Gregory Sivolapenko

Structural modifications of 99mTc-labelled bombesin-like peptides for optimizing pharmacokinetics in prostate tumor targeting

PURPOSE The main goal of the present study was to investigate the importance of the addition of a positively charged aa in the naturally occurring bombesin (BN) peptide for its utilization as radiodiagnostic agent, taking into consideration the biodistribution profile, the pharmacokinetic characteristics and the tumor targeting ability. METHODS Two BN-derivatives of the general structure [M-chelator]-(spacer)-BN(2-14)-NH(2), where M: (99m)Tc or (185/187)Re, chelator: Gly-Gly-Cys-, spacer: -(arginine)(3)-, M-BN-A; spacer: -(ornithine)(3)-, M-BN-O; have been prepared and evaluated as tumor imaging agents. RESULTS The peptides under study presented high radiolabelling efficiency (>98%), significant stability in human plasma (>60% intact radiolabelled peptide after 1h incubation) and comparable receptor binding affinity with the standard [(125)I-Tyr(4)]-BN. Their internalization rates in the prostate cancer PC-3 cells differed, although the amount of internalized peptide was the same. The biodistribution and the dynamic γ-camera imaging studies in normal and PC-3 tumor-bearing SCID mice have shown significant tumor uptake, combined with fast blood clearance, through the urinary pathway. CONCLUSION The addition of the charged aa spacer in the BN structure was advantageous for biodistribution, pharmacokinetics and tumor targeting ability, because it reduced the upper abdominal radioactivity levels and increased tumor/normal tissue contrast ratios.

6-mercaptopurine influences TPMT gene transcription in a TPMT gene promoter variable number of tandem repeats-dependent manner

AIM TPMT activity is characterized by a trimodal distribution, namely low, intermediate and high methylator. TPMT gene promoter contains a variable number of GC-rich tandem repeats (VNTRs), namely A, B and C, ranging from three to nine repeats in length in an A(n)B(m)C architecture. We have previously shown that the VNTR architecture in the TPMT gene promoter affects TPMT gene transcription. MATERIALS, METHODS & RESULTS: Here we demonstrate, using reporter assays, that 6-mercaptopurine (6-MP) treatment results in a VNTR architecture-dependent decrease of TPMT gene transcription, mediated by the binding of newly recruited protein complexes to the TPMT gene promoter, upon 6-MP treatment. We also show that acute lymphoblastic leukemia patients undergoing 6-MP treatment display a VNTR architecture-dependent response to 6-MP. CONCLUSION These data suggest that the TPMT gene promoter VNTR architecture can be potentially used as a pharmacogenomic marker to predict toxicity due to 6-MP treatment in acute lymphoblastic leukemia patients.

Antibody-drug conjugates: a mini-review. The synopsis of two approved medicines

Abstract Targeted drug delivery is a method of delivering bioactive compounds to a patient in a manner that increases the therapeutic index. The main goal of a targeted drug delivery system is to prolong, localize, target and have a protected drug interaction with the diseased tissue. Antibody-drug conjugates (ADC) represent an innovative therapeutic application that combines the unique properties of monoclonal antibodies with the potent cell killing activity of cytotoxic bioactive compounds. ADCs are complex molecules composed of an antibody linked, via a stable, chemical, linker with labile bonds, to a biological active cytotoxic (anticancer) payload or drug. The key components of ADC include a monoclonal antibody, a stable linker and a cytotoxic agent to target a variety of cancers. The present mini-review deals with the examination of clinical use and pharmacological properties, as well as the safety of antibody-drug conjugates that are marketed. Ado-trastuzumab emtasine and brenduximab vedotin were examined regarding their mechanism of action, pharmacology, clinical use and safety. These ADCs selectively deliver cargoes to tumor cells and provide clinical benefit by minimizing systemic toxicity.

Characterization and Biological Evaluation of Propolis from Poland

In this study, we assessed the therapeutic potential of propolis from Poland and performed chemical analysis by GC–MS, as well as determined its botanical origin. Chemical constituents typical for bud exudates of Populus nigra (section Aigeiros) were determined, however, glycerol esters of phenolic acids, as well as unusually high amounts of p-coumaric and ferulic acid and their benzyl esters, were also detected. These constituents are characteristic for buds of Populus tremula (section Leuce). We also evaluated the antiproliferative effect of propolis extracts against nine human cancer cell lines. Additionally, promising antibacterial activity of the dichloromethane extract (Minimal Inhibitory Concentration MIC values of 0.95–1.24 mg/mL), as well as a moderate antifungal activity (MIC values of 1.25–1.40 mg/mL), was noticed. Propolis from Poland appeared as a rich source of antibacterial and antiproliferative compounds and this confirmed that it is a valuable natural product with the potential to improve human health.

Advanced nanocarriers for an antitumor peptide

In this work, tigapotide (PCK3145) was incorporated into novel nanocarriers based on polymeric, lipidic, and dendrimeric components, in order to maximize the advantages of the drug delivery process and possibly its biological properties. PCK3145 was incorporated into lipidic nanocarriers composed of Egg phosphatidylcholine (EggPC) and dipalmytoylphosphatidylcholine (DPPC) (EggPC:PCK3145 and DPPC:PCK3145, 9:0.2 molar ratio), into cationic liposomes composed of EggPC:SA:PCK3145 and DPPC:SA:PCK3145 (9:1:0.2 molar ratio) into complexes with the block polyelectrolyte (quaternized poly[3,5-bis(dimethylaminomethylene)hydroxystyrene]-b-poly(ethylene oxide) (QNPHOSEO) and finally into dendrimeric structures (i.e., PAMAM G4). Light scattering techniques are used in order to examine the size, the size distribution and the Z-potential of the nanocarriers in aqueous and biological media. Fluorescence spectroscopy was utilized in an attempt to extract information on the internal nanostructure and microenvironment of polyelectrolyte/PCK3145 aggregates. Therefore, these studies could be a rational roadmap for producing various effective nanocarriers in order to ameliorate the pharmacokinetic behavior and safety issues of antitumor and anticancer biomolecules.

Development and validation of analytical methodology for the quantification of aldehydes in e-cigarette aerosols using UHPLC-UV

Aldehydes are produced in e-cigarette aerosols, as a result of the thermal decomposition of vegetable glycerin, propylene glycerol and flavorings in the atomizer. These aldehydes were collected with derivatization into 2,4-dinitrophenylhydrazine using impinger trapping. A new methodology for simultaneous quantitative analysis of aldehydes in base liquids was developed and validated. Chromatographic separation was carried out on a Jupiter Proteo 90A column, with the mobile phase consisting of 0.1% formic acid in water and acetonitrile, at a flow rate of 0.4 mL/min. Linearity was demonstrated over the range of concentrations 0.025-10 μg/mL, with correlation of determination ≥ 0.999. Intra- and inter-day % relative standard deviation and relative error were ≤ 10%. The lower limit of detection and quantification were 0.008 and 0.025 μg/mL, respectively. This method was further used for the quantification of aldehydes emitted by different base liquids. It has been shown that vegetable glycerin produces the highest percentage of aldehydes after thermal decomposition compared to propylene glycerol. We propose that it can be used for future e-liquids emissions studies.

Liposomal forms of anticancer agents beyond anthracyclines: present and future perspectives

Abstract Liposomes are widely used as delivery systems of cytotoxic drugs. The encapsulation into liposomes improves pharmacological properties and as a result therapeutic index and outcomes. To date, liposomal vincristine and cytarabine are approved and marketed for intravenous and intrathecal administration, respectively. The main goal of this review is to examine the clinical use and pharmacological properties, as well as the safety of liposomal forms of less widely used liposomal forms of anticancer agents compared to their conventional forms and to present data regarding clinical development of other liposomal agents. Liposomal forms of cytarabine and vincristine are less widely used and unknown compared to liposomal anthracyclines, because they are approved only for specific indications and only in the United States.

Development and validation of analytical methodologies for the quantification of PCK3145 and PEG-PCK3145 in mice

PCK3145 is an anti-metastatic synthetic peptide against prostate cancer. The objective of the study is to develop and validate novel and sensitive methods for the determination of PCK3145 and Pegylated PCK3145 in mouse plasma. An LC-MS/MS method was developed and validated for the determination of PCK3145 giving high sensitivity and linearity in the range of 0.125-4.0 μg/mL. PCK3145 characterised by short half-life, therefore, it was conjugated with the poly ethylene glycol (PEG). However, LC-MS/MS has been more difficult to apply for the quantitative analysis of PEGylated peptides due to the large size. A UHPLC-UV method was developed and validated for the determination of PEG-PCK3145, with linearity of 0.05-2.0 mg/mL. In order to further improve the sensitivity for the detection of PEG-PCK3145, an indirect ELISA method was used. It was found that this method was capable of detecting PCK3145 through the quantification of PEG with excellent sensitivity found at 0.132 ng/mL. The in vitro proteolytic stability of PCK3145 and PEG-PCK3145 in mouse plasma and whole blood was studied by LC-MS/MS and UHPLC, respectively. The LC-MS/MS and ELISA methods can be applied for monitoring levels of PCK3145 in mouse plasma for in vivo pharmacokinetic and bioavailability animal studies.

Development of a novel conjugatable sunitinib analogue validated through in vitro and in vivo preclinical settings

Sunitinib is an oral FDA/EMEA approved multi-targeted tyrosine kinase inhibitor. It possesses anti-angiogenic and antitumor activity against a variety of advanced solid tumors. However, its chemical core does not allow a potential linkage to tumor-homing elements that could eventually enhance its potency. Therefore, a novel linkable sunitinib derivative, designated SB1, was rationally designed and synthesized. The pharmaceutical profile of SB1 was explored both in vitro and in vivo. Mass spectrometry and NMR spectroscopy were utilized for characterization, while MTT assays and LC-MS/MS validated protocols were used to explore its antiproliferative effect and stability, respectively. Cytotoxicity evaluation in three glioma cells showed that SB1 preserved the antiproliferative effect of sunitinib. SB1 was stable in vitro after 24 h incubation in mouse plasma, while both agents exhibited bioequivalent pharmacokinetic characteristics after i.v. administration in Balb/c mice. To evaluate the levels of SB1 in mouse plasma, a novel analytical method was developed and validated in accordance to the US FDA and the EU EMA guidelines. We formulated a novel linkable sunitinib analog exhibiting similar antiproliferative and apoptotic properties with native sunitinib in glioma cell lines. Both SB1 and native sunitinib showed identical in vitro stability in mouse plasma and pharmacokinetics after i.v. administration in Balb/c mice.

Co-administration of succinylated gelatine with a 99mTc-bombesin analogue, effects on pharmacokinetics and tumor uptake

The bombesin analogue, [(99m)Tc-GGC]-(Ornithine)3-BN(2-14), (99m)Tc-BN-O, targeting gastrin releasing peptide receptors (GRPrs) on the surface of tumors, was pre-clinically investigated as potential imaging agent for single photon emission computed tomography (SPECT). In addition, the improvement of its pharmacokinetic profile (PK) was investigated through the co-administration of a succinylated gelatin plasma expander (Gelofusine), aiming to reduce its kidney accumulation and enhance its tumor-to-normal tissue contrast ratios. Biodistribution data were collected from normal mice and rats, and PC-3 tumor bearing mice, in reference to its PK, metabolism and tumor uptake. Imaging data were also collected from PC-3 tumor bearing mice. Biodistribution and imaging experiments showed that (99m)Tc-BN-O was able to efficiently localize the tumor (5.23 and 7.00% ID/g at 30 and 60min post injection, respectively), while at the same time it was rapidly cleared from the circulation through the kidneys. HPLC analysis of kidney samples, collected at 60min p.i. from normal mice and rats, showed that the majority of radioactivity detected was due to intact peptide i.e. 56% for mice and 73% for rats. Co-administration of (99m)Tc-BN-O with Gelo resulted in the reduction of kidney uptake in both animal models. The integrated area under the curve (AUC30-60 min) from the concentration-time plots of kidneys was decreased in both mice and rats by 25 and 50%, respectively. In PC-3 tumor bearing mice, an increase of tumor uptake (AUCtumor increased by 69%) was also observed with Gelo. An improvement in tumor-to-blood and tumor-to-normal tissue ratios was noted in all cases with the exception of the pancreas, which normally expresses GRPr. The results of this preclinical study may also be extended to other similar peptides, which are utilized in prostate cancer imaging and present similar PK profile.