Identification of two arylimides as cholinesterase inhibitors and testing of propranolol addition on impaired rat memory.

Alzheimer's disease (AD) is clearly linked to the decline of acetylcholine (ACh) effects in the brain. These effects are regulated by the hydrolytic action of acetylcholinesterase (AChE). Therefore, a central palliative treatment of AD is the administration of AChE inhibitors although additional mechanisms are currently described and tested for generating advantageous therapeutic strategies. In this work, we tested new arylamides and arylimides as potential inhibitors of AChE using in silico tools. Then, these compounds were tested in vitro, and two selected compounds, C7 and C8, as well as propranolol showed inhibition of AChE. In addition, they demonstrated an advantageous acute toxicity profile compared to that of galantamine as a reference AChE inhibitor. in vivo evaluation of memory performance enhancement was performed in an animal model of cognitive disturbance with each of these compounds and propranolol individually as well as each compound combined with propranolol. Memory improvement was observed in each case, but without a significant additive effect with the combinations.

Regeneration of the zebrafish retinal pigment epithelium after widespread genetic ablation.

The retinal pigment epithelium (RPE) is a specialized monolayer of pigmented cells within the eye that is critical for maintaining visual system function. Diseases affecting the RPE have dire consequences for vision, and the most prevalent of these is atrophic (dry) age-related macular degeneration (AMD), which is thought to result from RPE dysfunction and degeneration. An intriguing possibility for treating RPE degenerative diseases like atrophic AMD is the stimulation of endogenous RPE regeneration; however, very little is known about the mechanisms driving successful RPE regeneration in vivo. Here, we developed a zebrafish transgenic model (rpe65a:nfsB-eGFP) that enabled ablation of large swathes of mature RPE. RPE ablation resulted in rapid RPE degeneration, as well as degeneration of Bruch's membrane and underlying photoreceptors. Using this model, we demonstrate for the first time that zebrafish are capable of regenerating a functional RPE monolayer after RPE ablation. Regenerated RPE cells first appear at the periphery of the RPE, and regeneration proceeds in a peripheral-to-central fashion. RPE ablation elicits a robust proliferative response in the remaining RPE. Subsequently, proliferative cells move into the injury site and differentiate into RPE. BrdU incorporation assays demonstrate that the regenerated RPE is likely derived from remaining peripheral RPE cells. Pharmacological disruption using IWR-1, a Wnt signaling antagonist, significantly reduces cell proliferation in the RPE and impairs overall RPE recovery. These data demonstrate that the zebrafish RPE possesses a robust capacity for regeneration and highlight a potential mechanism through which endogenous RPE regenerate in vivo.

Efflux inhibition by IWR-1-endo confers sensitivity to doxorubicin effects in osteosarcoma cells.

Osteosarcoma is the most common bone tumor that affects children and young adults. Despite advances in the use of combination chemotherapy regimens, response to neoadjuvant chemotherapy in osteosarcoma remains a key determinant of patient outcome. Recently, highly potent small molecule inhibitors of canonical Wnt signaling through the poly(ADP-ribose) polymerase (PARP)-family enzymes, tankyrases 1 & 2 (Tnks1/2), have been considered as possible chemotherapy sensitizing agents. The goal of this study was to determine the ability of the highly specific Tnks1/2 inhibitor IWR-1-endo to sensitize chemotherapy-resistant osteosarcoma to doxorubicin. We found that IWR-1-endo significantly inhibited cellular efflux, as measured by cellular retention of Calcein AM and doxorubicin. In a model of doxorubicin resistant osteosarcoma, pre-treatment with IWR-1-endo strongly sensitized to doxorubicin. This sensitization reduced the doxorubicin IC50 in doxorubicin-resistant cells, but not in chemotherapy naïve cells and caused doxorubicin-treated cells to accumulate at the G2/M checkpoint. Further, we found that sensitization with IWR-1-endo produced increased γH2AX foci formation, indicating increased DNA damage by doxorubicin. Taken together, our findings show that IWR-1-endo increases cellular responses to doxorubicin, by blocking efflux transport in a drug-resistant model of osteosarcoma.

3B, a novel of photosensitizer, exhibited anti-tumor effects via mitochondrial apoptosis pathway in MCF-7 human breast carcinoma cells.

Photodynamic therapy (PDT) has been recognized as an innovated therapeutic modality for the treatment of various cancers. In this study, we evaluated the anticancer effect of a new photosensitizer 3B in breast cancer, which was considered one of the most common cancers in women worldwide. Here, we determined the effect of 3B not only on the cell growth, apoptosis, and Bcl-2 signal pathway in vitro but also on the anti-cancer effect in nude mice in vivo. Our results showed that 3B was primarily accumulated in mitochondria, increased the level of ROS, induced apoptotic cells death via Bcl-2 family, and its activity could be blocked by the caspase inhibitor (Z-VAD-FMK). In vivo study, 3B made a significant opening inhibition of tumor growth and showed drug toxicity hardly. TUNEL assay indicated that PDT group showed more positive cells (green) than other groups. These data supported that 3B might develop as potential therapeutic drug for the treatment of breast cancer.

Pharmacokinetics and tolerability of the new second-generation nonnucleoside reverse- transcriptase inhibitor KM-023 in healthy subjects.

BACKGROUND: KM-023 is a new second-generation nonnucleoside reverse-transcriptase inhibitor that is under development for the treatment of human immunodeficiency virus (HIV) type 1 infection. OBJECTIVE: This study determined KM-023 tolerability and pharmacokinetic characteristics in healthy subjects. MATERIALS AND METHODS: A randomized, double-blinded, placebo-controlled, dose-escalation study was conducted in 80 healthy South Korean male volunteers. The subjects were allocated to single- or multiple-dose (once daily for 7 days) groups that received 75, 150, 300, or 600 mg drug or placebo in a 4:1 ratio. Safety and pharmacokinetic assessments were performed during the study. Plasma and urine concentrations were quantified using liquid chromatography-tandem mass spectrometry. RESULTS: The average maximum concentration (Cmax) and area under the concentration-time curve from time 0 to infinity (AUC∞) values of KM-023 for the 75-600 mg doses in the single-dose study ranged from 440.2 ng/mL to 1,245.4 ng/mL and 11,142.4 ng · h/mL to 33,705.6 ng · h/mL, respectively. Values of the mean Cmax at a steady state and AUC within the dosing interval ranged from 385.1 ng/mL to 1,096.7 ng/mL and 3,698.9 ng · h/mL to 10,232.6 ng · h/mL, respectively, following 75-600 mg doses in the multiple-dose study. Dose proportionality was not observed for KM-023. KM-023 showed a 0.6-fold accumulation after multiple doses in the 600 mg dose group. The mean half-life values ranged between 20.7 and 31.2 hours. KM-023 was generally well tolerated without serious adverse events. CONCLUSION: KM-023 demonstrated dose- and time-dependent nonlinear pharmacokinetic characteristics after single or multiple doses over a dose range (75-600 mg) in healthy subjects. KM-023 showed favorable tolerability in this study. This Phase I clinical trial information can be used to design further clinical studies appropriately to evaluate KM-023 in patients with HIV-1 infection.

Glyconanosomes: disk-shaped nanomaterials for the water solubilization and delivery of hydrophobic molecules.

Herein, we describe the first report on a new class of disk-shaped and quite monodisperse water-soluble nanomaterials that we named glyconanosomes (GNS). GNSs were obtained by sliding out the cylindrical structures formed upon self-organization and photopolymerization of glycolipid 1 on single-walled carbon nanotube (SWCNT) sidewalls. GNSs present a sheltered hydrophobic inner cavity formed by the carbonated tails, surrounded by PEG and lactose moieties. The amphiphilic character of GNSs allows the water solubility of insoluble hydrophobic cargos such as a perylene-bisimide derivative, [60]fullerene, or the anti-carcinogenic drug camptothecin (CPT). GNS/C60 inclusion complexes are able to establish specific interactions between peanut agglutinin (PNA) lectin and the lactose moiety surrounding the complexes, while CPT solubilized by GNS shows higher cytotoxicity toward MCF7-type breast cancer cells than CPT alone. Thus, GNS represents an attractive extension of nanoparticle-based drug delivery systems.

Design and synthesis of novel DFG-out RAF/vascular endothelial growth factor receptor 2 (VEGFR2) inhibitors: 2. Synthesis and characterization of a novel imide-type prodrug for improving oral absorption.

As an alternative to the previously reported solid dispersion formulation for enhancing the oral absorption of thiazolo[5,4-b]pyridine 1, we investigated novel N-acyl imide prodrugs of 1 as RAF/vascular endothelial growth factor receptor 2 (VEGFR2) inhibitors. Introducing N-acyl promoieties at the benzanilide position gave chemically stable imides. N-tert-Butoxycarbonyl (Boc) introduced imide 6 was a promising prodrug, which was converted to the active compound 1 after its oral administration in mice. Cocrystals of 6 with AcOH (6b) possessed good physicochemical properties with moderate thermodynamic solubility (19µg/mL). This crystalline prodrug 6b was rapidly and enzymatically converted into 1 after its oral absorption in mice, rats, dogs, and monkeys. Prodrug 6b showed in vivo antitumor regressive efficacy (T/C=-6.4%) in an A375 melanoma xenograft model in rats. Hence, we selected 6b as a promising candidate and are performing further studies. Herein, we report the design, synthesis, and characterization of novel imide-type prodrugs.

Phase I dose-escalation study of the novel antiandrogen BMS-641988 in patients with castration-resistant prostate cancer.

PURPOSE: BMS-641988 is an androgen receptor antagonist with increased potency relative to bicalutamide in both in vitro and in vivo prostate cancer models. A first-in-man phase I study was conducted to define the safety and tolerability of oral BMS-641988 in patients with castration-resistant prostate cancer (CRPC). EXPERIMENTAL DESIGN: Doses were escalated from 5 to 150 mg based on discrete pharmacokinetic parameters in cohorts of three to six subjects. After establishing safety with 20 mg of BMS-641988 in the United States, a companion study was opened in Japan to assess differences in drug metabolism between populations. RESULTS: Sixty-one men with CRPC were treated with daily BMS-641988. The pharmacokinetics (PK) of BMS-641988 and its active metabolites were proportional to dose. One patient experienced an epileptic seizure at a dose of 60 mg administered twice. Despite achieving target drug exposures, antitumor activity was limited to one partial response. Seventeen of 23 evaluable patients (74%) exhibited stable disease on imaging (median 15 weeks; range 8-32), and 10 of 61 patients (16%) achieved a ≥ 30% decline in levels of prostate-specific antigen (PSA). Partial agonism was seen within the context of this study upon removal of the drug as evidenced by a decrease in PSA. CONCLUSIONS: Although the clinical outcomes of predominantly stable disease and partial agonism were similar to what was observed in the preclinical evaluation of the compound, the limited antitumor activity of BMS-641988 at therapeutic dose levels coupled with an episode of seizure activity led to study closure.

Statistical method for evaluation of dissolution stability in the formulation development of solid dosage forms: tablets of amonafide.

A statistical method for the evaluation of the dissolution stability results and for selecting the most stable formulation within a solid dosage form development is discussed. Three types of tablets of an antineoplastic drug, amonafide, stored at a relative humidities (RH), 45% and 75%, were used. The drug release from tablets was tested before and after storage. The experimental data were statistically fitted to empirical model equations. Furthermore, the best mathematical fit was the statistical comparison of the residuals. From the selected model equation, time-dependent dissolution (Q45 and DE45) and dissolved quantity-dependent parameters (t70, t100 and MDT) were calculated. A useful parameter to present and evaluate the results obtained in comparative stability studies was defined: the Modification Factor (MF). It allowed the selection of the most stable formulation in the easiest and fastest way: the most stable formulation should present the smallest modification of the studied characteristics, in other words, the smallest MF value. In this way, tablets II (manufactured by wet granulation and with Emcompress as main excipient) showed the greater dissolution stability of the three types of tablets studied. Amonafide tablets must be packaged in impermeable containers, since the environmental relative humidity strongly modifies their dissolution characteristics.

Phase II trial of amonafide in central nervous system tumors: a Southwest Oncology Group study.

Amonafide 300 mg/M2 was administered intravenously on a daily x 5 schedule to 27 eligible patients with recurrent or progressive central nervous system tumors. There were no objective responses. The most common toxicities were gastrointestinal, hematologic and neurologic. Further study of amonafide in patients with central nervous system malignancies is not indicated.

[The use of liposomal form of phenylimide of cis-aconitic acid in herpes therapy]. / Primenenie fenilimida tsisakonitovoi kisloty v liposomal'noi forme dlia terapii gerpesa.

The results of study of the antiviral activity and pharmacokinetics of phenylimide of cis-aconitic acid (PCAA) is presented. The 20% increase of the antiviral activity of PCAA incorporated into liposomes in comparison with the antiviral activity of the pure substance was shown. Liposomes with PCAA were tropic to lymphocytes and macrophages with maximum fluorescence being observed in the spleen, while empty liposomes were accumulated mainly in the liver. After the treatment with liposomal PCAA the symptoms of herpetic meningoencephalitis became less severe with 100% survival of the experimental animals. In the control group of rabbits 50% of the animals died, and in the surviving animals blindness or paralysis developed.

An ECOG phase II study of amonafide in unresectable or recurrent carcinoma of the head and neck (PB390). Eastern Cooperative Oncology Group.

The purpose of this study was to determine the efficacy and toxicity of amonafide in unresectable or recurrent head and neck cancer and to determine if the degree of toxicity with amonafide correlated with the acetylator phenotype of the patient. Thirty patients were registered on the study and received amonafide, 300 mg/m2, over two hours each day for five consecutive days every 21 days. There was one partial response (3%) which lasted four months. The dose-limiting toxicity was myelosuppression. Acetylator phenotype was determined prior to treatment using HPLC to quantitate caffeine metabolites in urine samples after administration of caffeine. This pharmacokinetic evaluation was performed in 21 patients and revealed that (17/21) 81% of the patients were slow acetylators and 19% of the patients were rapid acetylators. No association was found between acetylator phenotype and toxicity in our patient population. Based on this study, it appears that amonafide given at 300 mg/m2 for 5 consecutive days every 21 days is not active in squamous cell carcinoma of the head and neck, and that acetylator status does not correlate with toxicity.

Individualized dosing of amonafide based on a pharmacodynamic model incorporating acetylator phenotype and gender.

Amonafide is extensively metabolized, including conversion by N-acetylation to an active metabolite. Our previous studies have shown that fast acetylators of amonafide have increased toxicity, and we have recommended doses of 250 and 375 mg m-2 day-1 for 5 days, for fast and slow acetylators, respectively. Despite phenotype-specific dosing, significant variability in leukopenia persisted. The goal of this study was to construct and validate a pharmacodynamic model-based dosing strategy for amonafide, to try to further decrease inter-patient variability in leukopenia. The model was based on a training data set of 41 patients previously treated with amonafide. The first cycle nadir WBC was modelled as a function of dose, acetylator phenotype and baseline patient factors. This model was validated prospectively on patients similar to those in our previous studies. Based on the training data set, the optimal model was defined by three factors: acetylator phenotype, gender, and pretreatment WBC. Using this model and a target WBC nadir of 1700 microliters-1, six dosing strata were prospectively evaluated. A total of 24 fast acetylators received either 238 or 276 mg m-2 day-1 and 20 slow acetylators received between 345 and 485 mg m-2 day-1. The mean (+/- SE) error (deviation from target nadir) was 430 (+/- 240) cells microliters-1. Submaximal treatment (yielding grade 0-1 leukopenia) was limited to 20% of patients, while 55% experienced grade 2-3 toxicity. A complex dosing strategy for amonafide is feasible, employing prospective acetylator phenotyping, model-guided dosing, and adaptive control.

A phase II trial of amonafide, caracemide, and homoharringtonine in the treatment of patients with advanced renal cell cancer.

Forty-eight previously untreated, ambulatory patients with advanced or unresectable renal carcinoma were treated with either amonafide (17 patients), caracemide (17 patients), or homoharringtonine (14 patients). No objective responses were observed in any of the treatment cohorts. Amonafide and caracemide were well tolerated with no unexpected toxicities. One patient each died of pulmonary thromboembolism and sepsis with severe metabolic acidosis on the homoharringtonine arm. An additional 4 patients experienced grade 4 complications including myelosuppression, neurologic dysfunction, and respiratory failure. These severe and unexpected complications caused early termination of accrual to the homoharringtonine arm of the study. These agents have no activity in the treatment of advanced renal cell carcinoma.

Phase II study of mitonafide in non-small cell lung cancer (NSCLC).

BACKGROUND: The new intercalative agent Mitonafide was shown in early clinical trials to be toxic to the central nervous system when administered as a short intravenous infusion, but not when given as a 120-hour continuous infusion. Thus, clinical development in different tumor types was pursued using only this administration schedule. PATIENTS AND METHODS: Forty-nine patients with previously untreated non-small cell lung cancer (NSCLC) and at least one measurable site received Mitonafide as a 120-hour continuous (5 days) infusion every 3 weeks. The starting dose was 170 mg/m2/day x 5 in the first 26 patients and 200 mg/m2/day x 5 in the remainder. Patients were evaluated for toxicity after each course and for response every two courses and remained on treatment until excessive toxicity or disease progression were observed. A special test, the "Mini-mental state", was used to assess patients' cognitive functions. RESULTS: Of the 49 patients entered, 42 were evaluable for response and toxicity. Toxicity consisted mainly of myelosuppression and no neurologic side effects were observed. Only one patient presented a partial response. CONCLUSIONS: Although definitively safe with this schedule of administration, Mitonafide is not active in NSCLC.

Amonafide: An active agent in the treatment of previously untreated advanced breast cancer--a cancer and leukemia group B study (CALGB 8642).

Amonafide is a new imide derivative of naphthalic acid. The drug had demonstrated significant activity in preclinical studies and some activity in Phase I trials. The drug is extensively metabolized and detected in plasma and urine. Its toxicity has previously been correlated to the formation of an active metabolite, N-acetyl-amonafide. Amonafide was chosen for inclusion in the Cancer and Leukemia Group B (CALGB) master metastatic breast cancer protocol. CALGB 8642 randomizes previously untreated metastatic breast cancer patients either to one of several Phase II agents given for up to four cycles and then followed by standard cyclophosphamide-doxorubicin-5-fluorouracil, or to immediate treatment with standard cyclophosphamide-doxorubicin-5-fluorouracil. The end point of CALGB 8642 is to assess the difference in survival, toxicity, and overall response when limited exposure to Phase II agents precedes standard chemotherapy. This report deals only with amonafide as a Phase II agent. Comparisons with the cyclophosphamide-doxorubicin-5-fluorouracil arm will not be addressed. Patients had to have histologically documented measurable breast cancer and a performance status of 0-1. Patients could not have had prior chemotherapy for metastatic disease. Prior adjuvant chemotherapy was permitted. Patients could not have visceral crisis. Amonafide was given at 300 mg/m2/day i.v. for 5 days, and repeated at 21-day intervals for a maximum of four cycles. Escalation and reduction in dose was mandated dependent on hematotoxicity or lack thereof. Toxicity was primarily hematological and bimodal: 32% had grade 3 or 4 leukopenia and 24% had grade 3 or 4 thrombocytopenia; 22% had no leukopenia and 44% had no thrombocytopenia. The response rate was 18%, including one complete response. When response was analyzed by hematological toxicity, there was a 35.7% response if patients had leukopenia grade 3/4 (versus 8.3%, P = 0.08). There was a 50% response if patients had thrombocytopenia grade 3/4 (versus 7.1%, P = <0.01). We conclude that amonafide is somewhat active in previously untreated breast cancer patients. There may be a steep dose-response curve, based on the significant correlation between myelosuppression and response. Rates of responses in patients adequately dosed (i.e., with significant hematotoxicity) with amonafide ranged from 35 to 50%. Further studies will incorporate individualized dosing based on pretreatment acetylator phenotyping.

Phase II trial of gallium nitrate, amonafide and teniposide in metastatic non-small cell lung cancer. An Eastern Cooperative Oncology Group study (E2588).

Fifty-five patients with metastatic non-small cell lung cancer (NSCLC) were entered into this phase II randomized study for evaluating three new agents: gallium nitrate, amonafide and teniposide. The patients had to have ECOG performance status 0 or 1, no prior chemotherapy, and adequate hematological, hepatic and renal functions. Forty-seven patients were eligible and evaluable. Fourteen were randomized to receive gallium nitrate, 18 to amonafide and 15 to teniposide. Seventy-four percent of eligible patients were male. The majority of patients (89%) had an ECOG performance status 1. ECOG grade 4 toxicity occurred twice in patients on gallium nitrate, seven times on amonafide and 18 times on teniposide. The cause of death was attributed to amonafide in one patients (from sepsis) and to teniposide in two patients (due to infection and leukopenia). There was no objective response in all the patients entered. The overall survival times ranged from 2 weeks to 156 weeks with a medium of 23 weeks. There were no survival differences among the three treatment arms. We conclude that gallium nitrate, amonafide and teniposide are inactive in metastatic NSCLC and do not warrant any further testing in this disease.

Phase II trial of amonafide for the treatment of advanced, hormonally refractory carcinoma of the prostate. A Southwest Oncology Group study.

Amonafide (benzisoquinolinedione; NSC 308847) was subjected to a Phase II trial for the treatment of advanced hormone-refractory carcinoma of the prostate. In addition to adequate baseline organ functions, patients were required to have a favorable performance status, bidimensionally measurable disease and no prior chemotherapy. Amonafide was given at a dose of 225 mg/m2 intravenously daily for 5 days. Treatment cycles were repeated every 21 days. Dose escalation and reduction schema were used based upon toxicities from preceding cycles. Of 47 patients enrolled, 43 were evaluable. The most common toxicities were hematologic to include leukopenia (72%), granulocytopenia (32.6%), and thrombocytopenia (44.2%). There were no complete responses and only 5 partial responses for an overall response rate of 12% (95% confidence interval: 4-25%). The results indicate that Amonafide, in the dose and schedule tested, lacks sufficient activity against hormone-refractory prostate cancer to warrant further trials.

Pharmaceutical development of tablets of a new antineoplastic drug: mitonafide.

The pharmaceutical development of tablets of a 1,8 naphtalimide with antineoplastic activity is carried out in two steps. The preformulation step includes the study of those characteristics of the drug with special importance for the successful development of the formulation step. In this way, the low moisture, the low porosity and the good flowability of the drug as raw material allow direct compression to be taken into account, together with wet granulation as methods of tablet manufacture. As a result of the formulation studies, one formulation of tablets obtained by direct compression and another one obtained by wet granulation are selected. Both of them meet all the requirements imposed to a solid pharmaceutical form for oral administration.