 page 295, left-hand column, line 26 -right-hand column, line 2 page 314, left-hand column, line 3 - line 17 *
OLUMI ARIA F ET AL: "A novel coculture technique demonstrates that normal human prostatic fibroblasts contribute to tumor formation of LNCaP cells by retarding cell death." CANCER RESEARCH, vol. 58, no. 20, 15 October 1998 (1998-10-15), pages 4525-4530, XP002152739 ISSN: 0008-5472 *

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2005011729A1 (fr) * 2003-07-23 2005-02-10 Onyvax Limited Lignees cellulaires provenant de la prostate humaine, utilisees dans le traitement du cancer
GB2419530A (en) * 2003-07-23 2006-05-03 Onyvax Ltd Human prostate cell lines in cancer treatment
GB2419530B (en) * 2003-07-23 2008-01-02 Onyvax Ltd Human prostate cell lines in cancer treatment

Also Published As

Publication number Publication date
AU4935700A (en) 2000-12-12
WO2000071156A3 (fr) 2001-02-22

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