1 Three fully-de®ned a 1 -adrenoceptors (a 1A , a 1B and a 1D ) have been established in pharmacological and molecular studies. A fourth a 1 -adrenoceptor, the putative a 1L -adrenoceptor, has been de®ned in functional but not molecular studies, and has been proposed to mediate contraction of human lower urinary tract tissues; its relationship to the three fully characterized a 1 -adrenoceptors is not known. 2 In the present study, binding a nities were estimated by displacement of [ 3 H]-prazosin in membrane homogenates of Chinese hamster ovary (CHO-K1) cells stably expressing the human a 1A -, a 1B -and a 1D -adrenoceptors and were compared with a nity estimates obtained functionally in identical cells by measuring inhibition of noradrenaline (NA)-stimulated accumulation of [ 3 H]-inositol phosphates. 3 For the a 1A -adrenoceptor, binding studies revealed a pharmacological pro®le typical for the classically de®ned a 1A -adrenoceptor, such that prazosin, RS-17053, WB 4101, 5-methylurapidil, Rec 15/ 2739 and S-niguldipine all displayed subnanomolar a nity. A di erent pro®le of a nity estimates was obtained in inositol phosphates accumulation studies: prazosin, WB 4101, 5-methylurapidil, RS-17053 and S-niguldipine showed 10 to 40 fold lower a nity than in membrane binding. However, a nity estimates were not`frameshifted', as tamsulosin, indoramin and Rec 15/2739 yielded similar, high a nity estimates in binding and functional assays. 4 In contrast, results from human a 1B -and a 1D -adrenoceptors expressed in CHO-K1 cells gave antagonist a nity pro®les in binding and functional assays that were essentially identical. 5 A concordance of a nity estimates from the functional (inositol phosphates accumulation) studies of the a 1A -adrenoceptor in CHO-K1 cells was found with estimates published recently from contractile studies in human lower urinary tract tissues (putative a 1L -adrenoceptor). These data show that upon functional pharmacological analysis, the cloned a 1A -adrenoceptor displays pharmacological recognition properties consistent with those of the putative a 1L -adrenoceptor. Why this pro®le di ers from that obtained in membrane binding, and whether it explains the a 1L -adrenoceptor pharmacology observed in many native tissues, requires further investigation.

Alterations of the phosphoinositide-3 kinase (PI3K)/Akt signaling pathway occur broadly in cancer via multiple mechanisms including mutation of the PIK3CA gene, loss or mutation of phosphatase and tensin homolog (PTEN), and deregulation of mammalian target of rapamycin (mTOR) complexes. The dysregulation of this pathway has been implicated in tumor initiation, cell growth and survival, invasion and angiogenesis, thus, PI3K and mTOR are promising therapeutic targets for cancer. We discovered GDC-0980, a selective, potent, orally bioavailable inhibitor of Class I PI3 kinase and mTOR kinase (TORC1/2) with excellent pharmacokinetic and pharmaceutical properties. GDC-0980 potently inhibits signal transduction downstream of both PI3K and mTOR, as measured by pharmacodynamic (PD) biomarkers, thereby acting upon two key pathway nodes to produce the strongest attainable inhibition of signaling in the pathway. Correspondingly, GDC-0980 was potent across a broad panel of cancer cell lines, with the greatest potency in breast, prostate, and lung cancers and less activity in melanoma and pancreatic cancers, consistent with KRAS and BRAF acting as resistance markers. Treatment of cancer cell lines with GDC-0980 resulted in G1 cell-cycle arrest, and in contrast to mTOR inhibitors, GDC-0980 induced apoptosis in certain cancer cell lines, including those with direct pathway activation via PI3K and PTEN. Low doses of GDC-0980 potently inhibited tumor growth in xenograft models including those with activated PI3K, loss of LKB1 or PTEN, and elicited an exposure-related decrease in PD biomarkers. These preclinical data show that GDC-0980 is a potent and effective dual PI3K/mTOR inhibitor with promise for the clinic. Mol Cancer Ther; 10(12); 2426-36. Ó2011 AACR.

The discovery of 2 (GDC-0980), a class I PI3K and mTOR kinase inhibitor for oncology indications, is described. mTOR inhibition was added to the class I PI3K inhibitor 1 (GDC-0941) scaffold primarily through the substitution of the indazole in 1 for a 2-aminopyrimidine. This substitution also increased the microsomal stability and the free fraction of compounds as evidenced through a pairwise comparison of molecules that were otherwise identical. Highlighted in detail are analogues of an advanced compound 4 that were designed to improve solubility, resulting in 2. This compound, is potent across PI3K class I isoforms with IC(50)s of 5, 27, 7, and 14 nM for PI3Kα, β, δ, and γ, respectively, inhibits mTOR with a K(i) of 17 nM yet is highly selective versus a large panel of kinases including others in the PIKK family. On the basis of the cell potency, low clearance in mouse, and high free fraction, 2 demonstrated significant efficacy in mouse xenografts when dosed as low as 1 mg/kg orally and is currently in phase I clinical trials for cancer.

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The PI3K/AKT/mTOR pathway has been shown to play an important role in cancer. Starting with compounds 1 and 2 (GDC-0941) as templates, (thienopyrimidin-2-yl)aminopyrimidines were discovered as potent inhibitors of PI3K or both PI3K and mTOR. Structural information derived from PI3K gamma-ligand cocrystal structures of 1 and 2 were used to design inhibitors that maintained potency for PI3K yet improved metabolic stability and oral bioavailability relative to 1. The addition of a single methyl group to the optimized 5 resulted in 21, which had significantly reduced potency for mTOR. The lead compounds 5 (GNE-493) and 21 (GNE-490) have good pharmacokinetic (PK) parameters, are highly selective, demonstrate knock down of pathway markers in vivo, and are efficacious in xenograft models where the PI3K pathway is deregulated. Both compounds were compared in a PI3K alpha mutated MCF7.1 xenograft model and were found to have equivalent efficacy when normalized for exposure.