Therapeutics Targeting FGF Signaling Network in Human Diseases

TIPS 1376 No. of Pages 16

Review

Therapeutics Targeting FGF Signaling Network in Human Diseases Masaru Katoh1,* Fibroblast growth factor (FGF) signaling through its receptors, FGFR1, FGFR2, FGFR3, or FGFR4, regulates cell fate, angiogenesis, immunity, and metabolism. Dysregulated FGF signaling causes human diseases, such as breast cancer, chondrodysplasia, gastric cancer, lung cancer, and X-linked hypophosphatemic rickets. Recombinant FGFs are pro-FGF signaling therapeutics for tissue and/or wound repair, whereas FGF analogs and gene therapy are under development for the treatment of cardiovascular disease, diabetes, and osteoarthritis. FGF traps, anti-FGF/FGFR monoclonal antibodies (mAbs), and smallmolecule FGFR inhibitors are anti-FGF signaling therapeutics under development for the treatment of cancer, chondrodysplasia, and rickets. Here, I discuss the benefit–risk and cost-effectiveness issues of precision medicine targeting FGFRs, ALK, EGFR, and FLT3. FGFR-targeted therapy should be optimized for cancer treatment, focusing on genomic tests and recurrence.

Trends Gain- and loss-of-function alterations in FGF signaling molecules are involved in the pathogenesis of human diseases. The landscape of the genomic alterations in FGF signaling molecules in human diseases has been clarified by next-generation sequencing technologies. Small-molecule compounds, human mAbs, and FGF/FGFR analogs are under development as pro- or antiFGF signaling therapeutics for human diseases. Precision medicine, which utilizes large amounts of multilayered data to prevent and treat human diseases, is currently transforming medical practice. Diagnostics and therapeutics related to FGF signaling cascades are key components of precision medicine for the treatment of cancers, cardiovascular disease, and diabetes.

Overview of FGF Signaling FGFs are classified as either paracrine FGFs that bind to heparin-sulfate proteoglycans and FGFRs, or endocrine FGFs that bind to Klotho family proteins and FGFRs [1–3]. FGF induces the dimerization, activation, and tyrosine phosphorylation of FGFRs and the subsequent phosphorylation of FGFR substrate 2/ (FRS2/) and phospholipase Cg (PLCg), which leads to the activation of the RAS–extracellular signal-regulated kinase (ERK), phosphatidylinositol 3-kinase (PI3K)–AKT, inositol-1,4,5-trisphosphate (IP3)–Ca2+, and diacylglycerol (DAG)–protein kinase C (PKC) signaling cascades in a cellular context-dependent manner (Figure 1, Key Figure). FGFs regulate self-renewal, metabolism, survival, proliferation, the differentiation and epithelial-tomesenchymal transition (EMT) of their target cells, angiogenesis, and immunity in their microenvironment, and homeostasis in the whole body [1–3]. FGFRs were initially cloned and characterized as receptor tyrosine kinases (RTKs) that comprise extracellular ligand-binding domains, a transmembrane domain, and a cytoplasmic tyrosine kinase domain [4]. FGFRs have been discovered to have somatic alterations in human cancers [5], and germline mutations in craniosynostosis syndromes and skeletal dysplasias [6,7]. The role of FGF signaling cascades in angiogenesis and immunity are hot issues in translational oncology [8,9], because a ‘leaky’ hypoxic microenvironment, in which tumor vasculature accumulates more molecules in the bloodstream, and immune evasion, in which tumors are not detected by the immune system of the host, are involved in the therapeutic resistance and recurrence of cancers [10,11]. FGF signaling dysregulation is also implicated in other disorders; for example, it is involved in the progression of cardiovascular disease [12,13]. Thus, FGFs and FGFRs are emerging as promising targets for the treatment of cancers [1,2] and noncancerous diseases [14,15]. Here, I briefly summarize the genomic alterations of FGFRs and review recent

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

Increasing costs for diagnosis and prescription are emerging as serious issues for the implementation and maintenance of precision medicine.

1

Department of Omics Network, National Cancer Center, Tokyo 1040045, Japan

*Correspondence: [email protected] (M. Katoh).

http://dx.doi.org/10.1016/j.tips.2016.10.003 © 2016 Elsevier Ltd. All rights reserved.

1

TIPS 1376 No. of Pages 16

Key Figure

Overview of Fibroblast Growth Factor (FGF) Signaling and FGF Receptor (FGFR) Alterations in Human Diseases Germline LoF alteraons

Paracrine FGF

Endocrine FGF

FGFRs

FGFR1 Mut [21] HSPG

Hypogonadotropic hypogonadism 2 (HH2) (V102fs, S107X, E324X, Y585X, R622X, T657fs, R661X, etc.)

Klotho

Germline GoF alteraons FGFR1 Mut [6,16,17,19] Osteoglophonic dysplasia (Y372C) Pfeiffer syndrome (P252R) FGFR2 Mut [6,16,17,19] Antley-Bixler syndrome (S351C) Apert syndrome (S252W, P253R) Beare-stevenson syndrome (S372C, Y375C) Crouzon syndrome (C278F, W290R, Y340H/N, C342W/Y, G384R, etc.) Pfeiffer syndrome (Y340C, C342R/S/Y)

FGFR2 Mut [22] Lacrimo-auriculo-dento-digital (LADD) syndrome (A628T, A648T, R649S)

FGFR2 Del [25] FRS2α

PLCγ

10q26.1 Microdeleon syndrome

FGFR4 – NSD1 Del [27] Sotos syndrome associated with nephrocalcinosis or infanle hypercalcemia

RAS

PI3K

IP3

DAG

ERK

AKT

Ca2+

PKC

Somac LoF alteraons

FGFR3 Mut [7,18,19] Achondroplasia (G375C, G380R) Crouzon syndrome with acanthosis nigricans (A391E) Hypochondroplasia (N540K) Muenke syndrome (P250R) PLSD-SD (R248C, Y373C) SADDAN (K650M) Thanatophoric dysplasia (R248C, S249C, G370C, S371C, Y373C, K650E/M)

Mosaic GoF alteraons

FGFR2 Mut [40] Melanoma (D530N, I642V, A648T)

Self-renewal

Metabolism

Survival

EMT

Proliferaon

Angiogenesis

Differenaon

Immunity

Whole-body homeostasis

FGFR1 Mut [28] Encephalocraniocutaneous lipomatosis (ECCL) (N546K, K656E) FGFR2 Mut [29] Blaschko line acne on pre-existent hypomelanosis (P253R) FGFR3 Mut [30] Keranocyc epidermal naevus syndrome (KENS) (R248C, S249C)

Somac GoF alteraons FGFR1 Amp [1,2,31,32,34,35] Breast cancer (ER+) Gastric cancer Lung cancer (SCC, SC) Ovarian cancer Urothelial cancer

FGFR1 Fus [36] Glioblastoma (FGFR1-TACC1) MPN (BCR-FGFR1, CNTRL-FGFR1, ZMYM2-FGFR1, etc.) FGFR1 Mut [37,38] Ewing sarcoma (N546K) Glioblastoma (N546K, K656E)

FGFR2 Amp [1,2,34] Breast cancer (TNBC) Gastric cancer FGFR2 Fus [36] Breast cancer (FGFR2-AFF3, FGFR2-CASP7) Cholangiocarcinoma (FGFR2-BICC1, FGFR2-PPHLN1, etc.) Lung cancer (FGFR2-CIT)

FGFR2 Mut [37] Breast cancer (R203C, N549K, K659N) Endometrial cancer (S252W, P253R, N549K, K659E) Lung cancer (S252W, P253R, K659E)

FGFR3 Amp [2,33,35] Ovarian and urothelial cancers

FGFR4 Mut [37] Rhabdomyosarcoma (N535K, V550E)

FGFR3 Fus [36,44] Glioblastoma and lung cancer (FGFR3-TACC3) Lymphoma (ETV6-FGFR3) Mulple myeloma [t(4;14)(p16;q32)] Urothelial cancer (FGFR3-BAIAP2L1, FGFR3-JAKMIP1, FGFR3-TACC3)

FGFR3 Mut [37] Gallbladder cancer (R248C, S249C, G370C, Y373C, G380R, K650M) Lung cancer (R248C, S249C, G370C, K650E) Mulple myeloma (R248C, Y373C, K650E/M) Urothelial cancer (R248C, S249C, G370C, S371C, Y373C, N540S, K650E/M)

(See figure legend on the bottom of the next page.)

2

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

TIPS 1376 No. of Pages 16

Box 1. FGFR Splicing, EMT, and Noncoding RNAs Given the alternative splicing of mutually exclusive exons, the FGFR1, FGFR2, and FGFR3 genes encode two major isoforms with divergent ligand affinities. For example, FGFR2b is a high-affinity receptor for FGF3, FGF7, and FGF10, whereas FGFR2c is a receptor for FGF2, FGF4, and FGF20 [1–3]. ESRP1/2 and PTBP1/2 are RNA-binding proteins that are involved in epithelial FGFR2b splicing and mesenchymal FGFR2c splicing, respectively [96]. The loss-of-function ESRP2 variant (R353Q) in breast cancer inhibits FGFR2b splicing [[17_TD$IF]97], and derepression of PTBP2 due to miR-92ainduced RBM4 repression promotes FGFR2c splicing [[18_TD$IF]98]. FGFR2b is preferentially expressed on cells with an epithelial phenotype, whereas FGFR2c and FGFR1 are expressed on cells with a mesenchymal phenotype. The class-switch of FGFR and cadherin from an epithelial FGFR and E-cadherin to a mesenchymal FGFR and N-cadherin occurs during EMT [[19_TD$IF]99,100]. The mesenchymal FGFR promotes EMT, whereas the epithelial FGFR promotes the reverse process, mesenchymal-to-epithelial transition. The activation of FGFR1 signaling and EMT are associated with resistance to EGFR-targeted therapy [[20_TD$IF]88]. By contrast, the repression of FGFR1 by miR-573 leads to tumor suppression through the inhibition of EMT and metastasis [[21_TD$IF]101]. The class-switch of FGFR and EMT are closely connected with, and involved in, pathophysiological processes, such as tissue repair, cancer metastasis, and therapeutic resistance. miRNA and long noncoding RNA (lncRNA) are noncoding RNAs that regulate FGF signaling and a variety of cellular processes [[21_TD$IF]101–[2_TD$IF]106]. The FGFR3-TACC3 fusion mRNA is released from miR-99a-mediated repression following the loss of the miR-99a-binding motif in the 30 -untranslated region (UTR) of FGFR3 mRNA [[23_TD$IF]105]. Other type II FGFR fusion transcripts, such as FGFR1-TACC1, FGFR2-AFF3, FGFR2-BICC1, FGFR2-CASP7, FGFR2-CCAR2, FGFR2-CCDC6, FGFR2-CIT, FGFR2-OFD1, FGFR2-PPHLN1, FGFR3-BAIAP2L1, and FGFR3-JAKMIP1 [[24_TD$IF]36], are potentially derepressed following the loss of intrinsic miRNA-binding sites. The lncRNA UCA1 is upregulated in hepatocellular carcinoma and binds to miR-216b as an endogenous ‘miRNA sponge’. It promotes liver carcinogenesis by relieving FGFR1 from miR-126b-mediated repression and subsequently activating the FGFR1-ERK signaling cascade [[25_TD$IF]103]. The lncRNA asFGFR2 recruits the histone H3 lysine 27 (H3K27) methyltransferase EZH2 and the H3K36 demethylase KDM2A around the alternatively spliced exons of the FGFR2 gene in epithelial cells. This recruitment promotes FGFR2b splicing through the inhibition of MRG15-mediated PTBP1/2 recruitment [[26_TD$IF]107]. The lncRNA FGFR3-AS1 is upregulated in osteosarcoma and stabilizes FGFR3 mRNA [[27_TD$IF]108]. The IncRNA-mediated regulation of FGF signaling cascades in human diseases is an emerging topic in the field of FGF/FGFRrelated medicine.

advances in FGF/FGFR signaling, with an emphasis on noncoding RNAs (Box 1), angiogenesis, and immunity. Pro- and anti-FGF/FGFR therapies. as well as the benefit–risk and cost-effectiveness issues of FGF/FGFR-targeted therapy, are also discussed.

FGFR Alterations in Cancers and Noncancerous Diseases Germ-Line Mutations and Variations in FGFRs Congenital craniofacial and skeletal disorders are caused by gain-of-function mutations in the FGFR1, FGFR2, or FGFR3 genes [6,7,16–19] (summarized in Figure 1). Given that de novo mutations, such as Y340C, C342S, and P253R in FGFR2, and Y373C in FGFR3, contribute to the survival of mutant spermatogonia in the seminiferous tubules of aged human testes, the risks for Apert, Crouzon, and Pfeiffer syndromes and thanatophoric dysplasia are associated with

Figure 1. FGF signals are transduced to the RAS–extracellular signal-regulated kinase (ERK), phosphatidylinositol 3-kinase (PI3K)–AKT, inositol-1,4,5-trisphosphate (IP3)–Ca2+, and diacylglycerol (DAG)–protein kinase C (PKC) signaling cascades for the regulation of self-renewal, metabolism, survival, proliferation, differentiation, epithelial-to-mesenchymal transition (EMT), angiogenesis, immunity, and whole-body homeostasis. The amino-acid numbers of FGFR1, FGFR2, FGFR3, and FGFR4 are based on P11362, P21802, P22607, and P22455 in the UniProt database. Germline gain-of-function (GoF) mutations in the FGFR1, FGFR2, and FGFR3 genes occur in craniosynostosis, and such mutations in the FGFR3 gene occur in skeletal dysplasia. Postzygotic mosaic GoF mutations in the FGFR1, FGFR2, and FGFR3 genes are reported in encephalocraniocutaneous lipomatosis, Blaschko line acne on pre-existent hypomelanosis, and keratinocytic epidermal nevus syndrome, respectively. The FGFR1 gene is amplified in estrogen receptor-positive (ER+) breast cancer, squamous cell carcinoma (SCC), and small cell (SC) lung cancers, whereas the FGFR2 gene is amplified in triple-negative breast cancer (TNBC) and gastric cancer. FGFR1 fusions occur in glioblastoma and myeloproliferative neoplasia (MPN), FGFR2 fusions in breast cancer and cholangiocarcinoma, and FGFR3 fusions in glioblastoma and urothelial cancer. Somatic GoF mutations in the FGFR1, FGFR2, FGFR3, and FGFR4 genes are reported in Ewing sarcoma, endometrial cancer, urothelial cancer, and rhabdomyosarcoma, respectively. Germline loss-of-function (LoF) mutations in the FGFR1 and FGFR2 genes occur in hypogonadotropic hypogonadism 2 (HH2) and lacrimo-auriculo-dento-digital (LADD) syndrome, respectively, and somatic LoF mutations occur in the FGFR2 gene in melanoma. GoF and LoF FGFR alterations are involved in the pathogenesis of cancer and noncancerous diseases. Abbreviations: Amp, amplification; Del, deletion; Fus, fusion; HSPG, heparin-sulfate proteoglycan; Mut, mutation.

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

3

TIPS 1376 No. of Pages 16

advanced paternal age [20]. Craniosynostosis occurs due to the premature fusion of the skull bone sutures in approximately 1/2500 live births worldwide [6,16]. Fifty-two different mutations in the FGFR genes were detected in 187 of 630 probands of craniosynostosis, including P250R FGFR3 mutation in 49 cases of Muenke syndrome, S252W FGFR2 mutation in 34 cases of Apert syndrome, and disparate FGFR2 mutations in Crouzon, Pfeiffer, and other syndromes [17]. By contrast, FGFR3 mutations have been detected in skeletal dysplasias, such as achondroplasia (G375C and G380R), hypochondroplasia (N540K), and thanatophoric dysplasia (Y373C, K650E/M, etc.) [7,18]. Achondroplasia is the most common cause of dwarfism and occurs in approximately 1/20 000 live births, which is one of the FGFR-related congenital disorders amenable to therapeutic intervention, as discussed below. Loss-of-function mutations in FGFRs occur in other types of congenital disorder (summarized in Figure 1). V102fs, R622X, and other mutations throughout FGFR1 are loss-of-function mutations in patients with hypogonadotropic hypogonadism 2 (HH2) [21]. The A628T, A648T, and R649S loss-of-function mutations in FGFR2 are present in patients with LADD syndrome [22]. The rs2981578, rs35054928, and rs45631563 single nucleotide polymorphisms (SNPs) within intron 2 of the FGFR2 gene are associated with an increased risk for estrogen receptor-positive (ER+) breast cancer, and the rs2981578-C allele gives rise to a de novo binding site for FOXA1 [23]. Given that FOXA1 opens the chromatin for the binding of tissue-specific transcription factors, such as ER and androgen receptor (AR) [24], the rs2981578-C allele might be involved in a predisposition for breast cancer through the FOXA1/ER/-mediated transcriptional regulation of FGFR2 in mammary epithelial cells. The region encompassing the FGFR2 and TACC2 genes may be the critical region for 10q26.1 microdeletion syndrome, which presents with growth retardation, intellectual disability, and microcephaly [25], whereas the microdeletion of human chromosome 5q35, which encompasses the NSD1 gene, causes Sotos syndrome [26]. Nephrocalcinosis and infantile hypercalcemia can occur in patients with Sotos syndrome with an FGFR4 codeletion [27]. As a result of the development of next-generation sequencing (NGS) technologies, variations other than the common SNPs, such as rare SNPs, de novo SNPs, and microdeletions, can be identified and used to elucidate the heritability of the FGFR alterations. Postzygotic Mosaic Mutations in FGFRs Congenital disorders with asymmetric and focal malformations and without familial recurrence are caused by postzygotic mosaic mutations. Encephalocraniocutaneous lipomatosis [28], Blaschko line acne on pre-existent hypomelanosis [29], and keratinocytic epidermal nevus syndrome [30] are caused by postzygotic mosaic mutations in FGFRs. N546K and K656E mosaic mutations in FGFR1 lead to the activation of focal FGFR1 signaling in the brain, eye, and skin, which causes brain lipomas, ocular tumors, and cutaneous nevus, respectively. P253R mosaic mutations in FGFR2 cause Blaschko line acne on pre-existent hypomelanosis, whereas P253R germ-line mutations in FGFR2 cause Apert syndrome. Given that P253R mosaic mutations in FGFR2 are restricted to skin keratinocytes, craniosynostosis is absent in patients with Blaschko line acne on pre-existent hypomelanosis. Somatic Alterations in FGFRs Somatic gain-of-function alterations in FGFRs are mainly caused by gene amplification, gene fusion, and point mutations (summarized in Figure 1). The FGFR1 gene is amplified in ER+ breast and lung cancers; the FGFR2 gene is amplified in triple-negative breast and gastric cancers; and the FGFR3 gene is amplified in ovarian and urothelial cancers [1,2,31–35]. FGFR gene fusions are classified into type I fusions, which generate cytoplasmic chimeric FGFR receptors in hematological malignancies (BCR-FGFR1, CNTRL-FGFR1, ZMYM2-FGFR1, etc.), and type II

4

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

TIPS 1376 No. of Pages 16

fusions, which generate transmembrane-type FGFR receptors with C-terminal rearrangements in solid tumors (FGFR1-TACC1, FGFR2-BICC1, FGFR3-TACC3, etc.) [36]. N546K mutations in FGFR1, N549K mutations in FGFR2 (N550K FGFR2b), N540K/S mutations in FGFR3 (N542K/S FGFR3b), and N535K mutations in FGFR4 are located in the linker region between the /C helix and the b4 strand of FGFR kinases. These mutations, along with K656E, K659E (K660E FGFR2b), and K650E (K652E FGFR3b) gain-of-function point mutations in the activation loops of the FGFR1, FGFR2, and FGFR3 kinases, respectively, increase kinase activity [37–39]. FGFR1 amplification is one of the most common somatic alterations in the FGFR genes that are amenable to therapeutic intervention, as discussed below. Somatic loss-of-function alterations in FGFRs are caused by deletions, disruptive rearrangements, and point mutations. The loss of copy number in FGFR1 is observed in breast cancer [2], and loss-of-function point mutations in FGFR2 are observed in melanoma [40]. By contrast, FGFRs are overexpressed in other subsets of breast cancer [2] and melanoma [41] to drive carcinogenesis. FGFRs function as cancer drivers or tumor suppressors in a cellular contextdependent manner. FGFR1 is often co-overexpressed with other co-amplified genes, such as NSD3 in breast and lung cancers [42,43]. NSD3 is a SET-domain methyltransferase that is homologous to oncogenic NSD2 in multiple myeloma [26]. By contrast, NSD2 and FGFR3 at human chromosome 4p16.3 are overexpressed as cancer drivers in multiple myeloma with t(4;14)(p16;q32) [44]. The involvement of neighboring cancer-associated genes could lead to phenotypic diversity and altered therapeutic responses in human cancers with FGFR alterations. FGFR1 is overexpressed in lung cancer, malignant rhabdoid tumors, mesothelioma, and phosphaturic mesenchymal tumors in the absence of FGFR1 alterations [36,45–47]. FGFR2 is upregulated by the SYT-SSX2 cancer driver of synovial sarcoma [48], whereas FGFR4 is upregulated by the PAX3-FOXO1 cancer driver of rhabdomyosarcoma [24,37]. FGFRs are overexpressed as cancer drivers due to FGFR gene amplifications, altered distal FGFR enhancers, and other genetic alterations in FGFR trans-regulators.

FGF Signaling in Angiogenesis and Immunity FGF signaling dysregulation is caused by genetic alterations in FGF signaling molecules (Figure 1), genetic alterations that regulate transcription of FGF signaling molecules [36,37,45–48], and dysregulation of miRNAs, and long noncoding RNAs (lncRNAs) that regulate the expression of FGF signaling molecules (Box 1). Given that aberrant FGF signals directly cause human diseases, and also indirectly promote human diseases through dysregulation of angiogenesis and immunity, effects of FGF signaling on angiogenesis and immunity are highlighted in this section. Angiogenesis results from endothelial sprouting of pre-existing vessels and endothelial differentiation of endothelial progenitor cells (EPCs). Endothelial cells that coat the inner wall of blood and lymphatic vessels are involved in the formation and maintenance of vessels and the regulation of somatic stem cells and homeostasis [49]. FGF2, vascular endothelial growth factor (VEGF), and angiopoietin 2 (ANGPT2) signaling are involved in endothelial activation, whereas ANGPT1 and Notch signaling have a role in endothelial quiescence [36]. FGF2 promotes endothelial proliferation and migration following FGFR1/2 signaling and VEGF/ANGPT2 secretion [50]. Other FGFs, such as FGF5 and FGF18, also promote angiogenesis through FGFRs on endothelial cells [8,51]. FGFs are released from damaged tissues. FGF1 and FGF2 signaling through FGFR2 promote neutrophil chemotaxis to damaged tissues [52], whereas FGF23 signaling through FGFR2 impairs neutrophil recruitment [53]. Interferon g (IFN-g or IFNG) induces monocyte differentiation

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

5

TIPS 1376 No. of Pages 16

to M1 macrophages to promote inflammation, and FGF23 secreted from M1 macrophages upregulates tumor necrosis factor-/ (TNF-/ or TNF) to augment inflammation [54]. IFN-g and TNF-/ downregulate FGFR1 on endothelial cells and induce FGF resistance [55]. By contrast, IL4 and IL10 induce monocyte differentiation to M2 macrophages to mediate and resolve inflammation, and FGF2 secreted from the M2 macrophages is involved in angiogenesis and tissue repair [56]. Thus, FGF signaling dynamically regulates immunity and is regulated by immune cells during inflammation and tissue repair. Endothelial and immune cells work together in a variety of processes, and FGF signaling cascades have a key role in homeostasis and host defense through the regulation of angiogenesis and immunity. Therefore, FGF-dependent regulation of angiogenesis and immunity in physiological and pathological conditions is emerging as a hot issue in the field of translational medicine.

Pro- and Anti-FGF Signaling Therapy Loss- and gain-of-function alterations in FGF signaling cascades are both involved in human diseases due to the pleiotropic functions of FGFs (Figure 1). Therapeutics targeting FGF signaling cascades are largely classified into ‘pro-FGF signaling therapeutics’ for noncancerous diseases, such as cardiovascular disease, diabetes, oral mucositis, and skin ulcers, and ‘antiFGF signaling therapeutics’ for cancer and the noncancerous diseases of chondrodysplasia and rickets (Table 1). Pro-FGF Signaling Therapy Given that FGFs are involved in a variety of physiological processes, such as tissue repair, angiogenesis, and whole-body homeostasis [57–59], recombinant FGFs or FGF analogs, such as FGF1, FGF2 (trafermin), FGF7 (palifermin), FGF18 (sprifermin), FGF19 (NGM282), and FGF21 (LY2405319 and PF-05231023), were developed as first-generation pro-FGF signaling therapeutics (Table 1) as a way to augment beneficial effects of FGF. Trafermin, recombinant human FGF2, was approved in Japan for the treatment of patients with skin ulcers, and palifermin, a truncated recombinant human FGF7, was approved in the USA for the treatment of patients with oral mucositis. Given the potential risks, such as angiogenic dysregulation, immune dysregulation, metabolic dysregulation, and tumor proliferation, the clinical application of recombinant FGFs or FGF analogs is currently restricted to topical agents for tissue repair or wound healing. The application of proangiogenic FGFs for the treatment of vascular diseases has been a longstanding focus of FGF researchers. Cardiovascular disease, cerebrovascular disease, and critical limb ischemia are vascular pathologies that lead to unfavorable outcomes [60]. The intravenous injection of recombinant FGF2 failed to show clinical effectiveness in patients with coronary artery disease due to its short half-life. To overcome the short half-life of recombinant FGFs, a hydrogel-based slow-release form of recombinant FGFs and a vector-based delivery of the FGF genes have been developed as second-generation pro-FGF signaling therapeutics. NV1FGF is a nonviral vector-based FGF1 delivery system; SeV/dF-FGF2 (DVC1-0101) and Ad5. FGF4 are viral vector-based FGF2 and FGF4 delivery systems, respectively [58]. The intramuscular administration of NV1FGF to patients with critical limb ischemia improved pain and skin ulcers in Phase I and Phase II clinical trials, but failed to reduce amputation and death in a Phase III clinical trial [60]. The continuous and robust expression of FGF is sufficient to promote angiogenesis in experiments in preclinical models of acute ischemia, but was inefficient in patients with chronic ischemia, because chronic inflammation induces FGF resistance through IFN-g and TNF/ elevation, as mentioned above [55]. Alternatively, the aberrant activation of FGF signaling is involved in immature angiogenesis, which gives rise to a leaky and hypoxic microenvironment [36]. Given that a dynamic spatiotemporal network of the FGF, Hedgehog, and Notch signaling cascades is necessary for the physiological angiogenesis [14], further fine-tuning of the duration

6

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

TIPS 1376 No. of Pages 16

Table 1. Therapeutics Targeting FGFs and FGFRsa,b [7_TD$IF] Class Recombinant FGF or FGF analog

FGF trap

Monoclonal antibody (mAb)

Small-molecule FGFR inhibitor

Drug

Targets

Disease

Drug development

Refs

FGF2 (trafermin)

FGF2[10_TD$IF] receptor

Skin ulcers

Approved (Japan)

[59]

FGF7 (palifermin)

[9_TD$IF]FGF7[1_TD$IF] receptor

Oral mucositis

Approved (USA)

[59]

FGF18 (sprifermin)

[9_TD$IF]FGF18[12_TD$IF] receptor

Osteoarthritis

P2 (NCT01919164)

[59]

FGF19 (NGM282)

[9_TD$IF]FGF19[13_TD$IF] receptor

T2DM; PSC

P2 (NCT01943045) P2 (NCT02704364)

[57]

FGF21 (LY2405319)

[9_TD$IF]FGF21[14_TD$IF] receptor

T2DM

P1 (NCT01869959)

[57]

FGF21 (PF-05231023)

[9_TD$IF]FGF21[14_TD$IF] receptor

T2DM

P1 (NCT01285518)

[57]

FGF1

[9_TD$IF]FGF1[15_TD$IF] receptor

T2DM

Preclinical

[15]

FP-1039

FGFR1 ligands (FGF1, 2, 4, etc.)

Cancer

P1 (NCT01868022)

[63]

sFGFR3

FGFR3 ligands (FGF2, 9, 18, etc.)

Achondroplasia

Preclinical

[64]

KRN23

FGF23

XLH

P3 (NCT02537431)

[65]

BAY1179470

FGFR2

Cancer

P1 (NCT01881217)

NIHc[9_TD$IF]

FPA144

FGFR2b

Cancer

P1 (NCT02318329)

NIHc

MFGR1877S

FGFR3

Cancer

P1 (NCT01122875)

NIHc

AZD4547

FGFR1 FGFR2 FGFR3 FGFR4

(0.2 nM); (1.8 nM); (2.5 nM); (165 nM)

Cancer with FGFR alteration

P2 P2 P2 P2

(NCT01795768); (NCT02117167); (NCT02299999); (NCT02465060)

[66]

Dovitinib (TKI258)

FGFR1 (8.0 nM); FGFR2 (40 nM); FGFR3 (9.0 nM)

Cancer with FGFR alteration

P2 (NCT01719549); P2 (NCT01732107)

[67]

Erdafitinib (JNJ-493 or (JNJ-42756493)

FGFR1 FGFR2 FGFR3 FGFR4

(<1 nM); (<1 nM); (1.1 nM); (<1 nM)

Cancer with FGFR alteration

P2 (NCT02365597) P2 (NCT02699606)

[68]

Infigratinib (BGJ398)

FGFR1 FGFR2 FGFR3 FGFR4

(0.9 nM); (1.4 nM); (1.0 nM); (60 nM)

Cancer with FGFR alteration

P2 (NCT01975701); P2 (NCT02150967); P2 (NCT02160041)

[69]

Ponatinib (AP24534)

FGFR1 (2.2 nM); FGFR2 (1.6 nM); FGFR3 (18.2 nM); FGFR4 (7.7 nM)

Cancer with FGFR alteration

P2 (NCT02265341)

[70]

TAS-120

FGFR1 FGFR2 FGFR3 FGFR4

(3.9 nM); (1.3 nM); (1.6 nM); (8.3 nM)

Cancer with FGFR alteration

P2 (NCT02052778)

[71]

ASP5878

FGFR1 FGFR2 FGFR3 FGFR4

(0.5 nM); (0.6 nM); (0.7 nM); (3.5 nM)

Cancer

P1 (NCT02038673)

[72]

Debio 1347

FGFR1 FGFR2 FGFR3 FGFR4

(9.3 nM); (7.6 nM); (22 nM); (290 nM)

Cancer

P1 (NCT01948297)

[73]

E7090

FGFR1 FGFR2 FGFR3 FGFR4

(0.7 nM); (0.5 nM); (1.2 nM); (120 nM)

Cancer

P1 (NCT02275910)

[74]

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

7

TIPS 1376 No. of Pages 16

Table 1. (continued) Class

Drug

Targets

Disease

Drug development

Refs

LY2874455

FGFR1 FGFR2 FGFR3 FGFR4

(2.8 nM); (2.6 nM); (6.4 nM); (6.0 nM)

Cancer

P1 (NCT01212107)

[75]

FIIN-2

FGFR1 FGFR2 FGFR3 FGFR4

(3.1 nM); (4.3 nM); (27 nM); (45 nM)

Cancer

Preclinical

[76]

BLU9931

FGFR1 FGFR2 FGFR3 FGFR4

(591 nM); (493 nM); (150 nM); (3.0 nM)

Cancer

Preclinical

[77]

a

Effects of FGFR inhibitors on FGFRs are shown by IC50 values in parentheses. Abbreviations: P1, Phase I clinical trial; P2, Phase II clinical trial; P3, Phase III clinical trial; PSC, primary sclerosing cholangitis; T2DM, type 2 diabetes mellitus; XLH, X-linked hypophosphatemia. c https://clinicaltrials.gov. b

and level of FGF expression in combination with the regulation of other signaling cascades might be effective for the treatment of patients with vascular diseases. Metabolism and whole-body homeostasis are of interest to FGF researchers. The global epidemic of type 2 diabetes mellitus (T2DM), which is characterized by hyperglycemia and insulin resistance, is not only associated with obesity and chronic inflammation caused by excessive food intake and insufficient exercise, but also involves genetic factors, such as SNPs in the TCF7L2, ADCY5, and CCND2 genes, which are involved in the function and homeostasis of pancreatic b cells [61,62]. Insulin resistance in obese individuals is associated with the upregulation of IFN-g, IL6, and TNF-/, and M1 macrophage polarization, whereas insulin sensitivity is associated with IL10 upregulation and M2 macrophage polarization [61]. Paracrine FGF1 and endocrine FGF21 increasing the uptake of blood glucose in adipose tissues and endocrine FGF19 decreasing the supply of blood glucose from the liver induce insulin sensitivity and ameliorate dysregulated glucose metabolism in preclinical mouse models of insulin resistance [15,57]. Given that recombinant FGFs might promote the proliferation of covert tumors, stabilized FGF analogs devoid of mitogenic activity but retaining glucose homeostatic activity have been developed to treat insulin resistance in obese patients and those with diabetes. FGF19 or FGF21 analogs, including NGM282, LY2405319, and PF-05231023, are in clinical trials for the treatment of patients with T2DM (Table 1). Anti-FGF Signaling Therapy Soluble engineered proteins that adsorb multiple FGF ligands (FGF traps), FGF/FGFR-targeting human mAbs (anti-FGF mAb and anti-FGFR mAb) and FGFR-targeting small-molecule compounds (FGFR inhibitors) are under development as anti-FGF signaling therapeutics to treat human diseases caused by aberrant FGF signaling (Table 1). The FGF traps FP-1039 (GSK3052230) and sFGFR3 are soluble proteins that contain the extracellular regions of FGFR1 and FGFR3, respectively. The intraperitoneal injection of FP1039, which neutralizes FGFR1 ligands (FGF1, FGF2, FGF4, etc.), inhibited tumor formation that depended on the activation of paracrine FGF2-FGFR1 signaling in preclinical mouse models of tumorigenesis [63]. The subcutaneous injection of sFGFR3, which neutralizes FGFR3 ligands (FGF2, FGF9, FGF18, etc.), rescued chondrodysplasia phenotypes caused by the activation of FGFR3 signaling in a preclinical mouse model of achondroplasia [64]. A Phase I clinical trial of

8

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

TIPS 1376 No. of Pages 16

FP-1039 revealed that the intravenous injection of FP-1039 was well tolerated in patients with advanced solid tumors. Human mAb drugs, such as anti-FGF23 mAb (KRN23), anti-FGFR2 mAb (BAY1179470), antiFGFR2b mAb (FPA144), and anti-FGFR3 mAb (MFGR1877S), that target the FGF signaling cascades are in clinical trialsi. The elevated serum FGF23 in patients with loss-of-function PHEX mutations (X-linked hypophosphatemia, XLH) leads to rickets and osteomalacia due to hypophosphatemia and decreased serum 1,25-dihydroxy-vitamin D [1,25(OH)2D]. Given that a single dose of KRN23 was tolerable and corrected hypophosphatemia and decreased 1,25(OH)2D in a Phase I clinical trial for patients with XLH [65], KRN23 is now in Phase III clinical trials to evaluate its therapeutic effects on bone quality and osteomalacia in such patients. By contrast, because overexpressed FGFRs function as cancer drivers that promote tumor progression, FGFR mAbs have also been developed for cancer therapy. A Phase I clinical trial of FGFR3-targeting MFGR1877S was completed in 2012, and FGFR2-targeting BAY1179470 and FPA144 are currently in Phase I clinical trialsi. In July 2016, FPA144 obtained an Orphan Drug Designation from the US Food and Drug Administration (FDA) for the treatment of gastric cancer. The FGFR inhibitors in development for clinical application mainly target the cytoplasmic kinase domain [36,66–77], whereas a few FGF inhibitors in the preclinical stage target the extracellular ligand-binding region [78]. FGFR kinase inhibitors are as FGFR1/2/3 inhibitors (AZD4547, dovitinib/TKI258, infigratinib/BGJ398, etc.), selective FGFR4 inhibitors (BLU9931), or pan-FGFR inhibitors (erdafitinib/JNJ-493/JNJ-42756493, LY2874455, ponatinib/AP24534, etc.) due to the diversification of FGFR4 compared with the other FGFR family members [36]. FGFR kinase inhibitors are alternatively classified as FGFR family-restricted inhibitors (erdafitinib, infigratinib, LY2874455, etc.) or FGFR/CSF1R/VEGFR family inhibitors (AZD4547, dovitinib, ponatinib, etc.) due to the evolutionary conservation among the FGFR, CSF1R, and VEGFR family members [36]. FGFR inhibitors are summarized in Table 1, with a focus on their substrate specificities and drug-development stages. Patients with cancer with FGFR genetic alterations (Figure 1) are predicted to be appropriate subjects for the oral administration of an FGFR inhibitor and this theory is currently being tested in Phase II clinical trials (see below) (Table 1). In addition, patients with noncancerous diseases caused by aberrant FGF signaling activation are also potential subjects for FGFR-inhibitor administration. For example, the long-term oral administration of infigratinib (50 mg/kg body weight) ameliorated hypophosphatemia and skeletal phenotypes caused by aberrant FGF23 signaling in a mouse model of XLH [79], and the subcutaneous injection of low-dose infigratinib (2 mg/kg body weight) successfully ameliorated chondrodysplasia phenotypes caused by aberrant FGFR3 signaling in a mouse model of achondroplasia [80]. Therefore, clinical trials of FGFR inhibitors should be performed in patients with cancer or noncancerous diseases with aberrant FGF signaling activation.

FGFR-Targeted Therapeutics for Precision Medicine Gene amplification in FGFR1, point mutation in FGFR2, and genetic alterations in FGFR3 are relatively frequent in lung squamous cell carcinoma, uterine cancer, and bladder cancer, respectively [35,36,78]. However, because it is not clear at present which cancers or cancer subtypes and which genetic alterations are most likely to respond to FGFR-targeted therapy, the market size of FGFR inhibitors remains to be estimated. The benefit–risk and cost-effectiveness issues associated with FGFR-targeted cancer precision medicine are discussed below. Clinical tests of biopsied materials, which include NGS, fluorescence in situ hybridization (FISH), and immunohistochemistry (IHC), are essential for the molecular diagnosis of patients with cancer and therapeutic optimization (Figure 2). For example, FGFR gene amplification is

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

9

TIPS 1376 No. of Pages 16

Dormant epithelial CSC in normal epithelium

Normal epithelium Transformaon

Epithelial cancer at advanced stage

Progression

NGS, FISH, IHC, etc.

Targeted therapy for aconable mutaon

Progressive disease or stable disease

Complete response with dormant CSCs or paral response with residual CSCs

Complete response and cure

Recurrence

Direct effects of FGF signaling on tumor progression and recurrence (1) (2) (3) (4)

Maintenance of epithelial CSCs by mesenchymal FGFs Tumor progression by genec alteraon or overexpression of FGFRs Emergence of mesenchymal CSCs depending on FGFR1 signaling Recurrence of tumors that were inially independent of aberrant FGF signaling through EMT-based FGFR1 upregulaon or secondary FGFR alteraons

Figure 2. Tumor Development, Molecular Diagnosis, and Targeted Therapy. Cancer stem cells (CSCs) are generated via the oncogenic transformation of somatic stem or progenitor cells. De novo epithelial CSCs, maintained by an interaction with niche cells, are dormant in normal epithelia. Additional genetic alterations induce niche-independent survival and the proliferation of epithelial CSCs, which gives rise to overt cancer. Next-generation sequencing (NGS)-based genomic profiling, fluorescence in situ hybridization (FISH), and immunohistochemistry (IHC) are performed on biopsied materials for molecular diagnosis. The patient is then treated with targeted therapeutics for their actionable mutation. Targeted therapy induces a complete response and cures in a subset of patients (super-responders). Targeted therapy induces a complete response in another subset of patients with dormant CSCs or a partial response in patients with residual CSCs. Patients with cancer with dormant or residual CSCs eventually undergo recurrence. By contrast, targeted therapy is ineffective in other subsets of patients with progressive or stable disease. CSCs are heterogeneous populations of tumor-initiating cells that undergo evolution during cancer progression and in response to cancer therapy. The epithelial-to-mesenchymal transition (EMT) of CSCs promotes tumor invasion and metastasis, which results in a poor outcome.

10

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

TIPS 1376 No. of Pages 16

(A) Aconable RTK

First-line therapy

Resistance to first-line therapy

Second-line therapy

Resistance to second-line therapy

ALK Fus

Crizonib

Secondary ALK Mut (L1196M, G1269A, etc.)

Alecnib Cerinib

Terary ALK Mut (G1202R, etc.)

EGFR Mut

Afanib Erlonib Gefinib

Secondary EGFR Mut (T790M, etc.)

Osimernib

Terary EGFR Mut (C797S, etc.)

FGFR1 Amp

AZD4547 Infigranib

Secondary FGFR1 Mut (V561M, etc.)

FIIN-2 (Preclinical)

Terary FGFR1 Mut remaining unclear

FLT3 ITD

Quizarnib

Secondary FLT3 Mut (F691L, etc.)

Pexidarnib

Terary FLT3 Mut (M664I, etc.)

(B) Cancer with aconable mutaon

First-line targeted therapy (US$ 50 000)

CR with dormant CSCs/PR with residual CSCs

PD/SD

Cure

Recurrence

Second-line targeted therapy (US$ 50 000)

PD/SD

CR/PR

Cure

Immune checkpoint therapy ( $ 150 000) (US US$ 50 000

US$ 100 000

US$ 250 000

US$ 100 000

US$ 50 000

Figure 3. Inevitable Acquired Mutations and Prescription Costs of Targeted Therapy. (A) Receptor tyrosine kinase (RTK) inhibitors and acquired mutations. ALK, EGFR, fibroblast growth factor receptor 1 (FGFR1) and FLT3 are representative RTKs that are targeted by small-molecule inhibitors for cancer therapy. First-line RTK inhibitors are prescribed to target founding alterations, such as ALK fusion (Fus), EGFR mutations (Muts), FGFR1 amplification (Amp), and FLT3 internal tandem duplication (ITD). Secondary mutations induce resistance to first-line RTK inhibitors. Second-line RTK inhibitors are then prescribed to target the founding and secondary alterations. Tertiary mutations can then induce resistance to the second-line RTK inhibitors, leading to a vicious cycle of targeted therapy and tumor evolution. (B) Prescription costs of targeted therapy. RTKs are used as a first-line cancer therapy, and immune-checkpoint blockers are then used as second- or third-line cancer therapy. If the costs of RTK inhibitors and immune-checkpoint blockers are US $50 000 and US$150 000, respectively, the total prescription cost for the targeted therapies would be US$50 000[8_TD$IF]US $250 000. Abbreviations: CSCs, cancer stem cells; CR, complete response; PD, progressive disease; PR, partial response; SD, stable disease.

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

11

TIPS 1376 No. of Pages 16

detectable by NGS depending on the depth of sequencing read and validated by FISH, and FGFR overexpression is confirmed by IHC. In the NCI-MATCH trial (NCT02465060), the Oncomine Comprehensive Panel was utilized as a multiplex target-sequencing panel for the genomic profiling of patients with cancer and their enrollment into 24 arms of Phase II clinical trials, including treatment with an FGFR inhibitor ADZ4547 for patients with cancer with FGFR1/ 2/3 amplification, fusion, or mutationii. Directly targeted therapies are available for patients with actionable mutations and/or alterations in the genes encoding RTKs, such as ALK fusion [81,82], EGFR mutation [83,84], FGFR1 amplification [36], and FLT3 internal tandem duplication (ITD) [85,86] (Figure 3A). Targeted therapies initially showed remarkable antitumor effects and, in some cases, complete response; however, because of secondary mutations in the targeted RTK, bypassing activation of other RTKs and EMT-based phenotypic changes, recurrence after targeted therapy is inevitable except in hyper-responders (Figure 2).

Outstanding Questions

Given that EMT is associated with a class-switch of FGFRs from epithelial FGFR2b to mesenchymal FGFR1 (Box 1), FGFR2 on early-stage cancer stem cells (CSCs) with epithelial features is replaced by FGFR1 on late-stage CSCs with mesenchymal features [87]. The EMT-based paracrine FGFR1 signaling that occurs after treatment with other RTK inhibitors is involved in recurrence caused by aberrant FGF signaling activation [88]. A combination therapy using an FGFR inhibitor and an ALK or EGFR inhibitor might reduce recurrence and provide a cure.

Genomic alterations occur not only in tumor cells, but also in precancerous lesions and noncancerous mesenchymal cells. Are FGFR genetic alterations in precancerous lesions somatic mutations or postzygotic mosaic mutations? Do infrequent somatic mutations or postzygotic mosaic mutations in FGFRs contribute to the pathogenesis of cardiovascular disease and diabetes?

The decreases in tumor size responding to FGFR inhibitors have been observed in some patients with FGFR1-amplified breast cancers [89], FGFR2-amplified gastric cancers [90], and gliomas and urothelial cancers with FGFR3-TACC3 fusions [91,92]. The safety and feasibility of FGFR inhibitors have been demonstrated in clinical trials. The adverse events associated with FGFR inhibitors include asthenia and/or fatigue, gastrointestinal toxicities (nausea, vomiting, and diarrhea), liver toxicity, mucositis, and nail and/or skin toxicities; those associated with FGFR/VEGFR inhibitors include hypertension and proteinuria [67,93]. A sustained elevation of serum FGF23 occurs in patients who are prescribed FGFR inhibitors due to an FGF23-FGFR1 signaling blockade in the kidney and subsequent hyperphosphatemia. Given that pathological FGF23 elevation in patients with chronic kidney disease causes atherosclerosis, cardiac hypertrophy, and cardiac fibrosis, such patients undergoing long-term FGFRtargeted therapy must be monitored for these conditions [36]. Predictive biomarkers should be developed to maximize the efficacy and minimize the toxicity of the FGFR-targeted therapy.

Concluding Remarks Both gain- and loss-of-function alterations in the FGF–FGFR signaling cascades cause human diseases and, thus, anti- and pro-FGF signaling therapies are under development for the treatment of cancers and noncancerous diseases. The effectiveness of FGFR inhibitors has been demonstrated in a subset of patients with cancer with FGFR alterations, but monitoring for the cardiovascular toxicities caused by the secondary FGF23 elevation is necessary. Partial exome sequencing of a panel of cancer-associated genes cannot detect rare cis- and trans-regulatory alterations that promote tumorigenesis through FGFR overexpression. Wholegenome sequencing combined with transcriptome analysis should be carried out for the development of predictive biomarkers to maximize the efficacy and minimize the toxicity of FGFR-targeted therapy (see Outstanding Questions). Recurrence is inevitable for patients with cancer treated with RTK inhibitors due to acquired mutations or EMT. Given that EMT gives rise to mesenchymal CSCs that are characterized by FGFR1 upregulation and FGF2-dependent survival, combination therapy using FGFR inhibitors and EGF inhibitors is able to suppress recurrence of EGFR inhibitor-resistant tumors.

12

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

Genetic alterations in FGFRs and overexpression of FGFRs are involved in human carcinogenesis. Relatively inexpensive targeted sequencing detects genetic alterations in FGFRs themselves, but not rare cis- and trans-regulatory alterations that induce overexpression of FGFRs, whereas relatively expensive whole-genome sequencing combined with transcriptome analysis detects both. Which genomic test is necessary for the diagnosis of FGF signaling-dependent cancers?

CSCs are heterogeneous populations of tumor-initiating cells that undergo evolution. What are the best ways to monitor the signature of CSCs in patients to predict the evolutionary trajectory of CSCs and to precisely target CSCs? Do FGFR inhibitors in combination with other RTK inhibitors suppress recurrence in patients by targeting CSCs? Therapy blocking the immune checkpoint remains a promising strategy for a subset of patients with advanced cancer despite the recent failure of a Phase III clinical trial investigating the use of an anti-PD-1 mAb as a first-line therapy for non-small cell lung cancer. Anti-PD-1, anti-PD-L1, and anti-CTLA-4 antibodies ameliorate immune evasion in a subset of patients with cancer through the relief of cytotoxic T cells from exhaustion [94,95], whereas FGFR inhibitors reduced myeloid-derived suppressor cells in tumor microenvironment in preclinical model experiments [36]. How do we fine-tune combination therapy using FGFR inhibitors and immune-checkpoint blockers to treat patients with cancer?

TIPS 1376 No. of Pages 16

Box 2. Costs of Precision Medicine Precision medicine is in part advocated to reduce the total expenses of fully personalized medicine; however, it is ironic that increasing costs are predicted financial burdens for the implementation of precision medicine [[28_TD$IF]109]. The interim analyses of the NCI-MATCH trial that were released in May 2016 revealed that only 2.5% of patients with genotyped cancer were enrolled in 7 arms of Phase II trialsiii[16_TD$IF]. Considering the expansion of the NCI-MATCH trial to 25 arms and the practice of approved targeted therapeutics, approximately 10% of patients with genotyped cancer might be prescribed targeted therapeutics in the near future. If the cost of the NGS-based multiplex target sequencing panel test is US$1500, the initial diagnostic costs would be US$15 000 for a patient with cancer with successful enrollment in the targeted therapy. Although some patients might be cured with the first-line therapy, recurrence due to acquired mutations is inevitable (Figure 3A, main text). For example, the secondary ALK mutations L1196M and G1269A cause crizotinib resistance in patients with ALK-fused lung cancer [82]; the secondary EGFR mutation T790M causes resistance to afatinib, erlotinib, and gefitinib in patients with EGFR-mutated lung cancer [84]; the secondary FGFR1 mutation V561M causes resistance to AZD4547 and infigratinib [76]; and the secondary FLT3 mutation F691L causes quizartinib resistance in patients with FLT3-ITD leukemia [85]. A second-line ALK inhibitor (alectinib or ceritinib) [82], a second-line EGFR inhibitor (osimertinib) [84], or a second-line FLT3 inhibitor (pexidartinib) [86] will be prescribed to patients with relapsed cancer with secondary ALK, EGFR, or FLT3 mutations, respectively (Figure 3A, main text). However, re-recurrence due to tertiary mutations is unavoidable, and immune-checkpoint inhibitors, such as an anti-PD-1 mAb or an anti-CTLA-4 mAb [94,95], are prescribed for re-relapsed patients (Figure 3B, main text). If the costs of therapies targeting RTKs and the immune checkpoint are US$50 000 and US$150 000, respectively, the total prescription cost for the targeted therapies would be US$50 000US$250 000 (Figure 3B, main text). Together, these estimates indicate that the diagnostic and prescription costs are two major expenses for the future implementation of precision medicine. Developing an array of inexpensive therapeutics based on drug repositioning and genotype-dependent combination therapy targeting RTKs and dormant CSCs will contribute to the increased prescription rate and decreased recurrence rate, respectively, and will subsequently reduce the costs of precision medicine.

The diagnostic and prescription costs are two major expenses for the future implementation of precision medicine (Box 2). The development of predictive biomarkers and combination therapy targeting RTKs and dormant CSCs could reduce the diagnostic and precision costs for cancer precision medicine. [2_TD$IF]Acknowledgment This study was financially supported in part by a grant-in-aid for the Knowledgebase Project from the M Katoh's Fund.

Resources i

[1_TD$IF] https://clinicaltrials.gov

ii

www.cancer.gov/about-cancer/treatment/clinical-trials/nci-supported/nci-match

iii

http://ecog-acrin.org/nci-match-eay131/interim-analysis

[29_TD$IF]Supplemental [30_TD$IF]Information Supplemental [31_TD$IF]information [32_TD$IF]associated [3_TD$IF]with this article can be found[3_TD$IF] online[4_TD$IF] at doi:10.1016/j.tips.2016.10.003.

References 7.

Carter, E.P. et al. (2015) Careless talk costs lives: fibroblast growth factor receptor signalling and the consequences of pathway malfunction. Trends Cell Biol. 25, 221–233

Vajo, Z. et al. (2000) The molecular and genetic basis of fibroblast growth factor receptor 3 disorders: the achondroplasia family of skeletal dysplasias, Muenke craniosynostosis, and Crouzon syndrome with acanthosis nigricans. Endocr. Rev. 21, 23–39

8.

3.

Ornitz, D.M. and Itoh, N. (2015) The fibroblast growth factor signaling pathway. Wiley Interdiscip. Rev. Dev. Biol. 4, 215–266

Sonvilla, G. et al. (2008) FGF18 in colorectal tumour cells: autocrine and paracrine effects. Carcinogenesis 29, 15–24

9.

4.

Eswarakumar, V.P. et al. (2005) Cellular signaling by fibroblast growth factor receptors. Cytokine Growth Factor Rev. 16, 139–149

Holdman, X.B. et al. (2015) Upregulation of EGFR signaling is correlated with tumor stroma remodeling and tumor recurrence in FGFR1-driven breast cancer. Breast Cancer Res. 17, 141

5.

Grose, R. and Dickson, C. (2005) Fibroblast growth factor signaling in tumorigenesis. Cytokine Growth Factor Rev. 16, 179–186

10. Jain, R.K. (2013) Normalizing tumor microenvironment to treat cancer: bench to bedside to biomarkers. J. Clin. Oncol. 31, 2205–2218

6.

Wilkie, A.O. (2005) Bad bones, absent smell, selfish testes: the pleiotropic consequences of human FGF receptor mutations. Cytokine Growth Factor Rev. 16, 187–203

11. Vanneman, M. and Dranoff, G. (2012) Combining immunotherapy and targeted therapies in cancer treatment. Nat. Rev. Cancer 12, 237–251

1.

Katoh, M. and Nakagama, H. (2014) FGF receptors: cancer biology and therapeutics. Med. Res. Rev. 34, 280–300

2.

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

13

TIPS 1376 No. of Pages 16

12. Faul, C. et al. (2011) FGF23 induces left ventricular hypertrophy. J. Clin. Invest. 121, 4393–4408 13. Belperio, J. et al. (2016) Inflammatory mediators and clinical outcome in patients with advanced heart failure receiving cardiac resynchronization therapy. Am. J. Cardiol. 117, 617–625 14. Katoh, M. (2013) Therapeutics targeting angiogenesis: genetics and epigenetics, extracellular miRNAs and signaling networks. Int. J. Mol. Med. 32, 763–767 15. Suh, J.M. et al. (2014) Endocrinization of FGF1 produces a neomorphic and potent insulin sensitizer. Nature 513, 436–439 16. Melville, H. et al. (2010) Genetic basis of potential therapeutic strategies for craniosynostosis. Am. J. Med. Genet. 152A, 3007– 3015 17. Roscioli, T. et al. (2013) Genotype and clinical care correlations in craniosynostosis: findings from a cohort of 630 Australian and New Zealand patients. Am. J. Med. Genet. 163C, 259–270 18. Foldynova-Trantirkova, S. et al. (2012) Sixteen years and counting: the current understanding of fibroblast growth factor receptor 3 (FGFR3) signaling in skeletal dysplasias. Hum. Mutat. 33, 29–41 19. Helsten, T. et al. (2015) Fibroblast growth factor receptor signaling in hereditary and neoplastic disease: biologic and clinical implications. Cancer Metastasis Rev. 34, 479–496 20. Maher, G.J. et al. (2016) Visualizing the origins of selfish de novo mutations in individual seminiferous tubules of human testes. Proc. Natl. Acad. Sci. U. S. A. 113, 2454–2459 21. Kim, S.H. et al. (2007) Diversity in fibroblast growth factor receptor 1 regulation: learning from the investigation of Kallmann syndrome. J. Neuroendocrinol. 20, 141–163 22. Shams, I. et al. (2007) Lacrimo-auriculo-dento-digital syndrome is caused by reduced activity of the fibroblast growth factor 10 (FGF10)-FGF receptor 2 signaling pathway. Mol. Cell. Biol. 27, 6903–6912 23. Campbell, T.M. et al. (2016) FGFR2 risk SNPs confer breast cancer risk by augmenting estrogen responsiveness. Carcinogenesis 37, 741–750 24. Katoh, M. et al. (2013) Cancer genetics and genomics of human FOX family genes. Cancer Lett. 328, 198–206 25. Choucair, N. et al. (2015) 10q26.1 microdeletion: redefining the critical regions for microcephaly, genital anomalies. Am. J. Med. Genet. 167A, 2707–2713 26. Katoh, M. (2016) Mutation spectra of histone methyltransferases with canonical SET domains and EZH2-targeted therapy. Epigenomics 8, 285–305 27. Mutsaers, H.A. et al. (2014) Switch in FGFR3 and -4 expression profile during human renal development may account for transient hypercalcemia in patients with Sotos syndrome due to 5q35 microdeletions. J. Clin. Endocrinol. Metab. 99E, 1361– 1367 28. Bennett, J.T. et al. (2016) Mosaic activating mutations in FGFR1 cause encephalocraniocutaneous lipomatosis. Am. J. Hum. Genet. 98, 579–587 29. Kiritsi, D. et al. (2015) Blaschko line acne on pre-existent hypomelanosis reflecting a mosaic FGFR2 mutation. Br. J. Dermatol. 172, 1125–1127 30. Hafner, C. et al. (2006) Mosaicism of activating FGFR3 mutations in human skin causes epidermal nevi. J. Clin. Invest. 116, 2201– 2207 31. Schäfer, M.H. et al. (2015) Fibroblast growth factor receptor 1 gene amplification in gastric adenocarcinoma. Hum. Pathol. 46, 1488–1495 32. Ross, J.S. et al. (2014) Advanced urothelial carcinoma: nextgeneration sequencing reveals diverse genomic alterations and targets of therapy. Mod. Pathol. 27, 271–280 33. Cancer Genome Atlas Research Network (2014) Comprehensive molecular characterization of urothelial bladder carcinoma. Nature 507, 315–322 34. André, F. and Cortés, J. (2015) Rationale for targeting fibroblast growth factor receptor signaling in breast cancer. Breast Cancer Res. Treat. 150, 1–8

14

35. Helsten, T. et al. (2016) The FGFR landscape in cancer: analysis of 4,853 tumors by next-generation sequencing. Clin. Cancer Res. 22, 259–267 36. Katoh, M. (2016) FGFR inhibitors: effects on cancer cells, tumor microenvironment and whole-body homeostasis. Int. J. Mol. Med. 38, 3–15 37. Gallo, L.H. et al. (2015) Functions of fibroblast growth factor receptors in cancer defined by novel translocations and mutations. Cytokine Growth Factor Rev. 26, 425–449 38. Agelopoulos, K. et al. (2015) Deep sequencing in conjunction with expression and functional analyses reveals activation of FGFR1 in Ewing sarcoma. Clin. Cancer Res. 21, 4935–4946 39. Patani, H. et al. (2016) Landscape of activating cancer mutations in FGFR kinases and their differential responses to inhibitors in clinical use. Oncotarget 7, 24252–24268 40. Gartside, M.G. et al. (2009) Loss-of-function fibroblast growth factor receptor-2 mutations in melanoma. Mol. Cancer Res. 7, 41–54 41. Metzner, T. et al. (2011) Fibroblast growth factor receptors as therapeutic targets in human melanoma: synergism with BRAF inhibition. J. Invest. Dermatol. 131, 2087–2095 42. Gelsi-Boyer, V. et al. (2005) Comprehensive profiling of 8p11-12 amplification in breast cancer. Mol. Cancer Res. 3, 655–667 43. Rooney, C. et al. (2016) Characterization of FGFR1 locus in sqNSCLC reveals a broad and heterogeneous amplicon. PLoS One 11, e0149628 44. Kaiser, M.F. et al. (2013) A TC classification-based predictor for multiple myeloma using multiplexed real-time quantitative PCR. Leukemia 27, 1754–1757 45. Schelch, K. et al. (2014) Fibroblast growth factor receptor inhibition is active against mesothelioma and synergizes with radioand chemotherapy. Am. J. Respir. Crit. Care Med. 190, 763–772 46. Blackwell, C. et al. (2016) Inhibition of FGF/FGFR autocrine signaling in mesothelioma with the FGF ligand trap, FP-1039/ GSK3052230. Oncotarget. Published online May 20, 2016. http://dx.doi.org/10.18632/oncotarget.9515 47. Mok, Y. et al. (2016) From epistaxis to bone pain: report of two cases illustrating the clinicopathological spectrum of phosphaturic mesenchymal tumour with fibroblast growth factor receptor 1 immunohistochemical and cytogenetic analyses. Histopathology 68, 925–930 48. Garcia, C.B. et al. (2012) Reprogramming of mesenchymal stem cells by the synovial sarcoma-associated oncogene SYT–SSX2. Oncogene 31, 2323–2334 49. Rafii, S. et al. (2016) Angiocrine functions of organ-specific endothelial cells. Nature 529, 316–325 50. House, S.L. et al. (2016) Endothelial fibroblast growth factor receptor signaling is required for vascular remodeling following cardiac ischemia-reperfusion injury. Am. J. Physiol. Heart Circ. Physiol. 310H, 559–571 51. Allerstorfer, S. et al. (2008) FGF5 as an oncogenic factor in human glioblastoma multiforme: autocrine and paracrine activities. Oncogene 27, 4180–4190 52. Haddad, L.E. et al. (2011) Characterization of FGF receptor expression in human neutrophils, their contribution to chemotaxis. Am. J. Physiol. Cell Physiol. 301C, 1036–1045 53. Rossaint, J. et al. (2016) FGF23 signaling impairs neutrophil recruitment and host defense during CKD. J. Clin. Invest. 126, 962–974 54. Han, X. et al. (2016) Counter-regulatory paracrine actions of FGF23 and 1,25(OH)2 D in macrophages. FEBS Lett. 590, 53–67 55. Chen, P.Y. et al. (2012) FGF regulates TGF-b signaling and endothelial-to-mesenchymal transition via control of let-7 miRNA expression. Cell Rep. 2, 1684–1696 56. Jetten, N. et al. (2014) Anti-inflammatory M2, but not pro-inflammatory M1 macrophages promote angiogenesis in vivo. Angiogenesis 17, 109–118 57. Degirolamo, C. et al. (2016) Therapeutic potential of the endocrine fibroblast growth factors FGF19, FGF21 and FGF23. Nat. Rev. Drug Discov. 15, 51–69 58. El Agha, E. et al. (2016) Role of fibroblast growth factors in organ regeneration and repair. Semin. Cell Dev. Biol. 53, 76–84

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

TIPS 1376 No. of Pages 16

59. Zhang, J. and Li, Y. (2016) Therapeutic uses of FGFs. Semin. Cell Dev. Biol. 53, 144–154 60. Belch, J. et al. (2011) Effect of fibroblast growth factor NV1FGF on amputation and death: a randomised placebo-controlled trial of gene therapy in critical limb ischaemia. Lancet 377, 1929– 1937

83. Camidge, D.R. et al. (2014) Acquired resistance to TKIs in solid tumours: learning from lung cancer. Nat. Rev. Clin. Oncol. 11, 473–481

61. McNelis, J.C. and Olefsky, J.M. (2014) Macrophages, immunity, and metabolic disease. Immunity 41, 36–48

84. Costa, D.B. (2016) Kinase inhibitor-responsive genotypes in EGFR mutated lung adenocarcinomas: moving past common point mutations or indels into uncommon kinase domain duplications and rearrangements. Transl. Lung Cancer Res. 5, 331– 337

62. Fuchsberger, C. et al. (2016) The genetic architecture of type 2 diabetes. Nature 536, 41–47

85. Smith, C.C. et al. (2014) Crenolanib is a selective type I pan-FLT3 inhibitor. Proc. Natl. Acad. Sci. U. S. A. 111, 5319–5324

63. Tolcher, A.W. et al. (2016) A phase I, first in human study of FP1039 (GSK3052230), a novel FGF ligand trap, in patients with advanced solid tumors. Ann. Oncol. 27, 526–532

86. Smith, C.C. et al. (2015) Characterizing and overriding the structural mechanism of the quizartinib-resistant FLT3 ‘gatekeeper’ F691L mutation with PLX3397. Cancer Discov. 5, 668–679

64. Garcia, S. et al. (2013) Postnatal soluble FGFR3 therapy rescues achondroplasia symptoms and restores bone growth in mice. Sci. Transl. Med. 5, 203ra124 65. Carpenter, T.O. et al. (2014) Randomized trial of the anti-FGF23 antibody KRN23 in X-linked hypophosphatemia. J. Clin. Invest. 124, 1587–1597 66. Gavine, P.R. et al. (2012) AZD4547: an orally bioavailable, potent, and selective inhibitor of the fibroblast growth factor receptor tyrosine kinase family. Cancer Res. 72, 2045–2056 67. Porta, C. et al. (2015) Dovitinib (CHIR258, TKI258): structure, development and preclinical and clinical activity. Future Oncol. 11, 39–50 68. Verstraete, M. et al. (2015) In vitro and in vivo evaluation of the radiosensitizing effect of a selective FGFR inhibitor (JNJ42756493) for rectal cancer. BMC Cancer 15, 946 69. Guagnano, V. et al. (2012) FGFR genetic alterations predict for sensitivity to NVP-BGJ398, a selective pan-FGFR inhibitor. Cancer Discov. 2, 1118–1133 70. O’Hare, T. et al. (2009) AP24534, a pan-BCR-ABL inhibitor for chronic myeloid leukemia, potently inhibits the T315I mutant and overcomes mutation-based resistance. Cancer Cell 16, 401–412 71. Ochiiwa, H. et al. TAS-120, a highly potent and selective irreversible FGFR inhibitor, is effective in tumors harboring various FGFR gene abnormalities. Mol Cancer Ther 12 (11 Suppl.), Abstract nr A270. 72. Futami, T. et al. (2015) Preclinical antitumor activity of ASP5878, a novel inhibitor of FGFR1, 2, 3 and 4, in FGF19-expressing hepatocellular carcinoma. Mol Cancer Ther. 14 (Suppl. 2), Abstract nr A172 73. Nakanishi, Y. et al. (2014) The fibroblast growth factor receptor genetic status as a potential predictor of the sensitivity to CH5183284/Debio 1347, a novel selective FGFR inhibitor. Mol. Cancer Ther. 13, 2547–2558

87. da Silva-Diz, V. et al. (2016) Cancer stem-like cells act via distinct signaling pathways in promoting late stages of malignant progression. Cancer Res. 76, 1245–1259 88. Ware, K.E. et al. (2013) A mechanism of resistance to gefitinib mediated by cellular reprogramming and the acquisition of an FGF2–FGFR1 autocrine growth loop. Oncogenesis 2, e39 89. André, F. et al. (2013) Targeting FGFR with dovitinib (TKI258): preclinical and clinical data in breast cancer. Clin. Cancer Res. 19, 3693–3702 90. Pearson, A. et al. (2016) High-level clonal FGFR amplification and response to FGFR inhibition in a translational clinical trial. Cancer Discov. 6, 838–851 91. Di Stefano, A.L. et al. (2015) Detection, characterization, and inhibition of FGFR-TACC fusions in IDH wild-type glioma. Clin. Cancer Res. 21, 3307–3317 92. Tabernero, J. et al. (2015) Phase I dose-escalation study of JNJ42756493, an oral pan-fibroblast growth factor receptor inhibitor, in patients with advanced solid tumors. J. Clin. Oncol. 33, 3401–3408 93. Touat, M. et al. (2015) Targeting FGFR signaling in cancer. Clin. Cancer Res. 21, 2684–2694 94. Okazaki, T. and Honjo, T. (2007) PD-1 and PD-1 ligands: from discovery to clinical application. Int. Immunol. 19, 813–824 95. Sharma, P. and Allison, J.P. (2015) The future of immune checkpoint therapy. Science 348, 56–61 96. Warzecha, C.C. et al. (2009) ESRP1 and ESRP2 are epithelial cell-type-specific regulators of FGFR2 splicing. Mol. Cell 33, 591–601 97. Horvath, A. et al. (2013) Novel insights into breast cancer genetic variance through RNA sequencing. Sci. Rep. 3, 2256

74. Miyano, S. (2015) Characterization of E7090: a novel and selective FGFR inhibitor. Cancer Res. 76 (14 Suppl.), Abstract nr 3778

98. Liang, Y.C. et al. (2015) The impact of RNA binding motif protein 4-regulated splicing cascade on the progression and metabolism of colorectal cancer cells. Oncotarget 6, 38046–38060

75. Zhao, G. et al. (2011) A novel, selective inhibitor of fibroblast growth factor receptors that shows a potent broad spectrum of antitumor activity in several tumor xenograft models. Mol. Cancer Ther. 10, 2200–2210

99. Wendt, M.K. et al. (2014) Fibroblast growth factor receptor splice variants are stable markers of oncogenic transforming growth factor b1 signaling in metastatic breast cancers. Breast Cancer Res. 16, R24

76. Tan, L. et al. (2014) Development of covalent inhibitors that can overcome resistance to first-generation FGFR kinase inhibitors. Proc. Natl. Acad. Sci. U. S. A. 111, E4869–E4877

100. Ranieri, D. et al. (2015) HPV16 E5 expression induces switching from FGFR2b to FGFR2c and epithelial-mesenchymal transition. Int. J. Cancer 137, 61–72

77. Hagel, M. et al. (2015) First selective small molecule inhibitor of FGFR4 for the treatment of hepatocellular carcinomas with an activated FGFR4 signaling pathway. Cancer Discov. 5, 424–437

101. Wang, L. et al. (2015) miR-573 inhibits prostate cancer metastasis by regulating epithelial-mesenchymal transition. Oncotarget 6, 35978–35990

78. Tanner, Y. and Grose, R.P. (2016) Dysregulated FGF signaling in neoplastic disorders. Semin. Cell Dev. Biol. 53, 126–135

102. Katoh, M. (2014) Cardio-miRNAs and onco-miRNAs: circulating miRNA-based diagnostics for non–cancerous and cancerous diseases. Front. Cell Dev. Biol. 2, 61

79. Wöhrle, S. et al. (2013) Pharmacological inhibition of fibroblast growth factor (FGF) receptor signaling ameliorates FGF23mediated hypophosphatemic rickets. J. Bone Miner. Res. 28, 899–911 80. Komla-Ebri, D. et al. (2016) Tyrosine kinase inhibitor NVPBGJ398 functionally improves FGFR3-related dwarfism in mouse model. J. Clin. Invest. 126, 1871–1884

103. Wang, F. et al. (2015) Upregulated lncRNA-UCA1 contributes to progression of hepatocellular carcinoma through inhibition of miR-216b and activation of FGFR1/ERK signaling pathway. Oncotarget 6, 7899–7917

81. Mano, H. (2012) ALKoma: a cancer subtype with a shared target. Cancer Discov. 2, 495–502

104. Guan, X. et al. (2015) miR-223 regulates adipogenic and osteogenic differentiation of mesenchymal stem cells through a C/ EBPs/miR-223/FGFR2 regulatory feedback loop. Stem Cells 33, 1589–1600

82. Katayama, R. et al. (2015) Therapeutic targeting of anaplastic lymphoma kinase in lung cancer: a paradigm for precision cancer medicine. Clin. Cancer Res. 21, 2227–2235

105. Parker, B.C. et al. (2013) The tumorigenic FGFR3-TACC3 gene fusion escapes miR-99a regulation in glioblastoma. J. Clin. Invest. 123, 855–865

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy

15

TIPS 1376 No. of Pages 16

106. Xu, Q. et al. (2016) Long non-coding RNA regulation of epithelial– mesenchymal transition in cancer metastasis. Cell Death Dis. 7, e2254

108. Sun, J. et al. (2016) Long noncoding RNA FGFR3-AS1 promotes osteosarcoma growth through regulating its natural antisense transcript FGFR3. Mol. Biol. Rep. 43, 427–436

107. Gonzalez, I. et al. (2015) A lncRNA regulates alternative splicing via establishment of a splicing-specific chromatin signature. Nat. Struct. Mol. Biol. 22, 370–376

109. Doble, B. (2016) Budget impact and cost-effectiveness: can we afford precision medicine in oncology? Scand. J. Clin. Lab. Invest. 18, 1–6

16

Trends in Pharmacological Sciences, Month Year, Vol. xx, No. yy