The 2007 Herbert Tabor Journal of Biological Chemistry Lecture
Tony Hunter
Tyrosine phosphorylation: from discovery to the
kinome and beyond
ASBMB Annual Meeting
April 28, 2007
So far 43 JBC papers and one submitted!
The History of Protein Phosphorylation
Protein kinase
ATP
ADP
Protein
P.Protein
P
Protein phosphatase
The History of Protein Phosphorylation
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
The History of Protein Phosphorylation
J. Biol. Chem. 2:127 (1906)
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
The History of Protein Phosphorylation
J. Biol. Chem. 98:109 (1932)
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
The History of Protein Phosphorylation
J. Biol. Chem. 100:583 (1933)
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
The History of Protein Phosphorylation
J. Biol. Chem. 211:969 (1954)
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
The History of Protein Phosphorylation
Burnett and Kennedy, J. Biol. Chem. 211:969 (1954)
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
The History of Protein Phosphorylation
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
Historic moments in the discovery of phosphotyrosine
Pi
+
Pi
18/9/79
P.SER
P.SER
pH “1.9”
“X”
P.THR
P.TYR
P.THR
_
14/6/79
|
mT
acid
|
|
| |
LT
mT mT IgH/Src
in vivo acid protease
v-Src increases P.Tyr levels in transformed chick cells
uninfected
v-Src-transformed
+
pH 1.9
pH 3.5
+
32P-labeled control and RSV-transformed chick fibroblasts
Hunter and Sefton, PNAS 77:1311 (1980)
The History of Protein Phosphorylation
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
The History of Protein Phosphorylation
1900
1910
1920
Phosphoserine in
proteins
Protein
kinase
activity
Phosphotyrosine
in fly eggs
Src
tyrosine
kinase
1932
1954
1964
1979
1930
1940
1950
1960
1970
1980
1990
2000
Phospho
-protein
discovery
Phosphotyrosine
synthesis
Phosphorylase
kinase
cAMP
dependent PK
Gleevec
approved
for CML
1906
1933
1959
1968
2001
More than 29,000 papers on tyrosine
kinases have been published since 1979!
How many tyrosine kinases are there?
1. The finding that v-Src and c-Src phosphorylate tyrosine
gave us the first tyrosine kinase in 1979
2. By the end of 1980 four tyrosine kinases were known (Src,
Abl, EGF receptor, Fps/Fes)
3. By the end of 1990 over 50 tyrosine kinases had been
identified in vertebrates and equal numbers of tyrosine
kinases and serine kinases were known, leading to the
prediction that there might be several 100 tyrosine kinases
in a vertebrate genome and a total of over 1000 protein
kinases
4. The complete human genome sequence reported in 2001
reveals that there are 90 tyrosine kinases (all the tyrosine
kinases had been found by other means before the sequence
was completed), out of a total of 518 protein kinases
The History of Tyrosine Phosphorylation
SH2
domain
identified
PI3 kinase
associates
with MT
PLCγ
as RTK
substrate
STATs
are PTK
substrates
1986
1987
1989
1992
v-Erb is
v-Src MT + Src
derived
protein + tyrosine
from EGFR
kinase
kinase
1977/8
1975
1979
1980
1984
PTP1B
is first
PTPase
1988
1985
PTP1B
and IRK
structures
PTB
domain
binds pY
Human
kinome has
90 PTKs
1994
1995
2002
1990
1995
2000
2005
c-Src
gene
discovery
Abl +
EGFR
are PTKs
Bcr-Abl
fusion
in CML
SH2
domain
binds P.Tyr
SH2
domain
structure
Inactive
c-Src
structure
Gleevec
approved
for CML
1976
1980
1985
1990
1992
1997
2001
v-Src
kinase
sequence
v-Src ∆C
activation
mechanism
Cdc2 is
inhibited
by P.Tyr
Yersinia
encodes
PTPase
Grb2
SH2/SH3
adaptor
1980
1987
1989
1990
1992
Insulin receptor tyrosine kinase catalytic domain
Hubbard et al. Nature 372:746 (1994)
Receptor tyrosine kinases
?
EGFR
HER2
HER3
HER4
INSR PDGFRα FLT1
IGF1R PDGFRβ FLK1
FLT4
IRR
CSF1R
KIT
FLK2
LET-23 DAF-2
FGFR1
FGFR2
FGFR3
FGFR4
F59F3.1 EGL-15 KIN15
F59F3.5
KIN16
F40G9.13
TKR-1
C08H9.8
F59F5.3
Human RTKs
M01B2.1
R09D1.12
58 (20 classes)
R09D1.13
Worm RTKs
29 (11 classes)
11 Unclassified
CCK4 MET
RON
TRKA
TRKB
TRKC
AXL
MER
SKY
TIE
TEK
T01G5.1
T17A3.8
W04G5.6N
W04G5.6C
Y50D4B-4
ZK938.5
B0198.3
F54F7.5
(B0252.1, F11E6.8, F40A3.5, R151.4, T148.1, T22B11.3
Y38H6C.20, C24G6.2A, F08F1.1, F09A5.2, F09G2.1)
EphA1
EphA2
EphA3
EphA4
EphA5
EphA6
EphA7
EphA8
EphB1
EphB2
EphB3
EphB4
EphB5
EphB6
VAB-1
RYK
DDR1
DDR2
RET
C16B8.1 F11D5.3
C25F6.4
ROS
LTK ROR1 MuSK
ALK ROR2
C16D9.2
RTK106
CAM-1
T10H9.2
LMR1
LMR2
LMR3
Nonreceptor
protein-tyrosine
kinase
s
Nonreceptor
tyrosine
kinases
SH3 binding regio
Y
(Myr)
DNA BD
n
Actin BD
Abl
Y
Fes/Fe r
Y
Syk/Zap70
JEF domai n
Kinase-like domain
Y
Jak
PH domain
Y
Tec
Integrin-binding/JEF domain
Y
Focal adhesion-binding
Fak
Y
Cdc42-binding
Ack
Myr
Y
Y
Src
Csk
Y
Srm
PTK catalytic domain
Y
Y
Rak/Fr k
SH3 domain
Y
Brk/Si k
SH2 domain
Y
What is tyrosine phosphorylation used for?
1. Growth factor signaling (and oncogenesis)
2. Cell adhesion, spreading, migration and shape
3. Cell differentiation in development
4. Cell cycle control
5. Gene regulation and transcription
6. Endocytosis and exocytosis
7. Insulin stimulation of glucose uptake
8. Angiogenesis (formation of new blood vessels)
9. Regulation of ion channels in nerve transmission
Transmembranesignaling
signaling by tyrosine
phosphorylation
Transmembrane
by tyrosine
phosphorylation
EGF receptor
PDGF receptor
NH 2
Interferon receptor
T cell receptor
NH 2
Ligand binding
domain
Out
NH 2
Plasma
membrane
In
Catalytic domain
Transmembrane receptor
protein-tyrosine kinase
Bimolecular receptor
protein-tyrosine kinase
Receptor tyrosine kinase (RTK) signaling through
SH2 and PTB P.Tyr-binding domain proteins
Schlessinger, Cell 103:211 (2000)
Stone age bioinformatics!
Manual alignment - March 1985 (BB)
Eukaryotic protein kinases have related catalytic domains
Serine kinases
PKA-C
Cdc2
Tyrosine kinases
c-Src
EGFR
Catalytic domain ~300 aa
Protein kinase catalytic domain subdomains
Hanks, Quinn and Hunter, Science 241:42 (1988)
Structure of PKA catalytic subunit bound to PKI (5-24) and ATP
N-lobe
pT197
catalytic
cleft
C-lobe
Knighton et al. Science 253:414 (1991)
The birth of the kinome: a thousand and one protein kinases
Src
Phos K PKA
Hunter, Cell 50:823 (1987)
The first kinome
Hunter and Plowman, TiBS 22:18 (1997)
The first kinome tree
Hunter and Plowman, TiBS 22:18 (1997)
How many protein kinases are there?
• S. cerevisiae (6217 genes)
130 (116) protein kinases (2.1%) but no
bona fide tyrosine kinases
• S. pombe (4624 genes)
128 (114) protein kinases (2.8%) but no TKs
• C. elegans (19100 genes)
454 (434) protein kinases (2.4%) including
90 tyrosine kinases (20%)
• D. melanogaster (13600 genes)
239 (223) protein kinases (1.8%) including
32 tyrosine kinases (14%)
• H. sapiens (23,000 genes)
518 (478) protein kinases (2.2%) including
90 tyrosine kinases (16%) (chimpanzee kinome
is essentially identical)
• A. thaliana (26,800 genes)
1055 protein kinases (3.8%) but no
tyrosine kinases (>630 RLK)
M. brevicolli (unicellular choanoflagellate) has bona fide tyrosine kinases, SH2
domains and protein-tyrosine phosphatases (PTPs); the yeasts have PTPs, but no
tyrosine kinases or SH2 domains
~2% of all genes in eukaryotes encode protein kinases
Some more recent kinomes
•
Tetrahymena (27424 genes)
1069 PKs (3.8%) - no true TKs, but has
TKLs and 83 two-component HisK.
Has 630 PKs not assignable to known
families or subfamilies, with 37 novel
classes and 100s of unique PKs
•
Dictyostelium (12500 genes)
285 PKs (2.3%) 246 ePKs, including
TKLs but no true TKs, plus 26 aPKs
and 14 HisKs
•
Sea urchin (24000 genes)
(Strongylocentrotus)
353 PKs (1.5%) 329 ePKs, 24 aPKs
and 53 TKs, but no HisK. Lacks only
4/187 human kinase families
•
Mouse kinome is ~99% identical to human kinome - 540 mouse protein
kinase genes - 510 are orthologous to human protein kinases
•
One conclusion is that the tyrosine kinase-like kinases (TKLs) evolved
in unicellular organisms, perhaps serving as tyrosine (TK) progenitors,
and were secondarily eliminated from yeast. HisKs were lost during
evolution of metazoans, apparently replaced by TKLs and TKs
(Eisen et al. PLoS Biol 4:e286; Goldberg et al. PLoS Genet 2:e38; Bradham et al. Dev Biol 300:180)
ko
O
pi
st
ho
nt
ko
ni
U
nt
M
et
az
oa
n
C
oe
lo
m
at
e
D
eu
te
ro
st
om
e
The evolution of the kinome
+4
+9
+15
+76/-3
+6/-1
+27
53
80
+25/-4
+37
85
+6 / -28
158
+15 / -7
173
-11
Vertebrates
182
+3
Sea urchin
TKs
Drosophila
C. elegans
S. cerevisiae
Dictyostelium
TKLs
Tetrahymena
Gerard Manning
How many human protein kinases and phosphatases?
Protein kinases (the kinome)
518 protein kinases including: (not quite a 1001!)
478 conventional protein kinases (ePKs)
(16 have tandem catalytic domains)
388 protein-serine/threonine kinases
90 protein-tyrosine kinases
58 receptor protein-tyrosine kinases
32 non-receptor protein-tyrosine kinases
(~50 may lack catalytic activity; ~106 pseudogenes)
40 atypical protein kinases in 7 families (e.g. alpha kinases)
Manning et al. Science 208:1912 (2002) (http://www.kinase.com)
Protein phosphatases (the phosphatome)
Nature has invented several ways to remove
phosphate from proteins
1. Ser/Thr - phosphatases
DxH…DxxD…..N
2. Protein-tyrosine phosphatases
C(X)5R
Metal-containing enzymes
PP1, PP2A, PP2B (PPP); PP2C (PPM)
Protein-tyrosine phosphatases
Dual-specificity phosphatases
Low molecular weight phosphatases
Cdc25
3. RNA pol CTD phosphatase family Haloacid dehalogenase-related enzymes
DxDxT….GDxxxE
Eyes absent
Many others
How many human protein kinases and phosphatases?
Protein kinases (the kinome)
518 protein kinases including: (not quite a 1001!)
478 conventional protein kinases (ePKs)
(16 have tandem catalytic domains)
388 protein-serine/threonine kinases
90 protein-tyrosine kinases
~2.5% genes directly devoted to protein phosphorylation and
58 receptor protein-tyrosine kinases
dephosphorylation, and possibly up to 5%, if regulatory subunits,
32 non-receptor protein-tyrosine kinases
inhibitors and scaffolding/anchoring proteins are included.
(~50 may lack catalytic activity; ~106 pseudogenes)
40 atypical protein kinases in 7 families (e.g. alpha kinases)
There are as many tyrosine phosphatases as tyrosine kinases
Manning et al. Science 208:1912 (2002) (http://www.kinase.com)
Protein phosphatases (the phosphatome)
~140 protein phosphatases including:
38 protein-tyrosine phosphatases
38 serine/threonine phosphatases
(18 PP1/2A (PPP); 20 PP2C (PPM))
62 DSPs (e.g. MKPs, PTEN); 8 HADs (EyA/FCP)
Manning, Whyte, Martinez,
Hunter and Sudarsanam
Science 208:1912 (2002)
The beard kinome
A 1001 hairs?
Marc Bitensky (March, 1990)
Human
tyrosine
kinases
(90)
Alternative transcripts of mouse
protein kinases and phosphatases
Products
Number per locus
Transcripts
6.7
Polypeptides
3.7
Domain combinations
1.6
5’ exons
1.8
3’ exons
1.6
Most of the alternative coding products differ outside the catalytic domain,
but a significant number of kinases have variant catalytic domains
Forrest et al. Genome Biol 7:R5 (2006)
(variant.imb.uq.edu.au)
Global dynamics of protein phosphorylation
Analysis of global phosphorylation events using enrichment of
phosphopeptides by IMAC, TiO2, PAC or phosphoantibodies derived
by proteolytic digestion of cellular fractions followed by tandem MS
identification of phosphopeptides and phosphorylation sites
•
2,002 phosphorylation sites identified on 967 HeLa nuclear proteins
(Beausoleil, Gygi, PNAS 101:12130, 2004)
•
5,635 phosphorylation sites identified on 2,328 proteins from mouse liver
(Villen, Gygi, PNAS 104:1488, 2007)
•
6,600 phosphorylation sites identified on 2,244 proteins in HeLa cells;
~14% of these change >2 fold within 20 min of EGF (Olsen, Mann, Cell
127:635, 2006)
•
CST Phosphosite database has >48,400 phosphorylation sites, including
6,200 human proteins (N.B. Bodenmiller, Aebersold, Nat Meth 4:231, 2007)
These data suggest that the majority of intracellular proteins are
phosphorylated at one or more sites under the appropriate condition occupancy of many of these sites can change in response to stimuli
Functional categories of EGF-regulated phosphoproteins
QuickTime™ and a
TIFF (Uncompressed) decompressor
are needed to see this picture.
Olsen, Mann, Cell 127:635 (2006)
The human kinome
•
The completion of the human genome sequence has permitted a
complete cataloguing of all human protein kinases
•
There are 518 protein kinase genes, of which about 450 have
kinase activity and 90 are tyrosine kinases. This represents
about 2% of all human genes
•
There are more than 140 protein phosphatases, and a large
number of additional types of protein that recognize proteins
once they are phosphorylated through phosphobinding domains
•
Most of the proteins in a cell can be phosphorylated at one or
often multiple sites under the right conditions
•
Protein phosphorylation is a major mechanism of signal
transduction in eukaryotic cells and perturbation in
phosphorylation-driven processes can lead to disease
Protein kinases are implicated in diverse
diseases
•
>150/518 (~30%) of
human protein kinases
reportedly implicated in
various diseases
•
Many more are likely to
follow from expression,
sequencing, and SNP
analyses, genetics and
functional genomics
•
Kinases are tractable
drug targets with several
approved drugs and
huge development
efforts
Gerard Manning (CST “Kinase and Signaling Reference Guide”)
(www.cellsignal.com/reference/kinase disease.asp)
Human diseases caused by mutations in protein
kinases
DISEASE
KINASE
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
MYOTONIC MUSCULAR DYSTROPHY
X-LINKED AGAMMAGLOBULINEMIA
HIRSCHSPRUNG DISEASE, MEN
CD8 DEFICIENCY FORM OF SCID
X-LINKED SCID
CRANIOSYNOSTOSIS
PEUTZ-JEGHERS SYNDROME
COFFIN-LOWRY SYNDROME
ATAXIA-TELANGIELTASIA
LI-FRAUMENI SYNDROME
WILLIAMS SYNDROME
LEPRECHAUNISM, DIABETES
WOLFF-PARKINSON-WHITE SYNDROME
GORDON HYPERTENSION SYNDROME
WOLCOTT-RALLISON SYNDROME
HEREDITARY EARLY-ONSET PARKINSON’S DISEASE
HEREDITARY EARLY-ONSET PARKINSON’S DISEASE
PULMONARY HYPERTENSION
FAMILIAL ADVANCED SLEEP PHASE SYNDROME
RETT SYNDROME (NEURODEVELOPMENT DISORDER)
HYPORESPONSIVENESS TO BACTERIAL INFECTION
MYOTONIN PROTEIN KINASE
BRUTON TYROSINE KINASE (BTK)
RET
ZAP70
JAK3
FGF RECEPTOR KINASES
LKB1
RSK2
ATM
CHK2
LIMK1
INSULIN RECEPTOR
AMP-ACTIVATED KINASE (AMPK)
WNK1 AND WNK4
eIF2AK3/PEK
PINK1
LRRK2
TGFβ FAMILY RECEPTOR BMPR-II
CKIδ
CDKL5
IRAK4
•
•
•
•
•
•
•
•
POLYCYTHEMIA VERA
MELANOMA & OTHER SPORADIC CANCERSB-RAF
PAPILLARY RENAL CANCER
CHRONIC MYELOGENOUS LEUKEMIA
CHRONIC MYELOGENOUS LEUKEMIA
NON-HODGKINS LYMPHOMA
5-10% NONSMALL CELL LUNG CANCERS
COLON, BREAST & OTHER SPORADIC CANCERS
JAK2
MET RECEPTOR KINASE
TEL-PDGF RECEPTOR KINASE
BCR-ABL
ALK
EGF RECEPTOR KINASE
PI-3 KINASE (P110α SUBUNIT)
Protein kinases with disease connections
PKs associated
with human
diseases
Cell Signaling Technology, 2005
Human cancer genes
•
355 cancer genes implicated by mutation - ~1% of all human genes
•
90% cancer genes show somatic mutations in cancer, 20% show
germ line mutations and 10% show both
•
The most common mutation class among cancer genes are
chromosomal translocations that create chimeric genes or appose a
gene next a regulatory element of another gene
•
More cancer genes have been found in leukemias, lymphomas and
sarcomas than in other types of cancer despite the fact that they
represent only 10% all cancers
•
The protein kinase catalytic domain is the commonest domain
among cancer genes (30). Domains involved in DNA binding and
transcriptional regulation are also common
Futreal et al. Nat Rev Cancer 4:177 (2004)
Cancer and the kinome
Kinases control cancer pathways
•
120 kinases strongly implicated in cancer; more to come
•
Kinases control complex pathways and signal
transduction, implicated in all major steps/pathways of
transformation
Somatic mutations and kinases
•
Kinases as the test case for large scale resequencing of
tumor genes: efforts at Sanger Center, JHU, Venter,
Broad, elsewhere
•
Results to date are a mixture of stunning (BRAF in
melanoma), impressive (high frequency of PI3-kinase
pathway gene mutations in colon cancer - RTKs, IRS2,
PIK3CA, PTEN, PDK1, AKT2, PAK4 - as well as other
tyrosine kinase and PTP genes), and underwhelming
(breast cancer kinome)
(Davies et al. Nature 417:949; Bardelli et al. Science 300:949; Wang et al. Science
304:1164; Davies et al. Cancer Res 65:7591; Stephens et al. Nature Genet 37:590; Parsons
et al. Nature 436:792; Rand et al. PNAS 102:1434; Sjoblom et al. Science 314:268; Bignell
et al Genes Chr Cancer 45:42; Human Cancer Genome Project)
Protein Kinases/Phosphatases and Cancer
•
Over half of the 90 tyrosine kinases are implicated in human cancer
either through gain of function mutations (e.g. Bcr-Abl), gene
amplification (e.g. EGF receptor) or overexpression (e.g. c-Src) or as
tumor suppressors (e.g. Syk, c-Fes, Csk, EphB2, EphB3, EphB4)
•
Many serine kinases are also implicated in cancer through activating
mutations (e.g. B-Raf), overexpression (e.g. Aurora A), or loss of
function mutations (e.g. Lkb1)
•
•
164 protein kinase genes map to amplicons found in tumors
•
Inactivating and activating mutations in tyrosine/lipid and serine
phosphatases have also been implicated in cancer (e.g. PTEN, SHP-2,
PRL-3; RPTPβ, PTP-BAS, PEZ, RPTPγ, LAR, PTPH; Pr65/PP2A)
80 protein kinase genes map to chromosomal disease loci and these
are candidate genes for the causative mutation in hereditary disease
(e.g. activating mutations in the Ret and Met RTKs in predisposition
to cancer)
Protein kinases as drug targets in disease therapy
•
The involvement of protein kinases and altered phosphorylation in
cancer and other diseases has been well established
•
Protein kinases are often mutated or overexpressed in cancer. More
than 25 protein kinase genes are known to be mutated in human
cancer, and ~120 protein kinases are implicated in cancer. It seems
likely that additional currently uncharacterized protein kinases will
prove to play a role in cancer, and all these protein kinases make
potential drug targets for cancer therapy
•
•
Enzymes (e.g. protein kinases) generally make good drug targets
•
Selective protein kinase inhibitors have been developed and are
proving effective in cancer therapy - GleevecTM, TarcevaTM, SutentTM
Initial concerns that it would not be possible to make specific
inhibitors because most known kinase inhibitors bind to the conserved
ATP binding site and because high intracellular ATP concentrations
would compete for binding have proved groundless
Cancer drugs that act against tyrosine kinases
DRUG
Small molecule drugs
GleevecTM (imatinib)
IressaTM (gefitinib)
TarcevaTM (erlotinib)
SutentTM (sunitinib)
SprycelTM (dasatinib)
TykerbTM (lapatinib)
Many in trials
CANCER
TARGET
leukemia (CML)
lung cancer
lung cancer
GI stromal tumor/RCC
leukemia (CML)
breast cancer
Bcr-Abl tyrosine kinase
EGF receptor TK
EGF receptor TK
Kit receptor TK
Bcr-Abl tyrosine kinase
ErbB2 RTK
several types/AML
angiogenesis/Flt3 RTKs
Monoclonal antibody drugs
HerceptinTM (trastuzumab) breast cancer
breast/renal cancer
ErbituxTM (cetuximab)
AvastinTM (cevacizumab) colon cancer
ErbB2 RTK
EGF receptor TK
VEGF
>70 protein kinase inhibitors are in cancer clinical trials, including several directed against
serine/threonine kinases implicated in cancer (Mark Via, Cambridge Healthtech). The Raf inhibitor
sorafenib (NexavarTM) has recently been approved for treatment of renal carcinoma. Rapamycin, an
mTOR inhibitor, and analogues are also in clinical trials for several cancers
The long road to the GLEEVEC cancer drug
TM
1845
CML
1960
Chr 22∆
Ph Chr
1970
Ab-MuLV
1911
RSV
1970
v-src
gene
1952
polyoma
virus
1982
Bcr-Abl
(22:9)
1973
t(9:22)
translocation
1975
c-src
gene
1978
v-Abl
protein
1977
v-Src
protein
1977
mT
antigen
1978
v-Src
PK
1986
v-Kit
PTK
2000
STI571/Abl
structure
1988
First PTK
1979
inhibitors
v-Src
reported
tyrosine
(TKI)
kinase
1992
CGP57148
PDGFR/
Kit/Abl TKI
(Novartis)
1998
STI571
in CML
patients
1983
mT
associated
c-Src
1988
W is
c-kit
1986
HZ4-FeSV
1990
1996
Bcr-Abl causes STI571 inhibits
CML in mice CML cell growth
and v-Abl tumors
1980
v-Abl
tyrosine
kinase
1979
mT
associated
PTK activity
1927
W mutant
mouse
1984
Bcr-Abl
PTK
activity
Gleevec
1996
c-kit gof
leukemia
mutants
TM
1998
c-kit gof
GIST
mutants
2000
STI571
in GIST
patients
2001
NEJM
papers
reporting
STI571
efficacy
in CML
and GIST
approved by the FDA May 10, 2001
Some outstanding questions
•
•
•
•
•
How/when did tyrosine kinases and phosphatases evolve?
•
Where do critical tyrosine phosphorylation and
dephosphorylation events occur in the cell?
•
Can we develop more specific inhibitors or inhibitors that target a
desired set of tyrosine kinases or phosphatases, or block P.Tyr
interactions for disease therapy?
Are there additional functions for tyrosine phosphorylation?
Are there additional types of P.Tyr-binding domain?
How large is the P.Tyr phosphoproteome?
How are the actions of the tyrosine kinases and phosphatases
coordinated and do they act in combinations? How fast do
phosphates turn over at a particular site?
Kinomics
Gerard Manning (Sugen/Salk)
Greg Plowman
Sucha Sudarsanam
Sean Caenepeel
Science 208:1912
TiBS 27:514
PNAS 101:11707
http://www.kinase.com
THANKS GO TO
AND OF COURSE THE OLD BUFFER!
Walter Eckhart
Mary Anne Hutchinson
Bart Sefton
Jon Cooper
Karen Beemon