Glycolysis in the African Trypanosome: Targeting Enzymes and Their Subcellular Compartments for Therapeutic Development

Subspecies of the African trypanosome, Trypanosoma brucei, which cause human African trypanosomiasis, are transmitted by the tsetse fly, with transmission-essential lifecycle stages occurring in both the insect vector and human host. During infection of the human host, the parasite is limited to using glycolysis of host sugar for ATP production. This dependence on glucose breakdown presents a series of targets for potential therapeutic development, many of which have been explored and validated as therapeutic targets experimentally. These include enzymes directly involved in glucose metabolism (e.g., the trypanosome hexokinases), as well as cellular components required for development and maintenance of the essential subcellular compartments that house the major part of the pathway, the glycosomes

A target-based high throughput screen yields Trypanosoma brucei hexokinase small molecule inhibitors with antiparasitic activity. PLoS Negl Trop. Dis

Abstract Background: The parasitic protozoan Trypanosoma brucei utilizes glycolysis exclusively for ATP production during infection of the mammalian host. The first step in this metabolic pathway is mediated by hexokinase (TbHK), an enzyme essential to the parasite that transfers the c-phospho of ATP to a hexose. Here we describe the identification and confirmation of novel small molecule inhibitors of bacterially expressed TbHK1, one of two TbHKs expressed by T. brucei, using a high throughput screening assay

A Target-Based High Throughput Screen Yields Trypanosoma brucei Hexokinase Small Molecule Inhibitors with Antiparasitic Activity

African sleeping sickness is a disease found in sub-Saharan Africa that is caused by the single-celled parasite Trypanosoma brucei. The drugs used widely now to treat infections are 50 years old and notable for their toxicity, emphasizing the need for development of new therapeutics. In the search for potential drug targets, researchers typically focus on enzymes or proteins that are essential to the survival of the infectious agent while being distinct enough from the host to avoid accidental targeting of the host enzyme. This work describes our research on one such trypanosome enzyme, hexokinase, which is a protein that the parasite requires to make energy. Here we describe the results of our search for inhibitors of the parasite enzyme. By screening 220,223 compounds for anti-hexokinase activity, we have identified new inhibitors of the parasite enzyme. Some of these are toxic to trypanosomes while having no effect on mammalian cells, suggesting that they may hold promise for the development of new anti-parasitic compounds

Association of Accelerometry-Measured Physical Activity and Cardiovascular Events in Mobility-Limited Older Adults: The LIFE (Lifestyle Interventions and Independence for Elders) Study.

BACKGROUND:Data are sparse regarding the value of physical activity (PA) surveillance among older adults-particularly among those with mobility limitations. The objective of this study was to examine longitudinal associations between objectively measured daily PA and the incidence of cardiovascular events among older adults in the LIFE (Lifestyle Interventions and Independence for Elders) study. METHODS AND RESULTS:Cardiovascular events were adjudicated based on medical records review, and cardiovascular risk factors were controlled for in the analysis. Home-based activity data were collected by hip-worn accelerometers at baseline and at 6, 12, and 24 months postrandomization to either a physical activity or health education intervention. LIFE study participants (n=1590; age 78.9±5.2 [SD] years; 67.2% women) at baseline had an 11% lower incidence of experiencing a subsequent cardiovascular event per 500 steps taken per day based on activity data (hazard ratio, 0.89; 95% confidence interval, 0.84-0.96; P=0.001). At baseline, every 30 minutes spent performing activities ≥500 counts per minute (hazard ratio, 0.75; confidence interval, 0.65-0.89 [P=0.001]) were also associated with a lower incidence of cardiovascular events. Throughout follow-up (6, 12, and 24 months), both the number of steps per day (per 500 steps; hazard ratio, 0.90, confidence interval, 0.85-0.96 [P=0.001]) and duration of activity ≥500 counts per minute (per 30 minutes; hazard ratio, 0.76; confidence interval, 0.63-0.90 [P=0.002]) were significantly associated with lower cardiovascular event rates. CONCLUSIONS:Objective measurements of physical activity via accelerometry were associated with cardiovascular events among older adults with limited mobility (summary score >10 on the Short Physical Performance Battery) both using baseline and longitudinal data. CLINICAL TRIAL REGISTRATION:URL: http://www.clinicaltrials.gov. Unique identifier: NCT01072500

Homologous recombination DNA repair deficiency and PARP inhibition activity in primary triple negative breast cancer

Funder: Breast Cancer Now (BCN); doi: https://doi.org/10.13039/100009794Abstract: Triple negative breast cancer (TNBC) encompasses molecularly different subgroups, with a subgroup harboring evidence of defective homologous recombination (HR) DNA repair. Here, within a phase 2 window clinical trial, RIO trial (EudraCT 2014-003319-12), we investigate the activity of PARP inhibitors in 43 patients with untreated TNBC. The primary end point, decreased Ki67, occured in 12% of TNBC. In secondary end point analyses, HR deficiency was identified in 69% of TNBC with the mutational-signature-based HRDetect assay. Cancers with HRDetect mutational signatures of HR deficiency had a functional defect in HR, assessed by impaired RAD51 foci formation on end of treatment biopsy. Following rucaparib treatment there was no association of Ki67 change with HR deficiency. In contrast, early circulating tumor DNA dynamics identified activity of rucaparib, with end of treatment ctDNA levels suppressed by rucaparib in mutation-signature HR-deficient cancers. In ad hoc analysis, rucaparib induced expression of interferon response genes in HR-deficient cancers. The majority of TNBCs have a defect in DNA repair, identifiable by mutational signature analysis, that may be targetable with PARP inhibitors

Homologous recombination DNA repair deficiency and PARP inhibition activity in primary triple negative breast cancer

Funder: Breast Cancer Now (BCN); doi: https://doi.org/10.13039/100009794Abstract: Triple negative breast cancer (TNBC) encompasses molecularly different subgroups, with a subgroup harboring evidence of defective homologous recombination (HR) DNA repair. Here, within a phase 2 window clinical trial, RIO trial (EudraCT 2014-003319-12), we investigate the activity of PARP inhibitors in 43 patients with untreated TNBC. The primary end point, decreased Ki67, occured in 12% of TNBC. In secondary end point analyses, HR deficiency was identified in 69% of TNBC with the mutational-signature-based HRDetect assay. Cancers with HRDetect mutational signatures of HR deficiency had a functional defect in HR, assessed by impaired RAD51 foci formation on end of treatment biopsy. Following rucaparib treatment there was no association of Ki67 change with HR deficiency. In contrast, early circulating tumor DNA dynamics identified activity of rucaparib, with end of treatment ctDNA levels suppressed by rucaparib in mutation-signature HR-deficient cancers. In ad hoc analysis, rucaparib induced expression of interferon response genes in HR-deficient cancers. The majority of TNBCs have a defect in DNA repair, identifiable by mutational signature analysis, that may be targetable with PARP inhibitors

Identification of Inhibitors of \u3cem\u3eTrypanosoma brucei\u3c/em\u3e Hexokinases

The unicellular eukaryote Trypanosoma brucei (T. brucei) is the causative agent of human African trypanosomiasis (HAT), a disease that annually infects ~500,000 people in sub-Saharan Africa resulting in 50,000 – 70,000 deaths per year. Without treatment, HAT is fatal. Unfortunately, current treatments are limited in availability, have toxic side effects, are difficult to administer and are not well characterized in terms of their mechanism of action. Thus, the lack of affordable, safe, and effective therapies for those with African trypanosomiasis makes the identification of molecular target-specific chemotypes a priority in our effort to understand the mechanisms involved with parasite growth and viability, as well as for future therapeutic development. The probe identified from this effort, ML205, is a stable, small molecule possessing submicromolar activity (IC50 = 0.98 μM) against a defined T. brucei hexokinase 1 (rTbHK1) target. The probe was not toxic to mammalian cells (IMR-90 cells, EC50 \u3e 25 μM) and mechanistic studies revealed that the probe operates with mixed inhibition with respect to ATP

Validation of the HTS by LOPAC screening.

<p>Plot of percent inhibition for duplicate screen of the 1280 LOPAC compounds. LOPAC compounds (blue), minimum control, which should equal ∼100% inhibition of signal readout (red), IC₅₀ control compounds (yellow), and maximum control compounds, which should equal ∼0% inhibition of signal readout (green), are indicated.</p