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Ovarian cancer remains a significant challenge in women worldwide. Tumors of the high-grade serous carcinoma (HGSC) type represent the most common form of the disease. Development of new therapies for HGSC has been hampered by a paucity of preclinical models in which new drugs could be tested for target engagement and anti-tumor efficacy. Here, we systematically assessed in vivo growth of ovarian cancer cells, including six validated HGSC cell lines, in highly immunocompromised NSG mice by varying the injection site. We found that, with the exception of OVCAR3, HGSC cell lines COV318, COV362, KURAMOCHI, OVCAR4, and OVSAHO, generally demonstrate poor growth as either subcutaneous or intraperitoneal xenografts. Intrabursal injections performed with KURAMOCHI and COV362 cells did not improve tumor growth in vivo. Additional analysis revealed that OVSAHO and COV362 express moderate levels of estrogen receptor (ERα), which translated into improved growth of xenografts in the presence of 17β-Estradiol. Surprisingly, we also found that the growth of the widely used non-HGSC ovarian cell line SKOV3 could be significantly improved by estrogen supplementation. By describing successful establishment of estrogen-sensitive HGSC xenograft models, OVSAHO and COV362, this work will enable testing of novel therapies for this aggressive form of ovarian cancer. [ABSTRACT FROM AUTHOR]

Background: The main role of the cell cycle is to enable error-free DNA replication, chromosome segregation and cytokinesis. One of the best characterised checkpoint pathways is the spindle assembly checkpoint, which prevents anaphase onset until the appropriate attachment and tension across kinetochores is achieved. MPS1 kinase activity is essential for the activation of the spindle assembly checkpoint and has been shown to be deregulated in human tumours with chromosomal instability and aneuploidy. Therefore, MPS1 inhibition represents an attractive strategy to target cancers.Methods: To evaluate CCT271850 cellular potency, two specific antibodies that recognise the activation sites of MPS1 were used and its antiproliferative activity was determined in 91 human cancer cell lines. DLD1 cells with induced GFP-MPS1 and HCT116 cells were used in in vivo studies to directly measure MPS1 inhibition and efficacy of CCT271850 treatment.Results: CCT271850 selectively and potently inhibits MPS1 kinase activity in biochemical and cellular assays and in in vivo models. Mechanistically, tumour cells treated with CCT271850 acquire aberrant numbers of chromosomes and the majority of cells divide their chromosomes without proper alignment because of abrogation of the mitotic checkpoint, leading to cell death. We demonstrated a moderate level of efficacy of CCT271850 as a single agent in a human colorectal carcinoma xenograft model.Conclusions: CCT271850 is a potent, selective and orally bioavailable MPS1 kinase inhibitor. On the basis of in vivo pharmacodynamic vs efficacy relationships, we predict that more than 80% inhibition of MPS1 activity for at least 24 h is required to achieve tumour stasis or regression by CCT271850. [ABSTRACT FROM AUTHOR]

There is unmet need for chemical tools to explore the role of the Mediator complex in human pathologies ranging from cancer to cardiovascular disease. Here we determine that CCT251545, a small-molecule inhibitor of the WNT pathway discovered through cell-based screening, is a potent and selective chemical probe for the human Mediator complex-associated protein kinases CDK8 and CDK19 with >100-fold selectivity over 291 other kinases. X-ray crystallography demonstrates a type 1 binding mode involving insertion of the CDK8 C terminus into the ligand binding site. In contrast to type II inhibitors of CDK8 and CDK19, CCT251545 displays potent cell-based activity. We show that CCT251545 and close analogs alter WNT pathway-regulated gene expression and other on-target effects of modulating CDK8 and CDK19, including expression of genes regulated by STAT1. Consistent with this, we find that phosphorylation of STAT1SER727 is a biomarker of CDK8 kinase activity in vitro and in vivo. Finally, we demonstrate in vivo activity of CCT251545 in WNT-dependent tumors. [ABSTRACT FROM AUTHOR]

To study the interactions of the heat shock protein 90 (HSP90) inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) and carboplatin in vitro and in vivo. The combination of 17-AAG and carboplatin on the growth inhibition of A2780, SKOV-3, IGROV-1 and HX62 human ovarian cancer cells was studied in vitro by MTT assays. The effect of the sequence of administration of both drugs was further investigated in A2780 cells by sulforhodamine B assays. The ability of 17-AAG to deplete HSP90 client proteins either alone or in combination with carboplatin was evaluated by western blotting. Tumor concentrations of 17-AAG and carboplatin alone or in combination in vivo were determined by validated liquid chromatography with ultraviolet detection and atomic absorption spectroscopy methods. The growth inhibitory effects of 17-AAG, carboplatin and the combination were studied in the A2780 xenograft model. The combination index (CI) at fu0.5 for 17-AAG plus carboplatin was 0.97 (±0.12 SD) when A2780 cells were exposed to carboplatin followed by 17-AAG indicating additivity. The addition of carboplatin did not alter the ability of 17-AAG to cause C-RAF, CDK4 and p-AKT depletion or HSP70 induction. Tumor 17-AAG and carboplatin concentrations were not significantly different in the single agent and combination arms. Tumor weights relative to controls on day 6 (T/C) were 67% for the carboplatin, 64% for the 17-AAG and 22% for the combination. In the specified sequences of drug exposure, 17-AAG and carboplatin have additive growth inhibitory effects in vitro and beneficial effects were seen with the combination in vivo. These findings form the basis for the possible evaluation of 17-AAG and carboplatin in a clinical trial. [ABSTRACT FROM AUTHOR]

Dolastatins 10 and 15 are small peptides isolated from the marine sea hare Dolabella auricularia that have been shown to interact with tubulin. Their growth-inhibitory properties were compared using panels of human ovarian and colon-carcinoma cell lines. Both agents were very potent inhibitors of cell growth, with dolastatin 10 being an average of 9.1-fold more potent than dolastatin 15 [mean 50% inhibitory concentrations (IC values) 2.3×10 and 2.1×10 M, respectively; P <0.05] and more potent than paclitaxel or vinblastine. While neither dolastatin exhibited marked cross-resistance in cisplatin- or etoposide-resistant cell lines, contrasting effects were observed using an acquired doxorubicin-resistant (CH1doxR, 100-fold resistant, P-glycoprotein overexpressing) cell line. Resistance was significantly higher to dolastatin 15 (12.7-fold) than to dolastatin 10 (only 3.2-fold; P <0.05) and was reversible in both cases by verapamil. In vivo, using a s. c. advanced-stage human ovarian carcinoma xenograft and equitoxic doses, greater activity was observed with dolastatin 10 (6.1-day growth delay) versus 0.4 days for dolastatin 15. A radioimmunoassay for dolastatin 10 (limit of detection in mouse plasma 5 ng/ml) was developed. The rabbit antiserum also cross-reacted by 65% with dolastatin 15. Comparative mouse pharmacokinetics following i. v. administration of 1 mg/kg showed that both compounds are rapidly eliminated, but with a shorter second-phase half-life ( t) being observed for dolastatin 15 (being detectable for only up to 4 h post-administration), the t being 3 times longer for dolastatin 10. In addition, areas under the plasma concentration-time curve (AUC values) were 1.6-fold higher for dolastatin 10 (333 versus 208 ng ml h). Plasma binding of dolastatin 10 exceeded 90%. The highly sensitive RIA will be useful for pharmacokinetic studies in conjunction with the planned phase I clinical trials of these novel, extremely potent, tubulin-binding agents, of which dolastatin 10 appears to possess the more promising preclinical features. [ABSTRACT FROM AUTHOR]

Resistance to the clinically used platinum-based drugs cisplatin and carboplatin represents a major limitation to their clinical effectiveness. Using cisplatin-sensitive and -resistant human ovarian carcinoma cell lines previously characterized in terms of their major underlying mechanisms of resistance, we attempted to potentiate the cytotoxic effects of cisplatin and carboplatin using the clinically used antifungal agent amphotericin B (AmB). Using non-toxic concentrations of AmB (up to 15 micrograms/ml) and continuous exposure to cisplatin, a concentration-dependent selective potentiation (maximum of 3.2-fold) of cisplatin cytotoxicity was observed in two cisplatin-resistant cell lines (41McisR6, acquired resistant, and HX/62, intrinsically resistant). In both these cisplatin-resistant cell lines, previous studies have shown resistance to be due primarily to reduced platinum uptake. Notably, no significant potentiation was observed in the parent 41M cell line, in the intrinsically resistant SKOV-3 cell line (where reduced drug accumulation plays only a partial role in determining resistance) or in a pair of cell lines (CH1 and its acquired-resistant variant CH1cisR6) were reduced drug uptake does not play any role in determining resistance. The potentiating effect of AmB was lower with carboplatin and not significant in all cell lines. Platinum uptake following a 2-h exposure of cells to cisplatin was enhanced 3.5-fold in 41McisR6 cells (producing platinum levels similar to those obtained in the parental line) and 1.7-fold in 41M cells by the concomitant exposure to AmB. These data indicate that the potentiation of cisplatin (and carboplatin) cytotoxicity by AmB is not due to a generalized membrane disruption, as effects were observed only in resistant lines where reduced drug transport was apparent. Moreover, AmB did not increase the cytotoxicity of JM216 [bis-acetatoammine(cyclohexylamine)dichloroplatinum (II)], a recently developed, more lipophilic orally active platinum drug, in the 41M/41McisR6 lines. JM216 has previously been shown to circumvent acquired cisplatin resistance due to decreased drug uptake. In vivo, however, using the HX/62 xenograft. AmB (at its maximum tolerated dose of 20 mg/kg; q7d x 4 schedule) did not enhance the antitumour effect of carboplatin (at its maximum tolerated dose of 80 mg/kg; q7d x 4 schedule. [ABSTRACT FROM AUTHOR]

A disease-oriented approach to the discovery of novel platinum anticancer drugs has been established through the setting up of parallel human ovarian-carcinoma cell lines and xenografts. The correlation between in vitro and in vivo antitumour activity was determined for four reference platinum agents (cisplatin, carboplatin, iproplatin and tetraplatin) in eight companion lines. Two methods of assessing antitumour effect were used in vitro (tritiated thymidine incorporation and sulforhodamine B staining) and three were applied in vivo [28-day treated/control (T/C) ratio, growth delay and specific growth delay]. In vitro, large differences in cytotoxicity across the cell lines were observed for each drug. This was also reflected in the xenografts for cisplatin and carboplatin and, to a lesser extent, for iproplatin. A correlation analysis of in vitro vs in vivo data revealed a high, statistically significant positive correlation for cisplatin and a strong positive correlation for carboplatin. However, for the two platinum(IV) drugs, the correlation was less good. In particular, tetraplatin was markedly less active in vivo (showing a general lack of activity against all of the tumour lines) than its in vitro potency against the cell lines predieted, resulting in poor correlation coefficients. These human tumour panels may be valuable for the elucidation of both cellular/molecular and corresponding in vivo pharmacological mechanisms of platinum drug resistance. Moreover, the HX/62 and SKOV-3 tumour lines, which exhibit a level of intrinsic resistance to the four reference agents both in vitro and in vivo (and which were derived from patients who had not received prior platinum therapy), represent particularly useful evaluation models for the discovery of novel broad-spectrum platinum drugs. [ABSTRACT FROM AUTHOR]

The corresponding author of this article has informed us of concerns about the immunoblots in Fig. 2 which were carried out in the collaborating laboratory of Professor Ann Jackman. [ABSTRACT FROM AUTHOR]

Binary combinations of cisplatin and the 5-HT3 antagonist, BRL 43694, have been used to treat both conventional mice bearing either L1210 leukaemia or ADJ/PC6 plasmacytoma and nude mice xenografted with a human ovarian carcinoma (HX/110). In no case was there evidence for antagonism by the antiemetic of the antitumour properties of cisplatin. [ABSTRACT FROM AUTHOR]

Diffuse intrinsic pontine glioma (DIPG) is a lethal childhood brainstem tumour, with a quarter of patients harbouring somatic mutations in ACVR1, encoding the serine/threonine kinase ALK2. Despite being an amenable drug target, little has been done to-date to systematically evaluate the role of ACVR1 in DIPG, nor to screen currently available inhibitors in patient-derived tumour models. Here we show the dependence of DIPG cells on the mutant receptor, and the preclinical efficacy of two distinct chemotypes of ALK2 inhibitor in vitro and in vivo. We demonstrate the pyrazolo[1,5-a]pyrimidine LDN-193189 and the pyridine LDN-214117 to be orally bioavailable and well-tolerated, with good brain penetration. Treatment of immunodeprived mice bearing orthotopic xenografts of H3.3K27M, ACVR1R206H mutant HSJD-DIPG-007 cells with 25 mg/kg LDN-193189 or LDN-214117 for 28 days extended survival compared with vehicle controls. Development of ALK2 inhibitors with improved potency, selectivity and advantageous pharmacokinetic properties may play an important role in therapy for DIPG patients. Diana Carvalho, Kathryn R. Taylor et al. show that ALK2 inhibitors induce apoptosis of ACVR1 mutant diffuse intrinsic pontine glioma cells in vitro. These compounds also increase the survival of mice carrying ACVR1 mutant brainstem xenografts, suggesting ALK2 as a potential pharmacological target against this lethal cancer. [ABSTRACT FROM AUTHOR]