Extensive evidence implicates activation of the lipid phosphatidylinositide 3-kinase (PI3K) pathway in the genesis and progression of various human cancers. PI3K inhibitors thus have considerable potential as molecular cancer therapeutics. Here, we detail the pharmacologic properties of a prototype of a new series of inhibitors of class I PI3K. PI103 is a potent inhibitor with low IC 50 values against recombinant PI3K isoforms p110A (2 nmol/L), p110B (3 nmol/L), p110D (3 nmol/L), and p110; (15 nmol/L). PI103 also inhibited TORC1 by 83.9% at 0.5 Mmol/L and exhibited an IC 50 of 14 nmol/L against DNA-PK. A high degree of selectivity for the PI3K family was shown by the lack of activity of PI103 in a panel of 70 protein kinases. PI103 potently inhibited proliferation and invasion of a wide variety of human cancer cells in vitro and showed biomarker modulation consistent with inhibition of PI3K signaling. PI103 was extensively metabolized, but distributed rapidly to tissues and tumors. This resulted in tumor growth delay in eight different human cancer xenograft models with various PI3K pathway abnormalities. Decreased phosphorylation of AKT was observed in U87MG gliomas, consistent with drug levels achieved. We also showed inhibition of invasion in orthotopic breast and ovarian cancer xenograft models and obtained evidence that PI103 has antiangiogenic potential. Despite its rapid in vivo metabolism, PI103 is a valuable tool compound for exploring the biological function of class I PI3K and importantly represents a lead for further optimization of this novel class of targeted molecular cancer therapeutic. [Cancer Res 2007;67(12):5840-50]

SUMMARY The ALKF1174L mutation is associated with intrinsic and acquired resistance to crizotinib and cosegregates with MYCN in neuroblastoma. In this study, we generated a mouse model overexpressing ALKF1174L in the neural crest. Compared to ALKF1174L and MYCN alone, coexpression of these two oncogenes led to the development of neuroblastomas with earlier onset, higher penetrance and enhanced lethality. ALKF1174L/MYCN tumors exhibited increased MYCN dosage due to ALKF1174L-induced activation of the PI3K/AKT/mTOR and MAPK pathways, coupled with suppression of MYCN pro-apoptotic effects. Combined treatment with the ATP-competitive mTOR inhibitor Torin2, overcame the resistance of ALKF1174L/MYCN tumors to crizotinib. Our findings demonstrate a pathogenic role for ALKF1174L in neuroblastomas overexpressing MYCN and suggest a strategy for improving targeted therapy for ALK-positive neuroblastoma.

SummaryWe undertook a comprehensive clinical and biological investigation of serial medulloblastoma biopsies obtained at diagnosis and relapse. Combined MYC family amplifications and P53 pathway defects commonly emerged at relapse, and all patients in this group died of rapidly progressive disease postrelapse. To study this interaction, we investigated a transgenic model of MYCN-driven medulloblastoma and found spontaneous development of Trp53 inactivating mutations. Abrogation of p53 function in this model produced aggressive tumors that mimicked characteristics of relapsed human tumors with combined P53-MYC dysfunction. Restoration of p53 activity and genetic and therapeutic suppression of MYCN all reduced tumor growth and prolonged survival. Our findings identify P53-MYC interactions at medulloblastoma relapse as biomarkers of clinically aggressive disease that may be targeted therapeutically.

Angiotensin II (ANGII) acting on ANGII type 1 (AT1) receptors in the solitary tract nucleus (NTS) depresses the baroreflex. Since ANGII stimulates the release of nitric oxide (NO), we tested whether the ANGII‐mediated depression of the baroreflex in the NTS depended on NO release. In a working heart‐brainstem preparation (WHBP) of rat NTS microinjection of either ANGII (500 fmol) or a NO donor (diethylamine nonoate, 500 pmol) both depressed baroreflex gain by ‐56 and ‐67 %, respectively (P < 0.01). In contrast, whilst ANGII potentiated the peripheral chemoreflex, the NO donor was without effect. NTS microinjection of non‐selective NO synthase (NOS) inhibitors (l‐NAME; 50 pmol) or (l‐NMMA; 200 pmol) prevented the ANGII‐induced baroreflex attenuation (P > 0.1). In contrast, a neurone‐specific NOS inhibitor, TRIM (50 pmol), was without effect. Using an adenoviral vector, a dominant negative mutant of endothelial NOS (TeNOS) was expressed bilaterally in the NTS. Expression of TeNOS affected neither baseline cardiovascular parameters nor baroreflex sensitivity. However, ANGII microinjected into the transfected region failed to affect the baroreflex. Immunostaining revealed that eNOS‐positive neurones were more numerous than those labelled for AT1 receptors. Neurones double labelled for both AT1 receptors and eNOS comprised 23 ± 5.4 % of the eNOS‐positive cells and 57 ± 9.2 % of the AT1 receptor‐positive cells. Endothelial cells were also double labelled for eNOS and AT1 receptors. We suggest that ANGII activates eNOS located in either neurones and/or endothelial cells to release NO, which acts selectively to depress the baroreflex.