CDK7 inhibitors as anticancer drugs

Abstract

Cyclin-dependent kinase 7 (CDK7), along with cyclin H and MAT1, forms the CDK-activating complex (CAK), which directs progression through the cell cycle via T-loop phosphorylation of cell cycle CDKs. CAK is also a component of the general transcription factor, TFIIH. CDK7-mediated phosphorylation of RNA polymerase II (Pol II) at active gene promoters permits transcription. Cell cycle dysregulation is an established hallmark of cancer, and aberrant control of transcriptional processes, through diverse mechanisms, is also common in many cancers. Furthermore, CDK7 levels are elevated in a number of cancer types and are associated with clinical outcomes, suggestive of greater dependence on CDK7 activity, compared with normal tissues. These findings identify CDK7 as a cancer therapeutic target, and several recent publications report selective CDK7 inhibitors (CDK7i) with activity against diverse cancer types. Preclinical studies have shown that CDK7i cause cell cycle arrest, apoptosis and repression of transcription, particularly of super-enhancer-associated genes in cancer, and have demonstrated their potential for overcoming resistance to cancer treatments. Moreover, combinations of CDK7i with other targeted cancer therapies, including BET inhibitors, BCL2 inhibitors and hormone therapies, have shown efficacy in model systems. Four CDK7i, ICEC0942 (CT7001), SY-1365, SY-5609 and LY3405105, have now progressed to Phase I/II clinical trials. Here we describe the work that has led to the development of selective CDK7i, the current status of the most advanced clinical candidates, and discuss their potential importance as cancer therapeutics, both as monotherapies and in combination settings. ClinicalTrials.gov Identifiers: NCT03363893; NCT03134638; NCT04247126; NCT03770494.

Full text

CDK7 inhibitors as anticancer drugs
Georgina P. Sava
1
&Hailing Fan
1
&R. Charles Coombes
1
&Lakjaya Buluwela
1
&Simak Ali
1
#The Author(s) 2020
Abstract
Cyclin-dependent kinase 7 (CDK7), along with cyclin H and MAT1, forms the CDK-activating complex (CAK), which directs
progression through the cell cycle via T-loop phosphorylation of cell cycle CDKs. CAK is also a component of the general transcription
factor, TFIIH. CDK7-mediated phosphorylation of RNA polymerase II (Pol II) at active gene promoters permits transcription. Cell
cycle dysregulation is an established hallmark of cancer, and aberrant control of transcriptional processes, through diverse mechanisms,
is also common in many cancers. Furthermore, CDK7 levels are elevated in a number of cancer types and are associated with clinical
outcomes, suggestive of greater dependence on CDK7 activity, compared with normal tissues. These findings identify CDK7 as a
cancer therapeutic target, and several recent publications report selective CDK7 inhibitors (CDK7i) with activity against diverse cancer
types. Preclinical studies have shown that CDK7i cause cell cycle arrest, apoptosis and repression of transcription, particularly of super-
enhancer-associated genes in cancer, and have demonstrated their potential for overcoming resistance to cancer treatments. Moreover,
combinations of CDK7i with other targeted cancer therapies, including BET inhibitors, BCL2 inhibitors and hormone therapies, have
shown efficacy in model systems. Four CDK7i, ICEC0942 (CT7001), SY-1365, SY-5609 and LY3405105, have now progressed to
Phase I/II clinical trials. Here we describe the work that has led to the development of selective CDK7i, the current status of the most
advanced clinical candidates, and discuss their potential importance as cancer therapeutics, both as monotherapies and in combination
settings. ClinicalTrials.gov Identifiers: NCT03363893; NCT03134638; NCT04247126; NCT03770494.
Keywords CDK7 .CDK inhibitors .Cell cycle .Transcription .Cancer therapy .Combination therapy
1 Introduction
Cyclin-dependent kinase 7 (CDK7), along with cyclin H and
MAT1, comprises the CDK-activating kinase (CAK), which
provides the T-loop phosphorylation required for activation of
CDKs 1,2, 4 and 6, which drive cell cycle progression
(Table 1, Fig. 1a)[1–4]. CAK also has a role in the regulation
of transcription, as a component of the general transcription
factor TFIIH. At active gene promoters, CDK7 phosphory-
lates the C-terminal domain (CTD) of RNA polymerase II
(Pol II), at serine 5 (Ser5), to facilitate transcription initiation
(Table 1,Fig.1b)[5–7]. CDK7 also phosphorylates CDK9,
which in turn phosphorylates the Pol II CTD at Ser2, to drive
transcription elongation [8]. The activities of a variety of tran-
scription factors, including p53 [9,10], retinoic acid receptor
[11–13], oestrogen receptor [14,15] and androgen receptor
[16,17], are also regulated by CDK7-mediated phosphoryla-
tion (Table 1).
Because of its dual role in regulating the cell cycle
and transcription, CDK7 has been studied as an antican-
cer drug target, and a number of selective inhibitors of
CDK7 have been developed and investigated as cancer
therapies. Preclinical studies have revealed that cancer
cells can be preferentially targeted by transcriptional in-
hibition, at least in part because they are more reliant
than normal cells on high levels of super-enhancer (SE)-
driven transcription [18,19] mediated by specific onco-
genic drivers, such as RUNX1 in acute lymphoblastic
lymphoma (ALL) [20] and N-MYC in neuroblastoma
[21]. To date, four selective CDK7 inhibitors,
ICEC0942 [22], SY-1365 [23], SY-5609 [24,25]and
LY340515 [26], have progressed to Phase I/II clinical
trial for the treatment of advanced solid malignancies.
In this review we outline the role of CDK7 in both normal
and tumour cells and the rationale for inhibiting CDK7 in
cancer. We also discuss the development of selective CDK7
inhibitors, their mechanism of action in cancer and their po-
tential for use in combination therapies.
*Simak Ali
simak.ali@imperial.ac.uk
1
Division of Cancer, Department of Surgery & Cancer, Imperial
College London, Hammersmith Hospital Campus, London, UK
https://doi.org/10.1007/s10555-020-09885-8
Published online: 8 May 2020
Cancer and Metastasis Reviews (2020) 39:805–823
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

P
P
P
P
P
P
P
P
CDK7
Cyclin H
MAT1
CAK
CDK4/6
Cyclin D
CDK2
Cyclin E
Cyclin A
CDK2
CDK1
Cyclin A
CDK1
Cyclin B
CDK7
Cyclin H
MAT1
TFIIH
Pol II
P
P
P
P
Pol II
P
P
P
P
PCDK9
Cyclin T
a
b
PKCι
CK2
CDK8
Cyclin C
P
P
P
CDK7
Cyclin H
MAT1
Fig. 1 Overview of the regulation of CAK and the role of CDK7 in
regulating the cell cycle (a) and transcription (b). CAK = CDK
activatingkinase, CDK = cyclin-dependent kinase, CK2 = protein kinase
CK2, G1 = gap phase 1, G2 = gap phase 2, M = mitosis, P = phosphate,
PKCι= protein kinase C iota, Pol II = RNA polymerase II, S = synthesis,
TFIIH = transcription factor II H
Table 1 CDK7 substrates
Substrate Residue(s) Possible role(s) Refs
Cell cycle CDK1 Threonine 161 T-loop activation and cyclin binding [1,2]
CDK2 Threonine 160 T-loop activation [1]
CDK4 Threonine 172 T-loop activation [3]
CDK6 Threonine 177 T-loop activation [3]
CDK9 Threonine 186 T-loop activation [4]
Basal transcription RNA Pol II Serine 5 and Serine 7 Transcription initiation (Ser5); Unknown (Ser7) [5–7]
TFIIB Serine 65 Promotion of transcription [8]
MED1 Threonine 1457 Recruitment to chromatin [9]
Transcription factors AR Serine 515 Activation and turnover [10,11]
E2F1 Serine 403 and Threonine 433 Degradation [12]
ER⍺Serine 118 Activation and turnover [13,14]
Ets1 Threonine 38 Recruitment of coactivators [15]
p53 Serine 33 and a residue between 311 and 393 Enhanced DNA binding (Ser33) [16,17]
PPAR⍺Serine 112 Activation [18]
PPARγ2 Serine 12/21 Activation [18]
RAR⍺Serine 77 Activation [19,20]
RARγSerine 77/79 Activation [21]
YAP/TAZ Serine 128/90 Prevention of degradation [22]
806 Cancer Metastasis Rev (2020) 39:805–823
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2 CDK7 function
2.1 CAK structure and regulation
CDK7 is a 346 amino acid kinase, having a predicted
molecular mass of 39 kDa, with an N-terminal cyclin
H-binding region and a C-terminal MAT1 binding re-
gion [27]. A single crystal structure has been reported
forCDK7boundtoATP,intheinactiveconformation,
the structure being similar to that of the inactive con-
formation of ATP-bound CDK2 [28]. Cyclin H binding
is obligatory for CDK7 kinase activity, whilst the addi-
tion of MAT1 stabilises the trimeric CAK complex and
anchors it to TFIIH [27]. In addition, cyclin H and
MAT1bindinghavebeenshowntoregulateCDK7sub-
strate specificity, with the trimeric CDK7-cyclin H-
MAT1 complex having greater kinase activity for Pol
II, in comparison to CDK7-cyclin H, which preferential-
ly phosphorylates CDK2 [27–29].
The T-loop of CDK7 can be phosphorylated at two posi-
tions, threonine 170 (Thr170) and Ser164, enhancing both its
kinase activity and ability to bind cyclin H [6]. Furthermore,
T-loop phosphorylation of CDK7 seems to direct substrate
specificity, with Thr170 phosphorylation stimulating activity
towards Pol II over CDK2 [29]. In vitro, CDK1 and CDK2
can phosphorylate CDK7 and as substrates of CDK7 them-
selves; this hints at the possibility of a reinforcement activa-
tion loop between these CDKs [30]. In addition, protein ki-
nase C iota (PKCι), acting downstream of PI3K signalling,
can phosphorylate CDK7 at Thr170 (Fig. 1a)[31–35].
Regulation of CAK activity may also be mediated through
phosphorylation of cyclin H. CK2 can activate CAK in vitro,
via phosphorylation of cyclin H at Thr315 (Fig. 1a)[36],
whereas CDK8 has been shown to negatively regulate tran-
scription initiation, via phosphorylation of cyclin H at Ser5
and Ser304 (Fig. 1a)[37]. Furthermore, CDK7 complexed
with cyclin H and/or the trimeric CAK can phosphorylate
cyclin H in vitro. This autophosphorylation reduces activity
of CDK7-cyclin H but has no apparent effect on CDK7-cyclin
H-MAT1 activity. This suggests that MAT1 binding aids
maintenance of the transcriptional activity of CAK by
preventing regulation by cyclin H phosphorylation [28].
An additional means of CDK7 regulation has been ob-
served in mouse neural progenitor cells, where the
microRNA (miRNA) miR-210 regulates cell cycle progres-
sion by modulating expression levels of CDK7 [38]. This
raises the possibility that there may be additional miRNAs
that regulate CAK expression and activity in other cellular
contexts. There is clearly more to be discovered with regard
to the regulation of CDK7 and CAK activity and the identifi-
cation of players acting upstream of CDK7 could potentially
provide additional means by which to manipulate CDK7
activity.
2.2 CDK7 in the cell cycle
CDK7 controls the cell cycle by phosphorylating the cell cycle
CDKs 1, 2, 4 and 6 in their T-loops, to promote their activities
(Fig. 1a)[1]. Both CDK1 and CDK2 are activated by CDK7-
mediated T-loop phosphorylation, at Thr161 and Thr160,
respectively (Table 1)[2,20–22,39]. Inhibiting CDK7 during
G1 prevents CDK2 activation and delays S phase, whilst in-
hibition of CDK7 during S/G2 prevents CDK1 activation and
mitotic entry [2,22]. Whilst CDK7 can phosphorylate CDK2
prior to its binding to cyclin, and is not strictly requiredfor the
formation of CDK2-cyclin complexes, CDK7 phosphorylates
CDK1 in concert with cyclin B binding and is required for the
stabilisation of CDK1-cyclin B complexes [2,40].
Full commitment to the cell cycle is controlled at the re-
striction point, through phosphorylation of retinoblastoma
(RB) by CDK4/6-cyclin D, in response to mitogens (Fig.
1a). CDK7 phosphorylates both CDK4 and CDK6 in their
T-loops, at Thr172 and Thr177 (Table 1), respectively, and
CDK7 inhibition prevents their RB kinase activity, halting
G1 progression [3,4]. Although expression levels of the
CAK components remain constant throughout the cell cycle,
T-loop phosphorylation of CDK7 increases when cells are
released from serum starvation [3]. Therefore, a mitogen-
induced cascade of CDK T-loop phosphorylation regulates
progression through G1 [3].
Unlike cyclin-bound CDK2, which remains phosphorylat-
ed for up to 12 hours after CDK7 inhibition, CDK4 and CDK6
activity is rapidly lost following CDK7 inhibition [3]. This
difference is likely due to structural differences between the
complexes; the T-loop of CDK2 is protected from dephos-
phorylation by cyclin binding, whereas the T-loops of cyclin
D-bound CDK4/6 remain exposed to phosphatases [3]. As a
result, CDK7 activity is required to maintain CDK4/6 activity
during G1 whilst being required only for initial activation of
CDK1 and CDK2 during S/G2 [3].
2.3 CDK7 in transcription
CDK7 regulates gene expression, as a component of the gen-
eral transcription factor complex, TFIIH (Fig. 1b). TFIIH is
composed of two distinct sub-complexes: the core complex,
which contains two DNA helicases, xeroderma pigmentosum
type B (XPB) and xeroderma pigmentosum type D (XPD),
along with five other structural and regulatory proteins, and
the CAK complex. CAK is recruited to the core TFIIH com-
plex via a reversible interaction between the ARCH domain of
XPD and the latch domain of MAT1 [41,42]. TFIIH is re-
cruited by TFIIE to active gene promoters, where it joins the
other assembled general transcription factors (TFs), and Pol II,
in the preinitiation complex (PIC) [27]. The composition of
TFIIH and the structure of the PIC have recently been
reviewed by Rimel and Taatjes [27].
807Cancer Metastasis Rev (2020) 39:805–823
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After DNA is unwound at the transcription start site (TSS)
by XPB [43], Pol II must be released from the PIC to initiate
transcription, in a CDK7-regulated process termed promoter
escape [5]. The CTD of mammalian RPB1, the largest subunit
of Pol II, contains 52 repeats of a heptad sequence,
conformingto the consensus Y1-S2-P3-T4-S5-P6-S7, the res-
idues of which can be sequentially phosphorylated to regulate
Pol II activity throughout the transcription cycle [44]. Whilst
unphosphorylated, Pol II remains anchored to the PIC, via an
interaction with the mediator complex (another PIC compo-
nent) [5]. CDK7 phosphorylates Ser5 and Ser7 of the Pol II
CTD at gene promoters [6,7]; Ser5 phosphorylation facilitates
the release of Pol II from mediator, allowing Pol II to escape
the PIC and initiate transcription (Table 1, Fig. 1b)[5,45]. The
precise function of CDK7-directed Ser7 phosphorylation is as
yet unclear, but evidence suggests that Ser7 phosphorylation
may promote the transcription and post-transcriptional pro-
cessing of small nuclear RNA transcripts, by facilitating an
interaction between the integrator complex and Pol II [46].
After promoter escape, Pol II generally generates a tran-
script of around 20–80 bases, before halting progress, in a
process known as promoter-proximal pausing, which likely
functions as a checkpoint to ensure the establishment of a
range of co-transcriptional processes [6,47]. CDK7 is re-
quired for the recruitment of two complexes, the DRB sensi-
tivity inducing factor (DSIF) and the negative elongation fac-
tor (NELF), both of which are required to establish the
promoter-proximal pause [6,8,48–50]. For the release of
paused Pol II and commencement of the productive elonga-
tion phase of transcription, the activity of CDK9, as a compo-
nent of the positive transcription elongation factor (P-TEFb),
is required [8]. Like the cell cycle CDKs, for full functionality,
CDK9 must undergo T-loop phosphorylation by CDK7
(Table 1, Fig. 1b)[8]. Therefore, CDK7 plays a role in both
establishing the promoter-proximal pause and in release from
the pause, and inhibition of CDK7 has been shown to increase
the amount of Pol II paused at promoter-proximal regions [6,
51]. Active CDK9 phosphorylates the Pol II CTD, on Ser2,
promoting transcriptional elongation [52]; therefore, there is
an indirect requirement for CDK7 activity after Pol II pause
release.
CDK7 also regulates further transcriptional processes; for
example, CTD phosphorylation by CDK7 allows the co-
transcriptional interaction of Pol II with enzymes that add
the 5′-monomethyl-guanosine cap to nascent RNA transcripts
[50]. Additionally, CDK7 is necessary for appropriate tran-
scription termination, with read-through transcription ob-
served upon CDK7 inhibition [6]. CDK12 and CDK13 are
also involved in regulating transcription by phosphorylating
the Pol II CTD during elongation [53]. In vitro, CDK12 can
phosphorylate Ser2, Ser5 and Ser7 [54], whereas CDK13 can
phosphorylate Ser2 and Ser5 [55]. Like the previously
discussed CDKs, T-loop phosphorylation is necessary for
CDK12/13 activation and is likely mediated by CDK7 [54,
56]; thus, it is probable that additional transcriptional sub-
strates of CDK7, and further roles in transcriptional regula-
tion, remain to be identified.
Genetic targeting of Mat1 or Cdk7 in mice is early embry-
onic lethal and cells cultured from embryos of these animals
fail to enter S phase [57,58]. The activities of Cdks 2, 4 and 6
are reduced in mouse embryonic fibroblasts (MEFs) with
Cdk7 knockout, indicating that Cdk7 has an essential role in
cell proliferation [58]. Cdk7 targeting in adult animals results
in phenotypically normal low-proliferating tissues, such as the
liver, kidney or cerebellum. However, in rapidly dividing ep-
ithelial tissues, Cdk7 expression is retained due to tissue re-
newal sustained by stem cells with incomplete Cdk7 knock-
out. This eventually leads to stem cell exhaustion and prema-
ture ageing [58]. Interestingly, MEFs lacking Cdk7 expression
have unaltered Pol II CTD Ser5 phosphorylation and a largely
unchanged gene expression program, indicating that Cdk7 is
dispensable for de novo transcription [58]. This raises the pos-
sibility that another Pol II CTD kinase can compensate for a
lack of Cdk7.
2.4 CDK7 as a regulator of transcription factor activity
Alongside its critical role in directing transcription by Pol II,
CDK7 phosphorylates a number of TFs, functioning to either
promote their activities and/or regulate their degradation
(Table 1). The activity of retinoic acid receptor ⍺(RAR⍺)is
promoted by XPD-dependent phosphorylation of Ser77 by
CDK7 [11,13]. Likewise, the activity of RARγis also mod-
ulated by phosphorylation by TFIIH-incorporated CDK7 [12].
CDK7, as part of TFIIH, mediates ligand-dependent phos-
phorylation of oestrogen receptor ⍺(ER⍺)atSer118[14,
15], regulating the activity and turnover of the TF [59,60].
Phosphorylation by CDK7, at Ser515 in the transcription ac-
tivation function of androgen receptor (AR), has also been
reported [16,17]. Additionally, CDK7 can phosphorylate
p53 in a MAT1-dependent fashion, at both the C-terminus
(between residues 311 and 393) [10] and the N-terminus, at
Ser33 [9], the former of which has been shown to stimulate
p53 binding to DNA. Evidence that CDK7 phosphorylates
Ets1 [61], peroxisome proliferator-activated receptors
(PPARs) [62] and E2F1 has also been demonstrated, the latter
functioning to trigger E2F1 degradation [63](Table1).
Recently, the stabilisation of the transcriptional regulators
YAP/TAZ was shown to be mediated by CDK7, with phos-
phorylation of YAP at Ser128 and TAZ at Ser90, preventing
their ubiquitination and degradation [64]. At present we have
an incomplete understanding of the role CDK7 plays in regu-
lating the activities of sequence-specific transcriptional regu-
lators. Further knowledge in this area may be helpful in
informing the use of CDK7 inhibitors in specific cellular
contexts.
808 Cancer Metastasis Rev (2020) 39:805–823
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2.5CDK7inDNArepair
TFIIH plays a key role in the nucleotide excision repair (NER)
pathway [27], which repairs single-stranded DNA damage,
particularly that caused by ultraviolet light. TFIIH is recruited
to damaged DNA, where the NER protein, xeroderma
pigmentosum group A (XPA), catalyses the release of CAK
from the core TFIIH complex, allowing NER to proceed [65].
After DNA repair, CAK reassociates with TFIIH, and the
complex resumes its role in transcription [65]. Inhibition of
CDK7 kinase activity improves NER efficiency, suggesting
that CDK7 negatively regulates NER, directly or indirectly,
via phosphorylation of an as yet unidentified substrate(s) [66].
3 CDK7 in cancer
3.1 CDK7 expression in tumours
Two decades ago, immunohistochemical analyses on a range
of tumour types indicated that CDK7 expression is elevated in
tumour cells compared with their normal counterparts [67].
Since then, numerous studies have provided support for this
finding [68–73]. In oestrogen receptor-positive (ER+) breast
cancer, CDK7, cyclinH and MAT1 are overexpressed and are
co-regulated at the mRNA level [68]. Expression of the CAK
components positively correlates with ER expression and
Ser118 phosphorylation, as well as with improved patient out-
comes [68]. Conversely, in triple-negative breast cancer
(TNBC), CDK7 expression is correlated with poor prognosis
[74]. In addition, associations between CDK7 and reduced
survival have been observed in gastric cancer [69,70], ovarian
cancer [75], oral squamous cell carcinoma (OSCC) [71], he-
patocellular carcinoma [72] and glioblastoma [73]. For
OSCC, animal studies have also revealed a potential role for
CDK7 in disease development [71].
These findings raise the possibility that tumours with in-
creased expression of CDK7 may be more sensitive to CDK7
inhibition, particularly in the case ofER+ breast cancer, where
the CDK7-activated nuclear receptor, ER⍺, drives tumour
progression.
3.2 Transcriptional addiction in cancer
Common molecular features of cancer, such as mutation, copy
number changes and genomic rearrangements, can either di-
rectly or indirectly impact gene expression profiles that drive
cancer. For instance, a BRAF mutation in melanoma causes a
cascade of signalling events that ultimately leads to an altered
transcriptional profile and a distinct gene expression signature
[76,77]. Mutations in TF genes are also common in cancer
[78,79]. Across all cancer types, the most frequently mutated
gene (TP53)encodesfortheTFp53[80], and the most
frequently amplified gene, MYC [81], also encodes for a TF.
Other TFs are critical in specific tumour types. For example,
ER⍺activity drives the majority of breast cancer, and thera-
pies that target ER⍺,liketamoxifen[82] and fulvestrant [83]
are used in the treatment of ER+ breast cancer. Mutation,
rearrangements and deregulated expression of genes encoding
chromatin remodelling and histone modification enzymes,
such as EZH2 and ARID1A, are also frequent in cancer [78,
79]. These aberrations alter the accessibility of gene regulato-
ry regions, ultimately leading to downstream changes in gene
expression.
Recently, clusters of enhancers, termed super-enhancers
(SE), that control the expression of genes integral for cell
identity and function have been defined [84]. Deregulation
of the SE landscape is common in cancer and leads to dramatic
changes in gene expression and high transcriptional outputs,
which maintain the oncogenic cell state (Fig. 2). As a result,
cancer cells become transcriptionally addicted, requiring
higher levels of transcription than normal cells to sustain
growth [19]. The phenomenon of transcriptional addiction
suggests that cancer cells may be more responsive than normal
cells to transcriptional inhibition and provides a strong
basis for targeting transcriptional kinases, including CDK7,
in cancer (Fig. 2)[18]. Furthermore, oncogenic TFs, like
MYC, have proven notoriously difficult to target directly with
small molecules; therefore, the ability to target the general
transcription machinery to reduce their transcriptional output
is an attractive prospect.
4 Development of CDK7 inhibitors
4.1 Pan-CDK inhibitors
Early efforts to develop CDK inhibitors yielded relatively un-
selective compounds, with activities against multiple CDKs,
often including CDK7 [85]. The first CDK inhibitor to enter
clinical trial was the semi-synthetic flavone derivative,
alvocidib (flavopiridol; Fig. 3a), which inhibits CDK1, 2, 4,
6,7and9(Table2)[87–90]. Between 2008 and 2014,
alvocidib was evaluated in more than 60 clinical trials for
numerous tumour types [91]. Limited clinical activity was
seen in the majority of trials, however, modest responses
against chronic lymphocytic leukaemia (CLL) [92,93]and
mantle cell lymphoma [94] were shown. Currently, alvocidib,
marketed as a CDK9 inhibitor, is being trialled by Tolero
Pharmaceuticals for the treatment of acute myeloid leukaemia
(AML) (Clinicaltrials.gov identifiers: NCT03298984;
NCT03969420; NCT02520011). Another early pan-CDK in-
hibitor, the purine-based seliciclib (roscovitine; Fig. 3a),
which inhibits CDK1, 2, 5, 7, and 9 (Table 2)[95–97], was
also assessed in clinical trials for a variety of tumour types but,
likewise, showed limited clinical activity [91,98].
809Cancer Metastasis Rev (2020) 39:805–823
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Attempts to develop CDK inhibitors with improved selec-
tivity for CDK1 and CDK2 led to a second generation of
multi-target CDK inhibitors, including the aminothiazole-
based compound, SNS-032 (Fig. 3a), which potently inhibits
CDK2, 7 and 9 (Table2)[85,91,99,100]. Although SNS-032
has been trialled for the treatment of advanced lymphoid [101]
and advanced solid malignancies [102], the drug has not
progressed further than Phase I [91].
The inability of these early CDK inhibitors to selectively
target individual CDK family members probably contributed
to their failure in the clinic. As several CDK proteins are
critical for the function of normal tissues, the promiscuity of
Alvocidib Seliciclib SNS-032
YKL-5-124
SY-1365
ICEC0942
LDC4297
LY3405105
BS-181
a
b
THZ2
THZ1
Fig. 3 Chemical structures of selected inhibitors that target CDK7. Chemical structures of non-specific inhibitors of CDK7 (a) and selective inhibitors of
CDK7 (b). (The chemical structures of QS1189 and SY-5609 have not been disclosed)
gene
mRNA
enhancer
Super-enhancer
Normal
CDK7iCDK7i
Cancer
CDK7i CDK7i
CDK7i
ab
Fig. 2 Super-enhancer-driven genederegulation in cancer can be targeted
by CDK7 inhibitors. The super-enhancer landscape in normal cells (a)
becomes deregulated in cancer (b), leading to altered gene expression.
CDK7 inhibitors preferentially reduce gene expression driven by super-
enhancers in cancer cells compared with normal cells (A and B). CDK7i
= CDK7 inhibitor
810 Cancer Metastasis Rev (2020) 39:805–823
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these compounds likely limits their ability to discern cancer
cells from normal cells, resulting in a narrow therapeutic win-
dow and associated toxicities, which include fatigue, diar-
rhoea, nausea and hyperglycaemia [91,102–105]. In addition,
their lack of specificity makes it difficult to decipher which
CDKs are inhibited in vivo, and which are most important for
their underlying mechanism of action [91]. This paucity of
knowledge limits the potential to develop these pan-CDK in-
hibitors further as targeted therapies.
More recent efforts have focused on further improving the
selectivity of CDK inhibitors, with selective CDK4/6 inhibi-
tors proving the biggest success story to date. Three CDK4/6
inhibitors have been approved for the treatment of hormone
receptor (HR)-positive metastatic breast cancer: palbociclib
(PD0332991; Ibrance), ribociclib (LEE011; Kisquali) and
abemaciclib (LY2835219; Verzenio), in combination with
aromatase inhibitors or fulvestrant [106]. More than one hun-
dred clinical trials for CDK4/6 inhibitors in breast, but also in
other cancers, including glioma, sarcoma, lung, pancreatic,
head and neck, colorectal, prostate and ovarian cancer, are
actively recruiting patients or about to initiate. The success
of these selective CDK4/6 inhibitors is encouraging and pro-
vides some confidence that selective inhibitors of other CDKs
may prove similarly successful.
4.2 CDK7-specific inhibitors
A number of selective small molecule inhibitors of CDK7
have been developed. These include the pyrazolopyrimidine
derivatives, BS-181 [39] and ICEC0942 [22,107], and the
pyrazolotriazine derivatives, LDC4297 [108] and QS1189
[109](Fig.3b,Table3). These are type I inhibitors that bind
reversibly to the ATP-binding site of CDK7. ATP-competitive
covalent inhibitors of CDK7 have also been developed, in-
cluding the pyrimidine based THZ1 [20] and SY-1365 [23]
and the pyrrolidinopyrazole based YKL-5-124 [130](Fig.3b,
Table 3).
The first example of a highly selective CDK7 inhibitor was
BS-181, which is structurally related to the pan-CDK inhibitor
roscovitine (Fig. 3b, Table 3)[39]. BS-181 reduced phosphor-
ylation of CDK7 targets and impaired cancer cell line and
xenograft tumour growth, establishing CDK7 as a putative
cancer drug target [39]. Although in vivo activity was demon-
strated, poor bioavailability and insufficient cell permeability
precluded the development of BS-181 as a clinical candidate
[39].
Efforts to develop BS-181 analogues which retain CDK7
selectivity, but have improved drug-like properties, led to the
first orally bioavailable CDK7 inhibitor, ICEC0942 (CT7001;
Fig. 3b, Table 3)[22,107]. Although crystal structures of
CDK7 bound to ICEC0942 could not be obtained, a crystal
structure of CDK2 in complex with ICEC0942 was solved
[107]. Using this structure as a starting point, modelling stud-
ies revealed aspartate 155 (Asp155) as a residue that is likely
key in determining the selective binding of ICEC0942 to
CDK7 [107]. ICEC0942 potently inhibited the growth of a
panel of cancer cell lines and of ER+ breast cancer xenografts,
and its favourable absorption, distribution, metabolism, and
excretion (ADME) and pharmacokinetic (PK) properties
made ICEC0942 a promising clinical candidate [22]. The drug
was licenced to Carrick Therapeutics and is now in Phase I/II
clinical trial for advanced solid malignancies, with focused
cohorts of breast and prostate cancer patients (Table 4).
A number of covalent CDK7 inhibitors have also been
developed, the first being THZ1 (Fig. 3b, Table 3), which
targets a cysteine residue (Cys312) on a C-terminal extension
just outside the ATP-binding site of CDK7 [20,132] and has
strong activity in many cancer types (Table 3)[20,21,71,75,
114–124]. However, THZ1 also covalently links to CDK12
and CDK13, at Cys1039 and Cys1017, respectively,
inhibiting their activity [20,132]. THZ1 has been widely
employed as a tool to interrogate CDK7 function [50,51];
however, it was recently shown that its anti-transcriptional
and antitumour activities are reliant on inhibition of CDK12
Table 2 Characteristics of selected multi-target CDK7 inhibitors (see ref [86])
Name(s) Company
a
IC
50
(nM)
b
Development
phase reached
Alvocidib (flavopiridol) Tolero Pharmaceuticals
(Sanofi-Aventis)
CDK1-CycB= 41; CDK2-CycA = 100; CDK4-CycD= 65;
CDK6-CycD= ~100; CDK7-CycH = ~300; CDK9-CycT = 6
Phase II
Seliciclib (roscovotine;
CYC202)
Cyclacel (ManRos
Therapeutics)
CDK1-CycB= 2700; CDK2-CycE = 100; CDK4-CycD1 > 10,000;
CDK6-CycD1> 100,000; CDK7-CycH = 490; CDK9-CycT = 600
Phase II
SNS-032 Sunesis (Bristol-Myers
Squibb)
CDK1-CycB= 480; CDK2-CycA = 38; CDK4-CycD= 92;5
CDK6-CycD>1000; CDK7-CycH = 62; CDK9-CycT = 4
Phase I
a
Current developer (previous developer in brackets)
b
Data from in vitro kinase assays for CDKs 1, 2, 4, 6, 7, 9 and 12 have been listed,where available. Where data are available for a CDK in multiple cyclin
complexes, the complex with the lowest IC
50
is presented
811Cancer Metastasis Rev (2020) 39:805–823
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

Table 3 Characteristics of key CDK7 inhibitors in development
Name(s) Company Type of
inhibitor
IC
50
(nM)
a
Activity in preclinical models Combination agents tested Current development
phase (clinical Trial
ID)
BS-181 [39] - Non-covalent CDK1-CycB = 8100; CDK2-CycE = 880;
CDK4-CycD1 = 33,000; CDK6-CycD1 = 47,000;
CDK7-CycH-Mat1 = 21; CDK9-CycT = 4200
ER+ breast cancer [39], gastric cancer
[110], papillary thyroid cancer [111]
-–
ICEC0942 [22]
(CT7001)
CarrickTherapeutics Non-covalent CDK1-CycA1 = 1800; CDK2-CycA1 = 620;
CDK4-CycD1 = 49,000; CDK6-CycD1 = 34,000;
CDK7-CycH-MAT1= 40; CDK9-CycT1 = 1200
ER+ breast cancer [22], AML [112] Fulvestrant, tamoxifen [22]Phase
I/II (NCT03363893)
LY3405105 [26] Eli Lilly and Company - CDK1-CycB1 = 20,000; CDK2-CycE1 = 20,000;
CDK4-CycD1 = 2830; CDK6-CycD1 = 8079;
CDK7-CycH-Mat1 = 92.8; CDK9-CycT1 = 6320;
CDK12-CycK = 14,780
- - Phase I (NCT03770494)
LDC4297 [108] Lead Discovery Center
GmbH
Non-covalent CDK1-CycB = 54; CDK2-CycE = 6.4; CDK4-CycD ≥1000;
CDK6-CycD > 1000; CDK7-CycH-MAT1 < 5;
CDK9-CycT = 1711
HCMV antiviral activity [113]- -
QS1189 [109] Qurient Therapeutics Non-covalent CDK1-CycE1 = 690; CDK2-CycE1 = 270;
CDK4-CycD1 = 3700; CDK6-CycD1 = 6200;
CDK7-CycH-MAT1= 15; CDK9-CycK = 710;
CDK12-CycK = 570
Mantle cell lymphoma, Burkitt’s
lymphoma, DLBCL [109]
--
SY-5609 [24] Syros Pharmaceuticals Non-covalent CDK2-CycE1 = 2900
c
; CDK7-CycH-MAT1 = 0.06
b
;
CDK9-CycT1 = 970
c
; CDK12-CycK = 770 nM
c
ER+ breast cancer [25], ovarian cancer
[24], TNBC [24]
Fulvestrant [25]-
SY-1365 [23] SyrosPharmaceuticals Covalent CDK2-CycE1 = 2117; CDK7-CycH-MAT1 = 84;
CDK9-CycT1 = 914; CDK12-CycK = 204
AML [23] Venetoclax [23] Phase I (NCT03134638)
THZ1 [85] (SY-079) Syros Pharmaceuticals Covalent CDK7-CycH = 3.2 T-ALL [20], neuroblastoma [21], SCLC
[114], OSCC [71], PTCL [115],
ovarian cancer [75], DIPG [116], HGG
[117], melanoma [118], hepatocellular
carcinoma [119], thyroid cancer [120],
pancreatic cancer [121], cervical cancer
[122], TNBC [123], multiple myeloma
[124]
Fulvestrant [125], JQ1 [75,116,126],
panobinostat [116],
carfilzomib/bortezomib [124],
venetoclax [124]/navitoclax [127], 5-
-fluorouracil, nutlin-3 [128]
-
THZ2 [123] Syros Pharmaceuticals Covalent CDK1-CycB = 97; CDK2-CycA = 222; CDK7-CycH = 14;
CDK9-CycT = 194
TNBC [123], gastric cancer [129]- -
YKL-5-124 [130] Syros Pharmaceuticals Covalent CDK7-CycH-MAT1 = 9.7; CDK2-CycA = 1300;
CDK9-CycT1 = 3020
Mantle cell lymphoma [130] anti-PD-1+chemotherapy [131]-
a
Data from in vitro kinase assays for CDKs 1, 2, 4, 6, 7, 9 and 12 have been listed, where available. Where data are available for CDKs in complex with multiple cyclins, the complex with the lowest IC
50
is
presented.
b
K
d
determined by SPR.
c
K
i
determined by activity assay
812 Cancer Metastasis Rev (2020) 39:805–823
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Table 4 Summary of clinical trials investigating CDK7 inhibitors in cancer
Drug name(s) Clinical trial
ID
Dates Administration Trial type Trial design No. of
patients
Combination
agent
Results
ICEC0942
(CT7001)
NCT03363893 Nov
2017–March
2021
Orally once daily Modular
Phase I/II
Module
1A
Dose-escalation/safety advanced solid tumours 39 MBAD = 120 mg
once daily
Module
1B
Refine dose-escalation/safety—up to 4 cohorts: MTD =360 mg
once daily
Locally advanced or metastatic TNBC Up to 50
Castrate-resistant prostate cancer Up to 25
Additional cohorts (may include ovarian and SCLC) Up to 25
Module
2
Phase Ib/II safety and efficacy
Locally advanced or metastatic HR+HER2—breast
cancer
Up to 75 Fulvestrant
LY3405105 NCT03770494 Jan 2019–May
2022
Orally Phase Ia/Ib Safety advanced or metastatic solid tumours Up to 215
SY-1365 NCT03134638 May 2017–Nov
2019
Intravenously
once/twice weekly
Phase I
(2 parts)
Part 1 Dose-escalation/safety advanced solid tumours ~ 35
Part 2 Refine safety and test efficacy—5 cohorts:
Ovarian cancer treated with ≥3 prior lines of therapy ~ 24
Relapsed ovarian cancer with previous platinum
therapy
~ 24 Carboplatin
Primary platinum refractory ovarian cancer ~ 12
Biopsy-accessible advanced solid tumours 20–30
HR+metastatic breast cancer post CDK4/6 + aroma-
tase inhibitor treatment
~ 12 Fulvestrant
SY-5609 NCT04247126 Jan 2020–Jun
2021
Orally Phase I Dose-escalation select advanced solid tumours 60
HER2 human epidermal growth factor receptor 2, HR hormone receptor, MBAD minimum biologically active dose, MTD maximum tolerated dose, SCLC small cell lung cancer, TNBC triple-negative
breast cancer
813Cancer Metastasis Rev (2020) 39:805–823
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

and CDK13, in addition to CDK7 [130]. Consequently, YKL-
5-124 was developed, with a strategy that combined the co-
valent warhead of THZ1 with the pyrrolidinopyrazole core of
the PAK4 inhibitor, PF-3758309 [130]. Like THZ1, YKL-5-
124 covalently links to Cys312 of CDK7 but does not affect
the activities of CDK12 and 13 (Fig. 3b, Table 3)[130]. An
analogue of THZ1, with altered regiochemistry of the acryl-
amide and increased in vivo stability has also been developed
and was designated THZ2 (Fig. 3b, Table 3)[123].
To improve on the potency, selectivity and metabolic sta-
bility of THZ1, the THZ1-derived CDK7 inhibitor, SY-1365,
was developed by Syros Pharmaceuticals as a candidate for
clinical development [23]. SY-1365 entered Phase I clinical
trial for the treatment of advanced solid tumours, with planned
expansion cohorts focusing on ovarian cancer and breast can-
cer (Fig. 3b, Table 3and 4). However, Syros Pharmaceuticals
recently announced discontinuation of the clinical develop-
ment of SY-1365 and the prioritisation of a new, orally avail-
able CDK7 inhibitor, SY-5609 (Table 3), with greater selec-
tivity and potency for CDK7 [24,25]. SY-5609 has
antitumour activity in preclinical models of ovarian cancer
[24,25], TNBC [24,25] and ER+ breast cancer, in combina-
tion with fulvestrant [25], and sustained tumour regressions
were associated with alterations in the RB pathway [25]. A
Phase I trial, in patients with select advanced solid tumours,
began in early 2020 (Table 4).
Another CDK7 inhibitor, LY3405105, developed byEli Lilly,
is also undergoing clinical testing for advanced or metastatic
solid cancers [26](Fig.3b,Table3and 4). Little information
on LY3405105 has been released; however, selectivity data from
the corresponding patent (WO2019099298) is listed in Table 3.
5 Inhibiting CDK7 in cancer
Due to the importance of CDKs in regulating cell prolifera-
tion, and the deregulation of CDK pathways in many cancer
types, CDKs have long been considered important targets for
the design of cancer therapeutics [39]. The early pan-CDK
inhibitors, alvocidib and seliciclib, cause cell cycle arrest
and apoptosis, as well as altered expression of genes in these
pathways [85]. Seliciclib was also shown to reduce Pol II
CTD phosphorylation and Pol II-dependent transcription in
myeloma cells [133]. Whilst it is likely that some cellular
actions of these inhibitors are mediated through CDK7, their
lack of selectivity made it difficult to distinguish CDK7-
specific effects and ultimately led to the numerous side effects
that resulted in their failure in the clinic. The recently devel-
oped, highly specific inhibitors of CDK7 have been instru-
mental in revealing the potential of CDK7 as a cancer drug
target. Xenograft studies in mice showed that CDK7 inhibitors
are well tolerated and effective at reducing tumour growth
in vivo [21–23,39,114].
A number of reversible and covalent inhibitors of
CDK7 have been tested on large panels of cancer cell
lines [20,22,23,39].ScreeningofICEC0942against
the NCI-60 cancer cell line panel demonstrated a medi-
an GI
50
value of 250 nM [22]. Of over 1000 cancer cell
lines tested with THZ1, around half had a GI
50
value
under 200 nM [20]. Whilst CDK7 inhibitors are potent
at impairing the growth of many cancer cell lines,
representing a variety of tumour types, it is clear that
some cell lines respond more favourably than others.
Responses of 386 human cell lines, encompassing 26
cancer types, to the covalent CDK7 inhibitor SY-1365,
revealed varied responses ranging from cytostatic to
highly cytotoxic [23]. Expression levels of the anti-
apoptotic protein BCL-XL were predictive of SY-1365
response, with low BCL-XL expression associated with
high SY-1365 sensitivity [133]. It is clear that additional
features associated with response to CDK7 inhibition
remain to be discovered and this knowledge will likely
be beneficial for their future clinical success.
5.1 Effects on cell cycle progression
As CDK7 directs cell cycle progression, via the activation of
other CDK proteins, it is unsurprising that CDK7 inhibitors
reduce phosphorylation of cell cycle CDKs and consequently
cause cell cycle arrest [21,22,39,108,109,130]. CDK7
inhibition with ICEC0942 blocks progression at all stages of
the cell cycle and is associated with a reduction in phosphor-
ylation of CDK1, CDK2 and RB [22]. YKL-5-124 primarily
causes G1 arrest, with a reduction in CDK2 phosphorylation
[130], whereas THZ1 and QS1189 both arrest cells at G2/M
[21]. Interestingly, the extent and timing of cell cycle arrest
upon treatment with an individual CDK7 inhibitor can vary
among cell lines; A549 lung cancer cells, treated with
LDC4297, arrested in G1, whereas HCT116 colon cancer
cells exhibited a G2/M delay, only after an extended incuba-
tion period [108]. Again, it is apparent that factors influencing
the effect of CDK7 inhibition on cell cycle progression across
different cancers remain to be identified and these may have
an important impact on the clinical use of these inhibitors. In
addition to cell cycle arrest, apoptosis is observed following
CDK7 inhibition, in numerous cancer types, including solid
tumours [21–23,39,108,114] and haematological malignan-
cies [20,109]. YKL-5-124 is unique among the current CDK7
inhibitors, in that it causes cell cycle arrest at both G1 and G2/
M in the apparent absence of apoptosis [130].
5.2 Effects on transcription
The majority of CDK7 inhibitors, again, with YKL-5-124
being an exception, reduce Pol II CTD phosphorylation and
cause widespread alterations in Pol II-mediated transcription
814 Cancer Metastasis Rev (2020) 39:805–823
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

[20,22,23,39,108,109,130]. As CDK7 is dispensable for
global transcription [58], it makes sense that rather than reduc-
ing gene expression globally, only subsets of genes are down-
regulated by CDK7 inhibitors. Pathway analyses suggest that
cell cycle and DNA damage repair pathways are enriched in
gene sets altered by CDK7 inhibition [23,109,114].
Additionally, smaller subsets of genes are actually upregulated
following CDK7 inhibition [21,108]. For example, around
half of the 2% of genes whose expression was affected by
short-term LDC4297 treatment were upregulated [108].
mRNAs have differing half-lives, and it has been noted that
those with short half-lives are preferentially downregulated by
CDK7 inhibitors [108]. One explanation for the counterintui-
tive observation that some genes are upregulated following
CDK7 inhibition is that these represent a subset that are neg-
atively regulated by genes whose transcripts have short half-
lives. It is also possible that CDK7 plays a direct and crucial
role in regulating the transcription of specific subsets of genes,
possibly also repressing the expression of some genes, either
in a direct or an indirect fashion. One clue that may go some
way to explaining how the transcription of certain genes may
escape CDK7 inhibition comes from the work of Shandilya
et al. [134]. They showed that CDK7 phosphorylates the gen-
eral transcription factor, TFIIB (Table 1), and this is required
for the transcription of certain genes, but not for transcription
of p53 target genes, which escape CDK7 inhibition.
Numerous studies have shown that SE-associated genes
are preferentially downregulated in cancer cells treated
with CDK7 inhibitors (Fig. 2)[20,21,71–73,75,114,
116,118,123,135,136]. In T cell acute lymphoblastic
leukaemia (ALL), expression of the oncogenic TF,
RUNX1, is driven by a large SE and is disproportionately
repressedbyTHZ1treatment[20]. Similarly, neuroblasto-
mas driven by MYCN amplification, which promotes the
formation of aberrant SEs, are selectively sensitive to
CDK7 inhibition [21]. As MYC TFs are frequently upreg-
ulated in cancer but have proven difficult to target direct-
ly, blocking MYC expression and/or targeting the tran-
scription machinery downstream of MYC, by inhibiting
CDK7, is an attractive strategy. Similarly, oncogenic
ETS TFs, which are also notoriously difficult to target,
are reduced by THZ1 treatment in prostate cancer [137].
Other cancers in which THZ1-mediated downregulation
of SE-associated genes has been demonstrated include
ovarian cancer [75], melanoma [118] and small cell lung
cancer [114]. These studies have been integral for
uncovering the aforementioned phenomenon of transcrip-
tional addiction in cancer (Fig. 2). Furthermore, targeting
specific oncogenic transcriptional programs provides
some explanation for the observation that cancer cells
are more vulnerable to transcriptional inhibition than nor-
mal cells.
5.3 CDK7 inhibitors to treat drug-resistant cancers
CDK7 inhibition represents a novel strategy to treat cancers
with de novo or acquired resistance to other drugs, where
further treatment options are limited. Mutations in the ESR1
gene are common in advanced ER+ breast cancer, causing
oestrogen-independent receptor activation and hormone ther-
apy resistance [125]. CDK7 is an essential gene in both ER-
wild-type and ER-mutant breast cancer, and ER-mutant
MCF7 cells, that are partially resistant to anti-oestrogens, are
sensitive to CDK7 inhibition [125,138]. Furthermore, activat-
ing Ser118 phosphorylation of mutant ER is inhibited by
THZ1 [125,138]. This suggests that CDK7 inhibitors may
be effective at treating advanced, ER-mutant breast cancer.
THZ1 has also been shown to overcome HER2 inhibitor re-
sistance in breast cancer [139] and venetoclax resistance in
mantle cell lymphoma [109], and to inhibit castration-
resistant prostate cancer [137].
A number of CDK4/6 inhibitors are clinically approved for
use in combination with endocrine therapies for the treatment
of ER+ breast cancer, however, resistance to these drugs is an
emerging problem [140]. Breast cancer cells with acquired
resistance to palbociclib remain sensitive to THZ1 [140], sug-
gesting that CDK7 inhibitors may be useful following the
onset of resistance to drugs that target other CDKs.
Current clinical trials of CDK7 inhibitors are aimed at pa-
tients with advanced or metastatic cancer; therefore, a majority
of these will have received other lines of therapy prior to their
recruitment. The SY-1365 trial was designed to target specific
cohorts of patients with resistance to prior treatments, includ-
ing platinum resistance in ovarian cancer and CDK4/6 inhib-
itor plus aromatase inhibitor resistance in HR+ breast cancer
(Table 4). The prospect of using CDK7 inhibitors to overcome
resistance to prior treatments in cancer is exciting, and it is
crucial that information garnered from preclinical studies of
CDK7 inhibitors in the context of acquired drug-resistance
continues to be considered during clinical trial design.
5.4 Combination treatment strategies
Cancers are frequently treated with two or more therapeutic
agents simultaneously. Compared with single-agent treat-
ments, these combination therapies often have enhanced effi-
cacy, can delay the onset of resistance and may allow lower
doses of individual drugs to be used, thus reducing toxicity.
Multiple studies have investigated the potential of combining
CDK7 inhibitors with other anticancer drugs [22,23,116,
124–127].
As ER is the key transcriptional driver of ER+ breast cancer
and is activated by CDK7, CDK7 inhibitors have been
assessed in combination with anti-oestrogens in this context
(Table 3)[22,125]. In the ER+ breast cancer cell line, MCF7,
treatment with ICEC0942, plus either tamoxifen or
815Cancer Metastasis Rev (2020) 39:805–823
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

fulvestrant, caused greater growth inhibition than either agent
alone [22]. The combinatorial action of ICEC0942 with ta-
moxifen was also verified in mice bearing MCF7 xenografts
[22]. In addition, ER-mutant breast cancer cells are especially
sensitive to the combination treatment of THZ1 and
fulvestrant [125]. ICEC0942 is now being assessed in combi-
nation with fulvestrant in clinical trials (Table 4).
It has been noted that some CDK7 inhibitors reduce ex-
pression of the anti-apoptotic BCL2 family member, MCL1
[23,124,127]; therefore, it is plausible that CDK7 inhibitors
will work synergistically with apoptotic agents. CDK7 inhib-
itors have been investigated in combination with the
apoptosis-inducing BH3-mimetics, venetoclax (ABT-199)
[23,124] and navitoclax (ABT-263) (Table 3)[127].
Venetoclax, which inhibits anti-apoptotic BCL2 to cause ap-
optosis, is approved for the treatment of CLL, small lympho-
cytic leukaemia (SLL) and AML [23]. The combination of
SY-1365 and venetoclax is synergistic in AML cell lines and
xenografts [23]. Although the current clinical trials of CDK7
inhibitorsare focused on solid tumours, these results highlight
the potential for CDK7 inhibitors to combat blood cancers,
particularly if combined with BH3-mimetics. CDK7 inhibi-
tion also synergises with p53-activating agents, including the
chemotherapeutic, 5-fluorouracil, to induce apoptosis in colo-
rectal cancer cells [128].
BET inhibitors, compounds that target the
bromodomain extra-terminal (BET) family of proteins,
have garnered much recent interest as potential cancer ther-
apeutics and are being assessed clinically across a range of
cancers. The preclinical and clinical advancement of BET
inhibitors in cancer therapy has recently been reviewed
[141]. BET inhibitors are exemplified by the tool com-
pound JQ1, which, like THZ1, preferentially represses
SE-driven transcription and can be used to overcome on-
cogenic transcriptional addiction in cancer [19]. For this
reason, THZ1 has been tested alongside JQ1 (Table 3),
and this combination synergistically inhibits the growth
of diffuse intrinsic pontine glioma (DIPG) [116], ovarian
cancer [75] and neuroblastoma [126]. THZ1 has also been
investigated alongside the histone deacetylase (HDAC) in-
hibitor, panobinostat (Table 3), for the treatment of the
universally fatal paediatric cancer DIPG [116]. Like JQ1
and THZ1, panobinostat supresses transcription of SE-
associated genes in DIPG and acts synergistically with
CDK7 inhibition to suppress growth of DIPG-derived cell
lines [116]. However, the development of CDK7 inhibitors
with adequate brain penetrance would be required before
this combination therapy could be achieved in the clinic.
Recent work has highlighted the potential of CDK7 inhib-
itors to be used in combination with immunotherapies. The
CDK7 inhibitor, YKL-5-124, was shown to elicit immune
response signalling in small cell lung cancer (SCLC), activat-
ing anti-tumourigenic T cells [131]. In immunocompetent
mouse models of SCLC, the combination of YKL-5-124 and
anti-PD-1 immune checkpoint inhibition increased overall
survival in comparison with either treatment alone, and was
further enhanced by the addition of chemotherapeutics (cis-
platin and etoposide) [131].
Overall, there is much potential for CDK7 inhibitors to be
used in combination with other drugs for cancer therapy. We
anticipate that screening of CDK7 inhibitors in combination
with other compounds, and large-scale genomic perturbation
studies, may pave the way for the identification of additional
co-targeting strategies.
5.5 Resistance to CDK7 inhibitors
The emergence of resistance to cancer treatment, including
targeted therapies, remains a major issue, and it is possible
that even if CDK7 inhibitors prove a clinical success, resis-
tance may develop in some patients. To gain understanding of
potential resistance mechanisms, cell lines with acquired re-
sistance to both ICEC0942 and THZ1 have been developed
[142,143].
ATP-binding cassette transporters (ABC-transporters) are a
well-characterised mechanism of multidrug resistance in can-
cer and, when upregulated, can mediate the ATP-dependent
efflux of drugs that are substrates. Upregulation of ABCB1,
also known as p-glycoprotein, mediates resistance to THZ1 in
neuroblastoma and lung cancer [142] and resistance to both
THZ1 and ICEC0942 in breast cancer cell lines [143].
Increased expression of another ABC-transporter, ABCG2,
results in resistance to THZ1 [142], but not to ICEC0942
[143]. With no clinical data available as yet, it remains to be
seen whether ABC-transporter upregulation will arise in the
context of CDK7 inhibitor resistance in patients. Despite nu-
merous clinical trials, ABC-transport inhibitors have thus far
proven unsuccessful in the clinic, mainly due to issues with
potency and toxicity [144]. However, should future develop-
ments be made in this area, there may be scope to use
CDK7 inhibitors in combination with ABC-transport inhibi-
tors. Another way to overcome this mechanism of resistance is
the development of CDK7 inhibitors that are not substrates for
multidrug resistance transporters [145]. It is likely that other,
as yet unidentified, mechanisms of resistance will also be im-
portant in a clinical setting.
As clinical trials progress, analyses of tumour features
enriched in subsets of patients that are intrinsically
resistant to CDK7 inhibitors, or those who acquire resistance
after initially responding well, should help shed light on mech-
anisms of CDK7 inhibitor resistance. Alongside these, further
preclinical models of CDK7 inhibitor resistance, including 3D
cell culture and in vivo models, may prove informative.
Ultimately, the identification of mechanisms of CDK7 inhib-
itor resistance should aim to aid the identification of patients
816 Cancer Metastasis Rev (2020) 39:805–823
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

who will derive most benefit from these drugs, helping to
advance their clinical progress.
6 Conclusions
CDK7 has a dual role in driving the cell cycle and transcrip-
tion, is upregulated in a variety of cancers and has emerged as
a promising cancer therapeutic target. At least ten selective
inhibitors of CDK7, with activity against a wide range of
cancer types, have been developed, their antitumour action
likely mediated both through cell cycle arrest and inhibition
of oncogenic transcriptional programs. In the preclinical set-
ting, these inhibitors have demonstrated potential to overcome
treatment-resistant cancer, both asmonotherapies, and in com-
bination with other cancer drugs. To date, four CDK7 inhibi-
tors have progressed to Phase I/II clinical trial for the treatment
of advanced solid malignancies. Whilst ABC-transporters can
mediate resistance to some CDK7 inhibitors, additional fac-
tors that influence tumour response to CDK7 inhibition are yet
to be identified. Further efforts to elucidate mechanisms of
response, and to define patient selection strategies, will help
to facilitate the clinical utility of CDK7 inhibitors.
Funding information The authors’work is generously funded by Cancer
Research UK (grant C37/A18784). Additional support was provided by
the Imperial Experimental Cancer Medicine Centre, the Imperial NIHR
Biomedical Research Centre and the Cancer Research UK Imperial
Centre. The views expressed are those of the authors and not necessarily
those of the NHS, the NIHR or the Department of Health.
Compliance with ethical standards
Conflict of interest RCC and SA are named as inventors on CDK7
inhibitor patents and own shares in Carrick Therapeutics.
Open Access This article is licensed under a Creative Commons
Attribution 4.0 International License, which permits use, sharing, adap-
tation, distribution and reproduction in any medium or format, as long as
you give appropriate credit to the original author(s) and the source, pro-
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