The novel choline kinase inhibitor ICL-CCIC-0019 reprograms cellular metabolism and inhibits cancer cell growth

The glycerophospholipid phosphatidylcholine is the most abundant phospholipid species of eukaryotic membranes and essential for structural integrity and signaling function of cell membranes required for cancer cell growth. Inhibition of choline kinase alpha (CHKA), the first committed step to phosphatidylcholine synthesis, by the selective small-molecule ICL-CCIC-0019, potently suppressed growth of a panel of 60 cancer cell lines with median GI50 of 1.12 μM and inhibited tumor xenograft growth in mice. ICL-CCIC-0019 decreased phosphocholine levels and the fraction of labeled choline in lipids, and induced G1 arrest, endoplasmic reticulum stress and apoptosis. Changes in phosphocholine cellular levels following treatment could be detected non-invasively in tumor xenografts by [18F]-fluoromethyl-[1,2–2H4]-choline positron emission tomography. Herein, we reveal a previously unappreciated effect of choline metabolism on mitochondria function. Comparative metabolomics demonstrated that phosphatidylcholine pathway inhibition leads to a metabolically stressed phenotype analogous to mitochondria toxin treatment but without reactive oxygen species activation. Drug treatment decreased mitochondria function with associated reduction of citrate synthase expression and AMPK activation. Glucose and acetate uptake were increased in an attempt to overcome the metabolic stress. This study indicates that choline pathway pharmacological inhibition critically affects the metabolic function of the cell beyond reduced synthesis of phospholipids

The design of microfluidic reactors and their application to small molecule organic synthesis

Imperial Users onl

The design of microfluidic reactors and their application to small molecule organic synthesis

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Preclinical Assessment of Carboplatin Treatment Efficacy in Lung Cancer by ¹⁸F-ICMT-11-Positron Emission Tomography

Tumour response to therapy is assessed primarily in the clinic by monitoring reductions in tumour size. However, this approach lacks sensitivity since in many cases several weeks may elapse before there is evidence of tumour shrinkage. There is therefore a need to develop non-invasive imaging techniques for monitoring tumour treatment response in the clinic. Here, we assessed the pre-clinical utility of (18)F-ICMT-11 positron emission tomography--a method for detecting caspase 3/7 activation--in non-small cell lung cancer (NSCLC). (18)F-ICMT-11 uptake was compared to molecular biochemical measures of cell death in PC9 and A549 NSCLC cells following treatment with carboplatin in vitro and in vivo. Carboplatin-induced apoptosis in the ERCC1 low/mutant EGFR PC9 cells was characterised by time and dose-related increased caspase-3/7 activation, poly-ADP-ribose polymerase cleavage and Annexin V staining. 18F-ICMT-11 uptake was consequently increased up to 14-fold at 200 µM carboplatin compared to vehicle treated cells (P<0.01). In contrast, necrosis was the predominant death mechanism in ERCC1 high/wt EGFR A549 cells and no change in (18)F-ICMT-11 uptake was detected. In vivo, histological analysis of PC9 tumour xenografts indicated high pre-therapy necrosis. A 4.6-fold increase in cleaved caspase-3/7 was measured in non-necrotic regions of PC9 tumours at 48 h post carboplatin therapy. Average PET-derived tumour (18)F-ICMT-11 uptake was insensitive to changes in apoptosis in the presence of substantial pre-existing necrosis. PET-based voxel intensity sorting however, identified intra-tumoural regions of high (18)F-ICMT-11 uptake, enabling accurate assessment of apoptosis and therefore therapy response. In A549 tumours that lacked high pre-therapy necrosis, carboplatin induced growth inhibition that was only minimally associated with apoptosis and thus not detectable by (18)F-ICMT-11 PET

Differential responses to carboplatin treatment in PC9 and A549 cells.

<p>A: Carboplatin-induced growth inhibition in PC9 and A549 cells using a sulforhodamine B assay 72 h post treatment. B: Western blot analysis of the levels of uncleaved PARP, cleaved PARP and cleaved (active) caspase 3 72 h post carboplatin treatment (0–200 µM) in PC9 and A549 cells. Actin was used as a loading control. C, D: Flow cytometric analysis of PC9 (C) and A549 cells (D) treated with carboplatin (100 µM) or vehicle. Apoptotic cells were identified by Annexin V-Alexafluor488 (λ Ex/Em = 495/519 nm) and necrotic cells by 7-AAD (λ Ex/Em = 546/647 nm). Population Q4 represents viable cells, whereas population Q3 represents apoptotic cells that have low 7-AAD fluorescence and stain with Annexin V. Population Q2 represents secondary apoptotic/necrotic cells.</p