Publications by authors named "Paula Lewis"

Purpose: Targeting Bcl-2 family members upregulated in multiple cancers has emerged as an important area of cancer therapeutics. While venetoclax, a Bcl-2-selective inhibitor, has had success in the clinic, another family member, Bcl-x, has also emerged as an important target and as a mechanism of resistance. Therefore, we developed a dual Bcl-2/Bcl-x inhibitor that broadens the therapeutic activity while minimizing Bcl-x-mediated thrombocytopenia.

Experimental Design: We used structure-based chemistry to design a small-molecule inhibitor of Bcl-2 and Bcl-x and assessed the activity against cell lines, patient samples, and models. We applied pharmacokinetic/pharmacodynamic (PK/PD) modeling to integrate our understanding of on-target activity of the dual inhibitor in tumors and platelets across dose levels and over time.

Results: We discovered AZD4320, which has nanomolar affinity for Bcl-2 and Bcl-x, and mechanistically drives cell death through the mitochondrial apoptotic pathway. AZD4320 demonstrates activity in both Bcl-2- and Bcl-x-dependent hematologic cancer cell lines and enhanced activity in acute myeloid leukemia (AML) patient samples compared with the Bcl-2-selective agent venetoclax. A single intravenous bolus dose of AZD4320 induces tumor regression with transient thrombocytopenia, which recovers in less than a week, suggesting a clinical weekly schedule would enable targeting of Bcl-2/Bcl-x-dependent tumors without incurring dose-limiting thrombocytopenia. AZD4320 demonstrates monotherapy activity in patient-derived AML and venetoclax-resistant xenograft models.

Conclusions: AZD4320 is a potent molecule with manageable thrombocytopenia risk to explore the utility of a dual Bcl-2/Bcl-x inhibitor across a broad range of tumor types with dysregulation of Bcl-2 prosurvival proteins.

The intermediate mesoderm is located between the somites and the lateral plate mesoderm and gives rise to renal progenitors that contribute to the three mammalian kidney types (pronephros, mesonephros and metanephros). In this review, focusing largely on murine kidney development, we examine how the intermediate mesoderm forms during gastrulation/axis elongation and how it progressively gives rise to distinct renal progenitors along the rostro-caudal axis. We highlight some of the potential signalling cues and core transcription factor circuits that direct these processes, up to the point of early metanephric kidney formation.

With the advent of recent advances in molecular techniques and whole genome sequencing, we have come to know that the non-coding landscape (including non-coding RNAs, tRNAs and even telomeres) plays a major role in the regulation of cellular processes. Furthermore, the deregulation of this landscape has been found to contribute to and even bring about the pathogenesis of a large number of diseases. One of such diseases is diabetes mellitus (type 2 specifically) whose incidence rate and global burden is constantly increasing. Nutrition has been proven to be a key player in the development, onset and control of type 2 diabetes mellitus. Additionally, non-coding DNA based molecular markers are emerging as biomarkers of T2D, susceptibility, and perhaps dietary supplements can modulate non-coding DNA based markers expression and function in T2D management. In this review, we provide a brief overview of the developmental origins and genetics of type 2 diabetes mellitus, how each component of the non-coding landscape contributes to the development and progression of the disease and finally we discuss how dietary interventions modulate the non-coding landscape in T2D.

We report here a novel series of benzimidazole sulfonamides that act as antagonists of the S1P1 receptor, identified by exploiting an understanding of the pharmacophore of a high throughput screening (HTS)-derived series of compounds described previously. Lead compound 2 potently inhibits S1P-induced receptor internalization in a cell-based assay (EC50 = 0.05 μM), but has poor physical properties and metabolic stability. Evolution of this compound through structure-activity relationship development and property optimization led to in vivo probes such as 4. However, this compound was unexpectedly found to be a potent CYP3A inducer in human hepatocytes, and thus further chemistry efforts were directed at addressing this liability. By employing a pregnane X receptor (PXR) reporter gene assay to prioritize compounds for further testing in human hepatocytes, we identified lipophilicity as a key molecular property influencing the likelihood of P450 induction. Ultimately, we have identified compounds such as 46 and 47, which demonstrate the desired S1P1 antagonist activity while having greatly reduced risk of CYP3A induction in humans. These compounds have excellent oral bioavailability in preclinical species and exhibit pharmacodynamic effects of S1P1 antagonism in several in vivo models following oral dosing. Relatively modest antitumor activity was observed in multiple xenograft models, however, suggesting that selective S1P1 antagonists would have limited utility as anticancer therapeutics as single agents.

Structure-activity relationship analysis identified (+)-N-(3-aminopropyl)-N-[1-(5-benzyl-3-methyl-4-oxo-[1,2]thiazolo[5,4-d]pyrimidin-6-yl)-2-methylpropyl]-4-methylbenzamide (AZD4877), from a series of novel kinesin spindle protein (KSP) inhibitors, as exhibiting both excellent biochemical potency and pharmaceutical properties suitable for clinical development. The selected compound arrested cells in mitosis leading to the formation of the monopolar spindle phenotype characteristic of KSP inhibition and induction of cellular death. A favorable pharmacokinetic profile and notable in vivo efficacy supported the selection of this compound as a clinical candidate for the treatment of cancer.

The signals that promote regional growth and development of the brain are not well understood. Sonic hedgehog (Shh) is produced by Purkinje cells of the cerebellum and is a potent inducer of granule cell proliferation. Here, we demonstrate that Shh protein is present in the murine cerebellum during late stages of embryogenesis and is associated with Purkinje cell bodies and their processes. To better determine the role of Shh during cerebellar development, we genetically removed Shh activity specifically from Purkinje cells and the cerebellar anlage of the mouse embryo. We show that Shh is required for expansion of the granule neuron precursor population, but not for the subsequent differentiation of these cells. In addition, the loss of Shh activity influences Purkinje cell development and the formation of folia in the cerebellum. A role for Shh in compartmentalization of the cerebellum is also suggested by the more severe rostral defects observed in the absence of Hedgehog signaling. Together, these findings provide additional evidence for Shh's key regulatory role in controlling growth of the cerebellar primordium.

We have generated a transgenic line that expresses the Cre gene product under the regulation of a 12.5 kb upstream regulatory sequence from the Sox2 gene. Using a R26R reporter line, we show that this transgenic line induces recombination in all epiblast cells by embryonic day (E) 6.5 but little or no activity in other extraembryonic cell types at this time. When crossed to a conditional allele of the Sonic hedgehog gene (Shhc), all Sox2Cre;Shhn/Shhc embryos displayed a phenotype indistinguishable from that of the Shh null mutant. Sox2Cre functioned more efficiently in epiblast-mediated recombination than the Mox2Cre (MORE) transgenic line, which has also been shown to drive Cre-mediated recombination exclusively in the embryonic component of the early mouse embryo. Although most MORE; shhh/shhc embryos have a shh hull phenotype, 33% displayed a milder skeletal phenotype, most likely result of incomplete recombination at egg cylinder stages. In agreement with these findings, Sox2Cre was active earlier and Sox2Cre-mediated recombination was more advanced than MORE-mediated recombination at early gastrulation stages. The Sox2Cre line is likely to be more effective in generating complete, epiblast-specific removal of gene activity, and the mosaic activity of the MORE line will be helpful in generating partial loss-of-function phenotypes in the embryo-proper.

We have generated a transgenic line that expresses the Cre gene product under the regulation of a 12.5 kb upstream regulatory sequence from the Sox2 gene. Using a R26R reporter line, we show that this transgenic line induces recombination in all epiblast cells by embryonic day (E) 6.5 but little or no activity in other extraembryonic cell types at this time. When crossed to a conditional allele of the Sonic hedgehog gene (Shh(c)), all Sox2Cre;Shh(n)/Shh(c) embryos displayed a phenotype indistinguishable from that of the Shh null mutant. Sox2Cre functioned more efficiently in epiblast-mediated recombination than the Mox2Cre (MORE) transgenic line, which has also been shown to drive Cre-mediated recombination exclusively in the embryonic component of the early mouse embryo. Although most MORE; shh(h)/shh(c) embryos have a shh hull phenotype, 33% displayed a milder skeletal phenotype, most likely result of incomplete recombination at egg cylinder stages. In agreement with these findings, Sox2Cre was active earlier and Sox2Cre-mediated recombination was more advanced than MORE-mediated recombination at early gastrulation stages. The Sox2Cre line is likely to be more effective in generating complete, epiblast-specific removal of gene activity, and the mosaic activity of the MORE line will be helpful in generating partial loss-of-function phenotypes in the embryo-proper.

The adult retina is organized into three cellular layers--an outer photoreceptor, a middle interneuron and an inner retinal ganglion cell (RGC) layer. Although the retinal pigment epithelium (RPE) and Müller cells are important in the establishment and maintenance of this organization, the signals involved are unknown. Here we show that Sonic hedgehog signaling from RGCs is required for the normal laminar organization in the vertebrate retina.