Design and synthesis of novel human epidermal growth factor receptor 2 (HER2)/epidermal growth factor receptor (EGFR) dual inhibitors bearing a pyrrolo[3,2-d]pyrimidine scaffold

Dual inhibitors of human epidermal growth factor receptor 2 (HER2) and epidermal growth factor receptor (EGFR) have been investigated for breast, lung, gastric, prostate, and other cancers; one, lapatinib, is currently approved for breast cancer. To develop novel HER2/EGFR dual kinase inhibitors, we designed and synthesized pyrrolo[3,2-d]pyrimidine derivatives capable of fitting into the receptors' ATP binding site. Among the prepared compounds, 34e showed potent HER2 and EGFR (HER1) inhibitory activities as well as tumor growth inhibitory activity. The X-ray cocrystal structures of 34e with both HER2 and EGFR demonstrated that 34e interacts with the expected residues in their respective ATP pockets. Furthermore, reflecting its good oral bioavailability, 34e exhibited potent in vivo efficacy in HER2-overexpressing tumor xenograft models. On the basis of these findings, we report 34e (TAK-285) as a promising candidate for clinical development as a novel HER2/EGFR dual kinase inhibitor.     

Lapatinib ditosylate: expanding therapeutic options for receptor tyrosine-protein kinase erbB-2-positive breast cancer

Receptor tyrosine-protein kinase erbB-2 (HER2)-positive breast cancer is a specific entity with an aggressive behavior. Trastuzumab, a monoclonal antibody targeting erbB-2 (HER2) deeply transformed the outcome in patients. Nevertheless, resistance to trastuzumab is still a major concern. Lapatinib ditosylate is an orally available, small molecule targeting the tyrosine activity of the HER2 receptor. Lapatinib as a single agent and in combination therapy showed interesting activity in trastuzumab-resistant advanced tumors. In addition, lapatinib use seemed suitable in recurrent locally advanced inflammatory breast cancer and brain metastases. More recently, the Neo-ALTTO (NeoAdjuvant Lapatinib and/or Trastuzumab Treatment Optimisation) trial showed that lapatinib in combination with trastuzumab and paclitaxel significantly improved the pathological complete response in a neoadjuvant setting. Several clinical trials are still ongoing and data that may change current clinical practice are awaited with much interest.     

HER2基因突变晚期非小细胞肺癌治疗进展

肺癌是全球范围内一种高发病率和高死亡率的恶性肿瘤,非小细胞肺癌（NSCLC）是最常见的亚型,发生率约85%。人表皮生长因子受体2（HER2）是受体酪氨酸激酶（RTK）的重要成员之一,NSCLC中HER2基因主要表现为HER2突变、HER2扩增和HER2过表达三种形式。目前,靶向HER2突变的酪氨酸激酶抑制剂、单克隆抗体和抗体偶联药物在HER2突变的NSCLC中显示出一定的临床疗效,但相关免疫治疗疗效有限。本文就HER2突变相关NSCLC的治疗进展进行综述,以期为进一步完善HER2突变晚期NSCLC临床诊疗策略提供理论依据。     

Recent advances in novel targeted therapies for HER2-positive breast cancer

The monoclonal antibody trastuzumab has improved the outcomes of patients with breast cancer that overexpresses the human epidermal growth factor receptor 2 (HER2). However, despite this advancement, many tumors develop resistance and novel approaches are needed. Recently, a greater understanding of cellular biology has translated into the development of novel anti-HER2 agents with varying mechanisms of action. The small molecule tyrosine kinase inhibitor lapatinib has demonstrated activity in HER2-positive metastatic breast cancer (MBC) and in the preoperative setting. Pertuzumab is a monoclonal antibody with a distinct binding site from trastuzumab, which inhibits receptor dimerization. In recent studies, the addition of pertuzumab to combination therapy has led to improvements in progression-free survival in patients with HER2-positive MBC and higher response rates in the preoperative setting. An alternative approach is the use of novel antibody-drug conjugates such as trastuzumab-emtansine, which recently demonstrated activity in MBC. Neratinib, a pan-HER tyrosine kinase inhibitor, which irreversibly inhibits HER1 and HER2, also has proven activity in MBC. A range of compounds is being developed to attempt to overcome trastuzumab resistance by targeting heat shock protein 90, a molecular chaperone required for the stabilization of cellular proteins. Furthermore, agents are being developed to inhibit the mammalian target of rapamycin, a downstream component of the PTEN/PI3K pathway, which has been implicated in trastuzumab resistance. Finally, there are emerging data indicating that combinations of anti-HER2 agents may circumvent resistance mechanisms and improve patient outcomes. In this review, recent data on these emerging agents and novel combinations for HER2-positive breast cancer are discussed.     

Pharmacological strategies to overcome HER2 cross-talk and Trastuzumab resistance

Approximately 20-30% of breast cancers show increased expression of the HER2 receptor tyrosine kinase. Trastuzumab (Herceptin) is a clinically approved anti-HER2 monoclonal antibody. Many patients with HER2-overexpressing metastatic breast cancer respond to trastuzumab; however, a subset display primary drug resistance. In addition, many patients who initially respond to trastuzumab ultimately develop disease progression. Multiple molecular mechanisms contributing to trastuzumab resistance have been proposed in the literature. These mechanisms include cross-signaling from related HER/erbB receptors and compensatory signaling from receptors outside of the HER/erbB family, including receptors for insulin-like growth factor-I, vascular endothelial growth factor, and transforming growth factor beta. The major downstream signaling pathway activated by HER2 cross-talk is PI3K/mTOR, and a potential integrator of receptor cross-talk is Src-focal adhesion kinase (FAK) signaling. PI3K, Src, and FAK have independently been implicated in trastuzumab resistance. In this review, we will discuss pharmacological inhibition of HER2 cross-talk as a strategy to treat trastuzumab-refractory HER2-overexpresssing breast cancer.     

Novel Peptidomimetics for Inhibition of HER2:HER3 Heterodimerization in HER2‐Positive Breast Cancer

The current approach to treating HER2‐overexpressed breast cancer is the use of monoclonal antibodies or a combination of antibodies with traditional chemotherapeutic agents or kinase inhibitors. Our approach is to target clinically validated HER2 domain IV with peptidomimetics and inhibit the protein–protein interactions (PPI) of HERs. Unlike antibodies, peptidomimetics have advantages in terms of stability, modification, and molecular size. We have designed peptidomimetics (compounds 5 and 9 ) that bind to HER2 domain IV, inhibit protein–protein interactions, and decrease cell viability in breast cancer cells with HER2 overexpression. We have shown, using enzyme fragment complementation and proximity ligation assays, that peptidomimetics inhibit the PPI of HER2:HER3. Compounds 5 and 9 suppressed the tumor growth in a xenograft mouse model. Furthermore, we have shown that these compounds inhibit PPI of HER2:HER3 and phosphorylation of HER2 as compared to control in tissue samples derived from in vivo studies. The stability of the compounds was also investigated in mouse serum, and the compounds exhibited stability with a half‐life of up to 3 h. These results suggest that the novel peptidomimetics we have developed target the extracellular domain of HER2 protein and inhibit HER2:HER3 interaction, providing a novel method to treat HER2‐positive cancer.     

Recent advances in the design and discovery of small-molecule therapeutics targeting HER2/neu

The human epidermal growth factor receptor (HER) family is a highly explored and promising anticancer drug target. At present, several investigational agents targeted to the HER family of receptors are in various stages of development. Five drugs are already in the clinic for the treatment of cancers that overexpress HER family receptors. Two FDA-approved small-molecule drugs, gefitinib and erlotinib, inhibit HER1 tyrosine kinase activity. Two mAbs, cetuximab and panitumumab, target the extracellular domain of HER1, and another, trastuzumab, targets the extracellular domain of HER2. HER2 is a prominent member of the HER family of receptor tyrosine kinases and serves as a preferred dimerization partner for other HER family members. This paper reviews recently patented small-molecule inhibitors of HER2 receptor kinase activity, and inhibitors of HER2 expression and shedding. Apart from the well-explored quinazoline class of compounds (e.g., lapatinib), arylazole, benzodithiazole, pyrrolopyridazine, pyrrolotriazine and pyrrolopyrimidine classes of compounds were also claimed as HER2 tyrosine kinase inhibitors. Most of these compounds show considerable activity against all the HER family as well as members from different families of tyrosine kinases. It remains to be established how the combination of selective HER inhibitors compare with the single-agent pan-kinase inhibitors in disrupting HER family mediated signalling pathways. Such information is of paramount importance in the clinical development of HER-targeted inhibitors.     

Targeting EGFR in pancreatic cancer treatment

The prognosis of patients with pancreatic cancer is extremely poor, and current systemic therapies provide marginal survival benefits for treated patients. The era of targeted therapies has offered a new avenue to search for potentially more effective strategies. Epidermal growth factor receptor (EGFR) is a member of the erbB/human epidermal growth factor receptor family of tyrosine kinases, which includes erbB2/HER2, erbB3/HER3 and erbB4/HER4. Epidermal growth factor receptor overexpression may be detected in up to 90% of pancreatic tumors. Two pharmacologic approaches have been successfully used to inhibit epidermal growth factor receptor function in cancer treatment: neutralizing monoclonal antibodies and small molecule tyrosine inhibitors. The randomized trials studying the addition of EGFR targeted agents to gemcitabine compared with gemcitabine alone have been disappointing, although results with the EGFR tyrosine kinase inhibitor erlotinib were statistically significant but clinically of marginal benefit. In this article, we review the epidermal growth factor receptor signaling network in pancreatic cancer, the strategies to increase the effectiveness of epidermal growth factor receptor inhibitors, and the clinical trials of these inhibitors in pancreatic cancer.     

Tyrphostins. 2. Heterocyclic and alpha-substituted benzylidenemalononitrile tyrphostins as potent inhibitors of EGF receptor and ErbB2/neu tyrosine kinases.

We have previously described a novel series of low molecular weight protein tyrosine kinase inhibitors which we named tyrphostins. The characteristic active pharmacophore of these compounds was the hydroxy-cis-benzylidenemalononitrile moiety. In this article we describe three novel groups of tyrphostins: (i) one group has the phenolic moiety of the cis-benzylidenemalononitrile replaced either with other substituted benzenes or with heteroaromatic rings, (ii) another is a series of conformationally constrained derivatives of hydroxy-cis-benzylidenemalononitriles in which the malononitrile moiety is fixed relative to the aromatic ring, and (iii) two groups of compounds in which the position trans to the benzenemalononitrile has been substituted by ketones and amides. Among the novel tyrphostins examined we found inhibitors which discriminate between the highly homologous EGF receptor kinase (HER1) and ErbB2/neu kinase (HER2). These findings may lead to selective tyrosine kinase blockers for the treatment of diseases in which ErbB2/neu is involved.     

Drug combinations enhancing the antineoplastic effects of erlotinib in high-grade glioma

Glioblastoma multiforme (GBM) is the most common malignant brain tumor in adults. Especially in this disease, qualitative and quantitative aspects render the dysregulated epidermal growth factor receptor (HER1/EGFR) an outstanding therapeutic target. A variety of therapeutic compounds was developed to target HER1/EGFR among which the clinically most advanced agents are small molecule tyrosine kinase (TK) inhibitors. Unfortunately, clinical studies examining their therapeutic efficacy have so far failed to document a major therapeutic break-through in the setting of GBM. Thus, the targeted approach against HER1/EGFR likely requires a synergistic drug combination strategy to ultimately become successful in this disease. This patents review focuses on innovative therapeutic strategies combining HER1/EGFR-targeted TK inhibitors with novel agents which for the most part have not been evaluated for the treatment of GBM yet but which constitute interesting candidates for further evaluation in this setting.     

Computational‐based discovery of FAK FERM domain chemical probes that inhibit HER2‐FAK cancer signaling

The N‐terminal FERM domain of focal adhesion kinase (FAK) contributes to FAK scaffolding and interacts with HER2, an oncogene and receptor tyrosine kinase. The interaction between HER2 and FAK drives resistance to FAK‐kinase domain inhibitors through FAK Y397 transphosphorylation and FAK re‐activation upon inhibition. As such, FAK FERM remains an attractive drug discovery target. In this report, we detail an alternative approach to targeting FAK through virtual screening‐based discovery of chemical probes that target FAK FERM. We validated the binding interface between HER2 and FAK using site‐directed mutagenesis and GST pull‐down experiments. We assessed the ligandability of key‐binding residues of HER2 and FAK utilizing computational tools. We developed a virtual screening method to screen ~200,000 compounds against the FAK FERM domain, identifying 20 virtual chemical probes. We performed GST pull‐down screening on these compounds, discovering two hits, VS4 and VS14, with nanomolar IC₅₀s in disrupting HER2‐FAK. We performed further testing, including molecular docking, immunofluorescence, phosphorylation, and cellular invasion assays to evaluate the compounds’ biological effects. One probe, VS14, was identified with the ability to block both auto‐ and transphosphorylation of Y397. In all, these studies identify two new probes that target FAK FERM, enabling future investigation of this domain.     

Design and synthesis of pyrrolo[3,2-d]pyrimidine human epidermal growth factor receptor 2 (HER2)/epidermal growth factor receptor (EGFR) dual inhibitors: exploration of novel back-pocket binders

To develop novel human epidermal growth factor receptor 2 (HER2)/epidermal growth factor receptor (EGFR) kinase inhibitors, we explored pyrrolo[3,2-d]pyrimidine derivatives bearing bicyclic fused rings designed to fit the back pocket of the HER2/EGFR proteins. Among them, the 1,2-benzisothiazole (42m) ring was selected as a suitable back pocket binder because of its potent HER2/EGFR binding and cell growth inhibitory (GI) activities and pseudoirreversibility (PI) profile as well as good bioavailability (BA). Ultimately, we arrived at our preclinical candidate 51m by optimization of the N-5 side chain to improve CYP inhibition and metabolic stability profiles without a loss of potency (HER2/EGFR inhibitory activity, IC(50), 0.98/2.5 nM; and GI activity BT-474 cells, GI(50), 2.0 nM). Reflecting the strong in vitro activities, 51m exhibited potent tumor regressive efficacy against both HER2- and EGFR-overexpressing tumor (4-1ST and CAL27) xenograft models in mice at oral doses of 50 mg/kg and 100 mg/kg.     

Erlotinib in glioblastoma: lost in translation?

Glioblastoma represents the most common primary brain tumor in adults. Despite improvements of multimodal therapy, the prognosis of this disease remains unfavorable. Thus, great efforts have been made to identify therapeutic agents directed against those specific molecular targets whose presence was shown to be associated with worse clinical outcomes. The epidermal growth factor receptor (HER1/EGFR) has been identified as one such target, and different compounds were developed to inhibit HER1/EGFR and/ or its mutant form, EGFRvIII. However, clinical trials did not confirm the initial enthusiasm conveyed by promising results from experimental studies. Therefore, a therapeutic approach directed at inhibiting solely HER1/EGFR does not seem to translate into a clinical benefit. This review discusses the current therapeutic situation in the setting of glioblastoma while putting the spotlight on erlotinib, a HER1/EGFR-targeted small molecule tyrosine kinase inhibitor.     

Acute left ventricular dysfunction induced by a panHER and VEGFR tyrosine kinase inhibitor in a phase I trial

Tyrosine kinase inhibitors (TKI) or monoclonal antibodies targeting EGFR, HER2 or VEGFR receptors have demonstrated substantial clinical benefit in patients with advanced breast cancer, colon cancer, head and neck cancer, non-small cell lung cancer, and renal cell carcinoma. Nevertheless, these drugs have some target related adverse effects, particularly cardiovascular toxicities. We report here the case of a patient included in a phase I trial of a new compound, a tyrosine kinase inhibitor targeting HER1, HER2, HER4 and VEGFR2. The patient developed during this treatment an acute and transient left ventricular systolic dysfunction. Careful management of this adverse effect allowed the patient to continue therapy and to achieve a major partial response.     

EGFR和ErbB2双重酪氨酸激酶

拉帕替尼是一种人源2型表皮生长因子受体(HER2)和表皮生长因子受体(EGFR)双重酪氨酸激酶抑制剂,与卡培他滨联用治疗女性HER2阳性的乳腺癌。我们利用MEDLINE进行关键词为拉帕替尼的文献检索,对其药理作用、药动学、临床疗效及安全性,药物相互作用等进行综述。     

Foretinib (GSK1363089), a multi-kinase inhibitor of MET and VEGFRs, inhibits growth of gastric cancer cell lines by blocking inter-receptor tyrosine kinase networks

To explore the mechanism of action of foretinib (GSK1363089), an oral multi-kinase inhibitor known to target MET, RON, AXL, and vascular endothelial growth factor receptors (VEGFRs), in gastric cancer, we evaluated the effects of the agent on cell growth and cell signaling in the following panel of gastric cancer cell lines: KATO-III, MKN-1, MKN-7, MKN-45, and MKN-74. Of these, only MKN-45 and KATO-III, which harbor MET and fibroblast growth factor receptor 2 (FGFR2) amplification, respectively, were highly sensitive to foretinib. In MKN-45, 1 μM of foretinib or PHA665752, another MET kinase inhibitor, inhibited phosphorylation of MET and downstream signaling molecules as expected. In KATO-III, however, PHA665752 inhibited phosphorylation of MET independently of downstream molecules. Further, 1 μM of foretinib or PD173074, a selective FGFR kinase inhibitor, inhibited phosphorylation of FGFR2 and downstream molecules, suggesting that foretinib targets FGFR2 in KATO-III. We confirmed this novel activity of foretinib against FGFR2 in OCUM-2M, another FGFR2-amplified gastric cancer cell line. Using a phospho-receptor tyrosine kinase array, we found that foretinib inhibits phosphorylation of epidermal growth factor receptor (EGFR), HER3 and FGFR3 via MET inhibition in MKN-45, and EGFR, HER3 and MET via FGFR2 inhibition in KATO-III. Knockdown of HER3 and FGFR3 in MKN-45 with siRNA resulted in the partial inhibition of cell signaling and cell growth. In conclusion, foretinib appears effective against gastric cancer cells harboring not only MET but also FGFR2 amplification, and exerts its inhibitory effects by blocking inter-RTK signaling networks with MET or FGFR2 at their core.     

Development of HER2-specific humanized antibody Herceptin (trastuzumab)

HER2 is a member of the human epidermal growth factor receptor family, possessing protein kinase activity in its cytoplasmic domain. There were evidences indicating that (1) amplification of HER2/neu gene and HER2 protein over-expression in tumor cells was observed in 25-30% of human breast cancer and (2) amplification of HER2/neu correlated with poor prognosis, including shorter disease-free and overall survival. These evidences suggested HER2 was a promising candidate for novel molecular targets of breast cancer therapy. Herceptin is a recombinant humanized monoclonal antibody generated by Genentech, Inc. for the treatment of HER2 over-expressed/HER2 gene amplified metastatic breast cancer (MBC). Preclinical studies demonstrated that the antibody had anti-tumor activity in vivo and in vitro, and additive or synergistic enhancement of anti-tumor activity of the antibody was observed in combination with various anti-tumor agents in mouse models. In clinical studies, apparent extension of overall survival was observed in HER2 overexpressing MBC patients. Herceptin is the first anticancer drug whose use as a treatment for MBC patients is decided based on the status of the HER2 gene amplification/HER2 protein over-expression. The development and standardization of HER2 test were a key strategy in clinical development of this drug, since appropriate selection of patients with HER2 over-expression was the essential point for success.     

FASTlab Radiosynthesis of the 18F‐labelled HER2‐binding Affibody molecule [18F]GE‐226

An ¹⁸F‐labelled human epidermal growth factor receptor (HER2) receptor binding radiotracer is a potential tool to non‐invasively identify HER2 positive tumour lesions in subjects with recurrent metastatic breast cancer. Having explored the manual radiochemistry to conjugate the Affibody molecule Z_HER2:2891 with [¹⁸F]4‐fluorobenzaldehyde, we have developed and optimised a full protocol for the automated GE FASTlab synthesiser. Our chemometric model predicted the best radiochemical purity for a short conjugation time (2.8 minutes), a low temperature (65°C), and a medium Affibody molecule precursor amount (5.5 mg). Under these optimised conditions, [¹⁸F]GE‐226 was produced after solid‐phase extraction purification with activity yield of 30% ± 7 (n = 18) and a radiochemical purity of 94% ± 2 (n = 18). The synthesis and purification was complete after 43 minutes and provided apparent molar activities of 12 to 30 GBq/μmol (n = 12) at the end of synthesis.     

Targeting ErbB receptors in high-grade glioma

High-grade gliomas, including glioblastoma, are among the most malignant and treatment-refractory human neoplasms. The tumors show high levels of resistance to conventional therapies (i.e. surgery, irradiation, and chemotherapy), and despite treatment advances patient outcome remains poor. New therapeutic options are needed. An especially interesting idea is the rational development of new therapies targeting molecules in cancer specific signaling pathways, thereby ideally increasing treatment efficacy and minimizing toxicity. Clearly, rational design requires thorough understanding of the molecular pathogenesis and resistance mechanisms. One highly promising approach is the targeted inhibition of ErbB growth factor receptors, which are recognized as key signaling pathways in many types of human tumors, including high-grade glioma. The ErbB receptor family of tyrosine kinases comprises four members: epidermal growth factor receptor (EGFR/ErbB1/HER1), ErbB2 (HER2/neu), ErbB3 (HER3) and ErbB4 (HER4). Physiologically, signaling is induced by ligand initiated receptor homo- or heterodimerization, activating intracellular downstream signaling pathways and leading to increased cell proliferation, anti-apoptosis and migration. A truncated, constitutively activated mutant EGFR (EGFRvIII) is associated with poor survival in GBM. Thus, to date anti-ErbB approaches are mainly focused on EGFR. The two major classes of anti-ErbB therapeutics are monoclonal antibodies (e.g. cetuximab, panitumumab) and small molecule Tyrosine kinase inhibitors (TKI, e.g. gefitinib, erlotinib, lapatinib). Some compounds entered clinical trials already, but clinical efficacy needs to be enhanced. Here we review current therapeutic advances targeting ErbB receptors in high-grade gliomas, and give a concise overview on current understanding of ErbB biology in gliomas, paving the way to novel rational therapeutic development.