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The Food and Drug Administration (FDA) has approved duvelisib (DUV) for managing follicular lymphoma, small lymphocytic lymphoma, and relapsed or refractory chronic lymphocytic leukemia. Seliciclib (SEL) is a candidate drug for these cancers, neurodegenerative disorders, renal diseases, several viral infections, and chronic inflammation disorders. This work describes the development and validation of a 96-microwell-based spectrophotometric method (MW-SPM) and a high-performance liquid chromatography with fluorescence detection method (HPLC-FD) for the quantitation of DUV and SEL in their bulk forms and capsules. The MW-SPM is based on the formation of colored charge transfer complexes (CTCs) as products for the reactions of DUV and SEL, as n-electron donors, with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ), as a π-electron acceptor. The absorption intensity of the CTCs was measured by using an absorbance plate reader at 450 nm. The stoichiometric ratios of DUV:DDQ and SEL:DDQ were 1:1 and 1:2, respectively, and accordingly the reaction mechanisms were postulated. The HPLC-FD involved the chromatographic separation of DUV and SEL on a Hypersil™ Phenyl HPLC column (250 mm length × 4.6 mm i.d., 5 μm particle diameter) with a mobile phase composed of acetonitrile:acetate buffer, pH 4.5 (35:65, v/v) at a flow rate of 2.2 mL/min. DUV and SEL were detected at 370 nm after excitation at 280 nm. SEL was used as an internal standard (IS) for quantitation of DUV, and DUV was used as an IS for quantitation of SEL. Both MW-SPM and HPLC-FD were validated according to the guidelines of the International Council for Harmonization (ICH) for validation of analytical procedures. The linear ranges for both DUV and SEL were 14.52–200 µg/well (100 µL) and 0.12–3.2 µg/mL for MW-SPM and HPLC-FD, respectively. LOD values in MW-SPM for DUV and SEL were 4.4 and 3.17 µg/well, respectively; however, those for HPLC-FD were 0.03 and 0.05 µg/mL, respectively. The accuracy and precision of both methods were confirmed as the recovery values were ≥98.5% and the values of relative standard deviations (RSD) were ≤2.41%. Both methods were satisfactorily applied to the quantitation of DUV and SEL in their capsules; the mean recovery values were ≥99.2%. Both methods have simple procedures and high analytical throughput. Moreover, they consume a small volume of organic solvent; thus, they are economic and eco-friendly. Accordingly, the methods are valuable for routine use in quality control (QC) laboratories for quantitation of DUV and SEL in their bulk forms and capsules. [ABSTRACT FROM AUTHOR]

Aim: A model of progressively endocrine-resistant breast cancer was investigated to identify changes that can occur in signaling pathways after endocrine manipulation. Methods: The MCF7 breast cancer model is sensitive to estrogens and anti-estrogens while variant lines previously derived from wild-type MCF7 are either relatively 17β-estradiol (E2 )-insensitive (LCC1) or fully resistant to estrogen and anti-estrogens (LCC9). Results: In LCC1 and LCC9 cell lines, loss of estrogen sensitivity was accompanied by loss of growth response to transforming growth factor alpha (TGFα), heregulin-beta and pertuzumab. LCC1 and LCC9 cells had enhanced AKT phosphorylation relative to MCF7 which was reflected in downstream activation of phospho-mechanistic target of rapamycin (mTOR), phospho-S6, and phospho-estrogen receptor alpha Ser167 [ERα(Ser167)]. Both AKT2 and AKT3 were phosphorylated in the resistant cell lines, but small interfering RNA (siRNA) knockdown suggested that all three AKT isoforms contributed to growth response. ERα(Ser118) phosphorylation was increased by E2 and TGFα in MCF7, by E2 only in LCC1, but by neither in LCC9 cells. Multiple alterations in E2-mediated cell cycle control were identified in the endocrine-resistant cell lines including increased expression of MYC, cyclin A1, cyclin D1, cyclin-dependent kinase 1 (CDK1), CDK2, and hyperphosphorylated retinoblastoma protein (ppRb), whereas p21 and p27 were reduced. Estrogen modulated expression of these regulators in MCF7 and LCC1 cells but not in LCC9 cells. Seliciclib inhibited CDK2 activation in MCF7 cells but not in resistant variants; in all lines, it reduced ppRb, increased p53 associated responses including p21, p53 up-regulated modulator of apoptosis (PUMA), and p53 apoptosis-inducing protein 1 (p53AIP1), inhibited growth, and produced G2/M block and apoptosis. Conclusions: Multiple changes occur with progression of endocrine resistance in this model with AKT activation contributing to E2 insensitivity and loss of ERα(Ser118) phosphorylation being associated with full resistance. Cell cycle regulation is modified in endocrine-resistant breast cancer cells, and seliciclib is effective in both endocrine-sensitive and resistant diseases.

Ulcerative colitis (UC) is a chronic, recurrent, non-specific inflammatory disease, and the pathogenesis of the disease remains unclear. Ferroptosis is a form of programmed cell death characterized by the accumulation of iron-dependent lipid peroxides, which are simultaneously closely related to reactive oxygen species (ROS). Although seliciclib is highly effective against immune inflammation, its mechanism on colitis is unclear. This study demonstrated that seliciclib administration partially inhibited ferroptosis, alleviating symptoms and inflammation in experimental colitis. The mouse UC model was induced by 3.0 % dextran sodium sulfate (DSS) for 7 days and treated with seliciclib (10 mg/kg) for 5 days. In the in vitro model, LPS (100 μg/mL) was used for induction and seliciclib (10 μM) was applied for 2 h. Meanwhile, appropriate histopathology, inflammatory response, oxidative stress, and ferroptosis regulators were measured. This study primarily investigated the role of seliciclib in regulating ferroptosis in UC. Bioinformatics analysis indicated that Dual oxidase 2 (DUOX2) may serve a role involved in the ferroptosis of UC. The experimental findings demonstrated that seliciclib alleviates symptoms and inflammation in DSS-induced UC mice and partially mitigates the occurrence of ferroptosis both in vivo and in vitro, possibly through the modulation of DUOX2. Ferroptosis is strongly associated with the development of colitis, and seliciclib plays an essential role in ferroptosis and inflammation in UC. The suppression of ferroptosis in the intestinal epithelium could be a therapeutic approach for UC. [ABSTRACT FROM AUTHOR]

• A multicenter, double-blind, placebo-controlled, dose-ranging phase 2 study. • Designed to evaluate the safety and effects of roscovitine. • In Pseudomonas aeruginosa infected adult CF patients. • Carrying two CF causing mutations (at least one F508del-CFTR mutation). • And harboring a FEV1>40%. The orally available kinase inhibitor R-roscovitine has undergone clinical trials against various cancers and is currently under clinical evaluation against Cushing disease and rheumatoid arthritis. Roscovitine displays biological properties suggesting potential benefits in CF: it partially corrects F508del-CFTR trafficking, stimulates the bactericidal properties of CF alveolar macrophages, and displays anti-inflammatory properties and analgesic effects. A phase 2 trial study (ROSCO-CF) was launched to evaluate the safety and effects of roscovitine in Pseudomonas aeruginosa infected adult CF patients carrying two CF causing mutations (at least one F508del-CFTR mutation) and harboring a FEV1 ≥40%. ROSCO-CF was a multicenter, double-blind, placebo-controlled, dose-ranging study (200, 400, 800 mg roscovitine, orally administered daily for 4 days/week/4 weeks). Among the 34 volunteers enrolled, randomization assigned 11/8/8/7 to receive the 0 (placebo)/ 200/400/800 mg roscovitine doses, respectively. In these subjects with polypharmacy, roscovitine was relatively safe and well-tolerated, with no significant adverse effects (AEs) other than five serious AEs (SAEs) possibly related to roscovitine. Pharmacokinetics of roscovitine were rather variable among subjects. No significant efficacy, at the levels of inflammation, infection, spirometry, sweat chloride, pain and quality of life, was detected in roscovitine-treated groups compared to the placebo-treated group. Roscovitine was relatively safe and well-tolerated in CF patients especially at the 200 and 400 mg doses. However, there were 5 subject withdrawals due to SAEs in the roscovitine group and none in the placebo group. The lack of evidence for efficacy of roscovitine (despite encouraging cellular and animal results) may be due to high pharmacokinetics variability, short duration of treatment, and/or inappropriate dosing protocol. [ABSTRACT FROM AUTHOR]

The Food and Drug Administration (FDA) has approved duvelisib (DUV) for managing follicular lymphoma, small lymphocytic lymphoma, and relapsed or refractory chronic lymphocytic leukemia. Seliciclib (SEL) is a candidate drug for these cancers, neurodegenerative disorders, renal diseases, several viral infections, and chronic inflammation disorders. This work describes the development and validation of a 96-microwell-based spectrophotometric method (MW-SPM) and a high-performance liquid chromatography with fluorescence detection method (HPLC-FD) for the quantitation of DUV and SEL in their bulk forms and capsules. The MW-SPM is based on the formation of colored charge transfer complexes (CTCs) as products for the reactions of DUV and SEL, as n-electron donors, with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ), as a π-electron acceptor. The absorption intensity of the CTCs was measured by using an absorbance plate reader at 450 nm. The stoichiometric ratios of DUV:DDQ and SEL:DDQ were 1:1 and 1:2, respectively, and accordingly the reaction mechanisms were postulated. The HPLC-FD involved the chromatographic separation of DUV and SEL on a Hypersil™ Phenyl HPLC column (250 mm length × 4.6 mm i.d., 5 μm particle diameter) with a mobile phase composed of acetonitrile:acetate buffer, pH 4.5 (35:65, v/v) at a flow rate of 2.2 mL/min. DUV and SEL were detected at 370 nm after excitation at 280 nm. SEL was used as an internal standard (IS) for quantitation of DUV, and DUV was used as an IS for quantitation of SEL. Both MW-SPM and HPLC-FD were validated according to the guidelines of the International Council for Harmonization (ICH) for validation of analytical procedures. The linear ranges for both DUV and SEL were 14.52–200 µg/well (100 µL) and 0.12–3.2 µg/mL for MW-SPM and HPLC-FD, respectively. LOD values in MW-SPM for DUV and SEL were 4.4 and 3.17 µg/well, respectively; however, those for HPLC-FD were 0.03 and 0.05 µg/mL, respectively. The accuracy and precision of both methods were confirmed as the recovery values were ≥98.5% and the values of relative standard deviations (RSD) were ≤2.41%. Both methods were satisfactorily applied to the quantitation of DUV and SEL in their capsules; the mean recovery values were ≥99.2%. Both methods have simple procedures and high analytical throughput. Moreover, they consume a small volume of organic solvent; thus, they are economic and eco-friendly. Accordingly, the methods are valuable for routine use in quality control (QC) laboratories for quantitation of DUV and SEL in their bulk forms and capsules.

Deregulation of the cyclin‐dependent kinases (CDKs) has been implicated in the pathogenesis of multiple cancer types. Consequently, CDKs have garnered intense interest as therapeutic targets for the treatment of cancer. We describe herein the molecular and cellular effects of CCT068127, a novel inhibitor of CDK2 and CDK9. Optimized from the purine template of seliciclib, CCT068127 exhibits greater potency and selectivity against purified CDK2 and CDK9 and superior antiproliferative activity against human colon cancer and melanoma cell lines. X‐ray crystallography studies reveal that hydrogen bonding with the DFG motif of CDK2 is the likely mechanism of greater enzymatic potency. Commensurate with inhibition of CDK activity, CCT068127 treatment results in decreased retinoblastoma protein (RB) phosphorylation, reduced phosphorylation of RNA polymerase II, and induction of cell cycle arrest and apoptosis. The transcriptional signature of CCT068127 shows greatest similarity to other small‐molecule CDK and also HDAC inhibitors. CCT068127 caused a dramatic loss in expression of DUSP6 phosphatase, alongside elevated ERK phosphorylation and activation of MAPK pathway target genes. MCL1 protein levels are rapidly decreased by CCT068127 treatment and this associates with synergistic antiproliferative activity after combined treatment with CCT068127 and ABT263, a BCL2 family inhibitor. These findings support the rational combination of this series of CDK2/9 inhibitors and BCL2 family inhibitors for the treatment of human cancer.

Seliciclib, a broad cyclin-dependent kinases (CDKs) inhibitor, exerts its potential role in cancer therapy. For taking advantage of overexpressive transferrin receptor (TfR) on most cancer cells, T7 peptide, a TfR targeting ligand, was selected as a targeting ligand to facilitate nanoparticles (NPs) internalization in cancer cells. In this study, poly(d,l-lactide-co-glycolide) (PLGA) was conjugated with maleimide poly(ethylene glycol) amine (Mal-PEG-NH2) to form PLGA-PEG-maleimide copolymer. The synthesized copolymer was used to prepare NPs for encapsulation of seliciclib which was further decorated by T7 peptide. The result shows that the better cellular uptake was achieved by T7 peptide-modified NPs particularly in TfR-high expressed cancer cells in order of MDA-MB-231 breast cancer cells > SKOV-3 ovarian cancer cells > U87-MG glioma cells. Both SKOV-3 and U87-MG cells are more sensitive to encapsulated seliciclib in T7-decorated NPs than to free seliciclib, and that IC50 values were lowered for encapsulated seliciclib.

Roscovitine (Seliciclib), a new protein kinase inhibitor, was administered orally to adult patients with cystic fibrosis for the first time in the ROSCO-CF trial, a dose-escalation, phase IIa, randomized, controlled trial. Extensive pharmacokinetic sampling was performed up to 12 h after the first oral dose. Roscovitine and its main metabolite M3 were quantified by liquid chromatography coupled with tandem mass spectrometry. The pharmacokinetics analyses were performed by non-linear mixed effects modelling. Monte Carlo simulations were performed to assess the impact of dose on the pharmacokinetics of oral roscovitine. Twenty-three patients received oral doses ranging from 200 to 800 mg of roscovitine and 138 data points were available for both roscovitine and M3 concentrations. The pharmacokinetics was best described by a two-compartment parent-metabolite model, with a complex saturable absorption process modelled as the sum of Gaussian inverse density functions. The Monte Carlo simulations showed a dose-dependent and saturable first-pass effect leading to pre-systemic formation of M3. The treatment with proton-pump inhibitors reduced the rate of absorption of oral roscovitine. The pharmacokinetics of oral roscovitine in adult patients with cystic fibrosis was non-linear and showed significant inter-individual variability. A repeat-dose study will be required to assess the inter-occasional variability of its pharmacokinetics.

Deregulation of the cyclin‐dependent kinases (CDKs) has been implicated in the pathogenesis of multiple cancer types. Consequently, CDKs have garnered intense interest as therapeutic targets for the treatment of cancer. We describe herein the molecular and cellular effects of CCT068127, a novel inhibitor of CDK2 and CDK9. Optimized from the purine template of seliciclib, CCT068127 exhibits greater potency and selectivity against purified CDK2 and CDK9 and superior antiproliferative activity against human colon cancer and melanoma cell lines. X‐ray crystallography studies reveal that hydrogen bonding with the DFG motif of CDK2 is the likely mechanism of greater enzymatic potency. Commensurate with inhibition of CDK activity, CCT068127 treatment results in decreased retinoblastoma protein (RB) phosphorylation, reduced phosphorylation of RNA polymerase II, and induction of cell cycle arrest and apoptosis. The transcriptional signature of CCT068127 shows greatest similarity to other small‐molecule CDK and also HDAC inhibitors. CCT068127 caused a dramatic loss in expression of DUSP6 phosphatase, alongside elevated ERK phosphorylation and activation of MAPK pathway target genes. MCL1 protein levels are rapidly decreased by CCT068127 treatment and this associates with synergistic antiproliferative activity after combined treatment with CCT068127 and ABT263, a BCL2 family inhibitor. These findings support the rational combination of this series of CDK2/9 inhibitors and BCL2 family inhibitors for the treatment of human cancer. [ABSTRACT FROM AUTHOR]

This monograph reviews the discovery and development of the cyclin-dependent kinase inhibitor roscovitine (R-roscovitine, CYC202, Seliciclib). The authors summarise the in vitro and in vivo data that have formed the basis for clinical investigation of Seliciclib as an anti-cancer drug. Kinase selectivity, cellular effects and the pharmacological properties of the drug are discussed in addition to the clinical results of Seliciclib being reviewed. Novel results on the effect of the drug in cardiac hypertrophy are summarized and potential applications of Seliciclib in other therapeutic areas, including, inflammation, virology, glomerulonephritis and polycystic kidney disease, are discussed. Finally the authors argue that optimisation of the therapeutic effect of kinase inhibitors such as Seliciclib can be enhanced using a systems biology approach involving mathematical modelling of the molecular pathways regulating cell growth and division.

Seliciclib displayed limited brain exposure in vivo in adult rats with mature blood-brain barrier (BBB). Selicilib was shown to be a specific substrate of human ABCB1 in vitro . To demonstrate that ABCB1/Abcb1 can limit brain exposure in vivo in mice we are showing that seliciclib is a substrate of mouse Abcb1a, the murine ABCB1 ortholog expressed in the BBB as LLC-PK-Abcb1a cells displayed an efflux ratio (ER) of 15.31 ± 3.54 versus an ER of 1.44 ± 0.10 in LLC-PK1-mock cells. Additionally, in the presence of LY335979, an ABCB1/Abcb1a specific inhibitor, the observed ER for seliciclib in the LLC-PK1-mMdr1a cells decreased to 1.05 ± 0.25. To demonstrate in vivo relevance of seliciclib transport by Abcb1a mouse brain microdialysis experiments were carried out that showed that the AUC brain /AUC blood ratio of 0.143 in anesthetized mice increased about two-fold to 0.279 in the presence of PSC833 another ABCB1/Abcb1a specific inhibitor. PSC833 also increased the brain exposure (AUC brain ) of seliciclib close to 2-fold (136 vs 242) in awake mice. In sum, Abcb1a significantly decreases seliciclib permeability in vitro and is partly responsible for limited brain exposure of seliciclib in vivo in mice. [ABSTRACT FROM AUTHOR]

When mammalian tissues are infected by bacteria or fungi, inflammatory cytokines are released that cause circulating neutrophils to invade the infected tissue. The cytosolic tyrosine kinase, c-Abl, in these tissue neutrophils is activated by TNFα. c-Abl then phosphorylates STAT transcription factors, which results in production of the antiapoptotic protein Mcl-1. The normally short-lived tissue neutrophils are then unable to enter apoptosis. c-Abl also causes release of reactive oxygen species (ROS) from the mitochondria of the activated neutrophils. These ROS, and ROS generated by NADPH oxidase, are bactericidal agents of the innate immune system. In some inflammatory diseases, such as chronic obstructive pulmonary disease (COPD), the invading neutrophils become permanently activated, and the resulting ROS overproduction causes severe tissue damage. The cyclin-dependent kinase inhibitor, seliciclib, blocks transcription through inhibition of cdk9. This results in a relatively rapid decline of antiapoptotic Mcl-1 transcripts in activated neutrophils, an increase in neutrophil apoptosis, and less ROS leakage and oxidative damage. We present here a model of neutrophil kinetics that simulates the principal pathways of c-Abl signalling and use it to explore possible treatment options for inflammatory lung disease. [ABSTRACT FROM AUTHOR]

The outcome of bacterial meningitis critically depends on the rapid initiation of bactericidal antibiotic therapy and adequate management of septic shock. In community-acquired meningitis, the choice of an optimum initial empirical antibiotic regimen depends on the regional resistance patterns. Pathogens resistant to antibacterials prevail in nosocomial bacterial meningitis. Dexamethasone is recommended as adjunctive therapy for community-acquired meningitis in developed countries. In comatose patients, aggressive measures to lower intracranial pressure <20 mmHg (in particular, external ventriculostomy, osmotherapy and temporary hyperventilation) were effective in a case–control study. Although many experimental approaches were protective in animal models, none of them has been proven effective in patients. Antibiotics, which are bactericidal but do not lyse bacteria, and inhibitors of matrix metalloproteinases or complement factor C5 appear the most promising therapeutic options. At present, vaccination is the most efficient method to reduce disease burden. Palmitoylethanolamide appears promising to enhance the resistance of the brain to infections. [ABSTRACT FROM PUBLISHER]

Seliciclib (R-Roscovitine) was identified as an inhibitor of CDKs and has undergone drug development and clinical testing as an anticancer agent. In this review, the authors describe the discovery of Seliciclib and give a brief summary of the biology of the CDKs Seliciclib inhibits. An overview of the published in vitro and in vivo work supporting the development as an anti-cancer agent, from in vitro experiments to animal model studies ending with a summary of the clinical trial results and trials underway is presented. In addition some potential non-oncology applications are explored and the potential mode of action of Seliciclib in these areas is described. Finally the authors argue that optimisation of the therapeutic effects of kinase inhibitors such as Seliciclib could be enhanced using a systems biology approach involving mathematical modelling of the molecular pathways regulating cell growth and division. [ABSTRACT FROM AUTHOR]

Bacterial meningitis remains a disease with high mortality and long-term morbidity. Outcome critically depends on the rapid initiation of effective antibiotic therapy. Since a further increase of the incidence of pathogens resistant to antibacterials can be expected both in community-acquired and nosocomial bacterial meningitis, the choice of an optimum initial empirical antibiotic regimen will gain significance. In this context, the use of antibiotics which are bactericidal but do not lyse bacteria, may emerge as a therapeutic option. Conversely, the role of corticosteroids, which decrease the entry of hydrophilic antibacterials into the cerebrospinal fluid, as adjunctive therapy will probably decline as a consequence of the increasing antibiotic resistance of bacteria causing meningitis. Consequent vaccination of all children at present is the most efficient manner to reduce disease burden. [ABSTRACT FROM AUTHOR]

Low molecular weight cyclin E (LMW-E) plays an important oncogenic role in breast cancer. LMW-E, which is not found in normal tissue, can promote the formation of aggressive tumors and can lead to increased genomic instability and tumorigenesis. Additionally, breast cancer patients whose tumors express LMW-E have a very poor prognosis. Therefore, we investigated LMW-E as a potential specific target for treatment either alone or in combination therapy. We hypothesized that because LMW-E binds to CDK2 more efficiently than full length cyclin E, resulting in increased activity, CDK inhibitors could be used to target tumors with LMW-E bound to CDK2. To test the hypothesis, an inducible full length and LMW-E MCF7-Tet-On system was established. Cyclin E (full length (EL) or LMW-E) is only expressed upon induction of the transgene. The doubling times of cells were unchanged when the transgenes were induced. However, upon induction, the kinase activity associated with LMW-E was much higher than that in the EL induced cells or any of the uninduced cells. Additionally only the LMW-E induced cells underwent chromosome aberrations and increased polyploidy. By examining changes in proliferation and survival in cells with induced full length and LMW-E, CDK inhibitors alone were determined to be insufficient to specifically inhibit LMW-E expressing cells. However, in combination with Doxorubicin, the CDK inhibitor, Roscovitine (Seliciclib, CYC202), synergistically led to increased cell death in LMW-E expressing cells. Clinically, the combination of CDK inhibitors and chemotherapy such as Doxorubicin provides a viable personalized treatment strategy for those breast cancer patients whose tumors express the LMW-E. [ABSTRACT FROM AUTHOR]

Chemical inhibitors of cyclin-dependent kinase (CDK), like roscovitine, are promising drugs in the context of new cancer therapies. Roscovitine and related compounds, like seliciclib and olomoucine, are effective inducers of apoptosis in many proliferating cells in culture. These compounds are known to activate the intrinsic or mitochondrial pathway of apoptosis. In order to better characterize this intrinsic pathway, a transcriptional analysis was performed using the reverse transcriptase-multiplex ligation-dependent probe amplification procedure (RT-MLPA). In five cell lines, we detected an early and marked reduction of most transcripts, which is consistent with the disruption of transcription that results from the inhibition of CDK7 and CDK9. However, the mRNA of p53- upregulated modulator of apoptosis (PUMA) gene escaped from this transcription inhibition in neuroblastoma cells with a functional p53 protein. The increase of PUMA mRNA was not found in roscovitine-treated cell lines defective in p53, which underwent apoptosis like their p53 proficient counterparts. In addition, in SH-SY5Y cells, sublethal and lethal concentrations of roscovitine produced equivalent increases of PUMA mRNA and protein. In conclusion, the increased expression of PUMA was not associated with apoptosis induction. On the contrary, mRNA and protein depletion of MCL- 1 gene correlated the best with cell demise. Moreover, NOXA protein suffered a far minor decrease than MCL-1. Because of the selective neutralization of NOXA by MCL-1, we hypothesize that the disruption of this balance is a critical event in apoptosis induction by roscovitine and related compounds. [ABSTRACT FROM AUTHOR]

The cyclin-dependent kinase inhibitor, seliciclib (R-roscovitine, CYC202), has anti-proliferative activity through its inhibition of cyclin-dependent kinase 2. We hypothesized that treatment with seliciclib would reduce glomerular macrophage numbers and glomerular crescent formation in experimental crescentic glomerulonephritis even when treatment is started after onset of disease. Nephrotoxic nephritis (NTN) was induced in Wistar Kyoto rats. In experiment 1, seliciclib (150 mg/kg per day) was given by oral gavage from 1 h before induction of NTN and continued to day 14. In experiment 2, treatment was started on day 4 of NTN and continued to day 14 in order to examine the effect of seliciclib in established glomerulonephritis. In experiment 1, seliciclib reduced proteinuria (119.5 ± 13.9 vs 191.4 ± 18.8 mg/day, P < 0.01), serum creatinine (54.0 ± 3.0 vs 81.0 ± 2.5 µmol/L, P < 0.005) and glomerular crescent score (23.9 ± 2.1 vs 44.6 ± 2.2, P < 0.005) in comparison with controls. In experiment 2, seliciclib ameliorated established glomerulonephritis, with reduction in proteinuria (58 ± 16 vs 165 ± 13 mg/day, P < 0.005), serum creatinine (39 ± 3 vs 62 ± 5 µmol/L, P < 0.05), glomerular macrophage numbers (6.8 ± 2.5 vs 18.5 ± 1.2 ED1+ cells per glomerular cross section, P < 0.05), glomerular cell proliferation (1.2 ± 0.37 vs 4.2 ± 0.80 bromodeoxyuridine (BrdU)+ cells per glomerular section, P < 0.05) and crescent score (10.8 ± 1.6 vs 43.9 ± 1.4, P < 0.05), in comparison with the controls. Seliciclib is effective in both prevention and treatment of established crescentic glomerulonephritis in Wistar Kyoto rats, in association with a reduction in the number of glomerular macrophages. We suggest that seliciclib, or other cyclin-dependent kinase inhibitors, may represent a novel therapeutic approach for patients with proliferative glomerulonephritis. This study demonstrates the efficacy of a specific anti-proliferative therapy in a rat model of rapidly progressive crescentic glomerulonephritis. The beneficial effects are attributed to inhibition of glomerular macrophage accumulation, although anti-proliferative effects on other cell types involved in crescent formation may also contribute to amelioration of disease. [ABSTRACT FROM AUTHOR]

Abstract: Raman micro-spectroscopic analysis of cultured HCT116 colon cancer cells in the presence of roscovitine, [seliciclib, 2-(1-ethyl-2-hydroxy-ethylamino)-6-benzylamino-9-isopropylpurine], a promising drug candidate in cancer therapy, has been performed for the first time. The aim of this study was to investigate modulations in colon cancer cells induced by roscovitine. Raman spectra of the cultured HCT116 colon cancer cells treated with roscovitine at different concentrations (0, 5, 10, 25 and 50μM) were recorded in the range 400–1850cm−1. It was shown that the second derivative profile of the experimental spectrum gives valuable information about the wavenumbers and band widths of the vibrational modes of cell components, and it eliminates the appearance of false peaks arising from incorrect baseline corrections. In samples containing roscovitine, significant spectral changes were observed in the intensities of characteristic protein and DNA bands, which indicate roscovitine-induced apoptosis. Roscovitine-induced apoptosis was also assessed by flow cytometry analysis, and analysis of propidium iodide staining. We observed some modifications in amide I and III bands, which arise from alterations in the secondary structure of cell proteins caused by the presence of roscovitine. [Copyright &y& Elsevier]

Abstract: Aim: Phase I study of seliciclib (CYC202, R-roscovitine), an inhibitor of cyclin-dependent kinases 2, 7 and 9, causing cell cycle changes and apoptosis in cancer cells. Patients and methods: This phase I trial aimed at defining the toxicity profile, the maximum tolerated dose (MTD), the recommended phase II dose (RD) and the main pharmacokinetic and pharmacodynamic parameters of oral seliciclib. Three schedules were evaluated: seliciclib given twice daily for 5 consecutive days every 3weeks (schedule A), for 10 consecutive days followed by 2weeks off (schedule B) and for 3d every 2weeks (schedule C). Results: Fifty-six patients received a total of 218 cycles of seliciclib. Dose-Limiting Toxicities (DLT) consisting of nausea, vomiting, asthenia and hypokalaemia occurred at 1600mg bid for schedule A and in schedule C, DLT of hypokalaemia and asthenia occurred at 1800mg bid. The evaluation of longer treatment duration in schedule B was discontinued because of unacceptable toxicity at lower doses. Other adverse events included transient serum creatinine increases and liver dysfunctions. Pharmacokinetic data showed that exposure to seliciclib and its carboxylate metabolite increased with increasing dose. Soluble cytokeratin 18 fragments allowed monitoring of seliciclib-induced cell death in the blood of patients treated with seliciclib at doses above 800mg/d. One partial response in a patient with hepatocellular carcinoma and sustained tumour stabilisations were observed. Conclusions: The MTD and RD for seliciclib are 1250mg bid for 5d every 3weeks and 1600mg bid for 3d every 2weeks, respectively. [Copyright &y& Elsevier]

Circadian disruption accelerates malignant growth and shortens survival, both in experimental tumor models and cancer patients. In previous experiments, tumor circadian disruption was rescued with seliciclib, an inhibitor of cyclin-dependent kinases (CDKs). This effect occurred at a selective dosing time and was associated with improved antitumor activity. In the current study, seliciclib altered robust circadian mRNA expression of the clock genes Rev-erbα, Per2, and Bmal1 in mouse liver following dosing at zeitgeber time (ZT) 3 (i.e., 3 h after the onset of the 12 h light span), when mice start to rest, but not at ZT19, near the middle of the 12 h dark span, when mice are most active. However, liver exposure to seliciclib, as estimated by the liver area under the concentration×time curve (AUC), was ∼80% higher at ZT19 than at ZT3 (p = 0.049). Circadian clock disruption was associated with increased serum liver enzymes and modified glycogen distribution in hepatocytes, as revealed by biochemical determinations and optic and electronic microscopy. The extent of increase in liver enzymes was most pronounced following dosing at ZT3, as compared to ZT19 (p < 0.04). Seliciclib further up-regulated the transcriptional activity of c-Myc, a cell cycle gene that promotes cell cycle entry and G1-S transition (p < 0.001), and down-regulated that of Wee1, which gates cell cycle transition from G2 to M (p < 0.001). These effects did not depend upon drug dosing time. Overall, the results suggest the circadian time of seliciclib delivery is more critical than the amount of drug exposure in determining its effects on the circadian clock. Seliciclib-induced disruption of the liver molecular clock could account for liver toxicity through the resulting disruption of clock-controlled detoxification pathways. Modifications of cell cycle gene expression in the liver likely involve other mechanisms. Circadian clocks represent relevant targets to consider for optimization of therapeutic schedules of CDK inhibitors. [ABSTRACT FROM AUTHOR]

Abstract: Objectives. : Inhibition of cyclin-dependent-kinases (CDKs) represents an interesting approach in cancer therapy. We have explored this in cell lines of human uterine sarcoma-tumours associated with poor survival, chemo-unresponsiveness and deregulation of cell cycle components. We studied the effects of the CDK inhibitor seliciclib (CYC202, R-roscovitine) when used alone or in combination with cisplatin. Methods. : Cell lines used: SK-UT-1, SK-UT-1b and SK-LMS-1, the cytotoxicity of seliciclib and cisplatin was measured by the MTT assay. In combination with cisplatin the effects of seliciclib were examined by isobologram analysis. CDK2 levels were examined at mRNA and protein level by immunoblotting and PCR. We also looked at the effects of seliciclib on p53-dependent response of cells to seliciclib using immunoblotting. The effects of combination treatment were analysed using annexin V and PI staining by flow cytometric analysis. Results. : IC50 values for seliciclib were 10.5, 7.1 and 25.7 μM, for SK-UT-1, SK-UT-1b and SK-LMS-1 respectively, P53 in the SK-UT-1b (wild-type) and SK-LMS-1 lines (mutant) showed a wild-type response with induction seen with seliciclib treatment for 24 and 48 h. Seliciclib caused a decrease in CDK2 mRNA and protein over 72 h. A combination of cisplatin and seliciclib was synergistic in all three cell lines. Effects of combination treatment were an enhancement in apoptosis as judged by the emergence of a sub-G1 population in cell cycle analysis and a sub-G1 population with PI staining. Conclusions. : Our data demonstrate the effectiveness of seliciclib as a single agent and when used in combination with cisplatin where the effects are synergistic. [Copyright &y& Elsevier]

We have studied the role of cyclins and cyclin-dependent kinase (CDK) activity in apoptosis induced by camptothecin (CPT). In this model, 22% of the cells stain for annexin-V at 24 h and then proceed to be 93% positive by 72 h. This time window permits the analysis of cyclins in cells that are committed to apoptosis but not yet dead. We provide evidence that cyclin protein levels and then associated kinase levels increase after CPT treatment. Strikingly, cyclin B1 and cyclin E1 proteins are present at the same time in CPT treated HT29 cells. Although cyclin B1 and E1 CDK complexes are activated in CPT treated cells, only the cyclin B1 complex is required for apoptosis since reduction of cyclin B1 by RNAi or roscovitine treatment reduces the number of annexin-V-stained cells. We have detected poorly organized chromosomes and phosphorylated histone H3 epitopes at the time of maximum cyclin B1/CDK kinase activity in CPT-treated cells, which suggests that these cells enter a mitotic catastrophe. Understanding which CDKs are required for apoptosis may allow us to better adapt CDK inhibitors for use as anti-cancer compounds.Oncogene (2006) 25, 7361–7372. doi:10.1038/sj.onc.1209718; published online 19 June 2006 [ABSTRACT FROM AUTHOR]

Abstract: In this study, we have analyzed the consequences, on several neuroblastoma cell lines, of combined treatments with (R)-roscovitine (CYC202, Seliciclib), a CDK inhibitory drug, and nutlin-3, a p53 activating drug. Both compounds were found to synergize, causing significant levels of apoptosis in cultured cells when combined at sublethal concentrations. In SH-SY5Y cells, Bcl-XL protein overexpression protected from apoptosis induced by either nutlin-3 alone or the (R)-roscovitine plus nutlin-3 association but failed to prevent apoptosis triggered by (R)-roscovitine alone. Moreover, Western blot studies showed that (R)-roscovitine increased nutlin-3-mediated p53 stabilization. Therefore, we conclude the contribution of (R)-roscovitine to the synergism is basically the sensitization of SH-SY5Y cells to the action of nutlin-3 on p53. The relevance of this pharmacological synergism with respect to the treatment of neuroblastoma is discussed. [Copyright &y& Elsevier]

Abstract: Systems biology needs to show practical relevance to commercial biological challenges such as those of pharmaceutical development. The aim of this work is to design and validate some applications in anti-cancer therapeutic development. The test system was a group of novel cyclin-dependent kinase (CDK) inhibitors synthesised by Cyclacel Ltd. The measured in vitro IC50s of each compound were used as input data to a proprietary cell cycle model developed by Physiomics plc. The model was able to predict over three orders of magnitude the cytotoxicity of each compound without model adaptation to specific cancer cell types. This pattern matched the experimentally determined data. One class of compounds was predicted to cause an increase of the cell cycle length with a non-linear dose–response curve. Further work will use apoptosis and DNA replication simulations to look at overall cell effects. [Copyright &y& Elsevier]

Aim: A model of progressively endocrine-resistant breast cancer was investigated to identify changes that can occur in signaling pathways after endocrine manipulation., Methods: The MCF7 breast cancer model is sensitive to estrogens and anti-estrogens while variant lines previously derived from wild-type MCF7 are either relatively 17β-estradiol (E 2 )-insensitive (LCC1) or fully resistant to estrogen and anti-estrogens (LCC9)., Results: In LCC1 and LCC9 cell lines, loss of estrogen sensitivity was accompanied by loss of growth response to transforming growth factor alpha (TGFα), heregulin-beta and pertuzumab. LCC1 and LCC9 cells had enhanced AKT phosphorylation relative to MCF7 which was reflected in downstream activation of phospho-mechanistic target of rapamycin (mTOR), phospho-S6, and phospho-estrogen receptor alpha Ser167 [ERα(Ser167)]. Both AKT2 and AKT3 were phosphorylated in the resistant cell lines, but small interfering RNA (siRNA) knockdown suggested that all three AKT isoforms contributed to growth response. ERα(Ser118) phosphorylation was increased by E 2 and TGFα in MCF7, by E 2 only in LCC1, but by neither in LCC9 cells. Multiple alterations in E 2 -mediated cell cycle control were identified in the endocrine-resistant cell lines including increased expression of MYC, cyclin A1, cyclin D1, cyclin-dependent kinase 1 (CDK1), CDK2, and hyperphosphorylated retinoblastoma protein (ppRb), whereas p21 and p27 were reduced. Estrogen modulated expression of these regulators in MCF7 and LCC1 cells but not in LCC9 cells. Seliciclib inhibited CDK2 activation in MCF7 cells but not in resistant variants; in all lines, it reduced ppRb, increased p53 associated responses including p21, p53 up-regulated modulator of apoptosis (PUMA), and p53 apoptosis-inducing protein 1 (p53AIP1), inhibited growth, and produced G2/M block and apoptosis., Conclusions: Multiple changes occur with progression of endocrine resistance in this model with AKT activation contributing to E 2 insensitivity and loss of ERα(Ser118) phosphorylation being associated with full resistance. Cell cycle regulation is modified in endocrine-resistant breast cancer cells, and seliciclib is effective in both endocrine-sensitive and resistant diseases., Competing Interests: Conflicts of interest The authors declare that they have no conflicts of interest.

Seliciclib, a broad cyclin-dependent kinases (CDKs) inhibitor, exerts its potential role in cancer therapy. For taking advantage of overexpressive transferrin receptor (TfR) on most cancer cells, T7 peptide, a TfR targeting ligand, was selected as a targeting ligand to facilitate nanoparticles (NPs) internalization in cancer cells. In this study, poly(d,l-lactide-co-glycolide) (PLGA) was conjugated with maleimide poly(ethylene glycol) amine (Mal-PEG-NH2) to form PLGA-PEG-maleimide copolymer. The synthesized copolymer was used to prepare NPs for encapsulation of seliciclib which was further decorated by T7 peptide. The result shows that the better cellular uptake was achieved by T7 peptide-modified NPs particularly in TfR-high expressed cancer cells in order of MDA-MB-231 breast cancer cells > SKOV-3 ovarian cancer cells > U87-MG glioma cells. Both SKOV-3 and U87-MG cells are more sensitive to encapsulated seliciclib in T7-decorated NPs than to free seliciclib, and that IC50 values were lowered for encapsulated seliciclib. [ABSTRACT FROM AUTHOR]

Roscovitine (Seliciclib), a new protein kinase inhibitor, was administered orally to adult patients with cystic fibrosis for the first time in the ROSCO-CF trial, a dose-escalation, phase IIa, randomized, controlled trial. Extensive pharmacokinetic sampling was performed up to 12 h after the first oral dose. Roscovitine and its main metabolite M3 were quantified by liquid chromatography coupled with tandem mass spectrometry. The pharmacokinetics analyses were performed by non-linear mixed effects modelling. Monte Carlo simulations were performed to assess the impact of dose on the pharmacokinetics of oral roscovitine. Twenty-three patients received oral doses ranging from 200 to 800 mg of roscovitine and 138 data points were available for both roscovitine and M3 concentrations. The pharmacokinetics was best described by a two-compartment parent-metabolite model, with a complex saturable absorption process modelled as the sum of Gaussian inverse density functions. The Monte Carlo simulations showed a dose-dependent and saturable first-pass effect leading to pre-systemic formation of M3. The treatment with proton-pump inhibitors reduced the rate of absorption of oral roscovitine. The pharmacokinetics of oral roscovitine in adult patients with cystic fibrosis was non-linear and showed significant inter-individual variability. A repeat-dose study will be required to assess the inter-occasional variability of its pharmacokinetics. [ABSTRACT FROM AUTHOR]