2. 2 Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
Table 1.
Blanc et al.
Comparison Between the Different Types of Kinase Inhibitors
Type I
Type II
Type III
Type IV
Covalent Inhibitors
Type of binding
Reversible
Reversible
Reversible
Reversible
Irreversible
Binding site
ATP site
ATP site and DFG pocket
Allosteric (by ATP pocket)
Allosteric (substrate binding domain)
ATP site
ATP-competitive
Yes
No
No
No
No
Selectivity
Low
High
Very high
Very high
Low
O
N
N
N
O
O
HN
O
O
O
N
N
O
N
HN
N
Cl
H
N
S
HO
Dasatinib
Sprycel (BMS-2006)
Multitarget
Erlotinib
Tarceva (Genentech/Roche-2005)
ErbB-1
F
N
HN
Cl
Gefitinib
Iressa (Astrazeneca-2003)
ErbB-1
N
N
O
N
N
N
H
O
HN
S
O
HN S
O
F
N
NH 2
F
Cl
N
N
H
N
Ruxolitinib
Jakaf i (Incyte-2011)
JAK1/2
HN
N
Cl
N
N
F
N
H
NH2
Pazopanib
Votrient (GSK-2009)
VEGFR family, PDGFR, cKIT
N
N N
S
O
CN
O
Vemurafenib
Zelboraf (Roche-2011)
B-Raf
O
HN
Lapatinib
Tykerb (GSK-2007)
ErbB-1/ErbB-2
O
N
F
O
O
N
Cl
F
N
Sunitinib
Sutent (SUGEN/Pfizer-2006) H
VEGFR family, PDGFR
Cl
N
HN
N
H
O
N
N
N
O
Crizotinib
Xalkori (Pfizer-2011)
ALK/Met
N
O
N
O
CN
HN
Bosutinib
Pre-registration (Pfizer)
Bcr-Abl, Src
O
Cl
Cl
Fig. (1). Chemical structure of Type I kinase inhibitors approved by the FDA: generic name; brand name; company name; year of approval by the FDA; and
inhibited kinase/s.
However, the in vitro findings do not fully match with the
clinical data as in the case of dasatinib, who reported a high
increase in LDH levels in vitro [10], but exhibited a low level of
cardiopathy during clinical studies. On the other hand, lapatinib, a
compound presenting a high selectivity kinase inhibitor profile and
the lowest increase of LDH in vitro, was reported to cause left
ventricular ejection fraction (LVEF) depression in patients during
clinical studies [11]. Overall, the majority of the approved agents
are in fact well tolerated in monitored patients from a cardiac safety
perspective [9].
Other issues are yet to be discussed such as tolerance of the
compounds to mutations, since modifications in the ATP pocket are
likely to bring a decline in the activity of the inhibitor [12].
2.2. Type II Inhibitors
This type of molecule represents a non-ATP-competitor that
interacts with the catalytic site of the unphosphorilated inactive
conformation of kinases, exploiting new interactions inside the
lipophilic pocket derived from the change of conformation of the
phenylalanine residue of the DFG N-terminal loop (Fig. 2). The
inhibitor reversibly interacts with the kinase that results into the
formation of one, two or three hydrogen bonds with the protein in
the ‘hinge region’ and also causes extra interactions in the open
DFG-pocket. These new extra lipophilic interactions with the DFGpocket confer Type II inhibitors a high degree of selectivity towards
other undesired kinases. These interactions cannot occur in the
phosphorilated activated form of the kinase (Fig. 2: binding
comparison of dasatinib, Type I inhibitor, and imatinib, Type II
inhibitor, with Bcr-Abl).
The advantages and differentiation of the Type II inhibitors
have been further discussed in a later section (Section 3).
2.3. Type III or Allosteric Inhibitors
These molecules bind outside the catalytic domain of the
kinase, in regions that are involved in the regulatory catalytic
domain modulating the activity of the enzyme in an allosteric
manner. A high degree of kinase selectivity is exhibited because of
the exploitation of binding sites and regulatory mechanisms that are
unique to the target. Additionally, allosteric modulators can provide
subtle regulation of kinases controlled by multiple endogenous
factors, something not easily performed with ATP-competitors [15].
A new class of 4, 6-disubstituted pyrimidines (GNF-2
and GNF-5) that selectively inhibits Bcr-Abl dependent cell
proliferation was introduced by Gray and co-researchers. This is
particularly found to be significant for GNF-2 which does not
inhibit c-Abl kinase in vitro (Fig. 3). Docking, NMR, X-ray
3. Type II Kinase Inhibitors
Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
N
N
N
N
H
N
HN
HO
N
N
N
S
Dasatinib
Cl
H
N
N
H
N
O
Imatinib
O
3
N
N
Fig. (2). Overlay complex of dasatinib and imatinib with Bcr-Abl. Note the modification of conformation of the Phe382 of the DFG motif (highlighted with a
circle): the conformation of Phe382 (2GQG) in the complex of activated form of Bcr-Abl/dasatinib (black structure) would not allow the binding of imatinib
(grey structure). Conversely, dasatinib would be able to bind to the inactive form of the kinase due to the conformation of Phe382 (1IEP) [13, 14].
O
F3CO
NH
R
NH
N
N
R: -H;
GNF-2
-(CH 2)2OH; GNF-5
H
N
O
O
H
N
O
H
N
F
N
Selumetinib
(AZD6244)
N
HO
Cl
H
N
PD 0184352
(CI-1040)
HO
O
O
H
N
O
H
N
OH
F
I
HO
Cl
F
PD 0325901
F
O
H
N
O
H
N
N
Br
ARRY-509
F
I
F
HO
F
O
H
N
O
H
N
F
Br
I
O
G-894
HN N
Fig. (3). Allosteric inhibitors: GNF-2, GNF-5, PD 0184352 (CI-1040), PD 0325901, selumetinib (AZD6244), ARRY-509 and G-894.
crystallography, mutagenesis and hydrogen-exchange experiments
all show consistency with binding of these molecules to the
myristate-binding site located near the C-terminus of the kinase
domain, resulting in allosteric inhibition. This binding is thought to
induce a bent conformation of the -I helix that facilitates the
stabilization of an inactive form of Bcr-Abl. Mutations in the ATPpocket terribly affect the inhibitory activity of these molecules. In
contrast to imatinib, GNF-2 shows strong IL-3 reversible antiproliferative and apoptotic effect on mutants E255V and Y253H.
On the other hand, like imatinib, this family also does not exhibit
any activity towards cells expressing G250E, Q252H, F317L and
T315I Bcr-Abl mutant. Unsurprisingly, mutations at the myristate
pocket (A337N and A344L) report a detrimental impact on the
activity of GNF-2 and GNF-5. Encouragingly, combination therapy
of ATP and non-ATP competitors nilotinib and GNF-5 reported
complete disease remission in a T315I mutant murine bone-marrow
transplantation in vivo model [16, 17].
Scientists from Pfizer discovered a set of benzhydroxamate
allosteric MEK inhibitors that stabilize the kinase in the inactive
conformation of the enzyme (Fig. 3; PD 0184352 (CI-1040), and
PD 0325901) [18, 19]. These compounds were found to present a
higher selectivity towards MEK and non-ATP and ERK-competitive.
Crystal structures of MEK1 and MEK2 (closely related, dualspecific tyrosin/threonine protein kinases) with PD 0184352
reported the presence of a unique inhibitor-binding pocket adjacent
but not overlapping with the cMgATP-binding site within the
interlobal cleft of the kinase, explaining the non-competitivity with
ATP. A low sequence homology with other kinases, explaining
higher selectivity of these inhibitors was also observed at this
4. 4 Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
A)
Blanc et al.
B)
PD318088
Val127
I
Ser212
F
F
H
O
Lys97
O
Adenosine
O
P
O
O
O
P
O
O
O
O
P
Mg
N
F
Br
NH
HO
O
O
HO
Fig. (4). Mode of binding of PD318088 with MEK1 at the active site, close to the hinge. A) Model structure of the molecule with the kinase; B) schematic
drawing of the H-bond interactions of PD318088 with the amino acids of MEK1 [20].
region. SAR observations revealed the orthogonality of the aniline
ring towards the anthranilate ring, directing the aromatic ring
towards Phe209 in the hydrophobic pocket; the iodine was found
quite favourable to electrostatically interact with Val127; the
anthranilate phenyl ring was found to be present in less
hydrophobic pocket; and the 4-fluorine reported a formation of a
critical H-bond with Ser212 [3, 20, 21] (Fig. 4). PD 0184352
advanced into Phase II but could not present complete efficacy
because of the insufficient systemic exposure (oral bioavailability
in human: 5%) low solubility and rapid metabolism [19]. Better
physical properties were exhibited by PD 0325901 as compared to
its predecessor, and therefore having better systemic exposure (oral
bioavailability in human: >30%), but the compound was retrieved
from Phase I because of ocular toxicity concerns in patients.
Understanding the SAR and the chances offered by the
pharmacophore for MEK, has encouraged the development of new
series of compounds. Selumetinib (AZD6244) and ARRY-509 were
developed by the researchers of Array Biopharma with the replacement
of the central anthranilate phenyl ring by a benzimidazole and a
pyridine respectively [3]. Genentech replaced the central ring with
the help of a benzopyrazole (G-894) and other heteroaryls,
demonstrating the versatility of the pharmacophore [22].
2.4. Type IV or Substrate Directed Inhibitors
These kinase inhibitors are reported to be small molecules that
present a reversible interaction outside the ATP pocket, in the
kinase substrate binding site, but not competing with ATP. Since
this area being unique for the substrate, it forces this type of
compounds to potentially present a high degree of selectivity.
ON012380 is a potent non-ATP competitive inhibitor of BcrAbl (10.0 nM) (Fig. 5). This molecule is likely to target a site of the
natural substrate of the enzyme, such as Crk. Furthermore, imatinib
and ON012380 were found to synergistically inhibit wild-type
Bcr-Abl that suggests a binding of these two compounds to different
sites on the enzyme. ON012380 was found to be promising for
OMe
MeO
OMe
O O
S
OMe
NH
CO 2H
ON012380
Fig. (5). Substrate directed inhibitor ON012380.
inhibition of all the imatinib-resistant mutants of Bcr-Abl that were
tested such as T315I (7.5 nM) [23].
2.5. Covalent Inhibitors
These kinase inhibitors directly target a catalytic nucleophile
within the active site of the enzyme, and an irreversible covalent
bond is formed. This ‘suicide’ inhibition takes place via trapping of
a solvent-exposed cysteine residue either by SN2 displacement of a
good leaving group or by reacting with a Michael acceptor
incorporated within the inhibitor [4, 24, 25]. The structural
similarity of these compounds with Type I inhibitors, and the
irreversibility of the inhibition are found to cause the inhibition of
kinases with high Km values for ATP, by shifting the equilibrium
between the free and the inhibitor-bound fraction (Fig. 6). This
inhibitors exhibit a plethora of advantages as the one being the long
dissociation half-lifes, which maximizes the efficacy beyond the
clearance of the inhibitor, reduces the drug exposure and minimizes
off-target effects [26]. However, the potential irreversible modification
of on-target or off-target proteins and the potential lack of kinase
selectivity are yet to be defined. For addressing the latter,
researchers have targeted non-conserved cysteines in the kinome.
For refinement of this approach, a basic functionality is introduced
adjacent to the electrophilic centre to speed up bond formation by
activating the cysteine. Adjustments in the linker that position the
electrophilic centre close to the cysteine thiol, and modifications in
the original scaffold itself, can also play a key role in optimizing
selectivity and targeting inactivation rate. Irreversible inhibitors
generated from such an optimization program are known as rapid
kinase inhibitors that show a high degree of selectivity [27].
Recently, Winssinger and colleagues first time targeted a
cysteine that is found to appear at the time when the kinase adopts
the inactive DFG-out conformation, by arming the Type II inhibitor
imatinib with an electrophilic centre (Fig. 7). Among the kinases
that are likely to adopt the required conformation for accommodating
the imatinib pharmacophore, only cKIT and PDGFRs, possess a
suitably positioned cysteine residue at the beginning of the catalytic
loop (Cys 788 and Cys814 respectively). Interestingly, the authors
were able to discriminate with their inhibitors the selected kinases
amongst others, such as Bcr-Abl [26].
3. TYPE II INHIBITORS
While ATP-competitive kinase biochemical assays of highly
active recombinant kinase domains led the discovery of most early
Type I inhibitors, serendipity led to the discovery of a second type
of kinase inhibitors that was later shown to specifically bind an
inactive conformation of the kinase domain. In this so called ‘DFGout’ conformation, the side chain of the phenylalanine residue of
5. Type II Kinase Inhibitors
Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
5
O
O
N
O
N
N
H
O
N
CN
N
O
N
N
H
HN
Neratinib
O
O
N
CN
N
N
H
HN
Pelitinib
N
O
HN
Afatinib
F
F
Cl
Cl
Cl
O
N
O
N
O
N
O
N
N
HN
N
N
N
N
N
N
H
HN
O
Canertinib
O
N
HN
NH 2
Dacomitinib
F
F
Cl
Cl
PCI-32765
O
Fig. (6). Cysteine-targeted kinase inhibitors currently in clinical development.
N
Cl
1
H
N
O
H
N
Cl
N
H
N
O
N
N
R3
H
N
2
O
H
N
N
N
H
N
R2
N
H
N
O
3: R2 : -Me; R3: -H
4: R2 : -H; R3: -Me
O
H
N
N
N
Fig. (7). Compounds developed by Winssinger’s lab to target a cysteine that is available when the kinase adopts the inactive DFG-out conformation.
the DFG N-terminal motif of the kinase activation loop becomes
exposed and penetrates the ATP binding cleft, which results in the
stabilization of the inactive and unphosphorilated form of the
kinase, thereby, opening a hydrophobic pocket proximal to the
kinase “gatekeeper” residue. Since canonical ATP binding site of
activated kinases does not involve any such feature, this pocket is
conserved to a lesser extent across the kinome and hence promises
better prospects for the rational design of selective inhibitors
[7, 28].
The earliest and archetypal Type II kinase inhibitor drug imatinib
(STI-571) was classically optimized from a phenylaminopyrimidine
screening hit with broad spectrum kinase inhibitory activity into a
selective Bcr-Abl inhibitor using only kinase inhibition assay
information (Figs 8, 9 and 10) [29]. A posteriori from the X-ray
structure of an imatinib fragment in murine Bcr-Abl [30] was found
to be beneficial in deciphering the highly unusual binding mode
and was later confirmed with the full-size compound [31].
Approximately, six hydrogen bonds with the Bcr-Abl kinase
domain were formed by imatinib in the latter structure: one between
the pyridine nitrogen and the backbone NH of Met318 in the hinge
region; another between the anilino NH group and the side chain
hydroxyl of the “gatekeeper” residue Thr315; a pair of concerted Hbonds between the amide linker and both Glu286 of the C helix
and the Asp381 backbone NH of the DFG segment. Type II
inhibitors exhibit the high conservation of this distinctive H-bond
pattern between the inhibitor and the glutamic and aspartic acids of
the kinase. An interaction is found to be present between the
protonated methylpiperidine tail group of imatinib, while partially
solvent-exposed, and the backbone carbonyls of both Ile360 and
His361. Imatinib is also reported to involve in extensive
hydrophobic contacts with the Bcr-Abl kinase such as notable stacking interactions between the pyrimidine ring and both the
Tyr253 of the collapsed P-loop and Phe382 of the DFG segment. A
series of hydrophobic residues (Met290, Ile293, Leu298, Leu354,
Val379) line the DFG-out pocket that is occupied by the imatinib
benzamide group by replacing the displaced Phe382 side chain.
Interestingly, imatinib has also been shown to adopt a Type I
binding mode in Syk, acting as a weak inhibitor (IC50 > 10 M)
[32].
Solved X-ray structure of imatinib in Bcr-Abl has helped in the
formulations of general Type II kinase design guidelines. The
principal strategy, known as hybrid-design, is the combination of a
hinge binding group with a DFG-out pocket targeting hydrophobic
motif for exploring the high potency of some Type I platforms and
the selectivity potential promised by Type II inhibitor. Bond
between the hinge binding and DFG-out targeting motifs can
accurately be achieved by moieties that preserve the intricate Hbond network involving the conserved glutamic acid of the -C
helix and aspartic acid of the DFG segment as it plays a necessary
role in stabilizing the inactive kinase conformation. Thus, the
presence of an amide or urea in the molecule has been validated as
a hallmark of Type II inhibitors. In an additional approach, a
hydrophobic substitution may be introduced for occupying the
pocket formed by the shift of phenylalanine from the DFG motif
[7]. A recent work by Molteni and co-workers reported the
application of the hybrid-design method for discovering GNF-5837
(Fig. 9), a selective TRK inhibitor that exhibited its efficacy in
rodent models suffering from cancer tumour. A structure similar to
the Type I inhibitor sunitinib, has been combined with a tail portion
present in Type II inhibitors by the authors. This tail containing
urea in its centre is interacted with the kinase, and a terminal 2fluoro-5-trifluorophenyl hydrophobic moiety to occupy the place of
the phenylalanine of the DFG motif flipped into the ATP pocket,
for the stabilization of the inactive form of TRK kinase [34].
6. 6 Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
A)
Blanc et al.
B)
Imatinib (STI-571)
Ile360
His361
Met318
Thr315
Glu286
N
H
N
N
N
N
H
N
H
N
O
Asp381
Fig. (8). Mode of binding of imatinib with Bcr-Abl. A) Model structure of the molecule with the kinase; B) schematic drawing of the H-bond interactions of
imatinib with the amino acids of Bcr-Abl [30, 33].
Generally, Type II kinase inhibitors as compared to Type I
kinase inhibitors are more likely to present higher selectivity
towards target along with lower dissociation rate constant in
biochemical activity, and a profound impact on cellular activity.
Whereas the exploitation of the hydrophobic DFG-out pocket
results in the production of molecules with high MW and clogP,
which present detrimental consequences for druggability. Finally,
the increase in drug resistance that may be caused by the mutations
on the hinge region or on the “gatekeeper” of the targeted kinase is
reported to reduce the pharmacological effect of the molecule.
3.1. Selectivity
As discussed earlier, the identification of those molecules has
greatly been emphasized which are found to exhibit higher
selectivity towards a specific target for minimizing side effects and
toxicity. Type II inhibitors in comparison to those of Type I present
a higher selectivity because of their ability to recognize structurally
distinctive regions of the active cleft outside the highly conserved
ATP binding site that can only be reached in the inactive form of
the kinase. Moreover, subtle modifications of the regions of the
inhibitor interacting with the DFG-out pocket are found quite likely
to enhance this selectivity [35, 36]. A total of 72 kinase inhibitors
against 442 kinases have been introduced by Treiber and Zarrinkar,
covering more than the 80% of the human kinome. In comparison
to Type I inhibitors, those of Type II have been validated to be
more promising. This observation highlight a consistency with the
general assumption that the inactive conformation preferred by
Type II inhibitors is more kinase-specific than an active
conformation that can accommodate Type I inhibitors. However,
the data also reported a large number of other Type II inhibitors that
exhibit a lower degree of selectivity, and indeed a small number of
Type I inhibitors being quite selective. Therefore, it becomes clear
that selectivity of inhibitors does not depend on their type. A
common theme for the most selective compounds, regardless of
inhibitor type, is their structural features or kinase conformations
which are exploited for distinguishing the target kinase from other
kinases [8].
Another rationale for the observed high selectivity of some
Type II inhibitors, is that not all kinases have the appropriate
flexibility to adopt the DFG-out conformation [37]. Structural
studies with Aurora-A and Aurora-B demonstrated the extensive
state rearrangements observed by the protein during activation
probably caused because of the high degree of flexibility of these
enzymes. This property of the Aurora kinases may be exploited by
the inhibitors being able to stabilize the inactive conformation by
promoting hydrophobic collapse around the compound. These
conformation changes are considered unlikely to be tolerated in
other kinases lacking the same degree of flexibility [38].
Moreover, the selectivity profile of the desired molecule can be
tuned by the exploitation or avoiding of extra interactions around
the hinge. For example, masitinib is a molecule in which the central
pyrimidine of imatinib has been replaced by a thiazole (Fig. 9).
Interestingly, Hermine et al. observed a relative selectivity of
masitinib for cKIT versus Bcr-Abl 10 fold higher than for imatinib.
Docking of masitinib and imatinib in these kinases showed the
involvement of pyrimidine ring of imatinib in a hydrogen bond
network for conserving water molecules around the DFG motif of
Bcr-Abl interaction that was not observed in cKIT. This
observation helps to explain the selectivity of masitinib for cKIT,
avoiding the potential cardiotoxicity of imatinib related to Bcr-Abl
inhibition [39].
Whilst considering selectivity as a promising tool for reducing
side effects and toxicity, multiple kinase inhibitions have been
validated as therapeutically alternative approach. Thus, in multitarget drug discovery (MTDD), the approach of inhibiting two or
more targets simultaneously with one chemical agent to avoid
activation of alternative signalling pathways is considered to be
promising. The treatment of multi-kinase inhibitors (MKI) with a
multiple activity profile restricted to cancer-relevant protein kinases
is highly acclaimed for curing malignant disorders, by presenting a
complementary effect [37]. Moreover, multi-kinase inhibition of a
number of different kinases involved in cancer may be quite
effective against different kinds of tumours (Tables 2 and 3). For
instance, the kinases such as Bcr-Abl, cKIT and PDGF-R involved
in cancer are inhibited by imatinib. Imatinib since its approval by
the FDA in 2001, has received 10 different disease indications
(Table 2) [40].
Sorafenib is reported as a bis-aryl urea derivative that results
from an initial HTS for Raf-1 [41] (Table 2, Fig. 9) followed by
subsequent SAR development [42]. This oral anti-tumour agent
approved for RCC and HCC, is found to present a multikinase
activity profile (B-Raf, VEGFR family, PDGFR, cKIT and FLT-3)
(Table 2). Inhibition of these kinases results in a dual effect the one
as tumour suppression and the other being neovascularisation
inhibition [43, 44]. Regorafenib (Table 2, Fig. 9), a molecule
differed from sorafenib only because of the presence of a fluorine
atom in the centre phenyl ring, is also considered as a multitarget
kinase inhibitor (VEGFR family, PDGFR- , cKIT, B-Raf and
RET). This molecule having a complementary inhibitory profile is
found to be quite efficient for controlling tumour neo-angiogenesis,
vessel growth and metastasis [45], presenting a synergic effect on
cancer treatment [46]. In addition, combined inhibition of several
7. Type II Kinase Inhibitors
Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
CF3
N
N
H
N
N
H
N
O
O
O
N
N
H
N
Imatinib (STI-571)
CF3
O
Cl
N
N
H
O
Cl
N
H
7
O
N
H
F
O
N
H
N
N
H
Regoraf enib (BAY 73-4506)
Soraf enib (BAY 43-9006)
N
O
N
CF3
HN
O HN
O
HN
N
H
N
N
H
N
H
Motesanib (AMG-706)
O
O
N
H
N
O
N
N
H
N
N
S
O
N
H
N
N
O N
N
O
O
N
O
N
N
N
Tandutinib (MLN-518)
O
O
O
N
RAF265
F3C
AAL993
O
N
N
N
N
CF3
N
H
N
H
N
H
Quizartinib (AC220)
S
N
H
F
Masitinib (AB-1010)
O
N
N
H
Tivozanib (AV-951 or KRN951)
CF3
O
N
O
N
H
Cl
N
H
H2N
NH
N
N
H
O
O
Linif anib (ABT-869)
F
N
H
N
H
N
H
N
H
GNF-5837
O
O
N
N
N
N
H
O
N
H
Doramapimod (BIRB-796)
Fig. (9). Different Type II kinase inhibitors.
pro-angiogenic pathways may prevent resistance or prolong
progression-free survival. Indeed, adaptive responses by the tumour
and the vasculature to anti-VEGF therapy have been postulated
[47]. A similar approach for tivozanib (AV-951 or KRN951) [48,
49], and linifanib (ABT-869) has been adopted by other authors
[50].
AAL993 (Fig. 9) is known as a hybrid-design Type II inhibitor
derived from the Type I inhibitor PTK787 (vatalanib) [51]. It is
considered as a potent inhibitor of VEGFR family that was
identified after main optimization of an anthranilamide series [51,
52]. The lack of selectivity of AAL993 within the VEGFR family,
located in vascular endothelial cells (VEGFR-1 and -2) and
lymphatic vessels (VEGFR-3) [45], is found likely to present some
advantage. Inhibition of VEGFR-kinase is found quite promising to
suppress tumour growth, vascularisation and metastasis without
affecting normal tissue [52]. The kinase inhibitor motesanib was
identified after a subsequent lead optimization (Fig. 9) [53].
Motesanib is reported to inhibit the inactive state of five kinases
linked to the pathogenesis of several human cancers which are
named as VEGFR1, VEGFR2, and VEGFR3; cKIT and PDGFR.
The in vivo activity of motesanib is generally attributed to the broad
activity against all VEGFRs tested, such as VEGFR1, VEGFR2,
and VEGFR3. VEGFR1 has been shown to mediate the recruitment
of endothelial precursor cells to areas of active angiogenesis,
whereas VEGFR3 has been reported to play a remarkable role in
lymphangiogenesis. Moreover, the stabilization of nascent vessels
involves associations with pericytes, a process mediated by
PDGFR. This small-molecule multi-kinase inhibitor targeting
VEGFRs has shown promising clinical activity against various
solid tumours, including GIST, melanoma, and RCC [54].
Thus, desired selectivity may be refocused towards functional
selectivity, where the anticancer compound inhibits only those
kinases that are directly involved either in the pathological process
or in the pathological pathway of the disease to produce a beneficial
synergistic effect.
3.2. Activity: Biochemical vs Cellular
Type II inhibitors are quite likely to exhibit a low association
rate constant (kon), but a profoundly lower dissociation rate constant
(koff). This can result in a higher residence time compared with
Type I inhibitors, providing a potential benefit of extended kinase
inhibition. The phenomenon of low koff may be attributed to the
extra hydrogen bonding and higher lipophilicity of these molecules
may be the reason behind this phenomenon of low koff [55].
Doramapimod (Fig. 9), an inhibitor for p38- (serine/threonine
kinase [56]), exhibits a slow binding behaviour, whereby its activity
maximizes with the time (IC50 value reduces from 97 nM to 8 nM
after incubating for 2 hours) Moreover, the residence time for this
molecule is calculated to be 23 hours. The high contribution of the
very low koff value is reported to cause this high value. The
in vivo efficacy is affected by potential impact presented by this
slower dissociation [57].
Different binding affinities and residence time at each of the
different kinases inhibited can be presented by multikinase inhibitor
as an added complexity which is reported to affect selectivity and
8. 8 Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
Table 2.
Blanc et al.
Molecules that are Either FDA Approved or in Clinical Development
Generic
Name
Internal
Name
Brand Name
Company
Kinase Target
•
STI-571
Gleevec (USA)
Glivec (EU)
Novartis
International AG
•
cKIT
•
Approved by the FDA:
Bcr-Abl
PDGFR-
•
Imatinib
Indication Disease or Clinical Study Status for Cancer
PDGFR-
•
Nilotinib
AMN107
Tasigna
Bcr-Abl and mutants (except T315I).
cKIT
•
ASM associated with cKIT.
Approved by the FDA:
•
CML-CP and CML-AP, intolerant/resistant to imatinib.
Phase III clinical studies:
PDGFR-
•
MDS/MPD, HES/CEL associated with PDGFR- ;
•
•
GIST: associated with cKIT and PDGFR- ;
•
•
Novartis
International AG
CML and ALL associated with Brc-Abl;
•
PDGFR-
•
cKIT melanoma;
•
GIST: associated with cKIT and PDGFR- .
Approved by the FDA:
•
•
BAY
43-9006
Bayer Schering
Pharma AG
B-Raf and V600E mutant
•
RCC;
VEGFR family
•
HCC.
•
PDGFRcKIT
FLT-3
Phase II clinical studies:
•
VEGFR family/TIE2
Phase III clinical studies:
•
B-Raf and V600E mutant
•
PDGFR-
•
cKIT
•
VEGFR family
•
cKIT
•
RCC;
•
PDGFR
•
Breast cancer and CRC.
•
VEGFR
•
PDGFR
•
CSF1R
•
Nexavar
•
•
Sorafenib
cKIT, V559D, D816V and D814V
mutants
Phase III clinical studies:
•
•
Regorafenib
Tivozanib
Motesanib
BAY
73-4506
AV-951
KRN951
AMG 706
-
-
-
Bayer Schering
Pharma AG
AVEO
Pharmaceuticals,
Inc. and Astellas
Millennium
Pharmaceutical
(Takeda) and
Amgen, Inc.
•
NSCLC, thyroid cancer and breast cancer.
Breast cancer, ovarial/peritoneal cancer and CRC.
CRC and GIST.
Phase II clinical studies:
•
Cancer.
Phase III clinical studies:
Phase III clinical studies:
•
First-line non-small cell lung cancer.
Phase II clinical studies:
•
First-line breast cancer.
Phase III clinical studies:
Masitinib
AB-1010
-
AB Science
•
PDGFR
•
Pancreatic cancer;
•
GIST;
Phase II clinical studies:
•
drug efficacy in vivo. A total of 15 different kinases were inhibited
by Sorafenib. The longest residence time for Ckit was calculated to
be (811 minutes) followed by CDK8/CycC and B-Raf (576 and 568
minutes respectively). Medium residence times for DDR2 and
DDR1 were calculated to be (45 and 24 minutes respectively).
Residence times for other targets such as TAOK3 and TIE2 were
calculated to be lesser than 2 minutes. This difference in residence
time that is also reported to be the same for doramapimod can be
affective in vivo. Up to 50% of DDR1 and 100% of CDK8/CycC
are blocked by Sorafenib after 5 hours; after 7 hours, DDR1 is no
longer blocked while CDK8/CycC activity still gets inhibited by
90%. When the inhibition of DDR1 and cKIT is compared,
residence time starts to act even more strikingly. However DDR1 is
a high-affinity target and cKIT being a low-affinity target, both are
inhibited to an equal extent 4 hours post Cmax. After 7 h when,
DDR1 is no longer inhibited, the low-affinity target cKIT is still
blocked by 50%. In the same way, also the inhibition of
CDK8/CycC and cKIT becomes likely to draw level after 18 hours
[58].
Multiple myeloma.
Finally, Type II inhibitors while targeting the inactive DFG-out
state of the kinase, with a KM,ATP value higher than the
corresponding value for the active DFG-in state, have to face
weaker competition from cellular ATP, which may enhance activity
in vivo. Indeed, even though these compounds may be ATP
competitive, they might act primarily by locking the equilibrium
switch between conformational states in a way that prevents kinase
activation, rather than directly inhibiting it [36, 59].
3.3. The Effect of MW and logP on Solubility and Cell
Penetration
Many other selective and potent compounds can be identified if
the peculiarities of the hydropholic DFG-out pocket are exploited.
As described, however, this exploitation results into the production
of molecules with high MW and logP but exhibiting the potential
of limited solubility, cell penetration and PK properties. For oral
administration, problems can be created in late phases of drug
discovery by this drawbacks [60, 61]. However, evidence to date
suggests a slight difference between kinase inhibitors properties and
9. Type II Kinase Inhibitors
Table 3.
Generic
Name
Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
Molecules that are Either FDA Approved or in Clinical Development
Internal
Name
Brand
Name
-
Abbott
Indication Disease or Clinical Study Status for Cancer
PDGFR
Phase III clinical studies:
•
VEGFR family
•
AML;
•
cKIT
•
RCC;
•
ABT-869
Company
Kinase Target
•
Linifanib
9
FLT3
•
Breast cancer and CRC.
Phase II clinical studies:
Quizartinib
AC220
-
Ambit Biosciences
•
FLT3
•
•
cKIT
Phase I clinical studies:
•
AML.
GIST.
Approved by the FDA:
•
AP24534
Iclusig
ARIAD
Pharmaceuticals
•
•
cKIT
Phase I clinical studies:
•
PDGFR-
•
•
Ponatinib
Bcr-Abl and mutants (including
T315I).
cSRC, FGFR, VEGFR2 and Lyn
Preclinical studies:
CML and Ph+ALL associated with Brc-Abl
Pharmacological resistance.
AML.
Angiogenesis and solid tumours.
Phase II clinical studies:
•
•
Lyn
Phase I clinical studies:
PDGFR-
•
FLT3
•
cKIT
•
Bcr-Abl and mutants (including
T315I).
Phase I clinical studies:
•
CML, refractory/intolerant to imatinib/nilotinib.
•
FLT3, TIE2 and TRKA
•
ALL and AML.
B-Raf and V600E mutant
•
VEGFR2
Phase I clinical studies:
•
PDGFR-
•
•
CytRx Corporation
Bcr-Abl
•
-
•
•
Bafetinib
INNO-406
NS-187
cKIT
•
Tandutinib
-
-
Millennium
Pharmaceutical
MLN518
DCC-2036
RAF265
-
-
Deciphera
Pharmaceuticals
LLC
Novartis
International AG
the ones of “non-kinase-target” oral drugs. Thus small molecule
clinical compounds targeting kinases in comparison to other
compounds in the same phase of development are reported to
exhibit significantly higher MW and logP [62]. For oral
administration in oncology, compounds such as imatinib, nilotinib
(Fig. 10) and sorafenib can be purchased from the market available
at doses of 400 mg qd, 300 mg bid and 400 mg bid respectively.
These molecules have a high molecular weight and logP, especially
nilotinib (MW: 529.18; logP(octanol-water): 4.9, 5.0) [63].
Nevertheless, strategies continued to be directed towards
increasing solubility. A piperazinyl group in imatinib was
introduced by Kuriyan and co-authors to increase the solubility
compared with the parent compound. Target inhibition is not found
to be drastically altered by this new solubilising substituent. It is
likely to stand along a solvent accessible and partially hydrophobic
groove on the back of the kinase left unfilled by imatinib variant
[30]. The successor molecules such as ponatinib [64] or bafetinib
are also found to follow this same philosophy [65] (Fig. 10).
Alternative strategies may be utilized to introduce the solubilizing
substituent into the hinge region or the region adjacent to the ribose
pocket. This is the case for doramapimod (Fig. 9), where a
morpholino substituent is reported to interact with the hinge,
improving the physicochemical properties of the inhibitor for oral
dosing [57].
B-CLL and advanced prostate cancer
Brain cancer.
Phase I clinical studies:
•
Solid tumours.
Malignant melanoma.
In the development of tandutinib (Fig. 9), Pandey and coworkers report a link between a solubilising 7-piperidinepropoxy
group and a quinazoline derivative. This resulted in the production
of a potent compound exhibiting optimal pharmacokinetic
properties in the animal model, with an oral bioavailability of 50%.
This compound was found to suppress the progression of disease in
a FLT3-mediated leukemia mouse model, showing efficacy in a
nude mouse model for CML [66]. Bhagwat and co-workers have
also adopted a similar approach in the development of quizartinib
(Fig. 9). They introduced a solubilizing morpholinoethoxy group on
to the core ring interacting with the hinge. This led to the discovery
of a novel series of highly potent and selective compounds with a
significantly improved solubility and PK profile. Quizartinib was
identified as one of the most potent and selective FLT3 kinase
inhibitors of the series [67].
More drastic modifications have been explored. The presence
of an amide or urea in the molecule was found to be necessary for
the interaction of the glutamic and aspartic acid in the DFG-pocket.
On the other hand, these functional groups are reported to reduce
the solubility of the inhibitor. This drawback can be tackled by
substitutions of these functionalities presented by bio-isosteres [68].
Ramurthy and co-workers replaced the urea of sorafenib and
regorafenib by an aminoimidazole group, with the aim of
improving the physicochemical properties of the B-Raf inhibitor. A
10. 10 Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
Blanc et al.
First Generation Bcr-Abl inhibitors
N
N
H
N
N
H
N
O
N
N
Imatinib (STI-571)
Second Generation Bcr-Abl inhibitors
CF3
CF3
N
O
N
H
N
N
H
N
N
N
N
H
N
N
H
N
O
N
N
N
N
Bafetinib (INNO-406 or NS-187)
Nilotinib (AMN107)
Third Generation Bcr-Abl inhibitors
CF3
N
CF3
N
O
N
O
O
N
N
N
N
N
H
N
H
N
H
O
N
H
N
F
O
N
N
R
N
NH
O
Ponatinib (AP24534)
DCC-2036
N
GNF-6; R: H
N
GNF-7; R:
H
N
N
CF3
N
N
H
N
N
DSA8
H
N
N
O
H
N
O
O
N
N
N
N
N
O
N
H
NH
N
S
HG-7-85-01
Fig. (10). Different generations of Bcr-Abl inhibitors.
docking model of this bio-isostere showed a DFG-out induced
conformation [69, 70]. RAF265 (Fig. 9), a member of this series
has been brought to phase I/II clinical studies for melanoma, and its
efficacy in both wild type and mutated V600E B-Raf melanoma has
been validated by the recent data [71].
As with Type I kinase inhibitors, classical medicinal chemistry
structure modifications, such as addition of ionizable or polar
groups, reduction of logP or MW, addition of hydrogen bonds,
disruption of molecular planarity or construction of pro-drugs, have
helped to improve solubility of this type of the Type II compounds
[72].
3.4. Resistance for Type II Inhibitors
Natural selection by tumour cells may present drug resistance
during antineoplastic treatment. This selection can lead to a
predominant colony of cells likely to neutralize the effectiveness of
the treatment. Similar cancer cells can develop resistance
mechanisms against kinase inhibitors. For example, 33% of
imatinib patients are reported to develop resistance. Several
mechanisms are responsible for this resistance which could also
extend other kinase inhibitors:
• Over-expression of p-glycoprotein efflux transporters through
MDR-1 gene expression to increase active efflux across the cell
surface and to reduce intracellular concentration of the
compound [73].
• Over-expression of -1 acid glycoprotein to induce high
plasma protein binding of imatinib, and therefore, reducing
concentration of free fraction of the kinase inhibitor available to
the cancer cell [74, 75].
•
•
•
•
Over-expression of metabolic enzymes such as prostaglandinendoperoxide synthase 1/cyclooxygenase 1 (PTGS1/COX1)
which encodes the enzyme that metabolizes imatinib [76].
Activation of alternative biochemical signalling pathways to
bypass the effect of the kinase inhibitor [77].
Amplification of the oncogene Bcr-Abl, with the subsequent
increase of the production of Bcr-Abl [78].
Mutations in the primary structure of the kinase : These
mutations are reported to generate a “conformational escape”
[79], causing the destabilization in the equilibrium between the
phosphorylated forms and unphosphorylated ones of the
enzyme towards the active state [80]. Moreover, mutations
around the ATP pocket are reported to alter the binding
properties through which the inhibitor interacts, or introduces a
new steric restriction [12, 78]. These two phenomena are
reported to reduce the therapeutic activity of the inhibitor.
There are different locations where transformations in the
kinase can appear: Mutations in the P-loop and the activation
loop, destabilizing the inactive form of the kinase in favour of
the active state, the main consequence of which is reduced by
the activity of those inhibitors that target the DFG-out pocket
[81]; Mutations in the “gatekeeper”, the hinge and hydrophobic
pocket, are modified either by the binding interaction network
or by introducing steric clash. Such mutations in the “gatekeeper”
introduce a bulky hydrophobic residue (Ile or Met). Consequently,
a direct drop is observed on the binding strength, reducing the
inhibitory effect [78]. These mutations have thoroughly been
investigated in Bcr-Abl (T315I), cKIT (T670I), PDGFR(T674I), EGFR (T790M) and Src (T790M) [82, 83].
11. Type II Kinase Inhibitors
For suppressing mutant resistance, researchers have recently
come up with several strategies in drug design. These strategies
include synthesis of more potent compounds, overcoming the
impediments introduced by the mutations; stabilization of the
inactive conformation of the kinase with a Type II inhibitor; design
of new molecules by hybrid-design, or application of a combinatorial
therapy of different anti-neoplasics. In the subsequent section these
approaches are considered in turn [82].
3.4.1. Synthesis of More Potent Compounds
Nilotinib is considered as a Type II 2nd generation Bcr-Abl
inhibitor (Fig. 6) that has been developed by rational drug design
based on the crystal structure of a Bcr-Abl-imatinib complex to
override imatinib resistance [30, 31]. This inhibitor was achieved
for its high potency and selectivity towards Bcr-Abl, whilst
maintaining a good pharmacokinetic profile [84, 85].
Similar H-bond interactions are exhibited by nilotinib as the
ones presented by imatinib in the hinge and the “gatekeeper”
regions. An amide inversion helps to maintain the same interactions
as imatinib maintains with Glu286 and Asp381 of Bcr-Abl [86].
Crucially, the pendant N-methylpiperazine substituent present in
imatinib was replaced by 3-methylimidazole. The imidazole shows
less critical interactions with the C-terminal lobe as compared to the
ones presented by directional H-bond of the cationic Nmethylpiperazine of imatinib under physiological conditions. This
provides nilotinib with a less stringent induced-fit binding than the
predecessor [87]. In addition, this replacement, combined with the
introduction of a trifluoromethyl group, increases the clogP of the
molecule [63]. A displacement of the binding contribution from the
hinge region of imatinib to the lipophilic DFG-out pocket in
nilotinib is causes by new hydrophobic interactions. These
hydrophobic interactions are also reported to render greater
flexibility to the protein surface [88]. The chemical modifications
maximize the inhibitor activity towards wild type Bcr-Abl, and
exhibit a higher tolerance for point mutations than imatinib.
Moreover, nilotinib becomes more active against phosphorylated
[79]/unphosphorylated Bcr-Abl, and most of the known imatinibresistant Bcr-Abl mutants, with the exception of T315I. A loss of an
H-bond between Thr315 and the aniline; and a steric clash between
the mutated, bulky Ile315 and the methyl group of the same aniline
substituent can be the reason behind this latter exemption of T3I5I
[84].
The DSA library, derived from a Tie-2 kinase inhibitor library,
is a set of Type II 3rd generation inhibitors capable of inhibiting
Bcr-Abl, c-Src, and Hck [89]. As structure is concerned, this library
shows an acute similarity with imatinib. The presence of methoxy
aniline and the triazine in DSA8 (Fig. 10) may result in the formation
A)
Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
11
of extra H-bonds with the hinge region of Bcr-Abl by analogy with
imatinib, resulting in a stronger inhibition [90].
Regorafenib was developed from a novel discovery program
aiming to maximize the potency and drug-like properties within a
well-established urea class. The compound in comparison to
sorafenib is found to be more pharmacologically potent and is
obtained by introducing a fluorine atom onto the centre phenyl ring
(Fig. 5), leading to a similar but distinct biochemical profile [46].
3.4.2. Overcome the Steric Clash Introduced by Mutation
In the T315I mutation of Bcr-Abl, the incorporated isoleucine
introduces a bulky sec-butyl group close to the “gatekeeper”,
reducing the volume of the pocket around the “gatekeeper”, and as
a result brings a decline in the activity of imatinib and nilotinib.
Several approaches have been described to reset the activity by
reducing this steric clash.
Ponatinib is classified as a Type II 3rd generation inhibitor
(Fig. 11) that strongly inhibits Bcr-Abl and 14 related mutants,
including T315I [91]. The inhibitor was developed in an attempt to
increase the selectivity profile of a DFG-in library hit (AP23464)
[92]. By exploring the DFG-out pocket, the authors increased the
selectivity profile [93, 94]. Interestingly, introduction of an
acetylene linker was found to reduce the flexibility of the molecule.
The increase in rigidity diminishes the steric clash with the
“gatekeeper” T315I mutant, allowing more favourable Van der
Waals interactions with Ile315, and Phe382 of the DFG motif.
Moreover, activity is maximized due to the reduced entropy
imposed through rigidification [64]. Using the same linear
acetylene linker, Gray and co-workers synthesized a library of
different Type II 3rd generation inhibitors that also exhibited strong
activity against wt Bcr-Abl and T315I [95].
Conversely, in order to diminish the steric clash with the
“gatekeeper” produced by the T338M mutation in c-Src, Rauh et al.
introduced a more flexible 1,4-substituted phenyl element, being
able to freely rotate to avoid a collision with the bulky gatekeeper
side chain without disturbing the binding interactions formed by the
rest of the molecule [83].
Finally, in the DSA-library (Fig. 10), no interaction is observed
between the nitrogen of the 2-methylaniline and the “gatekeeper” of
Bcr-Abl or c-Src (T315 or T338) but there is rotation of the metadiaminophenyl ring in the library compounds relative to imatinib.
This leads to displacement of the linker amino group relative to the
position in Bcr-Abl·imatinib or c-Src·imatinib complexes. This
displacement avoids the steric clash initially expected to occur due
to the bulkier isoleucine side chain in the T315I and T338I
mutation. Loss of the H-bond to T338 in Src is compensated by
increasing interaction with the hinge [90].
Ponatinib (AP24534)
B)
Met318
Ile360
His361
Asp381
N
N
N
CF 3
Ile315
O
N
N
H
H
N
Glu286
Fig. (11). Mode of binding of ponatinib with Bcr-Abl (T315I mutant). A) Model structure of the molecule with the kinase; B) schematic drawing of the Hbond interactions of ponatinib with the amino acids of Bcr-Abl (T315I mutant) [64].
12. 12 Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
3.4.3. Design Drugs for the “Switch Control Pocket” Thereby,
Stabilizing the Inhibitor-bound Type II Conformation with the
Kinase in the Inactive Configuration, Even in the Face of
Phosphorylation or Mutations Such as T315I, that Otherwise
would Predispose the “Conformational Escape” of the Enzyme to
the Active Conformation
DCC-2036 (Fig. 10) is a Type II 3rd generation inhibitor,
capable of inhibiting both the phosphorylated and unphosphorylated
forms of Bcr-Abl wild type and T315I. The authors approached the
inhibitor resistance due to “conformational escape”, using “switch
control pocket” inhibition to stabilize the kinase in the inactive
form. The molecule provokes an interaction shift from residues
Arg386/Tyr393 present in the phosphorylated active state of BcrAbl, to a new interaction with the Arg386/Glu282 residues present
in the unphosphorylated inactive state of the kinase. To achieve
this, the quinoline nitrogen of DCC-2036 is reported to interact via
an H-bond with Glu282 that is stabilized by the close presence of
Arg386. The urea interacts with the Lys271-Glu286 salt bridge, and
Asp381; whilst the carboxamide-substituted pyridine ring results in
the formation of H-bonds with the hinge residue Met318. No
interaction or steric clash is reported between the molecule and the
“gatekeeper”, explaining the retention of potency against the T315I
mutant [79, 96].
3.4.4. Exploration of New Inhibitor Scaffolds
The application of hybrid-design is validated as a promising
approach for designing new inhibitor scaffolds [35], in which Type
I inhibitors are linked with Type II inhibitor tails known to interact
with the DFG-out pocket [7].
GNF-7 (Fig. 10) is classified as a compound derived from a set
of Type II 3rd generation inhibitors capable of inhibiting the T315I
“gatekeeper” mutant of Bcr-Abl [97]. On the basis of a hybriddesign, the authors combined derivatives of the Type I inhibitor
PD173955 as a core scaffold for interacting them with the hinge,
whilst exploring the DFG-out pocket through incorporating
substituents at the 3-position resembling Type II inhibitors nilotinib
and AAL993 [98, 99]. Co-crystallisation of another compound from
the same series, GNF-6, with Bcr-Abl confirmed the presence of a
pair of H-bonds in the hinge with Met318, and a pair of H-bonds
between the amide and Glu286 and Asp381 with no interaction
being observed with the “gatekeeper”.
HG-7-85-01 (Fig. 10) is another early example of Type II 3rd
generation inhibitor based on a hybrid-design. The compound
selectively inhibits several kinases involved in cancer: Bcr-Abl,
cKIT, PDGFR- and PDGFR- ; and the respective “gatekeeper”
mutants: T315I, T670I, T674I/M and T681I. An X-ray crystal
structure of the c-Src wt/HG-7-85-01 complex revealed two Hbonds in the hinge region with Met341 and Tyr340. Moreover, as
seen previously with imatinib, in the DFG-out pocket, two H-bonds
were found between the amide of the compound with Asp404 and
Glu310 of the kinase; and a further interaction between the
protonated nitrogen of the distal N-methylpiperazine and the
backbone carbonyls of Val383 and His384 [100].
3.4.5. Override the Interactions Affected by the Mutation
Both imatinib and nilotinib utilise aniline as an H-bond donor
for interacting with the T315 of the “gatekeeper” of the wild type
Bcr-Abl. This H-bond is lost if a T315I mutant is present, drastically
reducing the activity of the inhibitors towards the mutated kinase.
Compounds such as ponatinib [64], DSA-library [90] or HG-7-8501 [100], eliminate this interaction with the “gatekeeper” as
described above. To maintain high potent compounds, additional
interacting contributions are required to diminish the loss of this Hbond. In the future, the use of crystallography and fragment based
approaches may be quite promising for identifying such additional
interactions.
Blanc et al.
3.4.6. Combinatorial Therapies of Different Kinase Inhibitors
with Complementary Inhibition Scope, or other Classical Agents
to Obtain an Additive/Synergistic Effect
Combinatorial therapies or ‘cocktails’ of selective protein
kinase inhibitors, with either other targeted agents or conventional
chemotherapy, represent an emerging therapeutic concept for
preventing or overcoming resistance in human malignancies. These
combinatorial approaches are found to be more flexible in terms of
target selection and therapeutic design than multi-targeted protein
kinase inhibitors, because drugs with fundamentally different
biological modes of action can be co-administered at different ratios
relative to each other and according to variable time schedules.
However, more effort are needed to determine the optimal doses
that are both efficacious and well-tolerated by the treated patients
[82].
The Japan Adult Leukemia Study Group (JALSG) combined
imatinib with intensive traditional chemotherapy in a Phase II
study. The authors obtained complete remission for the majority of
patients that were diagnosed Bcr-Abl-positive ALL without an
increase in toxicity [101].
Another approach of drug cocktail is combination of
complementary Type I and Type II kinase inhibitors with an
overlapping profile of resistance mutations in vitro. Deininger and
co-workers compared dual combinations of imatinib, nilotinib, and
dasatinib, to determine the efficacy and resistance of the cocktail
against N-ethyl-N-nitrosourea-exposed Ba/F3-p210Bcr-Abl cells.
Interestingly, combination of two potent inhibitors, dasatinib (Type
I) and nilotinib (Type II), at different low concentration, resulted in
the elimination of mutations except the T315I mutant. Since the
nonhematologic side effects of nilotinib and dasatinib are not
identical, patients with intolerance to either agent could potentially
be managed with combinations at low doses, avoiding toxicity
while maintaining full anti-leukemic activity [102]. In the case of
T315I mutant treatment, this combination approach could be
applied in the presence of 3rd generation of Bcr-Abl inhibitors. On
the other hand, Bubnoff and co-workers proposed a combination of
kinase inhibitors with a non-overlapping profile such as sunitinib
(SU11248) (Type I) and sorafenib (Type II), to avoid resistance
against FLT3 mutations. They came to this conclusion after
analyzing individually these compounds against FLT3-ITD sitedirected mutagenesis and expressed in Ba/F3 cells [103].
4. FUTURE PERSPECTIVES
This review reports the development of the Type II kinase
inhibitors, despite some evident limitations in cancer therapeutics,
where they are emerging as components of standard-of-care
therapy. Moreover, the increasing knowledge about the effects and
efficacy, and about the existence and mechanistic basis for adaptive
evasive resistance and intrinsic indifference, puts forward an
exciting prospect for sustaining and improving the approach. For
overcoming resistance, significant advances have already been
made.
In the future, we speculate the development of rationally
designed inhibitors based on the Type II pharmacophore that will
allow the generation of high-affinity inhibitors stabilizing the DFGout conformation of many other kinases for which this
conformation has not yet been observed. In addition to serving as
drug discovery lead compounds and as tools to investigate
signalling pathways, these new Type II inhibitors will also facilitate
the exploitation of structural plasticity of the kinase active site. The
speed with which Type II inhibitors have been developed fuels
optimism regarding the achievement of the final goal of controlling
cancer.
13. Type II Kinase Inhibitors
5. CONFLICT OF INTEREST
The authors declare that there is no conflict of interest in this
review.
6. ACKNOWLEDGMENTS
The authors want to pay their gratitude to Dr. Stephen Fletcher,
Dr. Guy Van Lommen, Dr. Luc Van Rompaey and Dr. Laurent
Saniere for proof reading.
7. ABBREVIATIONS
ALK
= Anaplastic lymphoma kinase
ALL
= Acute Lymphoblastic Leukemia
AML
= Acute Myeloid Leukemia
ASM
= Aggressive Systemic Mastocytosis
ATP
= Adenosine Triphosphate
B-CLL
= B-cell Chronic Lymphocytic Leukemia
bid
= bis in die (Latin: twice a day)
CDK8
= Cyclin-Dependent Kinase 8
CEL
= Chronic Eosinophilic Leukemia
CHF
= Congestive Heart Failure
CML
= Chronic Myelogenous Leukemia
CML-AP
= Chronic Myelogenous Leukemia-Accelerated
Phase
CML-CP
= Chronic Myelogenous Leukemia-Chronic
Indolent Phase
CRC
= Colorectal Cancer
CSF1R
= Colony Stimulating Factor 1 Receptor
DDR1
= Discoidin Domain Receptor-1
DFG
= Aspartic acid-Phenylalanine-Glycine
DNA
= Deoxyribonucleic acid
ErbB-1/ErbB-2 = Subfamilies of Epidermal Growth Factor
Receptor (EGFR)
ERK
= Extracellular signal-regulated kinase
FDA
= Food and Drug Administration
FLT-3
= Fms-like Tyrosine Kinase 3
GIST
= Gastrointestinal Stromal Tumor
HCC
= Hepatocellular Carcinoma
HES
= Hypereosinophilic Syndrome
IC50
= Half Maximal Inhibitory Concentration
IL-3
= Interleukin 3
IP
= Intellectual Property
JAK
= Janus Kinase
JALSG
= Japan Adult Leukemia Study Group
LDH
= Lactate Dehydrogenase
LVEF
= Left ventricular ejection fraction
MDR-1
= Multidrug Resistant Protein 1
MDS
= Myelodysplastic Syndrome
MKI
= Multikinase Inhibitor
MPD
= Myeloproliferative Disorders
MTDD
= Multitarget Drug Discovery
MW
= Molecular Weight
NMR
= Nuclear Magnetic Resonance
NSCLC
= Non-Small Cell Lung Cancer
PDGFR
= Platelet-derived growth factor receptors
PK
= Pharmacokinetic
Anti-Cancer Agents in Medicinal Chemistry, 2013, Vol. 13, No. 0
PTGS1/COX1
=
qd
RCC
Syk
TRK
VEGFR
=
=
=
=
=
Prostaglandin-Endoperoxide Synthase
Cyclooxygenase 1
quaque die (Latin: once a day)
Renal Cell Carcinoma
Spleen Tyrosine Kinase
Tropomyosin Receptor Kinase
Vascular endothelial growth factor
13
1/
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Revised: October 09, 2011
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