Giorgio Caravatti

AEE788: a dual family epidermal growth factor receptor/ErbB2 and vascular endothelial growth factor receptor tyrosine kinase inhibitor with antitumor and antiangiogenic activity.

Aberrant epidermal growth factor receptor (EGFR) and ErbB2 expression are associated with advanced disease and poor patient prognosis in many tumor types (breast, lung, ovarian, prostate, glioma, gastric, and squamous carcinoma of head and neck). In addition, a constitutively active EGFR type III deletion mutant has been identified in non-small cell lung cancer, glioblastomas, and breast tumors. Hence, members of the EGFR family are viewed as promising therapeutic targets in the fight against cancer. In a similar vein, vascular endothelial growth factor (VEGF) receptor kinases are also promising targets in terms of an antiangiogenic treatment strategy. AEE788, obtained by optimization of the 7H-pyrrolo[2,3-d]pyrimidine lead scaffold, is a potent combined inhibitor of both epidermal growth factor (EGF) and VEGF receptor tyrosine kinase family members on the isolated enzyme level and in cellular systems. At the enzyme level, AEE788 inhibited EGFR and VEGF receptor tyrosine kinases in the nm range (IC(50)s: EGFR 2 nm, ErbB2 6 nm, KDR 77 nm, and Flt-1 59 nm). In cells, growth factor-induced EGFR and ErbB2 phosphorylation was also efficiently inhibited (IC(50)s: 11 and 220 nm, respectively). AEE788 demonstrated antiproliferative activity against a range of EGFR and ErbB2-overexpressing cell lines (including EGFRvIII-dependent lines) and inhibited the proliferation of epidermal growth factor- and VEGF-stimulated human umbilical vein endothelial cells. These properties, combined with a favorable pharmacokinetic profile, were associated with a potent antitumor activity in a number of animal models of cancer, including tumors that overexpress EGFR and or ErbB2. Oral administration of AEE788 to tumor-bearing mice resulted in high and persistent compound levels in tumor tissue. Moreover, AEE788 efficiently inhibited growth factor-induced EGFR and ErbB2 phosphorylation in tumors for >72 h, a phenomenon correlating with the antitumor efficacy of intermittent treatment schedules. Strikingly, AEE788 also inhibited VEGF-induced angiogenesis in a murine implant model. Antiangiogenic activity was also apparent by measurement of tumor vascular permeability and interstitial leakage space using dynamic contrast enhanced magnetic resonance imaging methodology. Taken together, these data indicate that AEE788 has potential as an anticancer agent targeting deregulated tumor cell proliferation as well as angiogenic parameters. Consequently, AEE788 is currently in Phase I clinical trials in oncology.Aberrant epidermal growth factor receptor (EGFR) and ErbB2 expression are associated with advanced disease and poor patient prognosis in many tumor types (breast, lung, ovarian, prostate, glioma, gastric, and squamous carcinoma of head and neck). In addition, a constitutively active EGFR type III deletion mutant has been identified in non-small cell lung cancer, glioblastomas, and breast tumors. Hence, members of the EGFR family are viewed as promising therapeutic targets in the fight against cancer. In a similar vein, vascular endothelial growth factor (VEGF) receptor kinases are also promising targets in terms of an antiangiogenic treatment strategy. AEE788, obtained by optimization of the 7H-pyrrolo[2,3-d]pyrimidine lead scaffold, is a potent combined inhibitor of both epidermal growth factor (EGF) and VEGF receptor tyrosine kinase family members on the isolated enzyme level and in cellular systems. At the enzyme level, AEE788 inhibited EGFR and VEGF receptor tyrosine kinases in the nm range (IC50s: EGFR 2 nm, ErbB2 6 nm, KDR 77 nm, and Flt-1 59 nm). In cells, growth factor-induced EGFR and ErbB2 phosphorylation was also efficiently inhibited (IC50s: 11 and 220 nm, respectively). AEE788 demonstrated antiproliferative activity against a range of EGFR and ErbB2-overexpressing cell lines (including EGFRvIII-dependent lines) and inhibited the proliferation of epidermal growth factor- and VEGF-stimulated human umbilical vein endothelial cells. These properties, combined with a favorable pharmacokinetic profile, were associated with a potent antitumor activity in a number of animal models of cancer, including tumors that overexpress EGFR and or ErbB2. Oral administration of AEE788 to tumor-bearing mice resulted in high and persistent compound levels in tumor tissue. Moreover, AEE788 efficiently inhibited growth factor-induced EGFR and ErbB2 phosphorylation in tumors for >72 h, a phenomenon correlating with the antitumor efficacy of intermittent treatment schedules. Strikingly, AEE788 also inhibited VEGF-induced angiogenesis in a murine implant model. Antiangiogenic activity was also apparent by measurement of tumor vascular permeability and interstitial leakage space using dynamic contrast enhanced magnetic resonance imaging methodology. Taken together, these data indicate that AEE788 has potential as an anticancer agent targeting deregulated tumor cell proliferation as well as angiogenic parameters. Consequently, AEE788 is currently in Phase I clinical trials in oncology.

Protein kinases as targets for anticancer agents: from inhibitors to useful drugs

Many components of mitogenic signaling pathways in normal and neoplastic cells have been identified, including the large family of protein kinases, which function as components of signal transduction pathways, playing a central role in diverse biological processes, such as control of cell growth, metabolism, differentiation, and apoptosis. The development of selective protein kinase inhibitors that can block or modulate diseases caused by abnormalities in these signaling pathways is widely considered a promising approach for drug development. Because of their deregulation in human cancers, protein kinases, such as Bcr-Abl, those in the epidermal growth factor-receptor (HER) family, the cell cycle regulating kinases such as the cyclin-dependent kinases, as well as the vascular endothelial growth factor-receptor kinases involved in the neo-vascularization of tumors, are among the protein kinases considered as prime targets for the development of selective inhibitors. These drug-discovery efforts have generated inhibitors and low-molecular weight therapeutics directed against the ATP-binding site of various protein kinases that are in various stages of development (up to Phase II/III clinical trials). Three examples of inhibitors of protein kinases are reviewed, including low-molecular weight compounds targeting the cell cycle kinases; a potent and selective inhibitor of the HER1/HER2 receptor tyrosine kinase, the pyrollopyrimidine PKI166; and the 2-phenyl-aminopyrimidine STI571 (Glivec(R), Gleevec) a targeted drug therapy directed toward Bcr-Abl, the key player in chronic leukemia (CML). Some members of the HER family of receptor tyrosine kinases, in particular HER1 and HER2, have been found to be overexpressed in a variety of human tumors, suggesting that inhibition of HER signaling would be a viable antiproliferative strategy. The pyrrolo-pyrimidine PKI166 was developed as an HER1/HER2 inhibitor with potent in vitro antiproliferative and in vivo antitumor activity. Based upon its clear association with disease, the Bcr-Abl tyrosine kinase in CML represents the ideal target to validate the clinical utility of protein kinase inhibitors as therapeutic agents. In a preclinical model, STI571 (Glivec(R), Gleevec) showed potent in vitro and in vivo antitumor activity that was selective for Abl, c-Kit, and the platelet-derived growth factor-receptor. Phase I/II studies demonstrated that STI571 is well tolerated, and that it showed promising hematological and cytogenetic responses in CML and clinical responses in the c-Kit-driven gastrointestinal tumors.

Characterization of the novel and specific PI3Kα inhibitor NVP-BYL719 and development of the patient stratification strategy for clinical trials.

Somatic PIK3CA mutations are frequently found in solid tumors, raising the hypothesis that selective inhibition of PI3Kα may have robust efficacy in PIK3CA-mutant cancers while sparing patients the side-effects associated with broader inhibition of the class I phosphoinositide 3-kinase (PI3K) family. Here, we report the biologic properties of the 2-aminothiazole derivative NVP-BYL719, a selective inhibitor of PI3Kα and its most common oncogenic mutant forms. The compound selectivity combined with excellent drug-like properties translates to dose- and time-dependent inhibition of PI3Kα signaling in vivo, resulting in robust therapeutic efficacy and tolerability in PIK3CA-dependent tumors. Novel targeted therapeutics such as NVP-BYL719, designed to modulate aberrant functions elicited by cancer-specific genetic alterations upon which the disease depends, require well-defined patient stratification strategies in order to maximize their therapeutic impact and benefit for the patients. Here, we also describe the application of the Cancer Cell Line Encyclopedia as a preclinical platform to refine the patient stratification strategy for NVP-BYL719 and found that PIK3CA mutation was the foremost positive predictor of sensitivity while revealing additional positive and negative associations such as PIK3CA amplification and PTEN mutation, respectively. These patient selection determinants are being assayed in the ongoing NVP-BYL719 clinical trials. Mol Cancer Ther; 13(5); 1117–29. ©2014 AACR.

Discovery of NVP-BYL719 a potent and selective phosphatidylinositol-3 kinase alpha inhibitor selected for clinical evaluation.

Phosphatidylinositol-3-kinase α (PI3Kα) is a therapeutic target of high interest in anticancer drug research. On the basis of a binding model rationalizing the high selectivity and potency of a particular series of 2-aminothiazole compounds in inhibiting PI3Kα, a medicinal chemistry program has led to the discovery of the clinical candidate NVP-BYL719.

Selective GRB2 SH2 inhibitors as anti-Ras therapy.

Given the key role of Ras in the mitogenic signaling by receptor tyrosine kinases, several targets upstream of Ras may prove to be excellent targets for drugs in the treatment of cancer caused by oncogenic tyrosine kinases. CGP78850 is a potent competitor of Grb2 SH2‐phosphopeptide interactions. This inhibitor has been obtained by rational drug design and is specific toward the Grb2 SH2 vs. other SH2 domains and the PTB domain of SHC in vitro. Accordingly, CGP78850 blocks epidermal growth factor receptor (EGFR)‐Grb2 and Shc‐Grb2 interactions in living cells. It also inhibits the growth of cells transformed by receptor tyrosine kinases, which transmit a proliferative signal through Grb2 to Ras, but not cells transformed by oncogenic Raf or cells that contain activating Ras mutations. Moreover, our results demonstrate that, in cells overexpressing receptor tyrosine kinases, such as the EGFR, Grb2 SH2 inhibitors induce expression of the cell cycle inhibitors p21Waf1/Cip1/CAP1 and p27Kip1 and reverse transformation. Int. J. Cancer 83:235–241, 1999.

Effect of Potent and Selective Inhibitors of the Grb2 SH2 Domain on Cell Motility

Cell motility has been correlated both with oncogenic invasiveness and metastatic potential. The development of selective inhibitors of motility has thus great potential importance. Grb2 is a SH2/SH3 domain-containing adaptor protein that links growth factor receptor tyrosine kinases to the Ras signaling pathway. We have developed specific small molecule inhibitors of the Grb2 SH2 domain as potential leads for drug discovery. Synthesis of the inhibitors and their effects on growth factor-induced growth in cells have been reported previously. In the current study, we establish that these inhibitors inhibit hepatocyte growth factor/scatter factor-induced A431 and Madin-Darby canine kidney cell motility and various cell motility-related events, including epidermal growth factor-induced ruffling of A431 cells and epidermal growth factor-induced translocation of the small GTPase Rac in these cells. We demonstrate for the first time a direct role for Grb2 in cell motility and indicate a new avenue for cancer therapeutics.

Synthesis and biological evaluation of highly potent analogues of epothilones B and D

A series of new epothilone B and D analogues incorporating fused hetero-aromatic side chains have been prepared. The synthetic strategy is based on olefin 3 as the common intermediate and allows variation of the side-chain structure in a highly convergent and stereoselective manner. Epothilone analogues 1a-d and 2a-d are more potent inhibitors of cancer cell proliferation than the corresponding parent epothilones B or D.

Modelling study of protein kinase inhibitors: Binding mode of staurosporine and origin of the selectivity of CGP 52411

SummaryA model for the binding mode of the potent protein kinase inhibitor staurosporine is proposed. Using the information provided by the crystal structure of the cyclic-AMP-dependent protein kinase, it is suggested that staurosporine, despite a seemingly unrelated chemical structure, exploits the same key hydrogen-bond interactions as ATP, the cofactor of the protein kinases, in its binding mode. The structure-activity relationships of the inhibitor and a docking analysis give strong support to this protein tyrosine kinase is rationalized on the basis of the model. It is proposed that this selectivity originates in the occupancy, by one of the anilino moieties of the inhibitor, of the region of the enzyme cleft that normally binds the ribose ring of ATP, which appears to possess a marked lipophilic character in this kinase.

Highly potent inhibitors of the Grb2-SH2 domain.

Highly potent inhibitors of the Grb2-SH2 domain have been synthesized. They share the common sequence: Ac-Pmp-Ac6c-Asn-NH-(3-indolyl-propyl). Different substituents at the 3-indolyl-propylamine C-terminal group were explored to further improve the activity. This is the first example of inhibitors of SH2 domains with sub-nanomolar affinity reported to date.

Entry into a new class of protein kinase inhibitors by pseudo ring design.

A pyrimidin-4-yl-urea motif forming a pseudo ring by intramolecular hydrogen bonding has been designed to mimic the pyrido[2,3-d]pyrimidin-7-one core structure of a well-established class of protein kinase inhibitors. Potent inhibition of a number of protein kinases was obtained with the first prototype compound synthesized to probe the design concept.