The Cancer Vaccine Company

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MONOCYTEAPHERESIS FOR GENERATING DENDRITIC CELL
VACINATION THERAPY WITH THE AMICUS SYSTEM
Volker Witt1), Elisabeth Beiglböck1),Gerhar
d Fritsch2), Thomas Felzmann2,3)1) St. Anna
Kinderspital, 2.) St. Anna Kinderkrebsforschung
3) Trimed, Vienna, Austria
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Coleys toxin
William B Coley (May 1893) The treatment of
malignant tumors by repeated inoculation of
erysipelas with a report of ten original cases.
American Journal of the Medical Sciences
Coleys toxin (heat-killed S. pyogenes S.
marrescens) remained the only known non-surgical
treatment for cancer until the 1930s. In
retrospect, it was the first effective
cancer-immune therapy! BCG treatment is a more
modern variation of this concept.
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Milestones of immune therapy
1796 Edward Jenner First vaccination against smallpox
1879 Louis Pasteur Cholera and rabies vaccines
1888 Emile Roux, Alexandre Yersin Tetanus and diphtheria toxins discovered
1890 Emil von Behring, Shibasato Kitasato Tetanus antitoxin
1890 William B. Coley First cancer vaccine, Coleys Toxin
1900 Paul Ehrlich Immune responses against cancer
1954 Jonas Salk, Albert Sabin Polio vaccine
Since 1960 Tumour antigens CEA (1965), AFP (1971)
Since 1970 Microorganismen BCG, CpG oligos (1990ies)
1973 Ralph Steinman Dendritic cells
1975 George Köhler, Cesar Milstein Monoclonal antibodies
1977 Small pox declared eradicated
1982 Monoclonal antibody treatment Rituxan (1997), Herceptin (1998)
Since 1986 Cytokines Interferons, interleukins
1995 Dendritic cell treatment First clinical applications
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DC cancer vaccine trials
Reference Disease Pts
Holtl, 2005 1882 Renal cell cancer 22
Yamanaka, 2005 1880 Glioma 24
Chang, 2005 1883 Lung cancer, 8
Liu, 2004 1884 Colorectal cancer, 10
Stift, 2004 1885 Thyroid cancer, (MTC) 10
Paczesny, 2004 1886 Melanoma 18
Avigan, 2004 1846 Breast cancer, renal cancer 32
Pandha, 2004 Renal cell cancer, prostata cancer 5, 11
Steiner, 2004 1888 Glioblastoma multiforme 23
Kikuchi, 2004 1889 Glioma 15
Yu, 2004 1890 Glioblastoma multiforme 14
Stift, 2004 1885 Gastro-intestinal cancer, 20
Gitlitz, 2003 1892 Renal cell cancer 14
O'Rourke, 2003 1898 Melanoma 17
Nagayama, 2003 1893 Melanoma 10
Slingluff, 2003 1894 Melanoma 26
Oosterwij, 2002 1895 Renal cell cancer 12
Marten, 2002 1896 Renal cell cancer 15
Panelli, 2000 1897 Melanoma 10
Nestle, 1998 410 Melanoma 16
Hsu, 1996 412 B lymphoma 4
Reference Disease Pts
Chakraborty, 1998 497 Melanoma 17
Tjoa, 1999 821 Prostate cancer 17
Höltl, 1999 762 Renal cell cancer 4
Mackensen, 2000 1425 Melanoma 14
Morse, 1999 1428 CEA expressing cancer 21
Yu, 2001 1357 Glioma 9
Thurner, 1999 843 Melanoma 13
Brossart, 2000 1160 Breast cancer, ovarian cancer 10
Geiger, 2000 1156 Solid pediatric tumors 10
Rains, 2001 1541 Colorectal cancer 15
Sadanaga, 2001 1542 Gastro-intestinal cancer 12
Burch, 2000 1527 Prostate cancer 13
Lau, 2001 1536 Melanoma 16
Panelli, 2000 1540 Melanoma 10
Banchereau, 2001 1525 Melanoma 18
Fong, 2001 1530 CEA expressing cancer 12
Kikuchi, 2001 1534 Glioma 8
Kugler, 2000 1141 Renal cell cancer 17
Triozzi, 2000 1545 Melanoma, breast cancer 7, 3
Heiser, 2002 1547 Prostate cancer 13
Dagher, 2002 1561 Sarcoma 16
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Cell-mediated anti-cancer immunity
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Generation of clinical grade dendritic cells

• Critical parameters for DC vaccines
• DC isolation and generation

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DC isolation and generation

• Islolation directly from blood or apheresis
  product
• low cell yields
• DC generation from proliferating CD34 progenitor
  cells
• variability in DC phenotype
• DC generation from monocytes after enrichment
• high cell yield from normal apheresis products
• GMP conformaty of the process

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DC manufacturing SOP development
Christina Eichstill, Dagmar Wagner, Sidrah
Ul-Haq, Doris Wimmer, Volker Witt
Leukocyte apheresis
DC yield
DC purity
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Generation of dendritic cells
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Our own expirience in St. Anna Kinderspital

• 51 aphereses
• from 49 aphereses the data were available
• 20 aphereses in healthy donors
• 29 aphereses in 27 patients
• median age 22.6 years (range 5 83)
• median bodyweight 58 kg (range 16 100)
• with prostata carcinoma (7), hypernephrom (7),
  osteosarcoma (5), adrenal carcinoma (1), Wilms
  Tumour (2), hepatocellular carcinoma (1),
  Ewingss sarcoma (1), Neuroblastoma (1), T-NHL (1)

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How old are our donors and patients?
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Parameter settings for the AMICUS system

• MNC offset 2.3
• Erythrocyte offset 6.8
• Cycle volume 1000 ml (800 1400 ml)
• Number of cycles 5-6 (2 12)
• Processed BV 6755 ml (2040 13342)
• Processed BV / kg bw 100 ml/kg (49 261)
• ACD-A whole blood ratio 1 12
• Time used 0243 (0155 0451)

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How many monocytes do we need?

• To perform a run with Elutra system, we need an
  absolute monocytes count of 109 cells
• A low granulocyte contamination
• A low red cell content

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What is the content of the product?
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Healthy donors and patients
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Reaching the cell yield of 110E09
monocytesdonor/patient bodyweight
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Monocytes in peripheral blood
Mono lt 450/µl Mono gt 450/µl
lt110E09 Mono 11 3 14
gt110E09 Mono 8 27 35
19 30 49
U-Test (Mann-Whitney) p 0.0106
Fishers exact test p 0.0005
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No statistically significant influence of
monocyte, lymphocyte, granulocyte, MNC or total
NC preapherese count, or percentage of Total NC
count to the collection efficiency of monocytes
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Linear regression monocytes preapheresis, whole
blood processed, and bodyweight
Monocyte yield 10E09 cells mono /µl0.0032
WB ml0,0003 bw kg0,0015 2.22
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Conclusion

• with the AMICUS system it is possible to obtain a
  sufficient number of monocytes for DC generation
  under GMP quality assurance
• monocyte preapheresis count gt 450/µl and
  bodyweight gt 30 kg are predictive markers to
  reach in one apheresis gt absolute 1109 monocytes

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Status of clinical development
Heike Hügel, Florian Wimpissinger, Volker Witt,
Philipp Funovics
Pilot trials
Childhood cancer Phase I completed, 20 patients treated, objectives achieved
Kidney cancer Phase I completed, 13 patients treated, objectives achieved
Orthopedic cancer Phase I in progress, 7 patients treated
Prostate cancer Phase I completed, 10 patients treated
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The consortium
Krankenanstalt Rudolfstiftung W Stackl, F Wimpissinger
Donauspital M Rauchenwald
Wilhelminenspital H Ludwig, N Zojer
Allgemeines Krankenhaus G Kramer
St. Anna Kinderspital V Witt
Krankenhaus Hietzing H Pflüger
Hanusch Krankenhaus F Stögermayer