Structure-based design, synthesis, and evaluation of imidazo[1,2-b]pyridazine and imidazo[1,2-a]pyridine derivatives as novel dual c-Met and VEGFR2 kinase inhibitors.

To identify compounds with potent antitumor efficacy for various human cancers, we aimed to synthesize compounds that could inhibit c-mesenchymal epithelial transition factor (c-Met) and vascular endothelial growth factor receptor 2 (VEGFR2) kinases. We designed para-substituted inhibitors by using co-crystal structural information from c-Met and VEGFR2 in complex with known inhibitors. This led to the identification of compounds 3a and 3b, which were capable of suppressing both c-Met and VEGFR2 kinase activities. Further optimization resulted in pyrazolone and pyridone derivatives, which could form intramolecular hydrogen bonds to enforce a rigid conformation, thereby producing potent inhibition. One compound of particular note was the imidazo[1,2-a]pyridine derivative (26) bearing a 6-methylpyridone ring, which strongly inhibited both c-Met and VEGFR2 enzyme activities (IC50=1.9, 2.2 nM), as well as proliferation of c-Met-addicted MKN45 cells and VEGF-stimulated human umbilical vein endothelial cells (IC50=5.0, 1.8 nM). Compound 26 exhibited dose-dependent antitumor efficacy in vivo in MKN45 (treated/control ratio [T/C]=4%, po, 5mg/kg, once-daily) and COLO205 (T/C=13%, po, 15 mg/kg, once-daily) mouse xenograft models.

Design, synthesis, and evaluation of novel VEGFR2 kinase inhibitors: discovery of [1,2,4]triazolo[1,5-a]pyridine derivatives with slow dissociation kinetics.

For the purpose of discovering novel type-II inhibitors of vascular endothelial growth factor receptor 2 (VEGFR2) kinase, we designed and synthesized 5,6-fused heterocyclic compounds bearing a anilide group. A co-crystal structure analysis of imidazo[1,2-b]pyridazine derivative 2 with VEGFR2 revealed that the N1-nitrogen of imidazo[1,2-b]pyridazine core interacts with the backbone NH group of Cys919. To retain this essential interaction, we designed a series of imidazo[1,2-a]pyridine, [1,2,4]triazolo[1,5-a]pyridine, thiazolo[5,4-b]pyridine, and 1,3-benzothiazole derivatives maintaining a ring nitrogen as hydrogen bond acceptor (HBA) at the corresponding position. All compounds thus designed displayed strong inhibitory activity against VEGFR2 kinase, and the [1,2,4]triazolo[1,5-a]pyridine 13d displayed favorable physicochemical properties. Furthermore, 13d inhibited VEGFR2 kinase with slow dissociation kinetics and also inhibited platelet-derived growth factor receptor (PDGFR) kinases. Oral administration of 13d showed potent anti-tumor efficacy in DU145 and A549 xenograft models in nude mice.

A novel inhibitor of c-Met and VEGF receptor tyrosine kinases with a broad spectrum of in vivo antitumor activities.

The c-Met receptor tyrosine kinase and its ligand, hepatocyte growth factor (HGF), are dysregulated in a wide variety of human cancers and are linked with tumorigenesis and metastatic progression. VEGF also plays a key role in tumor angiogenesis and progression by stimulating the proangiogenic signaling of endothelial cells via activation of VEGF receptor tyrosine kinases (VEGFR). Therefore, inhibiting both HGF/c-Met and VEGF/VEGFR signaling may provide a novel therapeutic approach for treating patients with a broad spectrum of tumors. Toward this goal, we generated and characterized T-1840383, a small-molecule kinase inhibitor that targets both c-Met and VEGFRs. T-1840383 inhibited HGF-induced c-Met phosphorylation and VEGF-induced VEGFR-2 phosphorylation in cancer epithelial cells and vascular endothelial cells, respectively. It also inhibited constitutively activated c-Met phosphorylation in c-met-amplified cancer cells, leading to suppression of cell proliferation. In addition, T-1840383 potently blocked VEGF-dependent proliferation and capillary tube formation of endothelial cells. Following oral administration, T-1840383 showed potent antitumor efficacy in a wide variety of human tumor xenograft mouse models, along with reduction of c-Met phosphorylation levels and microvessel density within tumor xenografts. These results suggest that the efficacy of T-1840383 is produced by direct effects on tumor cell growth and by an antiangiogenic mechanism. Furthermore, T-1840383 showed profound antitumor activity in a gastric tumor peritoneal dissemination model. Collectively, our findings indicate the therapeutic potential of targeting both c-Met and VEGFRs simultaneously with a single small-molecule inhibitor for the treatment of human cancers.

Design and synthesis of novel pyrimido[4,5-b]azepine derivatives as HER2/EGFR dual inhibitors.

A novel 7,6 fused bicyclic scaffold, pyrimido[4,5-b]azepine was designed to fit into the ATP binding site of the HER2/EGFR proteins. The synthesis of this scaffold was accomplished by an intramolecular Claisen-type condensation. As the results of optimization lead us to 4-anilino and 6-functional groups, we discovered 6-substituted amide derivative 19b, which has a 1-benzothiophen-4-yloxy group attached to the 4-anilino group. An X-ray co-crystal structure of 19b with EGFR demonstrated that the N-1 and N-3 nitrogens of the pyrimido[4,5-b]azepine scaffold make hydrogen-bonding interactions with the main chain NH of Met793 and the side chain of Thr854 via a water-mediated hydrogen bond network, respectively. In addition, the NH proton at the 9-position makes an additional hydrogen bond with the carbonyl group of Met793, as we expected. Compound 19b revealed potent HER2/EGFR kinase (IC50: 24/36 nM) and BT474 cell growth (GI50: 18 nM) inhibitory activities based on its pseudo-irreversible (PI) profile.

Discovery of N-[5-({2-[(cyclopropylcarbonyl)amino]imidazo[1,2-b]pyridazin-6-yl}oxy)-2-methylphenyl]-1,3-dimethyl-1H-pyrazole-5-carboxamide (TAK-593), a highly potent VEGFR2 kinase inhibitor.

Vascular endothelial growth factor (VEGF) plays important roles in tumor angiogenesis, and the inhibition of its signaling pathway is considered an effective therapeutic option for the treatment of cancer. In this study, we describe the design, synthesis, and biological evaluation of 2-acylamino-6-phenoxy-imidazo[1,2-b]pyridazine derivatives. Hybridization of two distinct imidazo[1,2-b]pyridazines 1 and 2, followed by optimization led to the discovery of N-[5-({2-[(cyclopropylcarbonyl)amino]imidazo[1,2-b]pyridazin-6-yl}oxy)-2-methylphenyl]-1,3-dimethyl-1H-pyrazole-5-carboxamide (23a, TAK-593) as a highly potent VEGF receptor 2 kinase inhibitor with an IC50 value of 0.95 nM. The compound 23a strongly suppressed proliferation of VEGF-stimulated human umbilical vein endothelial cells with an IC50 of 0.30 nM. Kinase selectivity profiling revealed that 23a inhibited platelet-derived growth factor receptor kinases as well as VEGF receptor kinases. Oral administration of 23a at 1 mg/kg bid potently inhibited tumor growth in a mouse xenograft model using human lung adenocarcinoma A549 cells (T/C=8%).

Structure-based discovery of cellular-active allosteric inhibitors of FAK.

In order to develop potent and selective focal adhesion kinase (FAK) inhibitors, synthetic studies on pyrazolo[4,3-c][2,1]benzothiazines targeted for the FAK allosteric site were carried out. Based on the X-ray structural analysis of the co-crystal of the lead compound, 8-(4-ethylphenyl)-5-methyl-1,5-dihydropyrazolo[4,3-c][2,1]benzothiazine 4,4-dioxide 1 with FAK, we designed and prepared 1,5-dimethyl-1,5-dihydropyrazolo[4,3-c][2,1]benzothiazin derivatives which selectively inhibited kinase activity of FAK without affecting seven other kinases. The optimized compound, N-(4-tert-butylbenzyl)-1,5-dimethyl-1,5-dihydropyrazolo[4,3-c][2,1]benzothiazin-8-amine 4,4-dioxide 30 possessed significant FAK kinase inhibitory activities both in cell-free (IC50=0.64µM) and in cellular assays (IC50=7.1µM). These results clearly demonstrated a potential of FAK allosteric inhibitors as antitumor agents.

Anti-angiogenic and anti-tumor effects of TAK-593, a potent and selective inhibitor of vascular endothelial growth factor and platelet-derived growth factor receptor tyrosine kinase.

We recently reported that TAK-593, a novel imidazo[1,2-b]pyridazine derivative, is a highly potent and selective inhibitor of the vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF) receptor tyrosine kinase families. Moreover, TAK-593 exhibits a uniquely long-acting inhibitory profile towards VEGF receptor 2 (VEGFR2) and PDGF receptor ß (PDGFRß). In this study, we demonstrated that TAK-593 potently inhibits VEGF- and PDGF-stimulated cellular phosphorylation and proliferation of human umbilical vein endothelial cells and human coronary artery smooth muscle cells. TAK-593 also potently inhibits VEGF-induced tube formation of endothelial cells co-cultured with fibroblasts. Oral administration of TAK-593 exhibited strong anti-tumor effects against various human cancer xenografts along with good tolerability despite a low level of plasma exposure. Even after the blood and tissue concentrations of TAK-593 decreased below the detectable limit, a pharmacodynamic marker (phospho VEGFR2) was almost completely suppressed, indicating that its long duration of enzyme inhibition might contribute to the potent activity of TAK-593. Immunohistochemical staining indicated that TAK-593 showed anti-proliferative and pro-apoptotic effects on tumors along with a decrease of vessel density and inhibition of pericyte recruitment to microvessels in vivo. Furthermore, dynamic contrast-enhanced magnetic resonance imaging revealed that TAK-593 reduced tumor vessel permeability prior to the onset of anti-tumor activity. In conclusion, TAK-593 is an extremely potent VEGFR/PDGFR kinase inhibitor whose potent anti-angiogenic activity suggests therapeutic potential for the treatment of solid tumors.

Structure-Based Approach for the Discovery of Pyrrolo[3,2-d]pyrimidine-Based EGFR T790M/L858R Mutant Inhibitors.

The epidermal growth factor receptor (EGFR) family plays a critical role in vital cellular processes and in various cancers. Known EGFR inhibitors exhibit distinct antitumor responses against the various EGFR mutants associated with nonsmall-cell lung cancer. The L858R mutation enhances clinical sensitivity to gefitinib and erlotinib as compared with wild type and reduces the relative sensitivity to lapatinib. In contrast, the T790M mutation confers drug resistance to gefitinib and erlotinib. We determined crystal structures of the wild-type and T790M/L858R double mutant EGFR kinases with reversible and irreversible pyrrolo[3,2-d]pyrimidine inhibitors based on analogues of TAK-285 and neratinib. In these structures, M790 adopts distinct conformations to accommodate different inhibitors, whereas R858 allows conformational variations of the activation loop. These results provide structural insights for understanding the structure-activity relationships that should contribute to the development of potent inhibitors against drug-sensitive or -resistant EGFR mutations.

Design, synthesis, and evaluation of imidazo[1,2-b]pyridazine derivatives having a benzamide unit as novel VEGFR2 kinase inhibitors.

The vascular endothelial growth factor (VEGF) signaling pathway has been implicated in tumor angiogenesis, and inhibition of the VEGF pathway is considered an efficacious method for treating cancer. Herein, we describe synthetic studies of imidazo[1,2-b]pyridazine derivatives as VEGF receptor 2 (VEGFR2) kinase inhibitors. The imidazo[1,2-b]pyridazine scaffold was designed and synthesized as a hinge binder according to the previously reported crystal structure of pyrrolo[3,2-d]pyrimidine 1 with VEGFR2. Structure-activity relationship studies revealed that meta-substituted 6-phenoxy-imidazo[1,2-b]pyridazine derivatives had potent affinity for VEGFR2. In particular, N-[3-(imidazo[1,2-b]pyridazin-6-yloxy)phenyl]-3-(trifluoromethyl)benzamide (6b) exhibited strong inhibitory activity against VEGFR2 with an IC(50) value of 7.1 nM, and it inhibited platelet-derived growth factor receptor ß kinase with an IC(50) value of 15 nM.

Design and synthesis of pyrrolo[3,2-d]pyrimidine HER2/EGFR dual inhibitors: improvement of the physicochemical and pharmacokinetic profiles for potent in vivo anti-tumor efficacy.

During the course of our studies on a novel HER2/EGFR dual inhibitor (TAK-285), we found an alternative potent pyrrolo[3,2-d]pyrimidine compound (1a). To enhance the pharmacokinetic (PK) profile of this compound, we conducted chemical modifications into its N-5 side chain and conversion of the chemically modified compounds into their salts. Among them, 2cb, the tosylate salt of compound 2c, showed potent HER2/EGFR kinase inhibitory activity (IC(50): 11/11 nM) and cellular growth inhibitory activity (BT-474 cell GI(50): 56 nM) with a good drug metabolism and PK (DMPK) profile. Furthermore, 2cb exhibited significant in vivo antitumor efficacy in both mouse and rat xenograft models with transplanted 4-1ST gastric cancer cell lines (mouse, T/C=0%, 2cb po bid at 100 mg/kg; rat, T/C: -1%, 2cb po bid at 25 mg/kg).

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.

A novel pyrrolo[3, 2-d]pyrimidine derivative, as a vascular endothelial growth factor receptor and platelet-derived growth factor receptor tyrosine kinase inhibitor, shows potent antitumor activity by suppression of tumor angiogenesis.

We recently reported that compound 20d (comp.20d), a novel pyrrolo[3, 2-d]pyrimidine derivative, is a potent and selective inhibitor of tumor angiogenesis-related kinases, including vascular endothelial growth factor receptor (VEGFR) and platelet-derived growth factor receptor (PDGFR). In this study, we show that comp.20d potently blocks the VEGF- and PDGF-stimulated cellular phosphorylation (IC(50) = 2.5 and 3.6 nM, respectively) and proliferation of HUVECs and human coronary artery smooth muscle cells with IC(50) values of 2.8 and 9.6 nM, respectively, and potently inhibits the VEGF-induced tube formation of endothelial cells cocultured with fibroblasts (IC(50) = 3.3 nM). Given orally twice daily, comp.20d at the doses of 1.5-6 mg/kg showed antitumor effects in mice bearing various human cancer xenografts. Consistent with the anti-angiogenic mechanism of action, histological examination of tumors from comp. 20d-treated mice indicated a decrease in microvessel density and inhibition of pericyte recruitment to microvessels, and these were concomitant with decreased interstitial fluid pressure that allowed for therapeutic intratumoral uptake of CPT-11 (irinotecan hydrochloride). In conclusion, comp.20d is an extremely potent inhibitor of VEGFR/PDGFR kinases whose activities suggest therapeutic potential for the treatment of solid tumors that rely on angiogenesis for their survival.

Orteronel (TAK-700), a novel non-steroidal 17,20-lyase inhibitor: effects on steroid synthesis in human and monkey adrenal cells and serum steroid levels in cynomolgus monkeys.

Surgical or pharmacologic methods to control gonadal androgen biosynthesis are effective approaches in the treatment of a variety of non-neoplastic and neoplastic diseases. For example, androgen ablation and its consequent reduction in circulating levels of testosterone is an effective therapy for advanced prostate cancers. Unfortunately, the therapeutic effectiveness of this approach is often temporary because of disease progression to the 'castration resistant' (CRPC) state, a situation for which there are limited treatment options. One mechanism thought to be responsible for the development of CRPC is extra-gonadal androgen synthesis and the resulting impact of these residual extra-gonadal androgens on prostate tumor cell proliferation. An important enzyme responsible for the synthesis of extra-gonadal androgens is CYP17A1 which possesses both 17,20-lyase and 17-hydroxylase catalytic activities with the 17,20-lyase activity being key in the androgen biosynthetic process. Orteronel (TAK-700), a novel, selective, and potent inhibitor of 17,20-lyase is under development as a drug to inhibit androgen synthesis. In this study, we quantified the inhibitory activity and specificity of orteronel for testicular and adrenal androgen production by evaluating its effects on CYP17A1 enzymatic activity, steroid production in monkey adrenal cells and human adrenal tumor cells, and serum levels of dehydroepiandrosterone (DHEA), cortisol, and testosterone after oral dosing in castrated and intact male cynomolgus monkeys. We report that orteronel potently suppresses androgen production in monkey adrenal cells but only weakly suppresses corticosterone and aldosterone production; the IC(50) value of orteronel for cortisol was ~3-fold higher than that for DHEA. After single oral dosing, serum levels of DHEA, cortisol, and testosterone were rapidly suppressed in intact cynomolgus monkeys. In castrated monkeys treated twice daily with orteronel, suppression of DHEA and testosterone persisted throughout the treatment period. In both in vivo models and in agreement with our in vitro data, suppression of serum cortisol levels following oral dosing was less than that seen for DHEA. In terms of human CYP17A1 and human adrenal tumor cells, orteronel inhibited 17,20-lyase activity 5.4 times more potently than 17-hydroxylase activity in cell-free enzyme assays and DHEA production 27 times more potently than cortisol production in human adrenal tumor cells, suggesting greater specificity of inhibition between 17,20-lyase and 17-hydroxylase activities in humans vs monkeys. In summary, orteronel potently inhibited the 17,20-lyase activity of monkey and human CYP17A1 and reduced serum androgen levels in vivo in monkeys. These findings suggest that orteronel may be an effective therapeutic option for diseases where androgen suppression is critical, such as androgen sensitive and CRPC.

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.

[A case of benign metastasizing leiomyoma medicating for 14 years].

We report a case of benign metastasizing leiomyoma medicated for 14 years after the diagnosis. A 47-year-old woman, who had undergone hysterectomy for uterine myoma at age 40 in 1989, was readmitted in 1996 because of abnormal shadows found on a chest X-ray film. Computed tomography (CT) and further chest X-ray films showed multiple nodules in bilateral lung fields. Open lung biopsy revealed leiomyomatous nodules histologically similar to those found at age 40. Tests for both estrogen and progesterone receptors in the biopsied specimen were positive. We diagnosed the lung nodules as benign metastasizing leiomyoma (BML) and gave her progesterone. Apart from 2 occasions when the patient elected to stop receiving medication, we obtained decreases in the size and number of tumors for 10 years from the start of treatment. However, despite continued administration of progesterone, the tumors continued to grow slowly during the next 4 years. We believe that the effectiveness of progesterone may have gradually decreased in this case, and thus are considering a change in treatment. BML is rare, and it must be carefully followed up long-term in post-menopausal patients.

Structural analysis of the mechanism of inhibition and allosteric activation of the kinase domain of HER2 protein.

Aberrant signaling of ErbB family members human epidermal growth factor 2 (HER2) and epidermal growth factor receptor (EGFR) is implicated in many human cancers, and HER2 expression is predictive of human disease recurrence and prognosis. Small molecule kinase inhibitors of EGFR and of both HER2 and EGFR have received approval for the treatment of cancer. We present the first high resolution crystal structure of the kinase domain of HER2 in complex with a selective inhibitor to understand protein activation, inhibition, and function at the molecular level. HER2 kinase domain crystallizes as a dimer and suggests evidence for an allosteric mechanism of activation comparable with previously reported activation mechanisms for EGFR and HER4. A unique Gly-rich region in HER2 following the α-helix C is responsible for increased conformational flexibility within the active site and could explain the low intrinsic catalytic activity previously reported for HER2. In addition, we solved the crystal structure of the kinase domain of EGFR in complex with a HER2/EGFR dual inhibitor (TAK-285). Comparison with previously reported inactive and active EGFR kinase domain structures gave insight into the mechanism of HER2 and EGFR inhibition and may help guide the design and development of new cancer drugs with improved potency and selectivity.

Biochemical characterization of TAK-593, a novel VEGFR/PDGFR inhibitor with a two-step slow binding mechanism.

Inhibition of tumor angiogenesis leads to a lack of oxygen and nutrients in the tumor and therefore has become a standards of care for many solid tumor therapies. Dual inhibition of vascular endothelial growth factor receptor (VEGFR) and platelet-derived growth factor receptor (PDGFR) protein kinase activities is a popular strategy for targeting tumor angiogenesis. We discovered that TAK-593, a novel imidazo[1,2-b]pyridazine derivative, potently inhibits tyrosine kinases from the VEGFR and PDGFR families. TAK-593 was highly selective for these families, with an IC(50) >1 µM when tested against more than 200 protein and lipid kinases. TAK-593 displayed competitive inhibition versus ATP. In addition, TAK-593 inhibited VEGFR2 and PDGFRß in a time-dependent manner, classifying it as a type II kinase inhibitor. Analysis of enzyme-inhibitor preincubation experiments revealed that the binding of TAK-593 to VEGFR2 and PDGFRß occurs via a two-step slow binding mechanism. Dissociation of TAK-593 from VEGFR2 was extremely slow (t(1/2) >17 h), and the affinity of TAK-593 at equilibrium (K(i)*) was less than 25 pM. Ligand displacement analysis with a fluorescent tracer confirmed the slow dissociation of TAK-593. The dissociation rate constants were in good agreement between the activity and ligand displacement data, and both analyses supported slow dissociation of TAK-593. The long residence time of TAK-593 may achieve an extended pharmacodynamic effect on VEGFR2 and PDGFRß kinases in vivo that differs substantially from its observed pharmacokinetic profile.

Design, synthesis, and evaluation of 5-methyl-4-phenoxy-5H-pyrrolo[3,2-d]pyrimidine derivatives: novel VEGFR2 kinase inhibitors binding to inactive kinase conformation.

We synthesized a series of pyrrolo[3,2-d]pyrimidine derivatives and evaluated their application as type-II inhibitors of vascular endothelial growth factor receptor 2 (VEGFR2) kinase. Incorporation of a diphenylurea moiety at the C4-position of the pyrrolo[3,2-d]pyrimidine core via an oxygen linker resulted in compounds that were potent inhibitors of VEGFR2 kinase. Of these derivatives, compound 20d showed the strongest inhibition of VEGF-stimulated proliferation of human umbilical vein endothelial cells (HUVEC). The co-crystal structure of 20d and VEGFR2 revealed that 20d binds to the inactive form of VEGFR2. Further studies indicated that 20d inhibited VEGFR2 kinase with slow dissociation kinetics and also inhibited PDGFR and Tie-2 kinases. Oral administration of the hydrochloride salt of 20d at 3mg/kg/day showed potent inhibition of tumor growth in a DU145 human prostate cancer cell xenograft nude mouse model.

[Successful reintroduction of mesylate imatinib after pneumonitis in two patients with gastrointestinal stromal tumor (GIST)].

We report 2 cases of successful reintroduction of mesylate imatinib for gastrointestinal stromal tumor (GIST) after drug-induced pneumonitis. Both of them were women in the fifth decade who had been medicated by mesylate imatinib about for 5 months previously, and had been given a diagnosis of imatinib mesylate-induced pneumonitis. After only cessation of that drug, symptoms and shadows on chest X-ray film improved. However, we had to reintroduce the drug because of the growth of primary tumor, so we gave half the previous dose of imatinib mesylate, with low dose prednisone. There has been no recurrence of drug related pneumonitis and effective control of the primary tumor was obtained. The evidence acquired from our cases suggests that it may be possible to reintroduce imatinib mesylate carefully by adjusting the dose with low dose prednisone in a GIST patient, without causing recurrence of drug-induced pneumonitis.

Macrophages participate in lymphangiogenesis in idiopathic diffuse alveolar damage through CCL19-CCR7 signal.

An abnormal healing process is believed to be involved in lung tissue damage in pulmonary fibrosis. Lymphangiogenesis has been determined to play a role in structural remodeling in diffuse alveolar damage. However, the mechanism of lymphangiogenesis remains unclear. The aim of this study is to investigate the cellular mechanisms of lymphangiogenesis in diffuse alveolar damage, focusing on the roles of macrophages. Formalin-fixed and paraffin-embedded lung tissues from 13 autopsy cases with idiopathic diffuse alveolar damage were used. Antibodies specific for D2-40 and CD68 were mainly used as the markers for lymphatics and macrophages, respectively, and immunohistochemical examinations and morphometric analyses were performed. Immunohistochemistry showed the aggregation of numerous CD68+ macrophages around newly formed lymphatics in the intraalveolar fibrotic lesions in the proliferative stage, and some of the CD68+ macrophages were colocalized with the lymphatic endothelium. These macrophages were characterized by the expression of vascular endothelial growth factor-C. Among the 3 stages of diffuse alveolar damage, the number of Ki-67+ cells on the lymphatic endothelium tended to increase in newly formed lymphatics of the proliferative stage. In addition, the number of CD68+ macrophages on the lymphatic endothelium was significantly increased in the newly formed lymphatics of the proliferative stage more than those of the fibrotic stage. The CD68+ macrophages around the newly formed lymphatics coexpressed CCR7. Dual immunostaining showed the coexpression of CCL19-a ligand for CCR7-on the lymphatic endothelium. Thus, macrophages may participate in lymphangiogenesis in diffuse alveolar damage, which is facilitated by CCL19 and CCR7.