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Italy was on the ‘front line’ in two senses during the Cold War. On the one hand, the southern end of Churchill’s ‘iron curtain’, Trieste, was on Italy’s redrawn frontier with Yugoslavia. It was still in Italian hands, but only just, and it continued to be disputed with Tito’s new communist regime until 1954. Even more significantly, it could be said that the ‘front line’ in the Cold War actually passed through Italy itself and that the war was ‘fought’ on Italian territory because Italy possessed the second largest Communist Party in the West. Allied in a ‘popular front’ with the still strongly marxian Italian Socialist Party, it presented a formidable challenge in the crucial 1948 general elections. Pope Pius XII and the Italian church hierarchy viewed this situation in apocalyptic terms, especially in 1948. Their response was, as the sociologist Gianfranco Poggi has described it, ‘one of maximum involvement and maximum commitment’ of the Church’s resources.1 The Catholic Action movement, with its massive presence in most areas of Italian civil society, was summoned to battle, a huge propaganda effort was launched in 1948, and in the longer term, popular devotions and the papal cult of the personality were employed against the ‘enemy’, along with the unleashing by the Holy Office in 1949 of the Church’s ultimate weapon — excommunication of all those involved in any way with the left.

The inclusion of Isodictyophorus Briq. in Plectranthus L’Her. is completed with the transfer of I. reticulatus (A. Chev.) J. K. Morton, for which a new combination, Plectranthus reticulatus B. J. Pollard, is made. Two IUCN conservation assessments are provided for P. reticulatus and P. defoliatus.

Background: Defects in the multifactorial DNA damage response (DDR) are common in cancer, making them vulnerable to inhibitors of other components of the DDR. ATR is a key component of the DDR, signalling ssDNA, arising from stalled replication forks, resected double strand breaks (DSBs) and NER intermediates to activate cell cycle arrest at the S/G2 checkpoints and initiate homologous recombination repair (HRR). Tumors often have a dysfunctional G1 checkpoint e.g. due to p53 mutations or other DDR defects. ATR inhibition of the S/G2 checkpoints and HRR has the potential to exploit these defects and selectively target cancer. VE-821 is a potent ATR inhibitor under advanced pre-clinical investigation Hypotheses: 1) Cells with dysfunctional p53 will be more sensitive to ATR inhibition. 2) Defects in other DDR components will sensitise cells to ATR inhibition. Methods: The survival of isogenic p53 mutant/null and wt/corrected human cell lines (HCT116 p53+/+ and p53-/- or U2OS p53WT and p53DN (dominant negative)) and a panel of repair defective Chinese hamster ovary and lung fibroblast cells was determined using colony formation after 24 hours exposure VE-821 alone or in the presence of gemcitabine or ionising radiation. Results: There was no significant difference in sensitivity to VE-821 between p53 wt or mutant/null cells in either HCT116 (LC50 = 2.13 and 4.56 µM, respectively) or U2OS pairs (LC50 = 2.54 and 3.34 µM, respectively). However, 1 µM VE-821 significantly potentiated the antimetabolite, gemcitabine, in HCT116 p53-/- (4.3-fold) but not p53+/+ cells (1.5-fold), and enhanced cell kill by 100 nM gemcitabine 40-fold in U2OS p53DN cells but only 3-fold in U2OS p53WT cells. Chinese hamster ovary cells defective in XRCC1 (single stranded break repair) were more sensitive to single agent VE-821 than the parental wt cells (35% and 55% survival at 10 µM, respectively). Data on other repair-defective cell lines will be presented. Conclusions: p53 status is not a determinant of sensitivity to VE-821 as a single agent but VE-821 preferentially sensitises cells lacking functional p53 to gemcitabine. This supports the hypothesis that p53 status may be predictive of cellular response to ATR inhibition in combination with certain chemotherapies. Furthermore, data using Chinese hamster cell lines indicate that DDR defects may be exploited by ATR inhibition. Citation Format: Fiona K. Middleton, Tao Chen, John R. Pollard, Nicola J. Curtin. Investigating p53 and other potential determinants of cell sensitivity to ATR inhibition by VE-821. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2418. doi:10.1158/1538-7445.AM2014-2418

The fly Drosophila melanogaster is one of the most intensively studied organisms in biology and serves as a model system for the investigation of many developmental and cellular processes common to higher eukaryotes, including humans. We have determined the nucleotide sequence of nearly all of the ∼120-megabase euchromatic portion of the Drosophila genome using a whole-genome shotgun sequencing strategy supported by extensive clone-based sequence and a high-quality bacterial artificial chromosome physical map. Efforts are under way to close the remaining gaps; however, the sequence is of sufficient accuracy and contiguity to be declared substantially complete and to support an initial analysis of genome structure and preliminary gene annotation and interpretation. The genome encodes ∼13,600 genes, somewhat fewer than the smaller Caenorhabditis elegans genome, but with comparable functional diversity.

The role which Catholicism played in the formation of the working class in Italy was profoundly different from that which it played in the US, whereas it presents many similarities with situations elswhere in Europe. Despite those similarities, however, the role which it played in Italy was predominantly determined by a number of factors peculiar to that country. The first was the nature of the working class itself: given the pattern of economic development in post-Unification Italy, the working class was essentially heterogeneous, consisting not only of an urban, industrial proletariat, not to mention a large residual artisan class, but a rural, agrarian proletariat of landless labourers, as well as various strata of peasant farmers. Indeed, it might make more sense to talk about the formation of the Italian working classes.

Introduction: The understanding of DNA repairs pathways and their interactions allowed for the development of several targeted therapies, which sensitize neoplastic cells to the effects of DNA damaging chemotherapy. Poly-ADP ribose polymerase (PARP) inhibitors can increase the cytotoxicity of several anticancer agents by inhibiting various DNA repair pathways including Rad51 related homologous recombinational repair (HRR). However, inhibition of PARP may result in compensatory activation of the ATR pathway partially limiting the sensitization of chemotherapeutic agents seen with PARP inhibitors. Recently, a specific ATR inhibitor, VE-821 has demonstrated excellent sensitization to various chemotherapeutic agents with preferential antitumor activity in tumor cells as compared to normal cells (Reaper PM et al Nature Chem Biol 7: 428-30, 2011) Colon cancer cell lines: HCT116/Lovo (p53 wild type) and HT29 (p53 mutated) were treated with SN38, the PARP inhibitor (ABT-888) and/or the ATR inhibitor (VE821) at different concentrations. The SRB cytotoxicity assay was used to determine the IC50 of each drug separately and in combination. DNA damage caused by these agents was quantified by phosphorylated-H2AX. Cell cycle alterations and protein levels (western analysis) were determined after drug treatment. Results: Nontoxic concentrations (0.5-1 μM) of ABT-888 or VE-821 demonstrated a two to three fold decrease in the IC50 of SN38 in the colon cancer cell lines. Significantly, the combination of both inhibitors at the same nontoxic concentrations resulted in a dramatic 4 to 25-fold decrease in the IC50 of SN38 [HCT116 (8.1 nM to 0.3 nM), HT29 (20.5 nM to 3.9 nM) and Lovo (13.5nM to 3 nM)]. Notably, the observed effect was dose-dependent. In the Lovo cell line, the combination of SN38/VE821/ABT888 increased G2/M cell cycle arrest compared to SN38/VE821, SN38/ABT888 and SN38 alone. Similarly, increased phosphorylated-H2AX and apoptosis were seen with SN38/VE821/ABT888 in the Lovo and HCT-116 cell lines. Preliminary data utilizing the annexin-5 apoptosis assay in the HCT116 cell line supports the increased cell killing by the combination of all three agents. In the HCT116 cell line, treated with the SN38/VE821/ABT888 combination as compared to cells treated with SN38 alone, increased expression of p21 along with decreased expression of phospho-Chk1 was observed. Conclusion: The SN38/ABT888/VE821 combination resulted in maximal synergy/chemosensitivity in all colon cancer cell lines, associated with increased DNA damage and apoptosis. Our data suggests that this combination may effectively overcome colon cancer resistance to irinotecan-based chemotherapy. Citation Format: Atlal Abu-Sanad, David Davidson, Yunzhe Wang, John Pollard, Raquel Aloyz, Lawrence Panasci. ATR and PARP inhibition enhances topoisomerase I-dependent DNA damage in colon cancer cell lines. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Synthetic Lethal Approaches to Cancer Vulnerabilities; May 17-20, 2013; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(5 Suppl):Abstract nr B13.

DNA damaging agents have been the cornerstone of cancer therapy for decades yet they provide only modest benefit for most patients. For example, standard of care for patients with non-small cell lung cancer (NSCLC) is dominated by the use of platinating drugs and ionizing radiation (IR), however outcome remains very poor with 5-year survival rates of VE-822 potently inhibits ATR in biochemical assays with Ki 90% of lines showed >3-fold shifts in IC50 for cisplatin in the presence of VE-822, with ~50% of lines showing >10-fold increases in cisplatin cytotoxicity. In contrast normal cells tolerate inhibition of ATR. In a panel of mouse xenograft models, derived from various primary human NSCLC tumor tissues, oral or IV administration of VE-822 strongly sensitized tumors to cisplatin treatment. In many cases, combinations including VE-822 led to tumor regression or extensive tumor growth delay. Inhibition of ATR activity and accumulation of DNA damage by VE-822 was observed coincident with efficacy. When administered alone or in combination with cisplatin VE-822 was well tolerated in mice at doses that block ATR activity. These data support the potential for ATR inhibitors to substantially increase the efficacy of standard-of-care agents in diseases such as NSCLC. Citation Format: Diane Boucher, Peter Charlton, Jean-Damien Charrier, Brinley Furey, Yong Gu, Amy Hall, Brian Hare, Howard Li, Sean Milton, Cheryl Murphy, Philip Reaper, Darin Takemoto, Taturo Udagawa, Yuxin Wang, Mark Wood, John Pollard. Comprehensive preclinical evaluation of VE-822, the first ATR-targeted drug candidate: a novel approach to transforming the efficacy of DNA damaging agents. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-299. doi:10.1158/1538-7445.AM2013-LB-299

DNA damaging agents, such as ionizing radiation (IR), have provided a basis for the treatment of solid tumors for decades, yet they provide only modest benefit for most patients. This reflects, in part, the efficient repair of DNA damage via a complex signaling and repair network known as the DNA damage response (DDR). The ATR, ATM and DNA-PK phosphoinositol 3-kinase-like serine/threonine protein kinases (PIKKs) are the key regulators of the DDR to IR-induced double-strand DNA breaks. This acts to detect the DNA lesions, enforce checkpoints to arrest cell cycle progression, and stimulate repair. Various components of the DDR have been shown to be commonly defective in cancer cells. These cells are widely hypothesized to become dependent on their remaining DDR pathways for survival from DNA damage. The tumor microenvironment has been reported to induce a DDR and in particular, ATR-mediated signaling in response to severe hypoxia. While inhibitors of a number of DDR enzymes, including ATM, DNA-PK, Chk1 and PARP, have been reported, there have been no reports of drug-like ATR inhibitors. Using the first potent and selective ATR inhibitor (VE-821), we now characterize the effects of ATR inhibition on the cellular response to IR and hypoxia. VE-821 sensitizes many cancer cell lines to IR in both clonogenic and short-term (MTS) assays. For example, VE-821 reduced the surviving fraction of MiaPaca-2 cells treated with 6 Gy IR by 3-fold. Consistent with this, ATR inhibition ablated both RAD51 focus formation and phosphorylation of Chk1 (but not Chk2) after IR treatment, delayed the IR-induced G2/M arrest analyzed by flow cytometry and slowed down DNA replication rates measured using DNA fiber technology. Furthermore, we provide evidence that a basis for the sensitization of cancer cell lines to IR with VE-821 is a synthetic lethal interaction between loss of ATM signaling (a frequent event in cancer resulting from loss of function of proteins such as ATM or p53) and ATR inhibition when cells are treated with IR. In keeping with these results, normal cells (with functional ATM) do not appear sensitized to IR by ATR inhibition. In this case a compensatory DDR is activated, which in turn leads to checkpoint arrest and a strong survival response. Importantly, ATR inhibition also sensitized cancer cells to IR under hypoxic conditions, and to anoxia alone. These studies show for the first time that a selective ATR inhibitor can preferentially sensitize cancer cells to ionizing radiation under both normoxic and hypoxic conditions and exploit a synthetic lethal interaction between ATM and ATR signaling. This underpins the broad potential of ATR inhibition as a highly promising new strategy to improve the efficacy of DNA damaging therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5501. doi:10.1158/1538-7445.AM2011-5501

DNA damaging agents have been the cornerstone of solid cancer therapy for decades yet they provide only modest benefit for patients with many tumor types. This reflects, in part, the efficient repair of DNA damage via a complex signaling and repair network known as the DNA damage response (DDR). Key regulators of the DDR are the phosphoinositol 3-kinase-like serine/threonine protein kinase (PIKK) family members ATR, ATM and DNA-PK. The DDR acts to detect DNA lesions, enforce checkpoints to halt cell cycle progression, and stimulate repair. Recent data have shown that elements of the DDR are commonly defective in cancer cells. It is widely believed that these cells become dependent on the remaining DDR pathways for survival from DNA damage. Inhibitors have been reported for a number of DDR enzymes, including ATM, DNA-PK, CHK1 and PARP, however there are no reports of drug-like ATR inhibitors. Here we disclose the in vitro characterization of a potent and highly selective ATR inhibitor (VE-821). This compound selectively blocks ATR signaling in cells (IC50 = 0.7 µM), but has little impact on ATM or DNA-PK signaling (IC50 >10 µM). Treatment with 10 µM VE-821 for 144 h causes little cell death in normal cell lines (5-11 %) but markedly higher death in cancer cell lines (28-46 %). VE-821 also dramatically sensitizes many cancer cells to multiple classes of genotoxic agents including antimetabolites, topoisomerase inhibitors and crosslinking agents; with over 10-fold increases genotoxic potency observed in some cases. In a panel of 36 lung cancer cell lines, VE-821 sensitized the cytotoxic effect of cisplatin to a far greater magnitude and over a broader subset of these lines than potent inhibitors of ATM, Chk1, or PARP. In over half of these cell lines, the IC50 of cisplatin was reduced by greater than 5 fold upon the addition of VE-821. We show that a basis for the cancer-selective effects of VE-821 is a synthetic lethal interaction between loss of ATM signaling (a frequent event in cancer resulting from loss of function of proteins such as ATM or p53) and ATR inhibition when cells encounter DNA damage. In keeping with this, ATR inhibition does not sensitize normal cells (with functional ATM) to the cytotoxic effects of genotoxic therapy. In this case a compensatory DDR is activated that is associated with marked activation of ATM, which in turn leads to reversible checkpoint arrest and a strong survival response. These studies show for the first time that a selective ATR inhibitor can preferentially sensitize cancer cells to genotoxic drugs by exploiting a synthetic lethal interaction between ATM and ATR signaling. This underpins the broad potential of ATR inhibition as a highly promising new strategy to improve the efficacy of genotoxic therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5491. doi:10.1158/1538-7445.AM2011-5491

The Costa del Sol is one of the world’s most developed coasts. Over the past 40 years much of the natural coastline has been destroyed and replaced with tourist-related urban facilities, including high rise apartments, condominiums and hotels. The urban shorefront has become a zone of conflict, as engineers have tried to stem progressive erosion through the construction of seawalls, bulkheads, groynes and breakwaters. The performance of these structures has ranged from satisfactory to poor, in some places exacerbating beach loss and shoreline recession. At Estepona and Marbella, groynes built in the 1960s have recently been removed to assist in stabilising the beach. The environmental stress on the physical resources of the coast may be traced to spiralling demands for water and aggregates. Low seasonal rainfall leads to large-scale water regulation in order to guarantee supplies for agriculture and the urban areas, and this in turn has reduced coastal stream discharge to the point where many rivers are dry for seven to nine months a year. Consequently, sediment delivery to the coast is minimised, and this has resulted in beach erosion, especially along the flanks of deltas. An additional problem arises from the extraction of sand and gravel for construction and agricultural purposes, further depleting beach levels. Also, the recent construction of marinas along the coast has encouraged remobilisation of beach sediment and facilitated its transport onto the shelf. The current phase of large-scale beach nourishment is serving to recycle these materials. There is growing awareness and, inevitably, concern about the future of the Costa del Sol environment, as embodied in recent national and regional directives aimed at reducing stress.

"Mortality data of the Federal Republic of Germany and the GDR [German Democratic Republic] are compared by age, sex and cause of death....Within the period from 1976 to 1986 life expectancy has risen in the Federal Republic of Germany, whereas in the GDR it increased to a minor extent only, so that the gap between the two parts of Germany has clearly enlarged. This enlarged gap in life expectancy largely depends on a higher mortality in the GDR due to diseases of the circular system and the respiratory organs, external influences and 'other' causes of death. The last-mentioned two groups also 'contain' the normally [separately] reported causes of death 'cirrhosis of the liver', 'suicide', 'accidents', and some diseases of the digestive system." (SUMMARY IN FRE)

Many factors need to be taken into account in setting the premium rate for an insurance contract. The more important ones, which are discussed in this chapter, are illustrated by the following breakdown of the office premium. risk premium: claim frequency claim size claims-handling expenses effect of a deductible effect of excess of loss reinsurance loading for contingencies adjustments for inflation adjustments for investment income expenses: proportional to premium per policy overheads profit loading market forces The principles involved are clearly demonstrated in the rating of the various personal lines of insurance (householders’ insurance, motor vehicle damage and so on), for which adequate statistics are usually available and claims are normally notified and settled with little delay. The same factors need to be considered and the same basic approach to rating adopted in respect of other classes of business, but, for many of these classes, other complications arise which cloud the basic rating routine. Long delays in notification and settlement of liability claims, for example, complicate the compilation of statistics on claim frequency and claim size and the estimation of the underlying distributions for the various liability classes.

MK-0457 is a pan-Aurora kinase inhibitor that has firstly shown clinical activity in BCR-ABL positive leukemia patients harboring the T315I mutation. Combining MK-0457 with ABL kinase inhibitors may provide several advantages, including enhanced efficacy and the potential to reduce the emergence of new resistant mutations. In the present study, we investigated the combined effects of MK-0457 and dasatinib in T315I mutant-expressing cells. Co-treatment with MK-0457and dasatinib caused significantly more inhibition of colony growth of primary leukemia cells and BaF3 cells expressing T315I BCR-ABL than either drug alone. In contrast, we did not observe the enhanced effects of MK-0457 and imatinib in T315I BCR-ABL-expressing cells. Treatment with 5 μM of MK-0457 for 48 hrs induced apoptosis in T315I BCR-ABL BaF3 cells, whereas, exposure to combination with 1.0 μM of MK-0457 and 50 nM of dasatinib exerted enhanced apoptotic effect. Combined treatment with MK-0457 and dasatinib also associated with more PARP cleavage, which is due to increased activation of caspase-3 and caspase-9 during apoptosis. Following co-treatment with MK-0457 and dasatinib caused more attenuation of the level of phospho-Stat5 and the downstream signal transducer, including Bcl-XL, Mcl-1, and cyclin D1. We also observed that combined treatment with MK-0457 and dasatinib enhanced the activation of p38MAP kinase and MAPKAP Kinase-2 (MK2). Pretreatment with p38MAP kinase or MP2 siRNA in T315I BCR-ABL BaF3 cells reduced the induction of apoptosis after MK-0457 and dasatinib exposure. These results suggest that p38MAP kinase and MK2 have a critical role for the induction of apoptosis after MK-0457 and dasatinib exposure. To assess the mechanism of combination effect between MK-0457 and dasatinib on T315I BCR-ABL-expressing cells, we used RNA interference to determine whether reduction of SRC-family kinases affected the growth inhibition. BaF3 cells expressing T315I BCR-ABL pretreated with Lyn or Hck siRNA showed enhanced growth inhibition with MK-0457. These results demonstrate that the enhanced growth inhibition by MK-0457 and dasatinib in T315I-expressing cells may be mediated by Lyn and Hck. To assess the in vivo efficacy of MK-0457 and dasatinib, athymic nude mice were injected i.v. with BaF3 cells expressing T315I mutant form of BCR-ABL. 24 hrs after injection, the mice were divided four groups (5 mice per group), with each group receiving either vehicle, MK-0457 (30mg/kg b.i.d.; ip for 5 days), dasatinib (10mg/kg q.d.; po for five days), MK-0457 (30mg/kg b.i.d.; ip for 5 days) + dasatinib (10mg/kg q.d.; po for five days). The treatment was repeated in every 3 weeks. The vehicle or dasatinib-treated mice died of a condition resembling acute leukemia by 28 days; the MK-0457 only-treated mice survived more than 56 days, and the combination of MK-0457 + dasatinib -treated mice survived more than 70 days. Histopathologic analysis of vehicle or dasatinib-treated mice revealed infiltration of the spleen and bone marrow with leukemic blasts. In contrast, histopathologic analysis of organs from MK-0457 plus dasatinib-treated mice demonstrated normal tissue architecture and no evidence of residual leukemia. Taken together, the present study shows that the combination of MK-0457 and dasatinib exhibits a desirable therapeutic index that can reduce the in vivo growth of T315I mutant form of BCR-ABL-expressing cells in an efficacious manner.

7050 Background: MK-0457 was designed as a small-molecule inhibitor of the Aurora kinases, AUR A, B, and C (Ki,app = 0.66–18 nM). Kinase screens indicate high selectivity of MK-0457 over other Ser/Thr kinases, but nM potency against a small set of wild type and mutant Tyr kinases, including BCR-ABL, JAK-2, and FLT-3. Consistent with this biochemical profile, MK-0457 has clinical activity in patients with chronic myeloid leukemia (CML) and acute lymphocytic leukemia (ALL) with the T315I BCR-ABL mutation and in patients with JAK-2 positive myeloproliferative diseases (MPD). This study compared the enzyme inhibitory profile of MK0457 with kinase inhibition profiles in cell models to further associate biological and clinical activity. Methods: MK-0457 was tested against a panel of ∼200 kinases (at Km for ATP). Viability of BaF3 cells transfected with wild type or mutant BCR-ABL or FLT3 (D845Y and FLT3-ITD mutants) was measured in MTS assays (72 hrs). Cell cycle profiles were determined by FACS analyses. Cellular inhibition of kinase activities was measured by Western Blots using phospho-substrate specific antibodies. Results: In enzyme profiling, MK-0457 (200 nM) shows = 50% inhibition of ABL, T315I ABL, ABL2, FLT-3, FLT-3 D835Y, BMX, JAK2, YES, LYN, and LCK kinases. Western blot analyses indicate nM IC50s for inhibition of phosphorylation of AUR B, Histone H3 (AUR B substrate), FLT-3, BCR-ABL, and STAT5 (substrate of BCR-ABL and FLT-3 mutants). In addition to AUR kinase inhibition, the Tyr kinases are expected to be inhibited at clinical doses of 24–32 mg/m2/hr (achieving plasma exposures of >1 μM in patients). Depsite its multi-kinase profile, MK-0457 induces similar cytotoxicity (IC50 ∼300 nM) and shows an AUR B-like inhibitory phenotype of G2/M arrest, endoreduplication and apoptosis in BaF3 cells transfected with ABL or FLT-3 (mutant and wild type) kinases. Conclusions: MK-0457 is a potent Aurora kinase inhibitor with cross-reactivity against a small set of Tyr kinases. Although MK-0457 displays an AUR inhibition phenotype in cells, inhibition of these Tyr kinases may contribute to its observed clinical activity in T315I mutant CML and ALL and in JAK-2 positive MPD. No significant financial relationships to disclose.

Background. MK-0457 (VX-680) is a small-molecule inhibitor of Aurora kinases A, B, and C, FLT3, and JAK-2 with nanomolar level broad spectrum pre-clinical anti-tumor activity. The T315I BCR-ABL mutation mediates high level resistance to imatinib, dasatinib and nilotinib. A Phase I study of MK-0457 is being conducted in patients with refractory hematologic malignancies. The study regime is a 5 day continuous IV regimen given every 2 to 3 weeks. Methods. Fifteen CML patients received MK-0457 dosed at 8, 12, 16, 20, 24, 28, and 32 mg/m2/hr (11/15 had detectable BCR-ABL T315I). Peripheral blood for biomarker studies was available from patients at the 12 mg/m2/hr dose and above. Specimens were drawn prior to initiation of the infusion and at the end of the infusion (Day 5) during Cycle 1. Patient specimens have been analyzed using flow cytometry assays to measure BCR-ABL inhibition (phospho-CRKL), FLT3 and JAK-2 inhibition (phospho-STAT5) and Aurora kinase inhibition (phospho-histone H3). Results. MK-0457 has in vitro activity against cells expressing wild-type or mutated BCR-ABL, including the T315I BCR-ABL mutation. In CML and ALL patients with the T315I BCR-ABL mutation, MK-0457 is capable of inhibiting BCR-ABL as measured by the phospho-CRKL assay. The observed decrease in BCR-ABL activity trends with plasma exposure to MK-0457. At dose levels less than 20 mg/m2/hr, plasma exposure of MK-0457 typically does not exceed 1 micromolar and inhibition of BCR-ABL has not been observed. At exposures that exceed 1 micromolar, MK-0457 inhibits BCR-ABL, with pCRKL abundance falling below the lower limit of detection in one patient dosed at 24 mg/m2/hr and decreasing compared to baseline in several other patients. There is also a trend correlating degree of BCR-ABL inhibition as measured by the pCRKL biomarker and clinical response to MK-0457. Conclusions. MK-0457 inhibits BCR-ABL activity in CML and ALL patients with the BCR-ABL T315I mutation. Objective responses have been observed in patients without demonstrable BCR-ABL inhibition, suggesting that inhibition of Aurora kinase may also contribute to the clinical efficacy of MK-0457 in CML and ALL patients with the BCR-ABL T315I mutation.

Multiple Myeloma (MM) remains an incurable plasma cell neoplasia, despite recent additions in the therapeutic arsenal for its management. Aurora kinases play integral roles in the orchestration of chromosomes and cytoskeletal mobility during the process of cell division. Aurora kinase activity has been implicated in several tumor types, including ovarian, colon, and prostate cancers. To determine whether inhibition of Aurora kinase activity could attenuate myeloma cell survival, we performed studies of the Aurora kinase inhibitor VE465 (Vertex Pharmaceuticals / Merck & Co., Inc.). VE465 inhibits all 3 Aurora isoforms (Aur A, B and C) with approximate Ki values of 1, 26, and 8.7 nM respectively. MTT colormetric survival assays (72–96hrs exposure) showed that VE465 is active against a wide panel of human MM cell lines: 26 of 38 MM cell lines had IC50 values at or < 100 nM, which are significantly lower than IC50 values for normal hematopoietic cells, e.g. unstimulated or PHA-stimatuled PBMCs. Importantly, VE465 was active in vitro against MM cell lines and/or primary MM tumor cells resistant to various anti-MM therapeutics, including dexamethasone, alkylating agents, anthracyclines, the proteasome inhibitor bortezomib, and/or immunomodulatory thalidomide derivatives (IMiDs). Moreover, VE465 maintained its activity despite the presence of protective bone marrow-derived cytokines (e.g. IL-6). PI cell cycle analyses showed that VE465 causes (even within 8 hrs of treatment) caused pronounced G2 arrest, followed by significant shift of MM cells to sub-G1 gate, consistent with cell death. Immunoblotting analyses confirmed that VE465 treatment induces cleavage of PARP, as well as cleavage of caspases-8 and -9, without significant changes in the expression levels of several key molecular effectors (e.g. Mcl-1, Bax, p53, hsp70, hsp90, hs27) which have been previously implicated in the mechanism of anti-MM activity of diverse other therapeutics. Screening of VE465-based combination regimens with other anti-MM agents showed additive effects of VE465 with the histone deacetylase inhibitor Vorinostat (SAHA) (Merck & Co., Inc). Ongoing studies in our Center are addressing the identification of specific molecular markers correlating with the degree of sensitivity of MM cells to VE465. Our in vitro evidence for induction of MM cell death and therapeutic window for the anti-MM effect of VE465, its ability to overcome protective effect of BM-derived cytokines, and the clearly distinct pattern of molecular sequelae of VE465 compared to several other agents in our current anti-MM therapeutic armamentarium, all suggest that Aurora kinase inhibition represents an intriguing novel targeted treatment strategy in MM. Importantly, these studies, particularly the identification of a sizeable subset of MM cell lines with higher sensitivity to VE465 than normal cells, provide the framework for in vivo VE465 studies in progress, alone and in combination with other anti-MM agents, to inform the design of potential clinical trials of this class of agents for MM.

MK-0457 (VX-680) is a potent small molecule inhibitor of the Aurora-family of protein kinases that inhibits cell proliferation through aberrant mitosis and failed cytokinesis. Affected cells are subsequently deleted by apoptosis. Tumor regression and increased survival have been reported in multiple in-vivo models and in January 2005 MK-0457 entered Phase I clinical trials as the first in class Aurora inhibitor. Although MK-0457 demonstrates significant selectivity over the vast majority of kinases tested, it has been shown to potently inhibit Flt-3 and Abl, both of which are targets for anti-cancer therapies. Most notably MK-0457 potently inhibits a series of mutant Abl kinases including the multi-drug resistant T315I form that is commonly expressed in Gleevec refractory CML and ALL. We report here the first MK-0457 / Aurora co-complex crystal structure and show that it adopts a closed inactive conformation. By comparison with other reported kinase structures we provide a structural basis for the cross-reactivity with wild type and drug resistant mutants of the Abl kinase. We also show that MK-0457 blocks both Aurora and Abl cellular kinase activity and displays potent cytotoxic activity against a range of leukemic cell lines, including Gleevec resistant cells that are reliant on mutant Abl for growth and survival. Similar observations have recently been made in the clinic, where new data show that MK-0457 has activity in patients with BCR-ABL T315I mutant CML and Philadelphia positive ALL.

The lowest oxidation potentials for 1,2-dithiins are in the range of 0.67-0.96 V in acetonitrile and 0.81-1.04 V in dichloromethane. These oxidation potentials are less anodic than expected based on the ionization potentials of 1,2-dithiin determined by photoelectron spectroscopy. Theoretical calculations suggest that the reason for this difference is a change in optimized geometry between 1,2-dithiin and its oxidized species.

Botanical inventories in western Cameroon (SW and NW provinces) have led to the publication of two comprehensive checklists (Cable & Cheek 1998; Cheek et al. 2000). In February 2003, during preparation of "The Plants of Mount Kupe and the Bakossi Mountains", the third in this series, the authors recognised a previously undescribed species of Bulbophyllum Thouars, described in this paper. Other new orchids from this area have been described recently (Cribb 1996; Cribb & Pollard 2002).

Summary. Two new varieties of Dorstenia poinsettiifolia Engl., var. achoundongiana Cheek & Bygrave and var. etugeana B. J. Pollard are described and illustrated from western Cameroon. In addition, Dorstenia astyanactis Ake Assi is newly reported from Cameroon, and notes on the distribution, ecology and conservation of all taxa are provided.

Two new orchids from Cameroon, Polystachya kupensis P. J. Cribb & B. J. Pollard and Angraecum sanfordii P. J. Cribb & B. J. Pollard are described and illustrated. Polystachya stauroglossa Kraenzl. is newly reported from Cameroon. Notes on their distribution, habitat and conservation are provided.