biomedical technology and increase in life expectancy · constantinescu et al., trends in...
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Biomedical technology and increase in life expectancy
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Biomedical technology and increase in life expectancy
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Biomedical technology and increase in life expectancy
Major successes:
Recombinant technology, production of recombinant proteins, such asEpo, IFN, Growth Hormone, Tpo mimetics, G-CSF and others
Treatments for anemia, viral infections, growth retardation, thrombocytopenia,neutropenia of chemotherapy
Replacement therapy for missing enzymes, such as glucocerebrosidases
Targeted therapy based on mutations/translocations: imatinib in CML
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• Epo is used in treatment of anemia of kidney disease and ofchemotherapy induced anemia in cancer- controversy!
• G-CSF is used to mobilize hematopoietic stem cells and for thetreatment of neutropenia in chemotherapy of for severe congenitalneutropenia.
• Thrombopoietin fusion proteins are used for the treatment ofthrombocythopenia in autoimmune thrombocytopenia purpuraand hepatitis C induced thrombocythopenia.
• Interferon alpha is used in leukemia/ cancer treatment, in the treatment of viral infections such as HCV, while interferon betais used in the treatment of relapsing remitting multiple sclerosis.
Proteins as Drugs: The Case of Cytokines
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Syed et. al., Nature 1998, 395, 511
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Stem cells CFU-GEMM Early BFU-E Late BFU-E CFU-E Red cell
IL3, SF,GM-CSF+/- Epo
IL3, SF,GM-CSF+/- Epo
3 Days
7-9 Days
Colonies from: CFU-GEMM BFU-E CFU-E
Constantinescu et al., Trends in Endocrinology and Metabolism 1999, 10, 18-23
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Development of Aranesp
A Novel Highly N-glycosylated Protein with Enhanced Stability- lessfrequent injections
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In vivo activity of EPO isoforms
Adapted from Egrie J, et al. Blood. 1997;90:56A. Abstract 243.
EPO isoform
In vivoactivity in mice
0 5
0
15 20 25 30Day of study
Incr
ease
in H
emat
ocr
it (%
)
Isoform 14Isoform 13rHuEPO (9-14)Isoform 12Isoform 11Isoform 10Isoform 9Isoform 8Placebo
15
10
25
10
Incr
easi
ng
seru
m h
alf-
life
Incr
easi
ng
rece
pto
r af
fini
ty
EPO = erythropoietin
5
20
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Effect of mutations on EPO bioactivity
>70% active
20%–70% active
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Aranesp development strategy
• Introduce N-linked glycosylation consensus sequences (Asn-Xxx-Thr/Ser) into r-HuEPO by site directed mutagenesis
• Identify individual variants that have the desired properties
• Test optimal combinations of variants
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Aranesp development questions
• Would the glycan addition be efficient?
• Would the molecules be properly folded and stable?
• Would the ability to stimulate erythropoiesis be retained?
• Would in-vivo activity be increased?
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Aranesp: molecular structure
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Biomedical technology and increase in life expectancy
Targeted therapy: the example of chronic myeloid leukemia,
HER2-positive breast cancer, EGFR-positive colon cancer
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Genomics Informed Medicine
NGS approaches Massive Parallel Sequencing - a very dynamic field-choice of test:
Whole Exome Sequencing WES- only coding regions explored,700 €/sample.
Whole Genome Sequencing WGS-entire genome explored, approx10,000 €/sample.
RNA-seq- determines sequence and levels of gene expression, 800 €/sample.
Chip-seq- determines DNA sequences bound to proteins,700 €/sample
FIRST GENOME SEQUENCING: SANGER METHOD, 1.5 BILLION US $
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Myeloproliferative Neoplasm(Bcr-Abl Positive)
First disease with chromosomal translocation tr 9:22, BCR-ABL
First disease with activated tyrosine kinase activity: ABL
First disease with successful targeted therapy: imatinib
First example of resistance emergence to inhibitors: ABL mutations, i.e. T315I
Blast crisis(>20% balsts in blood and marrow
Chronic Myeloid Leukemia
(Accelerated phase)10-20% blasts in blood and marrow
(Chronic leukemia=chronic phase)
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National Cancer Institute, USA SEER Fact sheets CML
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Platelets
Pro-megakaryocyte
BFU-E
CFU-Megamegakaryoblast
CFU-E
Erythrocyte
Reticulocyte
Proerythroblast
CFU-GM
CFU-Mmonoblast
CFU-Gmyeloblast
Pro-monocyteMeta-
megakaryocyte
*Hematopoietic stem cell
CFU-GEMM
peripheral blood
Tissue
Macrophage
Monocyte Segmented
neutrophil
N./E./B.band
Meta-myelocyte
Myelocyte
eosinophil basophil
Normoblast
Bone marrow
Tpo
Tpo
Tpo
Tpo
BCR-ABL*
**
Amplified in CML
*
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Y1294
Kinase Activation in BCR-ABL
Adapted from Smith KM, et al. Molecular Cell. 2003;12 :27-37.
ATP
Cat. domain
Y1294 PP
1. Phosphorylation
*Courtesy Prof. John GoldmanImperial College
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CGP57148B: 2-phenylaminopyrimidine derivative
N
N
N N
H
N
H
O
N
N
� Potent inhibition of Abl-K, c-kit and PDGF-R
� Salts are soluble in water
� Orally bioavailable
� Not mutagenic
Cellular permeability
No PKC inhibition
TK inhibitory activityStability to hydrolysis
Solubilisation
1992
Courtesy Prof. John Goldman
Imperial College
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Bcr-Abl
Y = TyrosineP = Phosphate
Bcr-Abl
ATP
Substrate
PPP
P
Y
Mechanism of action of imatinib – (2000)
Courtesy Prof. John GoldmanImperial College
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Substrate
Imatinib
Bcr-Abl
Y = TyrosineP = Phosphate
Bcr-Abl
ATP
Substrate
PPP
P
Y
Mechanism of action of imatinib – (2000)
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Cancer. Jun 15, 2012; 118(12): 3123–3127. Huang et al.
Estimations of the prevalence of CML due to success of therapy :
70,000 in 2010,
112,000 in 2020,
144,000 in 2030,
167,000 in 2040
181,000 in 2050.
Specific Inhibition of BCR-ABL in CML, KIT in GIST and PDGFR inLeukemias by Imatinib Derivatives Will Save Millions of Lives Worldwide
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Specific Inhibition of BCR-ABL in CML, KIT in GIST and PDGFR inLeukemias by Imatinib Derivatives Will Save Millions of Lives Worldwide
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BCR-ABL1 inhibitors (2011):Imatinib, nilotinib, dasatinib, bosutinib
Bosutinib(SK-606)
Nilotinib(AMN107)
Dasatinib Bosutinib(SK-606)
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Imatinib(Phos. IC 50)
PDGFR72 nM >
Kit99 nM >
BcrAbl221 nM >
Src>1000 nM
Nilotinib(Phos. IC 50)
BcrAbl20 nM >
PDGFR75 nM >
Kit209 nM >
Src>1000 nM
Dasatinib(Phos. IC 50)
Src0.1 nM >
BcrAbl1.8 nM >
PDGFR2.9 nM >
Kit18 nM
Bosutinib(Phos. IC 50)
Src3 nM >
BcrAbl85 nM >
PDGFR>3000 >
Kit>10000 nM
1. Manley PW, et al. Proc Am Assoc Cancer Res 2007;48:772.2. Weisberg E, et al. Cancer Cell 2005;7:1129.3. Remsing Rix LL, et al. Leukemia 2009;23:477.
Kinase targets of the inhibitors
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Threonine
Steric hindrance: Replacement of threonine by isoleucine at 315 (T315I)
Gorre et al, Science 2001
isoleucineThreonine
ImatinibImatinib Imatinib
Isoleucine
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Ponatinib (AP 24534)
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New Field: Precision Medicine
• Every patient is different : unique sequences.
• Clones are genetically different and co-exist in cancer
• Sub-clones emerge after treatment by selection or by new mutations
• Treatment efficacy depends on genetic make-up
• Sequencing-based prognosis can adjust severity of treatment
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Myeloproliferative Neoplasms
Polycythemia Vera(Vaquez 1892)
Essential Thrombocythemia (Epstein and Goedel, 1934)
Myelofibrosis(Heuck 1879)
These 3 syndromes are 5 fold more prevalent
than chronic myelogenous leukemia31
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32
The Homologous V617F Mutation Activates JAK1 and Tyk2
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JAK2 Inhibitors Also Target Other Kinases and May Induce Resistence
JAK2 Inhibitor Manufacturer Target Clinical Activity IC50, nMCurrent Stage of Clinical Development
INCB018424 (Ruxolitinib, Jakafi)
Incyte / NovartisJAK1, JAK2, JAK3, TYK2
Decreased spleen size, improved quality of life, decreased inflammatory cytokine levels. No significant effect on JAK2V617F allele burden. PhaseIII trial evidence of increased survival.
JAK1: 2.7 JAK2: 4.5 JAK3: 322
Approved FDA, EMEA
TG101348 SAR302503
TargeGen / SanofiJAK1, JAK2, JAK3
Dose-dependent reduction in spleen size and leukocytosis, no thrombocytopenia.
JAK1: 105 JAK2: 3 JAK3: 996
Phase 1/2, Phase 3
XL019 ExelixisJAK1, JAK2, JAK3, TYK2
Decreased spleen size only in patients with JAK2V617F or MPL mutations, decreased pruritus, decrease in circulating blasts in peripheral blood.
JAK1: 132 JAK2: 2 JAK3: 250
Development halted
CEP-701 (Lestaurtinib) Cephalon FLT3, JAK2 Decreased spleen size.JAK2: 1 JAK3: 3 Phase 2
SB1518 S*BIOJAK1, JAK2, JAK2
Reduction of leukocytosis, hepatosplenomegaly, and phospho-STAT5.
JAK1: 1276 JAK2: 22 JAK3: 1392
Phase 1
CYT387Cytopia / YM Biosciences / Gilead
JAK1, JAK2Decrease of spleen size, decrease of transfusion requirements (decrease in severity of anemia), broad anti-cytokine effects.
JAK2: 11 JAK1: 18 JAK3: 155
Phase 1/2
JAK2 Inhibitor Ruxolitinib Is Approved for the Treatment of Int 2 and High Risk Myelofibrosis and is Tested in Pancreatic Cancer and Autoimmune Allopecia
SC1
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Slide 33
SC1 Title for this slide?Sandipan Chatterjee; 06/06/2013
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JAK2 Inhibitor Cures Autoimmune Allopecia
Nature Medicine and Columbia University
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Guessing The Future...............
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The problem with the future is that:
It is No Longer What It Used to Be