William Avery

The Novel Combination of Chlorpromazine and Pentamidine Exerts Synergistic Antiproliferative Effects through Dual Mitotic Action

Combination therapy has proven successful in treating a wide variety of aggressive human cancers. Historically, combination treatments have been discovered through serendipity or lengthy trials using known anticancer agents with similar indications. We have used combination high-throughput screening to discover the unexpected synergistic combination of an antiparasitic agent, pentamidine, and a phenothiazine antipsychotic, chlorpromazine. This combination, CRx-026, inhibits the growth of tumor cell lines in vivo more effectively than either pentamidine or chlorpromazine alone. Here, we report that CRx-026 exerts its antiproliferative effect through synergistic dual mitotic action. Chlorpromazine is a potent and specific inhibitor of the mitotic kinesin KSP/Eg5 and inhibits tumor cell proliferation through mitotic arrest and accumulation of monopolar spindles. Pentamidine treatment results in chromosomal segregation defects and delayed progression through mitosis, consistent with inhibition of the phosphatase of regenerating liver family of phosphatases. We also show that CRx-026 synergizes in vitro and in vivo with the microtubule-binding agents paclitaxel and vinorelbine. These data support a model where dual action of pentamidine and chlorpromazine in mitosis results in synergistic antitumor effects and show the importance of systematic screening for combinations of targeted agents.

Selective amplification of glucocorticoid anti-inflammatory activity through synergistic multi-target action of a combination drug.

IntroductionGlucocorticoids are a mainstay of anti-inflammatory therapy, but significant adverse effects ultimately limit their utility. Previous efforts to design glucocorticoid structures with an increased therapeutic window have focused on dissociating anti-inflammatory transcriptional repression from adverse effects primarily driven by transcriptional activation. An alternative to this medicinal chemistry approach is a systems biology based strategy that seeks to amplify selectively the anti-inflammatory activity of very low dose glucocorticoid in immune cells without modulating alternative cellular networks that mediate glucocorticoid toxicity.MethodsThe combination of prednisolone and the antithrombotic drug dipyridamole was profiled using in vitro and in vivo models of anti-inflammatory activity and glucocorticoid-induced adverse effects to demonstrate a dissociated activity profile.ResultsThe combination synergistically suppresses release of proinflammatory mediators, including tumour necrosis factor-α, IL-6, chemokine (C-C motif) ligand 5 (RANTES), matrix metalloproteinase-9, and others, from human peripheral blood mononuclear cells and mouse macrophages. In rat models of acute lipopolysaccharide-induced endotoxemia and delayed-type hypersensitivity, and in chronic models of collagen-induced and adjuvant-induced arthritis, the combination produced anti-inflammatory activity that required only a subtherapeutic dose of prednisolone. The immune-specific amplification of prednisolone anti-inflammatory activity by dipyridamole did not extend to glucocorticoid-mediated adverse effects, including corticosterone suppression or increased expression of tyrosine aminotransferase, in vivo after repeat dosing in rats. After 8 weeks of oral dosing in mice, treatment with the combination did not alter prednisolone-induced reduction in osteocalcin and mid-femur bone density, which are markers of steroid-induced osteoporosis. Additionally, amplification was not observed in the cellular network of corticotroph AtT-20/D16v-F2 cells in vitro, as measured by pro-opiomelanocortin expression and adrenocorticotropic hormone secretion.ConclusionsThese data suggest that the multi-target mechanism of low-dose prednisolone and dipyridamole creates a dissociated activity profile with an increased therapeutic window through cellular network selective amplification of glucocorticoid-mediated anti-inflammatory signaling.

Infra‐Red and Raman Spectra of Polyatomic Molecules. IV. Allene

The infra‐red absorption spectrum and the Raman spectrum of allene were examined. In the infra‐red nine bands were observed at 4200, 2960, 2420, 1980, 1700, 1389, 1165, 1031 and 852 cm—1 and in the Raman effect there were ten individual shifts of 3062, 2992, 2858, 1956, 1684, 1430, 1069, 838, 705 and 353 cm—1. With the aid of heat capacity data it was possible to assign values to the frequencies of all the eleven fundamental modes of vibration of allene. The assignment accounted for all the features of the Raman and infra‐red spectra. Calculations were made of the free energy, the entropy and the heat capacity and expressions for the first two, as functions of temperature, derived.

Adenosine A2A and Beta-2 Adrenergic Receptor Agonists: Novel Selective and Synergistic Multiple Myeloma Targets Discovered through Systematic Combination Screening

The use of combination drug regimens has dramatically improved the clinical outcome for patients with multiple myeloma. However, to date, combination treatments have been limited to approved drugs and a small number of emerging agents. Using a systematic approach to identify synergistic drug combinations, combination high-throughput screening (cHTS) technology, adenosine A2A and β-2 adrenergic receptor (β2AR) agonists were shown to be highly synergistic, selective, and novel agents that enhance glucocorticoid activity in B-cell malignancies. Unexpectedly, A2A and β2AR agonists also synergize with melphalan, lenalidomide, bortezomib, and doxorubicin. An analysis of agonists, in combination with dexamethasone or melphalan in 83 cell lines, reveals substantial activity in multiple myeloma and diffuse large B-cell lymphoma cell lines. Combination effects are also observed with dexamethasone as well as bortezomib, using multiple myeloma patient samples and mouse multiple myeloma xenograft assays. Our results provide compelling evidence in support of development of A2A and β2AR agonists for use in multi-drug combination therapy for multiple myeloma. Furthermore, use of cHTS for the discovery and evaluation of new targets and combination therapies has the potential to improve cancer treatment paradigms and patient outcomes. Mol Cancer Ther; 11(7); 1432–42. ©2012 AACR.

Abstract 1524: M402, a novel heparan sulfate mimetic, inhibits pancreatic tumor growth and desmoplasia potentially via sonic hedgehog signaling in an orthotopic mouse model

One of the hallmarks of pancreatic ductal adenocarcinoma (PDAC) is extensive desmoplasia in the surrounding tumor microenvironment that prevents delivery of chemotherapeutics (such as gemcitabine). Dysregulation of sonic hedgehog (SHH, a heparin-binding morphogen), has been implicated in pancreatic cancer tumorigenesis. We have rationally designed a heparan sulfate mimetic, M402, that has previously been shown in animal studies to reduce metastatic seeding through disruption of several key heparin-binding growth factors (including FGF2, VEGF, SDF-1α and P-Selectin). We hypothesized that M402, alone or in combination with gemcitabine, could potentially modulate tumor-stroma interactions in an orthotopic pancreatic cancer model (Capan-2 cell line) which secretes SHH and displays a desmoplastic response in vivo. Capan-2 human adenocarcinoma cells (∼1x10 6 cells) were injected into the pancreata of immunodeficient mice (Nu/Nu CD-1). M402 (10-40 mg/kg/day) or saline treatment commenced on Day 4 or 32. Gemcitabine (30-60 mg/kg, twice weekly, IP) treatment commenced between Weeks 3-6. At the termination of each study (Week 8), gross anatomical observations were made of the primary pancreatic tumor and metastatic lesions in the surrounding tissues including the spleen, intestines and liver. Gemcitabine was increasingly less effective when started at later time points, but still reduced the primary tumor weight by 60-70% (at 30-45 mg/kg) when treatment was started at week 5. While the addition of M402 to gemcitabine showed some additive effect on primary tumor burden, metastasis, invasion, and surrounding fibrotic lesions appeared particularly impacted by the combination treatment. M402 was also effective as monotherapy showing a dose-dependent reduction in primary tumor weight and fibrotic lesions. Immunohistochemical and qPCR analyses showed reduced fibrosis and SHH signaling with M402 and gemcitabine combination treatment. These results demonstrate that M402 can modulate tumor-stroma interactions involved in the desmoplastic response in a murine model of pancreatic cancer and provide a rationale for the clinical investigation of M402 as a potential anti-desmoplastic agent in pancreatic cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1524. doi:1538-7445.AM2012-1524

Abstract 3113: M402, a novel heparan sulfate mimetic, modulates the desmoplastic response in an orthotopic pancreatic cancer model in mice

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Pancreatic ductal adenocarcinoma (PDAC), currently the fourth leading cause of cancer-related deaths in the US, is one of the most lethal cancers due in part to its resistance to or poor delivery of existing chemotherapeutics (such as gemcitabine). One of the hallmarks of this deadly disease is extensive desmoplasia in the surrounding tumor microenvironment. Recent advances in pancreatic cancer research implicate the involvement of several heparin-binding growth factors that control tumor-stroma interactions (including sonic hedgehog (shh), HB-EGF, TGFs, PDGF and HGF). We have rationally designed a heparan sulfate mimetic, M402, that has previously been shown in animal studies to reduce metastatic seeding through disruption of multiple heparin-binding growth factor-mediated pathways. We hypothesized that M402, alone or in combination with gemcitabine, could potentially modulate tumor-stroma interactions in an orthotopic pancreatic cancer model (Capan-2 cell line) which displays moderate desmoplasia in vivo. Capan-2 human adenocarcinoma cells (∼1×106 cells) were injected into the pancreases of immunodeficient mice (Nu/Nu CD-1, female, 8 weeks old). M402 (40 mg/kg/day) or saline treatment commenced on day 4 (via osmotic pump). Gemcitabine (30-60 mg/kg, twice weekly, IP) treatment commenced between weeks 3-6. At the termination of each study (week 8), gross anatomical observations were made of the primary pancreatic tumor and metastatic lesions in the surrounding tissues including the spleen, intestines and liver. The primary pancreatic tumor was weighed and further processed for immunohistochemistry and mRNA expression of fibrotic matrix markers (fibronectin and COLI) and shh signaling (shh and Gli1). Gemcitabine was increasingly less effective when started at later time points, but still reduced the primary tumor weight by 60-70% (30-45 mg/kg) when treatment was started at week 5. M402 was also effective as monotherapy. The addition of M402 to gemcitabine showed only marginally more activity on primary tumor burden; however, metastasis, invasion, and surrounding fibrotic lesions appeared more impacted by the combination treatment than either agent alone. Immunohistochemical and qPCR analyses showed reduced shh and Gli1 with M402 and gemcitabine combination treatment. These results show that M402 can modulate tumor-stroma interactions involved in the desmoplastic response in a murine model of pancreatic cancer. These results provide a rationale for the clinical investigation of M402 as a potential anti-desmoplastic agent in pancreatic cancer patients. 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 3113. doi:10.1158/1538-7445.AM2011-3113