Foretinib is effective therapy for metastatic sonic hedgehog medulloblastoma.

Medulloblastoma is the most common malignant pediatric brain tumor, with metastases present at diagnosis conferring a poor prognosis. Mechanisms of dissemination are poorly understood and metastatic lesions are genetically divergent from the matched primary tumor. Effective and less toxic therapies that target both compartments have yet to be identified. Here, we report that the analysis of several large nonoverlapping cohorts of patients with medulloblastoma reveals MET kinase as a marker of sonic hedgehog (SHH)-driven medulloblastoma. Immunohistochemical analysis of phosphorylated, active MET kinase in an independent patient cohort confirmed its correlation with increased tumor relapse and poor survival, suggesting that patients with SHH medulloblastoma may benefit from MET-targeted therapy. In support of this hypothesis, we found that the approved MET inhibitor foretinib could suppress MET activation, decrease tumor cell proliferation, and induce apoptosis in SHH medulloblastomas in vitro and in vivo. Foretinib penetrated the blood-brain barrier and was effective in both the primary and metastatic tumor compartments. In established mouse xenograft or transgenic models of metastatic SHH medulloblastoma, foretinib administration reduced the growth of the primary tumor, decreased the incidence of metastases, and increased host survival. Taken together, our results provide a strong rationale to clinically evaluate foretinib as an effective therapy for patients with SHH-driven medulloblastoma.

Alterations in eicosanoid levels during U937 bcl-xL tumour growth suppression and recovery in NU/NU mice in vivo-Involvement of phospholipase A2.

We report the effects of two anti-cancer drugs, PBT-4, an experimental antagonist to the pro-inflammatory hepoxilins, and Gleevec (STI-571), an anti-leukaemic drug, on eicosanoid tumour levels in immunodeficient mice (NU/NU) xenografted with the leukaemic cell line, U937 bcl-xL. After the tumours had grown to 80-100mm(3) volume, an 8-day treatment with the drugs was initiated and the animals were monitored for 28 days. On various days, tumours were removed for measurement of 24 omega-6 eicosanoids. The data show remarkable direct correlation between inhibition of tumour AA release and 12-LOX products (including 12-HETE and hepoxilins) during PBT or STI treatment with tumour growth suppression. These findings suggest that inhibition of AA release may represent a novel underlying mechanism of action of PBT-4 (and STI) in vivo in suppressing tumour growth. As the PBT wears off, AA and 12-LOX products rise rapidly (Day 18) leading to the observed tumour growth spurt.

Hepoxilin analogs, PBT-3 and PBT-4, cause apoptosis of Gleevec-resistant K562 cells in vitro.

The use of Gleevec in the treatment of leukemia has been widely accepted, although resistance to Gleevec is commonly observed. Gleevec represents a new direction in the development of target-focused chemotherapeutic agents in cancer. Gleevec inhibits the tyrosine kinase activity of Bcr-Abl, which is responsible for leukemic cell survival. We have previously shown that PBT-3 (racemic anti-10(R/S)-hydroxy-11, 12-cyclopropyl-eicosa-5Z, 8Z, 14Z-trienoic acid methyl ester) and PBT-4 (racemic syn- 10(R/S)-hydroxy- 11,12-cyclopropyleicosa-5Z 8Z, 14Z-trienoic acid methyl ester), stable analogs of the hepoxilins, caused apoptosis of the human leukemic K562 cell line in vitro and in vivo. We also showed that PBTs inhibited the growth of tumours derived from the inoculation of immunodeficient mice with K562 cells and that the effect of PBTs was synergistic with that of Gleevec. We now show that the effect of PBT-3 and of PBT-4 is independent of that of Gleevec, demonstrating that Gleevec-resistant K562 cells retain their responsiveness to PBT treatment, resulting in apoptosis. These findings provide important information suggesting that the two compounds, PBT and Gleevec, can be used together in the treatment of leukemia. The PBTs may provide a new platform for the development of apoptotic drugs in cancer.

Mechanotransduction of stretch-induced prostanoid release by fetal lung epithelial cells.

Mechanical ventilation is the primary supportive treatment for infants and adults suffering from severe respiratory failure. Adverse mechanical ventilation (overdistension of the lung) triggers a proinflammatory response. Along with cytokines, inflammatory mediators such as bioactive lipids are involved in the regulation of the inflammatory response. The arachidonic acid pathway is a key source of bioactive lipid mediators, including prostanoids. Although ventilation has been shown to influence the production of prostanoids in the lung, the mechanotransduction pathways are unknown. Herein, we established that cyclic stretch of fetal lung epithelial cells, but not fibroblasts, can evoke an extremely sensitive, rapid alteration in eicosanoid metabolism through a cyclooxygenase (COX)-2 dependent mechanism. Cyclic stretch significantly increased PGI(2), PGF(2alpha), PGD(2), PGE(2), and thromboxane B(2) levels in the media of epithelial cells, but did not alter leukotriene B(4) or 12-hydroxyeicosatetraenoic acid levels. Inhibition of COX-2, but not COX-1, attenuated the cyclic stretch-induced PG increase in the media, suggesting that cyclic stretch primarily affected PG synthesis. Substrate (free arachidonic acid) availability for PG generation was increased because of a cyclic stretch-induced activation of cytosolic phospholipase A(2) (cPLA(2)) via an influx of extracellular calcium and phosphorylation by mitogen-activated protein kinase, p44/42MAPK. The data are compatible with cPLA(2) and COX-2 being intimately involved in regulating the injury response to adverse mechanical ventilation.

PBT-3, a hepoxilin stable analog, causes long term inhibition of growth of K562 solid tumours in vivo.

We demonstrate herein that daily administration of PBT-3 for 8 days to NU/NU mice bearing solid tumours derived from the s.c. administration of the leukemic cell line K562 results in inhibition of growth of the tumours in vivo, and this inhibition lasts for 60 days after stopping treatment with PBT-3 before recovery of tumour growth is re-established. Similar findings were observed when the mice were treated with Gleevec (STI-571). These results provide new evidence that PBT-3 is effective in controlling solid tumour growth in vivo and suggest that the PBT family may be useful in the development of new drugs in cancer therapy.

The hepoxilin analog, PBT-3, inhibits growth of K-562 CML solid tumours in vivo in nude mice.

PBT-3 is one of a family of stable chemical analogs of the hepoxilins, products derived from arachidonic acid. We previously showed that PBT-3 caused apoptosis in the chronic myelogenous leukemia (CML) cell line K-562 in vitro (Anti-cancer Res 23: 3617-3622, 2003). It was as effective as Gleevec, a novel agent that blocks tyrosine kinase activity during treatment of CML. We describe, herein, the growth inhibiting effects of PBT-3 in nude mice into which K-562 cells were transplanted subcutaneously. Groups of mice were treated with vehicle as control, or PBT-3, or Gleevec. PBT-3 was effective during the 8-day treatment protocol in inhibiting the growth of the tumours in vivo as was Gleevec. Analysis of the tumours demonstrated the presence of apoptosis (DNA laddering and TUNEL assay) in both the PBT-3- and Gleevec-treated groups. These results demonstrate that PBT-3 is effective in vivo in controlling tumour growth and provides a novel platform for the therapeutic control of cancer.

The hepoxilin analog PBT-3 induces apoptosis in BCR-ABL-positive K562 leukemia cells.

BACKGROUND: Leukemia is a heterogeneous disease characterized by malignant proliferation of cells of the hematopoietic system. The use of chemotherapeutic agents is still the mainstay of anti-leukemia therapy. Despite this, significant morbidity and mortality still occurs. We describe herein novel apoptotic effects of PBT-3, one of a family of stable analogs of the Hepoxilins, natural products derived from arachidonic acid. MATERIALS AND METHODS: Inhibition of [3H]-thymidine incorporation, nuclear fragmentation, DNA laddering, FACS analysis as well as Annexin V binding were assessed. RESULTS: PBT-3 dose-dependently causes apoptosis of the CML cell line, K562, in vitro. PBT-3 acts by increasing cytochrome c release into the cytoplasm and by activation of caspase-3 degradation. The effects of PBT-3 compare favorably with those of STI571 (Gleevec), while thromboxane agonists and antagonists are without effect. CONCLUSION: These results suggest that PBT analogs may provide a new platform for the development of apoptotic drugs in leukemia.

A new family of thromboxane receptor antagonists with secondary thromboxane synthase inhibition.

We report herein a novel class of thromboxane receptor (TP receptor) antagonists modeled on unstable natural lipids that we identified several years ago, the hepoxilins. These antagonists have been rendered chemically and biologically more stable than the natural compounds through structural modification by chemical synthesis. We demonstrate that the analogs inhibit the aggregation of human platelets in vitro evoked by the thromboxane receptor agonists, I-BOP ([1S-[1alpha,2alpha(Z),3beta(1E,3S*),4alpha]]-7-[3-[3-hydroxy-4-(4-iodophenoxy)-1-butenyl]-7-oxabi-cyclo[2.2.1]hept-2-yl]5-heptenoic acid) and U46619 (9,11-dideoxy-9alpha,11alpha-methanoepoxy-prosta-5Z,13E-dien-1-oic acid). The most potent of the analogs described, PBT-3 [10(S)-hydroxy-11,12-cyclopropyl-eicosa-5Z,8Z,14Z-trienoic acid methyl ester], has an IC(50) versus aggregation by I-BOP = 0.6 x 10(-7) M and versus U46619 = 7 x 10(-7) M, representing one of the most potent anti-aggregating substances so far described. PBT-3 also inhibits thromboxane formation and aggregation evoked by collagen with an IC(50) = 8 x 10(-7) M. Other PBT (hepoxilin cyclopropane) analogs so far tested were 5- to 10-fold less active, and the native hepoxilins were about 500-fold less active. Neither PBT-3 nor the other analogs inhibited 12-lipoxygenase, phospholipase A(2), or cyclooxygenase 1 or 2, and weakly stimulated adenyl cyclase (threshold stimulation at 10(-7) M and little selectivity for each of the PBT compounds). TP antagonism by PBT-3 was further demonstrated in receptor binding studies through use of (125)I-BOP, where the IC(50) for PBT-3 was 8 x 10(-9) M, approximately 16-fold less than for I-BOP itself. These findings identify a new mode of action of PBT-3 and other related analogs as primarily TP antagonists. These studies identify a new family of compounds useful in further development as novel therapeutics for thromboxane-mediated diseases.