Strategic pre-clinical development of Riminophenazines as resistance circumventing anticancer agents

Cancer is responsible for upward of 13% of human deaths. Contemporary chemotherapy of disseminated cancer is often thwarted by dose limiting systemic toxicity and by multi-drug resistance (MDR). Riminophenazines are a novel class of potential anticancer agents that possess a potent multi-mechanistic antineoplastic action. Apart from their broad action against intrinsic, non-classical resistance, Riminophenazines inhibit the action of Pgp and hypothetically all ABC transporters demonstrating their great utility against classical MDR. Considering that combination chemotherapy is the norm, the vision directing R&D efforts was that Riminophenazines could be used with benefit within many standard chemotherapeutic regimes. The strategic intent of this project was to attain improved therapeutic benefit for patients through gains in both pharmaco dynamic and pharmacokinetic specificity for cancer cells over what is currently available. Tactically, this was driven through the use of synergistic Fixed-Ratio Drug Combinations (FRDC) encapsulated within tumour-targeting Nanoparticulate Drug Delivery Systems (NDDS). Long-term aims of this R&D project were to: 1) Screen FRDC of clofazimine (B663) and the lead derivative (B4125) with etoposide, paclitaxel and vinblastine for synergistic drug interactions in vitro. 2) Design, assemble and characterize a novel nanoparticulate, synergistic, anticancer co-formulation. 3) Evaluate the in vivo safety and efficacy of the developed product/s in accordance with international regulatory guidelines. Using the median effect and combination index equations, impressive in vitro synergistic drug interactions (CI<1) were shown for various FRDC of the three standard chemotherapeutics tested (etoposide, paclitaxel and vinblastine) in combination with either B663 or B4125 against MDR neoplastic cell cultures. Considering in vitro results and with the view to advance quickly to clinical studies, the already approved clofazimine (B663) was elected as the combination partner for paclitaxel (PTX). Considering the potency and wide action of PTX, a novel coformulation (designed to circumvent drug resistance) has the potential to greatly impact upon virtually all cancer types, particularly if selectively delivered through innovative delivery systems and loco-regional administration. A passively tumour targeting, micellular NDDS system called Riminocelles™ that encapsulates a synergistic FRDC of B663 and PTX has been designed, assembled using thin film hydration methods and characterized in terms of drug loading, particle size, zeta potential, CMC and drug retention under sink conditions. An acute toxicity and a GLP repeat dose toxicity study confirmed Riminocelles to be well tolerated and safe at clinically relevant dosages whilst Taxol® (QDx7) produced statistically significant (P<0.05) weight loss within 14 days. The same study demonstrated statistically significant (P<0.05) tumour growth delays superior to that of Taxol at an equivalent PTX dosage of 10 mg/kg. Importantly, all components (amphiphiles and drugs) used in assembly of Riminocelles are already individually approved for medicinal use - this promotes accelerated development towards advanced clinical trials and successful registration. Although these results are very promising (outperforming Taxol), this system was however found in a pharmacokinetic study to suffer from in vivo thermodynamic instability due to the high concentration (abundance) of albumin present in plasma. For this reason, in vivo longevity within circulation, permitting passive tumour accumulation was not fully realized. A second NDDS called the RiminoPLUS™ imaging system was additionally developed. This lipopolymeric nanoemulsion system has successfully entrapped Lipiodol® Ultra fluid (an oil based contrast agent) within the hydrophobic core of a monodisperse particle population with a size of roughly 100 nm and a stability of one week. This formulation is therefore thought capable of CT imaging of tumour tissue and drug targeting after either intravenous or loco-regional injection. In vivo proof of the imaging concept is warranted. The results of this study serve to highlight the great potential of in vitro optimized synergistic FRDC against drug resistant cancers. Lipopolymeric micelles are an effective way to formulate multiple hydrophobic drugs for intravenous administration and present a means by which cancer can be readily targeted; provided that the delivery system possess the prerequisite in vivo stability and surface attributes. Further experiments exploring synergistic drug and biological combinations as well as “intelligent” NDDS actively guided through specific molecular recognition are called for.Thesis (PhD)--University of Pretoria, 2012.Pharmacologyunrestricte

Anticancer efficacy and toxicokinetics of a novel paclitaxel-clofazimine nanoparticulate co-formulation

Contemporary chemotherapy is limited by disseminated, resistant cancer. Targeting nanoparticulate drug delivery systems that encapsulate synergistic drug combinations are a rational means to increase the therapeutic index of chemotherapeutics. A lipopolymeric micelle co-encapsulating an in vitro optimized, synergistic fixed-ratio combination of paclitaxel (PTX) and clofazimine (B663) has been developed and called Riminocelles™. The present pre-clinical study investigated the acute toxicity, systemic exposure, repeat dose toxicity and efficacy of Riminocelles in parallel to Taxol® at an equivalent PTX dose of 10 mg/kg. Daily and weekly dosing schedules were evaluated against Pgp-expressing human colon adenocarcinoma (HCT-15) xenografts implanted subcutaneously in athymic mice. Riminocelles produced statistically significant (p<0.05) tumor growth delays of 3.2 and 2.7 days for the respective schedules in contrast to Taxol delaying growth by 0.5 and 0.6 days. Using the control tumor doubling time of 4.2 days, tumor-cell-kill values of 0.23 for Riminocelles and 0.04 for Taxol following daily schedules were calculated. A significant weight loss of 5.7 % after 14 days (p<0.05) relative to the control group (n=8) was observed for the daily Taxol group whereas Riminocelles did not incur significant weight loss neither were blood markers of toxicity elevated after acute administration (n= 3). The safety and efficacy of Riminocelles is statistically superior to Taxol. However, passive tumor targeting was not achieved and the tumor burden progressed quickly. Prior to further animal studies, the in vivo thermodynamic instability of the simple lipopolymeric micellular delivery system requires improvement so as to maintain and selectively deliver the fixed-ratio drug combination.BioPAD (BPH 004) and the Department of Pharmacology, University of Pretoria.http://link.springer.com/journal/133462016-06-30hb201

Potential for identifying plant-based toxins on San hunter-gatherer arrowheads

The antiquity of the use of hunting poisons has received much attention in recent years. In this paper we present the results of a pilot study designed to detect the presence of organic compounds, typically of less than 1200 Da, from poisonous plants that may have been used as hunting poisons in the past. We used ultra-performance liquid chromatography connected to a Synapt G2 high-resolution MS-QTOF mass spectrometer (UPLC-QTOF-MS) to provisionally identify plant-based toxins present in (1) extracts of fresh plant material, (2) a blind control recipe consisting of three plant ingredients and (3) a Hei||om arrow poison of unknown ingredients. Although not all expected toxic compounds were identified, those that were identified compared favourably with those reported in the literature and confirmed through databases, specifically the Dictionary of Natural Products and ChemSpider. MS/MS fragmentation patterns and accurate mass were used for tentative identification of compounds because archaeological residues usually contain insufficient material for unambiguous identification using nuclear magnetic resonance. We highlight the potential of this method for accurately identifying plant-based toxins present on archaeological artefacts and unique (albeit non-toxic) chemical markers that may allow one to infer the presence of toxic plant ingredients in arrow poisons. Any chemical study of archaeological material should consider the unique environmental degradative factors and be sensitive to the oxidative by-products of toxic compounds

Preliminary screening of polybrominated diphenyl ethers (PBDEs), hexabromocyclododecane (HBCDD) and tetrabromobisphenol A (TBBPA) flame retardants in landfill leachate

The occurrence of selected brominated flame retardants, including nine polybrominated diphenyl ether (PBDE) congeners, hexabromocyclododecane (HBCDD) and tetrabromobisphenol A (TBBPA) in leachate samples from eight landfill sites in South Africa, were investigated. In addition, the possible influences of dissolved organic carbon on their levels were also evaluated. Filtered leachate samples were subjected to solid-phase extraction to isolate the various target compounds. PBDEs with six bromine substituents and above, as well as α-HBCDD, β-HBCDD and TBBPA, were generally found below the detection limit. However, the mean value of the total lower PBDE congeners ranged between 0.04 and 0.48 μg L−1, and the concentrations of γ-HBCDD ranged from not detectable (ND) to 0.05 μg L−1. No significant correlation was observed between the target compounds and dissolved organic carbon, although weak to moderate correlations were mostly observed for the lower PBDEs.https://link.springer.com/journal/106612018-08-30hj2017Chemistr

Potential for identifying plant-based toxins on San hunter-gatherer arrowheads

The antiquity of the use of hunting poisons has received much attention in recent years. In this paper we present the results of a pilot study designed to detect the presence of organic compounds, typically of less than 1200 Da, from poisonous plants that may have been used as hunting poisons in the past. We used ultra-performance liquid chromatography connected to a Synapt G2 high-resolution MS-QTOF mass spectrometer (UPLC-QTOF-MS) to provisionally identify plant-based toxins present in (1) extracts of fresh plant material, (2) a blind control recipe consisting of three plant ingredients and (3) a Hei||om arrow poison of unknown ingredients. Although not all expected toxic compounds were identified, those that were identified compared favourably with those reported in the literature and confirmed through databases, specifically the Dictionary of Natural Products and ChemSpider. MS/MS fragmentation patterns and accurate mass were used for tentative identification of compounds because archaeological residues usually contain insufficient material for unambiguous identification using nuclear magnetic resonance. We highlight the potential of this method for accurately identifying plant-based toxins present on archaeological artefacts and unique (albeit non-toxic) chemical markers that may allow one to infer the presence of toxic plant ingredients in arrow poisons. Any chemical study of archaeological material should consider the unique environmental degradative factors and be sensitive to the oxidative by-products of toxic compounds. Significance:&nbsp; Methodology is presented for the identification of ancient plant-based arrow poisons