Robert W. Tuman

JNJ-28312141, a novel orally active colony-stimulating factor-1 receptor/FMS-related receptor tyrosine kinase-3 receptor tyrosine kinase inhibitor with potential utility in solid tumors, bone metastases, and acute myeloid leukemia

There is increasing evidence that tumor-associated macrophages promote the malignancy of some cancers. Colony-stimulating factor-1 (CSF-1) is expressed by many tumors and is a growth factor for macrophages and mediates osteoclast differentiation. Herein, we report the efficacy of a novel orally active CSF-1 receptor (CSF-1R) kinase inhibitor, JNJ-28312141, in proof of concept studies of solid tumor growth and tumor-induced bone erosion. H460 lung adenocarcinoma cells did not express CSF-1R and were not growth inhibited by JNJ-28312141 in vitro. Nevertheless, daily p.o. administration of JNJ-28312141 caused dose-dependent suppression of H460 tumor growth in nude mice that correlated with marked reductions in F4/80+ tumor-associated macrophages and with increased plasma CSF-1, a possible biomarker of CSF-1R inhibition. Furthermore, the tumor microvasculature was reduced in JNJ-28312141–treated mice, consistent with a role for macrophages in tumor angiogenesis. In separate studies, JNJ-28312141 was compared with zoledronate in a model in which MRMT-1 mammary carcinoma cells inoculated into the tibias of rats led to severe cortical and trabecular bone lesions. Both agents reduced tumor growth and preserved bone. However, JNJ-28312141 reduced the number of tumor-associated osteoclasts superior to zoledronate. JNJ-28312141 exhibited additional activity against FMS-related receptor tyrosine kinase-3 (FLT3). To more fully define the therapeutic potential of this new agent, JNJ-28312141 was evaluated in a FLT3-dependent acute myeloid leukemia tumor xenograft model and caused tumor regression. In summary, this novel CSF-1R/FLT3 inhibitor represents a new agent with potential therapeutic activity in acute myeloid leukemia and in settings where CSF-1–dependent macrophages and osteoclasts contribute to tumor growth and skeletal events. [Mol Cancer Ther 2009;8(11):3151–61]

Synthesis and anti-angiogenic activity of 6-(1,2,4-thiadiazol-5-yl)-3-amino pyridazine derivatives

General screening for inhibitors of microvessel growth in vitro in the rat aortic ring assay led to the discovery of a novel series of thiadiazole pyridazine compounds with potential anti-angiogenic activity. Chemical optimization produced orally active compounds with potent in vitro and in vivo anti-angiogenesis and anti-tumor activities.

Validation of in vivo pharmacodynamic activity of a novel PDGF receptor tyrosine kinase inhibitor using immunohistochemistry and quantitative image analysis

With the advent of agents directed against specific molecular targets in drug discovery, it has become imperative to show a compounds cellular impact on the intended biomolecule in vivo. The objective of the present study was to determine if we could develop an assay to validate the in vivo effects of a compound. Hence, we investigated the in vivo pharmacodynamic activity of JNJ-10198409, a relatively selective inhibitor of platelet-derived growth factor receptor tyrosine kinase (PDGF-RTK), in tumor tissues after administering the compound orally in a nude mouse xenograft model of human LoVo colon cancer. We developed a novel assay to quantify the in vivo anti-PDGF-RTK activity of the inhibitor in tumor tissue by determining the phosphorylation status of phospholipase Cγ1 (PLCγ1), a key downstream cellular molecule in the PDGF-RTK signaling cascade. We used two antibodies, one specific for the total (phosphorylated and unphosphorylated forms) PLCγ1 (pan-PLCγ1) and the other, specific for phosphorylated form of PLCγ1 (ph-PLCγ1) to immunohistochemically detect their expression in tumor tissues. Computer-assisted image analysis was then used to directly compare the ratio of ph-PLCγ1 to pan-PLCγ1 immunolabeling intensities in serial sections (5 μm) of tumors obtained from vehicle- and JNJ-10198409-treated tumor-bearing mice. Our data showed statistically significant, dose-dependent differences in the ph-PLC/pan-PLC ratio among the four treatment groups (vehicle, 25, 50, and 100 mg/kg b.i.d.). These results confirmed this compounds ability to suppress PDGF-RTK downstream signaling in tumor tissues in vivo. In addition to this specific application of this in vivo validation approach to those targets that use PLCγ as a downstream signaling partner, these methods may also benefit other drug discovery targets.

Effect of the fatty acid oxidation inhibitor 2-tetradecylglycidic acid (TDGA) on glucose and fatty acid oxidation in isolated rat soleus muscle

1. The effect of 2-tetradecylglycidic acid (TDGA), a potent, specific inhibitor of long-chain fatty acid oxidation, on fatty acid and glucose oxidation by isolated rat soleus muscle was studied. 2. TDGA inhibited [1-14C]palmitate oxidation by soleus muscle in a concentration-dependent manner. 3. TDGA inhibited the activity of soleus muscle mitochondrial carnitine palmitoyltransferase A (CPT-A). 4. Added palmitate (0.5 mM) significantly inhibited D-[U-14C]glucose oxidation and, under conditions where TDGA inhibited palmitate oxidation, the oxidation of D-[U-14C]glucose by isolated soleus muscle was significantly stimulated. 5. TDGA stimulation of glucose oxidation was reversed by octanoate, a medium-chain fatty acid whose oxidation is not inhibited by TDGA. 6. When nondiabetic rats were treated with TDGA (10 mg/kg p.o./day x 3 days), fasting plasma glucose was significantly lowered and the ability of isolated contralateral soleus muscles to oxidize palmitate was inhibited while glucose oxidation was significantly stimulated.

Hypoglycaemic and hypoketonaemic effects of single and repeated oral doses of methyl palmoxirate (methyl 2-tetradecylglycidate) in streptozotocin/alloxan-induced diabetic dogs

1 The hypoglycaemic and hypoketonaemic effects of orally administered methyl palmoxirate were studied in streptozotocin/alloxan‐induced diabetic dogs. 2 Single oral 50 mg doses (≅7.5 mg kg−1) of methyl palmoxirate produced statistically significant reductions of plasma glucose (32 ± 6% maximum reduction from baseline) and ketones (74 ± 12% maximum reduction from baseline), with the peak effect on plasma ketones (3.5 h) preceding that for plasma glucose (6.0 h). 3 Lower doses (0.7‐2.0 mg kg−1 daily) of methyl palmoxirate given repeatedly for seven days produced reductions of blood glucose and ketones equivalent to those produced with the higher single dose. Maximal reductions of plasma ketones were generally observed following the first dose of drug, whereas significant lowering of plasma glucose required several days of continuous dosing. 4 Repeated daily doses of methyl palmoxirate markedly reduced the overnight fasting ketone levels but not glucose levels of diabetic dogs. 5 In conclusion, administration of the fatty acid oxidation inhibitor methyl palmoxirate, in the absence of concomitant insulin therapy, was able to lower the plasma glucose and ketone levels of insulin‐deficient streptozotocin/alloxan diabetic dogs. Only the plasma ketones were decreased to normal by this treatment.

Chapter 20. Mechanism of Action of Insulin, Glucagon and Somatostatin

Publisher Summary This chapter discusses certain aspects of insulin, glucagon, and somatostatin (SRIF) hormones actions that have undergone rapid and important new advances. In case of insulin, these advances have been mainly in the areas of insulins second messenger and effects on membrane structure and function. For glucagon, effects on the enzyme systems are emphasized and, as for SRIF, it produces a wide variety of anti-secretory effects, including the inhibition of insulin and glucagon secretion. The metabolic effects of insulin and glucagon involve binding to specific membrane receptor sites with subsequent modulation of plasma membrane functions and numerous intracellular catabolic and anabolic processes. SRIF mainly alters metabolism indirectly by affecting the secretion of insulin and glucagon. These hormones affect several major target tissues, such as liver, muscle, fat, GI tract, etc. Insulin interacts with specific high affinity (Kd-lnM) cell surface membrane receptors. The number of these receptors is small and can be decreased by insulin exposure in vitro or hyperinsulinemia in vivo . Only minimal receptor occupancy by insulin incurs maximal physiological response, although exceptions have been reported. Insulin binding to receptors generally yields noncompetitive dissociation kinetics and curvilinear scatchard plots that have been attributed to negative cooperativity or multiple classes of independent receptors, whereas glucagon regulates the direction and magnitude of cellular carbon flux by influencing the activity of three allosteric enzymes.