Small molecules in targeted cancer therapy: advances, challenges, and future perspectives

Due to the advantages in efficacy and safety compared with traditional chemotherapy drugs, targeted therapeutic drugs have become mainstream cancer treatments. Since the first tyrosine kinase inhibitor imatinib was approved to enter the market by the US Food and Drug Administration (FDA) in 2001, an increasing number of small-molecule targeted drugs have been developed for the treatment of malignancies. By December 2020, 89 small-molecule targeted antitumor drugs have been approved by the US FDA and the National Medical Products Administration (NMPA) of China. Despite great progress, small-molecule targeted anti-cancer drugs still face many challenges, such as a low response rate and drug resistance. To better promote the development of targeted anti-cancer drugs, we conducted a comprehensive review of small-molecule targeted anti-cancer drugs according to the target classification. We present all the approved drugs as well as important drug candidates in clinical trials for each target, discuss the current challenges, and provide insights and perspectives for the research and development of anti-cancer drugs.

Protein kinase is a kind of enzyme that catalyzes the transfer of γ-phosphate group from ATP to protein residues containing hydroxyl groups. It has an important role in cell growth, proliferation, and differentiation (Fig. 2 ). 5 The human kinome comprises ~535 protein kinases. 6 According to the substrate residues, protein kinases can be classified as tyrosine kinases (including both receptor and non-receptor tyrosine kinases), serine/threonine kinases, and tyrosine kinase-like enzymes. Dysregulation of protein kinases is linked to various diseases, particularly cancer. Protein kinases are the most widely studied tumor therapeutic targets. Currently, a large number of protein kinase inhibitors have been reported. These kinase inhibitors can be classified into different categories by using many ways. Here we adopted an integrated classification system proposed by Roskoski, which is one of the most widely used methods. 7 According to this classification system, protein kinase inhibitors are classified into six types (Type-I–VI). Type-I inhibitors bind to the active conformation of the kinase (DFG-Asp in, αC-helix in). Type-I½ inhibitors bind to a DFG-Asp in inactive kinase conformation with αC-helix out, while type-II inhibitors bind to a DFG-Asp out inactive conformation. These types of inhibitors occupy part of the adenine binding pocket and form hydrogen bonds with the hinge region connecting the small and large lobes of the enzyme. Among them, type-I½ and type-II antagonists can be further divided into A and B subtypes. Type A inhibitors extend past the Sh2 gatekeeper residue into the back cleft, while type B inhibitors fail to extend into the back cleft. The possible importance of this difference is that type A inhibitors have longer residence times compared with type B inhibitors when binding to their targets. Type III and type IV kinase inhibitors are allosteric in nature. Type III inhibitors restrain kinase activity by binding to an allosteric site, which is in the cleft between the small and large kinase lobes adjacent to the ATP-binding pocket. Contrariwise, type IV inhibitors bind outside of the cleft. Moreover, the bivalent molecules that span two distinct regions of the kinase domain are type V inhibitors. Type-I–V inhibitors are all reversible. In contrast, compounds that bind covalently with the kinase active site are called type VI inhibitors (irreversible kinase inhibitors). Fig. 2 Activation of different protein kinase-dependent pathways. The set of RTKs influences a small number of intermediaries, such as phosphoinositide 3-kinase (PI3K) and mitogen-activated protein kinases (MAPK), thereby activating the complex signaling networks that are related to cell proliferation, differentiation, adhesion, apoptosis, and migration. The aggregation, activation, and depolymerization of the periodic CDK-cyclin complex are critical events driving cell cycle turnover. Figure created with BioRender.com

Cellular-mesenchymal-epithelial transition factor (c-Met), also known as hepatocyte growth factor receptor (HGFR), is encoded by the MET proto-oncogene located on chromosome 7q21-31. 41 , 42 Under normal physiology, the binding of c-Met to its sole ligand HGF initiates the activation of the HGF/c-Met signaling pathway, which further activates several downstream signals including the PI3K/AKT, MAPK, STAT, and NF-κB pathways, and has a central role in a variety of cytoplasmic and nuclear processes, such as cell proliferation, survival, invasion, motility, scattering, angiogenesis, and epidermal–mesenchymal transition. 42 – 44 These normal regulatory functions mainly occur during embryonic development, wound healing, and post-injury tissue regeneration. However, aberrant activation of c-Met signaling caused by MET amplification, mutation, inadequate degradation, transcriptional deregulation, or aberrant HGF autocrine or paracrine has been implicated in the development of various solid tumors. 44 , 45 c-Met overexpression has also been reported to be related to poor prognosis and resistance to cytotoxic and molecular targeted therapy, especially for patients treated with EGFR inhibitors, in which MET amplification accounts for ~20% of resistant cases. 46 Activating MET mutations usually occur in the semaphoring domain (e.g., E168D) and juxtamembrane domain (e.g., T1010I, P1009S, skipping mutation) of exons 14, 18, and 19. Of these, c-Met exon 14 skipping mutations promote its oncogenic activity by suppressing c-Met receptor degradation. 43 , 45 , 47 These mutations are rare in patients with primary tumors but common in advanced cancers with metastases, especially in lung adenocarcinoma, brain gliomas, and renal cell carcinoma (RCC). 47 , 48 During the last decade, great progress has been made in antitumor therapy targeting the HGF/c-Met signaling pathway. The early developed c-Met inhibitors were multikinase inhibitors. As early as 2011 and 2012, two multitarget c-Met inhibitors, crizotinib and cabozantinib, were approved for the treatment of NSCLC and medullary thyroid cancer (MTC) as well as RCC, respectively. 49 , 50 However, the indications are not based on their ability to target c-Met but are due to the inhibitory effect of crizotinib on the ALK fusion protein and the multikinase inhibitory activity of cabozantinib. The development of selective c-Met inhibitors has progressed rapidly in recent years, and two highly selective c-Met inhibitors, capmatinib and tepotinib, were approved in the first half of 2020 (Table 1 ). Capmatinib (INCB28060) is an oral competitive c-Met inhibitor with ≥10,000-fold selectivity for c-Met compared with other kinases and potently inhibits c-Met activity at picomolar concentrations. 51 In the GEOMETRY mono-1 trial ( NCT02414139 ) conducted in patients with MET exon 14 skipping mutations, capmatinib exhibited a high objective response rate (ORR) and relatively durable responses in both previously treated and newly diagnosed patients, including those with brain metastases. 52 Combination therapy of capmatinib and gefitinib was also evaluated in a phase II trial in NSCLC patients with disease progression after gefitinib treatment ( NCT01610336 ). A disease control rate of 80% was achieved in 65 subjects, and more responses were observed in patients with MET amplification. 53 Similar results were reported in the combination therapy of capmatinib with other EGFR inhibitors, such as erlotinib. 54 Due to its significant efficacy compared to existing therapies, capmatinib granted a breakthrough therapy designation by the FDA for NSCLC patients harboring MET exon 14 skipping mutations in 2019 and was approved for this indication on May 6, 2020. Tepotinib (EMD1214063), developed by Merck, has more than 1000-fold selectivity for c-Met. 55 Clinical trials of tepotinib ( NCT04647838 and NCT03940703 ) also showed significant effectiveness in the treatment of cancer patients harboring MET mutations and in combination therapy with EGFR TKIs. 56 It has been approved by the Ministry of Health, Labour and Welfare (MHLW) of Japan for the treatment of unresectable, advanced, or recurrent NSCLC in patients with skipping mutations in MET exon 14. 57 There are also many small-molecule c-Met inhibitors at different stages of clinical trials. Representative multikinase c-Met inhibitors include foretinib (XL880/GSK1363089), glesatinib (MGCD265), BMS-777607, and S49076 , which target c-Met/RON/VEGFR-2/KIT/TIE2/PDGFR, c-Met/TIE2/RON/VEGFR-1/2/3, c-Met/RON/AXL, and c-Met/AXL/MER/FGFR, respectively. 58 – 61 Several clinical trials were carried out to test their efficacy for cancer therapy, but some of the results have not been disclosed. In a phase I trial for NSCLC patients who progressed after chemotherapy ( NCT01068587 ), combined foretinib with erlotinib could achieve a response rate of 17.8% in the evaluated patients, and the clinical response was closely associated with baseline c-Met expression. 58 I

Fms-like tyrosine kinase 3 (FLT3), which is widely expressed in hematopoietic stem and progenitor cells, is a transmembrane protein encoded by the proto-oncogene FLT3 . It belongs to the type III RTK family, which also includes PDGFR, FMS, and KIT. All of them consist of an extracellular ligand-binding domain, a single transmembrane hydrophobic alpha helix region, and an intracellular kinase domain. FLT3 is activated by binding to the ligands, which results in its dimerization and conformational changes. Subsequent autophosphorylation of FLT3 triggers signal transduction, activating intracellular signaling cascades such as PI3K/AKT/mTOR, RAS/RAF/MAPK, and JAK/STAT, 98 , 99 which are closely related to cell proliferation, differentiation, survival, and apoptosis. FLT3 is widely overexpressed in patients with acute myeloid leukemia (AML), and its mutations lead to the constitutive activation of downstream signals. 100 , 101 Internal tandem duplication (ITD) mutations in FLT3 (FLT3-ITD) are detected in ~25% of AML patients, and point mutations in the tyrosine kinase domain (TKD) are observed in 7–10% of patients. 102 These mutations have been identified to be involved in the occurrence of leukemia. Due to the established pathogenetic and prognostic roles of FLT3-ITD and FLT3-TKD in AML, several FLT3 inhibitors have been developed for AML therapy. The first-generation FLT3 inhibitors, including sorafenib, sunitinib, midostaurin, tandutinib, and lestaurtinib, are multikinase inhibitors. 103 – 105 They are not specific for FLT3 and have inhibitory activity against various other RTKs, such as PDGFR, KIT, VEGFR, RAF, or JAK2. The clinical efficacy of most of these inhibitors as monotherapy for AML was unimpressive, and their off-target inhibition also increased adverse events. 106 Therefore, clinical studies on first-generation FLT3 inhibitors for AML monotherapy were discontinued except for midostaurin. A randomized phase III trial (RATIFY study, NCT00651261 ) showed that the addition of midostaurin to cytarabine chemotherapy significantly improved overall survival (OS) for FLT3-mutated AML patients. 104 Based on the beneficial results of the RATIFY study, midostaurin was approved by the US FDA for combination therapy with standard chemotherapy in 2017 (Table 1 ). Distinguishingly, pexidartinib (Turalio) is also an orally bioavailable multitarget inhibitor, with IC 50 values of 9, 12, and 17 nM against FLT3-ITD, c-Kit, and colony-stimulating factor 1 receptor (CSF1R), respectively. However, it is not used clinically for AML therapy but approved for the treatment of adult patients with tenosynovial giant cell tumors (TGCTs). This indication is based on its inhibitory effect on CSF1R, which is frequently overexpressed in TGCTs. 107 Second-generation FLT3 inhibitors developed by rational drug design are more potent and specific and have less toxicity related to off-target effects. Gilteritinib is the first approved second-generation FLT3 inhibitor and is also the first effective FLT3 inhibitor for AML monotherapy (Table 1 ). 108 A randomized open-label phase III trial (ADMIRAL study, NCT02421939 ) showed that the median OS was significantly longer in the gilteritinib monotherapy group (9.3 months) than in conventional chemotherapy-treated patients (5.6 months) ( p < 0.001). It was approved for the treatment of relapsed or refractory AML patients with FLT3 mutations in 2018. Quizartinib was screened by the KinomeScan technique to improve the affinity and specificity to FLT3 kinase, and it showed strong activity and selectivity against FLT3-ITD but not TKD. 109 The clinical efficacy of quizartinib was also superior to conventional chemotherapy ( NCT00989261 ); therefore, it was approved for relapsed or refractory AML patients with FLT3-ITD mutations in 2019. 110 Currently, many promising FLT3 inhibitors are still under clinical evaluation. Crenolanib, originally developed as an inhibitor of PDGFR, is also a second-generation FLT3 inhibitor with inhibitory activity against both FLT3-ITD and FLT3 D835 mutations. 111 Crenolanib development focused on assessing the combination effects of this drug with conventional chemotherapy in terms of first-line and relapse treatment. Several clinical trials are underway to evaluate the clinical efficacy of crenolanib, including a randomized phase III trial evaluating the potency of crenolanib in combination with induction chemotherapy for relapsed or refractory FLT3-mutated AML patients ( NCT02298166 ) and a multicentre phase III trial comparing the effects of crenolanib with midostaurin during induction chemotherapy and consolidation therapy for newly diagnosed FLT3-mutated AML patients ( NCT03258931 ). SKLB-1028 is a multitarget inhibitor with FLT3 inhibitory activity. 112 A phase I trial ( NCT02859948 ) was conducted to evaluate the safety, tolerability and pharmacokinetic characteristics of SKLB-1028 in FLT3 mutant AML subjects. 113 Moreover, it has been reported that SKLB-102

The tropomyosin receptor kinase (TRK) family is composed of three members, TRKA, TRKB, and TRKC, which are encoded by the neurotrophic tyrosine receptor kinase (NTRK) genes NTRK1 , NTRK2 , and NTRK3 , respectively. 225 To activate TRK receptors, neurotrophins (TRK ligands) bind to the extracellular domain of the receptors, stimulating homodimerization and autophosphorylation of TRK proteins, thereby activating downstream signaling pathways, such as RAS/MAPK/ERK, PI3K/AKT, and PLCγ. NTRK gene rearrangements containing a kinase domain of one of the three TRKs and a dimerization domain of another gene generate fusion proteins and result in aberrant activation of TRKs, which have been identified as oncogenic drivers of various cancers. 226 – 228 Therefore, TRKs are emerging as important targets for cancer therapy. The rearrangements of NTRK genes occur in only 1% of all malignancies; they have been widely detected at low frequencies in some common cancers, such as lung cancer, thyroid carcinoma, glioblastoma, and colorectal cancer. However, in several rare pediatric and adult cancer types, including infantile fibrosarcoma, secretory breast carcinoma, and salivary gland secretory carcinoma, NTRK gene rearrangements are common. The discovery of TRK inhibitors renewed interest in NTRK gene rearrangements as oncogenes. Currently, two first-generation TRK inhibitors are available for clinical cancer treatment (Table 1 ). Larotrectinib is the first approved selective oral pan-TRK inhibitor with high potency against TRKA, TRKB, and TRKC. 229 Entrectinib (RXDX-101/NMS-E628) is a potent multikinase inhibitor targeting TRKA/B/C, ROS1, and ALK. 230 Both agents received the FDA breakthrough therapy identification; this breakthrough designation highlights the efficacy of TRK inhibitors in various cancers that have the same mutation, regardless of cancer type and patient age. Based on the tumor-agnostic efficacy of the “basket trail” conducted in diverse NTRK fusion-positive cancers, larotrectinib and entrectinib granted FDA approval for the treatment of adult and pediatric patients with TRK fusion solid tumors. In clinical use, NTRK gene fusions should be diagnosed to select patients for targeted TRK therapy. Remarkably, both larotrectinib and entrectinib displayed activity against CNS tumors with NTRK fusions, indicating the ability for BBB penetration. 231 , 232 When referring to the adverse events of first-generation TRK inhibitors, it should be noted that both agents have favorable overall safety profiles compared to other small-molecule TKIs. These drugs are generally well-tolerated in patients, with low incidences of dose reductions, discontinuations, and grade 3–4 adverse events. 225 Recently, there have been several small-molecule TRK inhibitors in different stages of clinical research, some of which target multiple kinases, such as cabozantinib (targeting c-Met, RET, VEGFR-2, ROS1, ALK, and TRK), 233 merestinib (targeting c-Met, TEK, ROS1, and TRK), 234 belizatinib (targeting ALK and TRK), 235 sitravatinib (targeting c-Met, RET, AXL, and TRK), 236 altiratinib (targeting c-Met, TIE2, VEGFR-2, FLT3, and TRK), 237 and DS-6051b (targeting ROS and TRK). 238 These inhibitors displayed varying degrees of inhibitory activity against TRK. Some of them have been approved for indications other than TRK fusion tumors; for example, cabozantinib was approved as an anti-angiogenic inhibitor for the treatment of patients with advanced RCC in 2016, and data are limited on its efficacy against NTRK fusions. However, with the increasing interest in TRK as a cancer therapy target, an increasing number of clinical trials have been performed to evaluate the effects of these inhibitors in patients with TRK fusion-positive tumors. In addition, a phase I study was carried out to assess the safety, PKs, and PDs of the selective TRK inhibitor PLX7486 as a single agent in patients with any histological solid tumors with activating NTRK point or NTRK fusion mutations ( NCT01804530 ). 239 However, the results were not disclosed. Acquired resistance to TKI treatment can be mediated by on-target mutations or off-target (bypass activation) mechanisms. Until now, on-target mutations in the kinase domain of NTRK fusion have been the only resistance mechanism of first-generation TRK inhibitors, which can result in amino acid substitutions of the solvent front, activation loop xDFG motif, and gatekeeper residues in the kinase domain of TRK fusion proteins, interfering with TRK inhibitor binding. 225 The first resistance case to TRK inhibition was discovered in a colorectal cancer patient treated with entrectinib, and two acquired resistance mutations, TRKA G595R and TRKA G667C, were detected in the plasma cfDNA of this patient. 240 Several other resistant mutations were subsequently identified in patients resistant to larotrectinib and entrectinib, including the acquired TRKC G623R substitution, A608D mutation and gatekeeper F589L substitution in TRKA

The B-cell receptor (BCR) pathway has a key role in the progression of a variety of B-cell malignancies. 270 Abnormal activation of BCR signaling has been identified in multiple heterogeneous hematologic malignancies, including B-cell non-Hodgkin’s lymphoma (NHL), chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), mantle cell lymphoma (MCL), marginal zone lymphoma (MZL), follicular lymphoma, Waldenstrom’s macroglobulinemia (WM), and DLBCL. 271 Bruton’s agammaglobulinemia tyrosine kinase (BTK), a crucial component of the BCR pathway, belongs to the non-receptor tyrosine kinase of the TEC family, which contains four other members: tyrosine kinase expressed in hepatocellular carcinoma (TEC), interleukin-2-inducible T-cell kinase (ITK), resting lymphocyte kinase (RLK/TXK), and bone marrow expressed kinase (BMX). 272 BTK is abundantly expressed in B-cell leukemias and lymphomas and functions as a vital regulator of cell proliferation and survival in various B-cell malignancies. 273 Inhibiting BTK is considered an effective therapeutic strategy for some hematologic malignancies. 274 Ibrutinib is the first-generation BTK inhibitor and has been proven to be superior to standard chemotherapy in multiple studies including older patients with significant comorbidity. It is an irreversible small-molecule inhibitor that covalently binds to Cys-481 within the ATP-binding pocket of BTK. Based on the high response rates and durable responses of its monotherapy 275 or in combination with anti-CD20 antibody, 276 ibrutinib has been approved by the FDA for the treatment of MCL, CLL, WM, SLL, and MZL between 2013 and 2017 (Table 2 ). 277 – 279 The clinical efficacy of ibrutinib in the treatment of DLBCL, refractory/recurrent primary central nervous system lymphoma, and secondary central nervous system lymphoma is still undergoing evaluation to expand its indications. 280 Despite the clinical achievement of ibrutinib, side effects including arthralgia, atrial fibrillation, pneumonitis and rash have also been reported and limit its clinical use. Most of the toxicity of ibrutinib is due to its off-target activities against four other TEC family kinases, EGFR, HER2, and Janus kinase 3 (JAK3). 281 Particularly, in combination therapy with the CD20 antibody rituximab, off-target inhibition of ITK by ibrutinib led to an antagonistic effect on antibody-dependent cell-mediated cytotoxicity, influencing the combined effects. 282 The off-target activity of ibrutinib triggered the development of more selective second-generation BTK inhibitors. Acalabrutinib 283 and zanubrutinib 284 are currently approved second-generation BTK inhibitors (Table 2 ). Similar to ibrutinib, they are irreversible inhibitors and form covalent bonds with the Cys-481 residue of the BTK active site. 284 And their selectivity is significantly improved. Acalabrutinib inhibited BTK with an IC 50 of 3 nM and had less off-target activity on EGFR, ITK, or TEC; 285 zanubrutinib had similar inhibitory activity to ibrutinib against BTK, but its IC 50 s on TEC, ITK, EGFR, HER2, and JAK3 were 2–70 times higher than those of ibrutinib. 286 Currently, the FDA has approved them for the treatment of adult MCL patients who have received at least one prior therapy. 287 Both of them are still evaluated in the clinic for the treatment of some other malignancies, such as NHL, 288 multiple myeloma (MM), 289 and ovarian cancer. 290 Several promising BTK irreversible inhibitors are under clinical evaluation. 291 Tirabrutinib (ONO-4059) is a highly selective covalent inhibitor of BTK with an IC 50 of 2.2 nM. In a phase I trial conducted in patients with B-cell malignancies ( NCT02457559 ), tirabrutinib showed significant potency on patients in the CLL group. Ninety-six percent of CLL patients responded to tirabrutinib, and all of the evaluated CLL patients harboring del 17p or TP53 mutations without del 17p responded. 292 Moreover, a phase II trial ( NCT02968563 ) is underway to assess the efficacy and safety of tirabrutinib in combination with the PI3K inhibitor idelalisib and the anti-CD20 antibody obinutuzumab. 293 Spebrutinib (CC-292/AVL-292) is also a second-generation BTK inhibitor and inhibits BTK activity with an IC 50 of 0.5 nM. 294 The results of phase I studies ( NCT01692184 , NCT01732861 , and NCT01351935 ) showed that spebrutinib was safe and well-tolerated following once-daily administration in patients with relapsed or refractory CLL/SLL, WM, and NHL. 295 Despite its high in vitro activity, spebrutinib exhibited inferior clinical efficacy compared with the approved BTK inhibitors. The reasons for the suboptimal effect are not fully understood, but the highly variable PK and pharmacodynamics (PD) seem to limit spebrutinib to continuously reach the in vivo targets. 296 In addition, due to the critical role of BTK in the development and function of B cells, BTK has also been confirmed as a potential therapeutic target for autoimmune disorders. Several BTK in

BRAF/MEK/ERK inhibitors Rat sarcoma virus (RAS)-rapidly accelerated fibrosarcoma (RAF)-mitogen-activated protein kinase (MAPK)-extracellular signal-regulated kinase (ERK) signaling is a well-established pathway that controls cell growth, proliferation, and survival in normal cells and cancer cells. 331 , 332 Core components of RAS-RAF-MAPK-ERK signaling include the small GTPase RAS, the serine/threonine kinase RAF, the protein kinases MEK1/2 and ERK1/2. RAF, including ARAF, BRAF, and CRAF, is the direct downstream of RAS, which serves as a transducer of extracellular stimuli. 332 RAF activates the dual-specificity protein kinase MEK1/2, which subsequently phosphorylates ERK1/2. Activated ERK phosphorylates multiple substrates in the cytosol and nucleus, and then promote cell proliferation and survival. 332 Dysregulation of RAS/RAF/MEK/ERK signaling can be observed in a large number of cancers, which is most commonly due to mutations in RAS or BRAF. 332 , 333 BRAF mutations (mostly BRAF V600E) have been identified in ~40–50% of melanomas and 6% of other malignancies, which results in several-fold hyperactivation of BRAF and continuous activation of downstream MEK and ERK. 333 , 334 Due to the picomolar affinities of RAS to GTP, RAS was taken as an undruggable target until the appearance of irreversible small-molecule inhibitors of KRAS G12C in recent years. 335 Therefore, BRAF and the downstream kinases MEK1/2 and ERK are considered attractive therapeutic targets for malignant tumors, especially melanoma. Small-molecule BRAF inhibitors can be classified into two types. Type-I inhibitors stabilize the kinase in its active (DFG-in) conformation by occupying the ATP-binding pocket. 336 The FDA has approved three type-I competitive BRAF inhibitors for the treatment of non-resectable BRAF V600E/K/D mutant melanoma as single agents or combined with MEK inhibitors including vemurafenib in 2011, 337 dabrafenib in 2013, 338 and encorafenib in 2018 (Table 3 ). 339 Type-II BRAF inhibitors bind to the adjacent hydrophobic sites of ATP-binding pocket and stabilize the kinase in its inactive (DFG-out) conformation. 336 Sorafenib, a representative type-II pan-RAF inhibitor, is a multikinase inhibitor. This agent was initially developed as a RAF kinase inhibitor; however, it also showed inhibitory activity against VEGFR-1/2/3, PDGFR, FLT1, KIT, and RET. It is currently approved by FDA for the treatment of unresectable HCC, advanced RCC, and thyroid carcinoma refractory to radioactive iodine, but not for melanoma due to the unsatisfactory results of clinical trials. 340 Table 3 Properties of approved small-molecule inhibitors of serine/theonine kinases Chemical structure Name Targets Approved indications (year) Corporation Vemurafenib (Zelboraf) BRAF Melanoma (2011) Roche/Genentech Dabrafenib (Tafinlar) BRAF/CRAF Melanoma (2013) NSCLC (2017) ATC (2018) Novartis/GlaxoSmithkline Encorafenib (Braftovi) BRAF Melanoma (2018) Array Trametinib (Mekinist) MEK1/2 Melanoma (2013) NSCLC (2017) ATC (2018) Novartis/GlaxoSmithkline Cobimetinib (Cotellic) MEK1/2 Melanoma (2015) Roche/Genentech/ Exelixis Binimetinib (Balimek) MEK1/2 Melanoma (2018) Array Selumetinib (Koselugo) MEK1/2 Neurofibromatosis type 1 (2020) Plexiform neurofibromas (2020) AstraZeneca/Merck Palbociclib (Aiboxin) CDK4/6 Breast cancer (2015) Pfizer Ribociclib (Kisqali) CDK4/6 Breast cancer (2017) Novartis Abemaciclib (Verzenio) CDK4/6 Breast cancer (2017) Lilly Idelalisib (Zydelig) PI3Kδ CLL (2014) Follicular lymphoma (2014) Gilead Copanlisib (Aliqopa) Pan-PI3K Follicular lymphoma (2017) Bayer Duvelisib (Copiktra) PI3Kγ/δ CLL (2018) SLL (2018) Follicular lymphoma (2018) Verastem Alpelisib (Piqray) PI3Kα Breast cancer (2019) Novartis Temsirolimus (Torisel) mTOR RCC (2007) Pfizer Everolimus (Afinitor) mTOR RCC (2009) Pancreatic cancer (2011) Breast cancer (2012) Novartis Sirolimus (Rapamune) mTOR LAM (2015) Pfizer The discovery and use of BRAF inhibitors opened up an avenue for the development of inhibitors of MEK1/2 and ERK, downstream targets of RAS/RAF/MEK/ERK signaling. Three MEK1/2 inhibitors have been approved in combination with BRAF inhibitors for the treatment of unresectable or metastatic melanoma harboring BRAF V600E/K mutation including reversible allosteric inhibitors trametinib and binimetinib as well as cobimetinib, a highly selective inhibitor that restrains the catalytic activity of MEK1/2 (Table 3 ); trametinib is also administered as a monotherapy. 341 The FDA-approved combination regimens of BRAF and MEK inhibitors (vemurafenib plus cobimetinib, dabrafenib plus trametinib, and encorafenib plus binimetinib) can achieve a longer PFS in patients with melanoma compared with BRAF inhibitor monotherapy in clinical trials, and encorafenib-binimetinib combination had the best toxicity profile. 342 The combination of dabrafenib and trametinib has also been approved for the treatment of NSCLC or anaplastic thyroid cancer (ATC) patients harboring BRAF V600E mutat

The phosphatidylinositol 3-kinase (PI3K)/V-AKT murine thymoma viral oncogene homolog (AKT)/mammalian target of rapamycin (mTOR) signaling pathway has an important role in cell growth, proliferation, survival, apoptosis, and motility and is frequently activated in human cancer. 353 PI3Ks are a family of lipid kinases that catalyze the phosphorylation of phosphatidylinositol D3. According to the structural characteristics of the subunits and substrates, PI3Ks can be divided into three classes. Of these, class I PI3K is the major isoform implicated in cancer and can be further subdivided into class IA and class IB, which are activated by RTKs and GPCRs, respectively. Class IA consists of PI3Kα, PI3Kβ, and PI3Kδ encoded by PIK3CA , PIK3CB , and PIK3CD genes, respectively. Class IB comprises only the PI3Kγ subtype encoded by PIK3CG. 386 , 387 PI3K/AKT/mTOR pathway is activated by various mechanisms in oncogenesis and progression. PIK3CA gene is frequently dysregulated in multiple cancers, both by point mutations and amplification. The tumor suppressor PTEN negatively regulates the PI3K pathway by dephosphorylating PIP3 to PIP2. It is also mutated frequently and results in loss of function in human cancers, which upregulates PIP3 levels and leads to constitutive activation of AKT and downstream components. Moreover, as the immediate downstream effector of PI3K, amplification and activating mutations of AKT are often observed in solid tumors. 388 Hyperactivation of PI3K/AKT/mTOR signaling is not only common in a variety of tumors but also closely related to drug resistance (e.g., resistance mechanism of EGFR inhibitors); thus, this pathway has become an attractive target for developing antitumor targeted drugs. Many small-molecule inhibitors of PI3K, AKT, and mTOR have been developed in the past few years. However, only several PI3K and mTOR inhibitors have been approved for cancer treatment. The early developed PI3K inhibitors are mostly pan-PI3K inhibitors that are capable of binding to all class I PI3Ks, such as wortmannin and LY294002. However, due to their poor PK properties, they are not ultimately approved for clinical use. The second-generation isoform-selective PI3K inhibitors are highly selective for the four isoforms of the class I PI3K catalytic subunit p110 (α, β, γ, and δ). As indicated in Table 3 , idelalisib is the first approved selective PI3Kδ inhibitor based on its efficacy in the treatment of relapsed or refractory CLL patients. It is also recommended for the treatment of lymphocytic lymphoma patients who have received at least two prior systemic therapies or patients with relapsed follicular B-cell NHL. 389 Copanlisib, approved in September 2017, is a pan-PI3K inhibitor with IC 50 values of 0.5, 3.7, 6.4, and 0.7 nM against class I PI3K-α, β, γ, and δ isoforms, respectively. 390 It is used clinically for the treatment of adult patients with relapsed follicular lymphoma who have received at least two prior systemic therapies. Duvelisib is an oral dual PI3Kγ and PI3Kδ inhibitor with IC 50 values of 27 and 2.5 nM, respectively. 391 It prevents the activation of PI3K-γ and δ isoforms by competitively and reversibly binding to the ATP-binding pocket of the p110 subunit. 392 The FDA has approved duvelisib for the treatment of adult patients with relapsed or refractory CLL, SLL, and follicular lymphoma after at least two prior therapies. The newly approved alpelisib is a selective inhibitor targeting the α-isoform of class I PI3K, with an in vitro IC 50 of 4.6 nM. It is indicated in combination with fulvestrant for the treatment of postmenopausal women and men with HR-positive, HER2-negative, PIK3CA -mutated, advanced, or metastatic breast cancer as detected by an FDA-approved test following progression on or after an endocrine-based regimen. 393 This is also the first PI3K inhibitor approved for clinical treatment of breast cancer. In addition, there are also a variety of PI3K inhibitors undergoing clinical evaluation, such as the pan-class I PI3K inhibitors XL147 and ZSTK474, PI3Kγ inhibitor IPI-549, and PI3Kα inhibitor serabelisib. 394 – 397 Some of them have progressed to phase II or III trials and have good prospects for approval. In addition, the promising efficacy of PI3K inhibitors raised the question of whether the combined inhibition of multiple pathway components could further improve the efficacy without excessive side effects. Developing dual PI3K/mTOR inhibitors is a predominant strategy due to the high homology between the ATP-binding domain of p110 and the catalytic site of mTOR. Several dual PI3K/mTOR inhibitors have entered clinical trials, such as bimiralisib, dactolisib (BEZ235), GDC-0084 (RG7666), and gedatolisib (PKI-587/PF-05212384). 398 – 400 Current clinical trials focus on combining them with a range of other antitumor agents. Notably, GDC-0084 is orally administered and can penetrate the BBB. It is currently specifically developed for patients with glioma or brain metastase

Histone deacetylases (HDACs) are important epigenetic regulators that remove the acetyl groups from the N-acetylated lysine residues of histones and various non-histone substrates. To date, 18 HDACs have been identified in mammals and are characterized into 4 subfamilies: class I HDACs (HDACs 1, 2, 3, 8), class II HDACs (class IIa: HDACs 4, 5, 7, 9; class IIb: HDACs 6, 10), class III HDACs (Sirt1-7) and class IV HDAC (HDAC11). 441 , 442 Class I, II, and IV HDACs are Zn 2+ -dependent histone deacetylases and can be restrained by inhibitors (such as TSA and SAHA) that occupy the catalytic core of the zinc-binding site. Class III HDACs require nicotinamide adenine dinucleotide (NAD + ) for their activity. 441 – 443 Aberrant upregulation of HDACs has been reported in various types of cancers. Such changes alter the transcription of oncogenes and tumor suppressor genes, which are closely associated with cell proliferation, apoptosis, differentiation, migration, and cancer angiogenesis. 443 , 444 Inhibition of HDACs has proven to be an effective strategy for cancer therapy. To date, a variety of HDAC inhibitors have been approved by the FDA (Table 4 ) or are undergoing clinical trials. Based on chemical structural differences, HDAC inhibitors can be divided into four categories: hydroxamates, cyclic tetrapeptides, benzamides, and aliphatic acids. 445 Hydroxamide HDAC inhibitors inhibit HDAC activity through coordination with Zn 2+ . TSA was the first natural hydroxamate HDAC inhibitor. Its structural analog vorinostat (SAHA) is a pan-inhibitor of classical classes of HDACs (I, II, and IV) and was approved in October 2006 for the treatment of cutaneous T-cell lymphoma (CTCL) (Table 4 ). 446 Belinostat and panobinostat are two other hydroxamate pan-HDAC inhibitors that are approved for the treatment of relapsed or refractory peripheral T-cell lymphomas (PTCL) and drug-resistant MM, respectively. 447 , 448 Both of them are derivatives of M-carboxycinnamic acid bishydroxamate. Cyclic tetrapeptides are a class of HDAC inhibitors with complex structures. Romidepsin isolated from Chromobacterium violaceum is the only such inhibitor granted US FDA approval. Its indications were initially only CTCL and expanded to PTCL in November 2011, and the objective response rate was 34% in CTCL patients and 25% in patients with PTCL. 449 Moreover, tucidinostat (chidamide), containing pyridine and N -(2-amino-5-fluorophenyl)-benzamide groups, is a benzamide inhibitor of HDAC1/2/3/10. It was developed wholly in China and approved for the treatment of refractory or relapsed PTCL in 2015 by the NMPA. 450 Aliphatic acids, such as valproic acid and phenylbutyrate, show relatively weak inhibitory activity against class I and class II HDACs, and both of these aliphatic acid HDAC inhibitors have already been approved for some non-oncological uses in the clinic and are recently under clinical evaluation for cancer therapy. 443 Sirtuin (class III HDAC) inhibitors include the allosteric non-competitive pan-inhibitor nicotinamide and Sirt-specific inhibitors such as EX-527, sirtinol, and cambinol. 451 Although Sirt inhibitors have been reported to be useful for cancer treatment, so far no such inhibitors have been approved for clinical use. Moreover, more than 10 other HDAC inhibitors have undergone or are undergoing clinical trials as monotherapy or in combination therapy in patients with hematologic malignancies or solid tumors. 452 , 453 For example, the benzamide inhibitor entinostat is currently assessed in phase III trials ( NCT02115282 and NCT03538171 ) for the clinical benefit in patients with HR-positive, HER2-negative, locally advanced, or metastatic breast cancer. A phase I dose-escalation multicentre trial ( NCT00741234 ) demonstrated that the hydroxamate HDAC inhibitor pracinostat was safe, with modest single-agent activity in patients with advanced hematological malignancies. 454 Currently, the clinical activity of HDAC inhibitors as monotherapy is largely restricted to hematological malignancies, including lymphomas, leukemia, and MM. In solid tumors, HDAC inhibitors just showed limited single-agent activity, which may be attributed to the non-specific blocking of angiogenesis and inflammation as well as the poor PK properties of some agents. 443 , 455 Inhibition of tumor angiogenesis hinders drug delivery in solid tumors. The anti-inflammatory effect may induce apoptosis of tumor-fighting immune cells. Another obstacle limiting the clinical use of HDAC inhibitors is their side effects. The common side effects associated with vorinostat, belinostat, and romidepsin were nausea, anorexia, fatigue, and vomiting, which are mostly manageable, but some agents may cause more serious toxicities. 455 Currently, major efforts to overcome these obstacles are focused on the development of small-molecule isoforms or class-selective HDAC inhibitors. Selective inhibitors can also be used as chemical probes to explore the epigenetic effects and b

The B-cell lymphoma 2 (BCL-2) family of proteins consists of more than 20 members that regulate the intrinsic apoptosis pathway, and fall into three subfamilies (anti-apoptotic proteins, pro-apoptotic proteins, and the cell death mediators) based on their structure and function. 500 Anti-apoptotic proteins such as BCL-2 and the closely related proteins BCL-XL, BCL-W, MCL-1, and A1/BFL-1 have four tandem BCL-2 homology (BH) domains and promote cell survival. 500 , 501 While the multi-region pro-apoptotic effector proteins (BAX and BAK) and BCL-2 homology 3 (BH3)-only pro-apoptotic proteins (BIM, PUMA, BAD, or NOXA) promote cell apoptosis upon diverse cellular stresses by the alternation of mitochondrial outer membrane permeabilization and release of cytochrome C from the mitochondria (Fig. 4 ). 501 , 502 The interaction between the anti-apoptotic and pro-apoptotic BCL-2 family of proteins regulates the apoptotic state of cells. Dysregulation of the apoptosis pathway is common in malignant tumors, especially in hematologic malignancies. 503 – 505 BCL-2 was first discovered as an inhibitor of cellular apoptosis, and the in-depth understanding of this target promotes the development and application of BCL-2 inhibitors in cancer treatment. 506 , 507 Fig. 4 Schematic illustration of extrinsic and intrinsic pathways of apoptosis. In healthy cells, anti-apoptotic BCL-2 proteins (BCL-2, BCL-XL, BCL-W, MCL-1, and A1/BFL-1) bind to and inhibit activators (BH3-only proteins) and effectors (BAX and BAK). Treatment with BCL-2 inhibitors releases the inhibitory effects of anti-apoptotic BCL-2 proteins on activators and effectors. The subsequent activation and oligomerization of the pro-apoptotic proteins BAK and BAX result in the formation of mitochondrial outer membrane permeabilization (MOMP) and the release of cytochrome C as well as a second mitochondria-derived activator of caspase (SMAC) from the mitochondria. Cytochrome C can form a complex with procaspase 9 and apoptosis protease-activating factor 1 (APAF1), thereby activating caspase 9. Caspase 9 then activates procaspase 3 and procaspase 7, resulting in cell apoptosis. Figure created with BioRender.com Several early efforts have been made to target the anti-apoptotic BCL-2 family members including antisense oligonucleotide drug oblimersen, the natural product gossypol, and its synthetic derivatives. 508 A major breakthrough in targeting BCL-2 is the development of small-molecule BH3 mimetics ABT-737 and ABT-263 (navitoclax). 509 , 510 These agents can mimic the physiologic inhibitors of anti-apoptotic BCL-2 and its relatives, thereby promoting apoptosis. ABT-737 is the first synthetic BH3 mimetic discovered by structure-activity relationship and nuclear magnetic resonance screening. It binds to BCL-2, BCL-XL, and BCL-W with high affinity, and exhibits 500-fold weaker affinity for MCL1 and BFL-1/A1. 509 However, ABT-737 has poor bioavailability and requires continuous parenteral administration, which hindered its clinical development. Navitoclax is a structurally related molecule of ABT-737 with high oral bioavailability (~50% in dogs) and entered clinical trials in 2006. 511 Although the efficacy of navitoclax was observed in preclinical and clinical studies, dose-dependent thrombocytopenia caused by BCL-XL suppression is the major obstacle restricting the clinical application of navitoclax. 510 Initial efforts on the BH3 mimetics ABT-737 and navitoclax facilitated the successful development of ABT-199 (venetoclax), a highly selective BH3 mimetic with greater affinity for BCL-2 but a much lower affinity for BCL-XL and BCL-W. 512 In a single-arm phase II trial of venetoclax monotherapy, 79.4% of patients with CLL with the 17p deletion achieved an objective response over a median of 12 months among 107 enrolled patients, and complete remission was achieved in a median of 8.2 months ( NCT01889186 ). 513 Due to its remarkable efficacy and safety, the FDA-approved venetoclax for the treatment of CLL patients with 17p deletion in April 2016 (Table 5 ). This indication was subsequently expanded in June 2018 to include patients with CLL or SLL, with or without 17p deletion, who have received at least one prior therapy. Moreover, in November 2018, venetoclax was approved in combination with standard chemotherapy agents, including azacitidine, decitabine, and low-dose cytarabine, for the treatment of newly diagnosed AML in adults who are 75 years of age or older, or who have comorbidities that preclude the use of intensive induction chemotherapy; this approval was based on the impressive results of two clinical studies conducted in this population. 514 , 515 As a BCL-2-specific inhibitor, venetoclax is also under assessment for the treatment of many other malignancies in the clinic, including solid tumors, and most of the clinical trials have tested its role in combination therapies. 516 Table 5 Properties of approved small-molecule inhibitors of BCL-2, hedgehog pathway,

Proteasomes are large multicatalytic enzyme complexes that are expressed in the nucleus and cytoplasm of all eukaryotic cells and are responsible for more than 80% protein degradation in human cells. 548 The ubiquitin–proteasome system (UPS) has an important role in maintaining cellular protein homeostasis and regulating numerous biological processes, such as cell survival, signal transduction, DNA repair, and antigen presentation. 549 Most misfolded, unassembled, or damaged proteins that could otherwise form potentially toxic aggregates are degraded via UPS, in which proteins are tagged by ubiquitin and then recognized and degraded into small peptides by the proteasome complex (Fig. 6 ). Structurally, all proteasomes contain a common core, referred to as the 20S proteasome. The 20S core consists of a cylinder made of four stacked rings: 2 identical outer α-rings and 2 identical inner β-rings, each containing 7 distinct but related subunits. 548 , 550 The specificity of the 20S proteasome for substrate action depends on the peptide bond on the N2 terminal threonine residue of the β1, β2, β5 subunits. Dysfunction of the UPS is related to multiple human diseases, such as cancers, autoimmune diseases, and genetic diseases; 550 , 551 thus, much work has been conducted by targeting the UPS as a potential treatment strategy. Fig. 6 Proteasome inhibition acts through multiple mechanisms to induce cell apoptosis. Proteasome inhibition leads to NF-κB deactivation, thereby downregulating multiple pro-neoplastic pathways associated with cell proliferation, invasion, metastasis, and angiogenesis. Inhibition of proteasome activates the JNK signaling pathway and results in programmed cell death via caspase 3 and 7. Additionally, proteasome inhibition can indirectly cause apoptosis by preventing the degradation of pro-apoptotic family proteins such as BAX, BID, BIK, and BIM as well as NOXA. Inhibition of proteasome prevents the degradation of ubiquitinated proteins, which can increase endoplasmic reticulum (ER) stress and activate the UPR, cell cycle arrest, and subsequent apoptosis. Figure created with BioRender.com Multiple myeloma (MM) cells produce excessive paraproteins, and their growth is dependent on proteasome-regulated signaling pathways. Therefore, MM cells are particularly susceptible to proteasome inhibition, and proteasome inhibitors (PIs) have become the backbone of MM clinical therapy. 552 Bortezomib is the first approved PI for the treatment of relapsed or refractory MM (Table 5 ). It is a peptide boronic acid and reversibly acts on the β5 catalytic subunit of the proteasome. 553 The clinical application of bortezomib significantly improves the long-term outcomes for MM patients. Moreover, Bortezomib has also been approved for the treatment of MCL. Currently, there are more than 200 clinical trials related to bortezomib that focus on its combination with other agents, efficacy in other cancers, and even other noncancer applications such as graft-versus-host disease. 554 However, bortezomib treatment has several limitations including primary resistance in MM and MCL patients, relapse in many initially-responding patients, and induction of dose-limiting peripheral neuropathy (PN). 554 , 555 To conquer these limitations, the second-generation PIs have been developed, and many of them are derived from synthetic and natural products. 552 , 556 Carfilzomib approved by the FDA in 2012 is a second-generation PI and is derived from the natural product epoxomicin (Table 5 ). 557 Unlike bortezomib, carfilzomib is an irreversible inhibitor that contains an epoxyketone warhead, which could covalently bind to the N-terminal threonine-containing active sites of the 20S proteasome. 558 The irreversible nature of carfilzomib contributes to its efficacy even in MM patients relapsed from or refractory to bortezomib. In addition, the carfilzomib-containing regimens exhibit significantly reduced peripheral neurotoxicity, while cardiovascular events were observed in MM patients treated with carfilzomib. 557 Another second-generation PI ixazomib was approved in 2015 in combination with lenalidomide and dexamethasone for MM patients who have received at least one prior therapy (Table 5 ). 559 Ixazomib is an N-capped dipeptidyl leucine boronic acid that reversibly inhibits the CT-L proteolytic (β5) site of the 20S proteasome. 560 Ixazomib shares the same pharmacophore boronic acid residue with bortezomib, however, the elimination half-life of the former is much shorter than that of the latter (18 vs. 110 min), which may contribute to the improved safety profiles of xazomib over bortezomib. 559 , 560 Patients resistant to bortezomib can still benefit from ixazomib treatment. Moreover, it is worth mentioning that both bortezomib and carfilzomib require parenteral (intravenous or subcutaneous) administration, whereas ixazomib is the first oral PI and is a prodrug. 561 Ixazomib is rapidly hydrolyzed to the active form (MLN2238) under

With the evolution of modern molecular biology and the application of some advanced technologies such as computer-aided drug design, structure biology, and combinatorial chemistry, small-molecule targeted anti-cancer drugs have entered a rapid development stage. To date, 89 small-molecule targeted drugs have been approved by the FDA and/or NMPA to treat various cancers (Fig. 1 ). Thousands of targeted agents are undergoing clinical trials for cancer treatment (Supplementary Fig. S1 ). Among them, a large number of promising agents have advanced to phase III trials (Supplementary Tables S2 – S5 ). According to the prediction of the Business Research Company, the global anti-cancer drug market size will reach 200 billion dollars in 2021, among which targeted drugs are the “main force”. Despite the significant progress achieved, there are still some challenges that small-molecule targeted anti-cancer drugs face. The first major challenge is drug resistance. Almost all targeted anti-cancer drugs come across resistance after a period of time of clinical use. Drug resistance has been linked to many mechanisms, including gene mutation, amplification, CSCs, efflux transporters, apoptosis dysregulation, and autophagy, etc (Fig. 8 ). 593 – 599 Gene mutation is the main reason leading to anti-cancer drug resistance. There are two different views regarding drug-resistant gene mutations. One is that the gene mutations are induced by drugs. The other one is that the drug-resistant mutations have already existed. In the early stage of treatment, cancer cells with drug-sensitive mutations dominate and suppress the proliferation of cells containing drug-resistant mutations. After cells with sensitive mutations are killed, the resistant mutant cells become the mainstream and show resistance. Amplification of other genes is another common reason for anti-cancer drug resistance. For example, MET amplification accounts for about 20% of EGFR inhibitor-resistant cases. 46 CSCs are also thought to be an important reason for drug resistance and recurrence. 595 , 596 The CSC theory proposes that the different cells within a tumor, as well as metastasis deriving from it, originate from a single subpopulation of cells with self-renewal and differentiation capacities, which are similar to stem cells. Overexpression of efflux transporters, such as multidrug resistance transporter proteins, especially P-glycoprotein, which renders the resistance to chemotherapeutic drugs, also has a role in the targeted anti-cancer drug resistance. 597 , 598 In addition to these causes, apoptosis dysregulation and autophagy also could be responsible for anti-cancer drug resistance. 593 , 599 Fig. 8 Mechanisms and insights in drug resistance of small-molecule targeted anti-cancer agents. Figure created with BioRender.com Low efficiency is another major challenge for targeted anti-cancer drugs. As mentioned many times before, targeted anti-cancer drugs are effective only in a limited number of patients. For example, less than 20% of patients with NSCLC are sensitive to EGFR inhibitors (e.g., gefitinib and erlotinib). Patients that are sensitive to these EGFR inhibitors are found to harbor EGFR-activating mutations (for example, exon 19 deletion or exon 21 L858R point mutations). 74 TRK inhibitors larotrectinib and entrectinib have been approved for the treatment of patients with NTRK gene rearrangements, regardless of cancer type and patient age. These rearrangements can be found at high frequencies (up to 90%) in certain types of cancer, such as infantile fibrosarcoma (a rare disease), but the incidence is only 1% of all malignancies. 225 These highlight the importance of the identification of predictive biomarkers for response to targeted anti-cancer drugs. Currently, in order to deal with the major challenges of targeted anti-cancer drugs, many strategies have been applied, such as new generation anti-cancer drugs against drug resistance mutations, multitarget drugs, combination therapy, and drugs targeting CSCs. 594 , 600 , 601 In addition to these, several new research trends in this area deserve attention. The first one is the drug discovery against new type cancer targets. For example, in the past years, some new epigenetic regulatory proteins have garnered increasing attention, such as RNA m6A methylation-related proteins (METTL3/14, FTO, ALKBH5, WTAP, and YTHDFs). 602 , 603 microRNAs (miRNAs) are another new type of cancer targets, which are frequently dysregulated in cancers and may serve as promising targets for cancer therapy. Currently, some effects have been paid to discover small-molecule inhibitors against miRNAs, such as miR-21 inhibitor AC1MMYR2 (also known as NSC211332), Lin28-let-7 inhibitors 6-hydroxy-DL-DOPA, SB/ZW/0065, and KCB3602. 604 Furthermore, some proteins that are previously thought undruggable may also be attractive anti-cancer targets. A typical example is KRAS , a most frequently mutated isoform of RAS proto-oncogene, whic