birinapant and Colorectal-Neoplasms

Apoptosis resistance worsens treatment response in cancer and is associated with poor prognosis. Inhibition of anti-apoptotic proteins can restore cell death and improve treatment efficacy. cIAP1, cIAP2, and XIAP belong to the inhibitor of apoptosis protein (IAP) family and block apoptosis. Targeting IAPs with peptides or peptidomimetics mimicking the IAP-antagonizing activity of the cell's endogenous IAP antagonist SMAC (SMAC mimetics) showed promising results and fueled development of novel compounds. ASTX660 belongs to the recently introduced class of non-peptidomimetic IAP antagonists and successfully completed phase I clinical trials. However, ASTX660 has thus far only been evaluated in few cancer entities. Here, we demonstrate that ASTX660 has cell death-promoting activity in colorectal cancer and provide a head-to-head comparison with birinapant, the clinically most advanced peptidomimetic IAP antagonist. ASTX660 facilitates activation of the extrinsic apoptosis pathway upon stimulation with the death ligands TNF and TRAIL and boosts effector caspase activation and subsequent apoptosis. Mechanistically, ASTX660 enhances amplification of death receptor-generated apoptotic signals in a mitochondria-dependent manner. Failure to activate the mitochondria-associated (intrinsic) apoptosis pathway attenuated the apoptosis-promoting effect of ASTX660. Further clinical studies are warranted to highlight the therapeutic potential of ASTX660 in colorectal cancer.

Topics: Cell Line, Tumor; Cell Proliferation; Cell Survival; Colorectal Neoplasms; Dipeptides; Down-Regulation; Drug Synergism; Gene Expression Regulation, Neoplastic; HCT116 Cells; HT29 Cells; Humans; Indoles; Inhibitor of Apoptosis Proteins; Morpholines; Piperazines; Pyrroles; Signal Transduction; TNF-Related Apoptosis-Inducing Ligand

Resistance to chemotherapy-induced cell death is a major barrier to effective treatment of solid tumours such as colorectal cancer, CRC. Herein, we present a study aimed at developing a proteomics-based predictor of response to standard-of-care (SoC) chemotherapy in combination with antagonists of IAPs (inhibitors of apoptosis proteins), which have been implicated as mediators of drug resistance in CRC. We quantified the absolute expression of 19 key apoptotic proteins at baseline in a panel of 12 CRC cell lines representative of the genetic diversity seen in this disease to identify which proteins promote resistance or sensitivity to a model IAP antagonist [birinapant (Bir)] alone and in combination with SoC chemotherapy (5FU plus oxaliplatin). Quantitative western blotting demonstrated heterogeneous expression of IAP interactome proteins across the CRC cell line panel, and cell death analyses revealed a widely varied response to Bir/chemotherapy combinations. Baseline protein expression of cIAP1, caspase-8 and RIPK1 expression robustly correlated with response to Bir/chemotherapy combinations. Classifying cell lines into 'responsive', 'intermediate' and 'resistant' groups and using linear discriminant analysis (LDA) enabled the identification of a 12-protein signature that separated responders to Bir/chemotherapy combinations in the CRC cell line panel with 100% accuracy. Moreover, the LDA model was able to predict response accurately when cells were cocultured with Tumour necrosis factor-alpha to mimic a pro-inflammatory tumour microenvironment. Thus, our study provides the starting point for a proteomics-based companion diagnostic that predicts response to IAP antagonist/SoC chemotherapy combinations in CRC.

Topics: Alkaline Phosphatase; Antineoplastic Agents; Apoptosis; Caspase 8; Cell Line, Tumor; Colorectal Neoplasms; Dipeptides; Drug Resistance, Neoplasm; Fluorouracil; Gene Expression Regulation, Neoplastic; GPI-Linked Proteins; Humans; Indoles; Inhibitor of Apoptosis Proteins; Neoplasm Proteins; Oxaliplatin; Proteomics; Receptor-Interacting Protein Serine-Threonine Kinases; Transcriptome; Tumor Microenvironment

High expression levels of Inhibitors of Apoptosis Proteins (IAPs) have been correlated with poor cancer prognosis and block the cell death pathway by interfering with caspase activation. SMAC-mimetics are small-molecule inhibitors of IAPs that mimic the endogenous SMAC and promote the induction of cell death by neutralizing IAPs.. In this study, anti-tumour activity of new SMAC-mimetics Birinapant and AT-406 is evaluated against colorectal adenocarcinoma cells and IAP cross-talk with either oncogenic BRAF or BCL-2, or with the TRAIL are further exploited towards rational combined protocols.. It is shown that pre-treatment of SMAC-mimetics followed by their combined treatment with BRAF inhibitors can decrease cell viability, migration and can very efficiently sensitize colorectal tumour cells to apoptosis. Moreover, co-treatment of TRAIL with SMAC-mimetics can efficiently sensitize resistant tumour cells to apoptosis synergistically, as shown by median effect analysis. Finally, Birinapant and AT-406 can synergise with BCL-2 inhibitor ABT-199 to reduce viability of adenocarcinoma cells with high BCL-2 expression.. Proposed synergistic rational anticancer combined protocols of IAP antagonists Birinapant and AT-406 in 2D and 3D cultures can be later further exploited in vivo, from precision tumour biology to precision medical oncology.

Topics: Antineoplastic Agents; Azocines; Benzhydryl Compounds; Bridged Bicyclo Compounds, Heterocyclic; Caco-2 Cells; Cell Culture Techniques; Cell Line, Tumor; Cell Proliferation; Cell Survival; Colorectal Neoplasms; Dipeptides; Drug Resistance, Neoplasm; Drug Synergism; Gene Expression Regulation, Neoplastic; HCT116 Cells; HT29 Cells; Humans; Indoles; Mutation; Proto-Oncogene Proteins B-raf; Sulfonamides; TNF-Related Apoptosis-Inducing Ligand

Non-invasive serial imaging is desirable to detect processes such as necrotic and apoptotic cell death in cancer patients undergoing treatment. This study investigated the use of diffusion-weighted (DW-) magnetic resonance imaging (MRI) for imaging cell death induced by either a cytotoxic drug (irinotecan), or the apoptosis-inducing agent birinapant, in human tumour xenografts in vivo.. Nude mice bearing human SW620 colon carcinoma xenografts were treated with vehicle, irinotecan (50 mg kg(-1)) or birinapant (30 mg kg(-1)) for up to 5 days. DW-MRI was performed prior to and on days 1, 3 and 5 during treatment. Assessment of tumour apoptosis and necrosis ex vivo was used to validate the imaging findings.. Both irinotecan and birinapant induced significant tumour growth delay. Irinotecan induced a small increase in the tumour apparent diffusion coefficient (ADC) after 1 day, with a 20 and 30% increase at days 3 and 5 respectively. ADC was unchanged in the vehicle- and birinapant-treated tumours despite a growth delay in the latter. Histological analysis showed that irinotecan increased necrosis at days 3 and 5, and induced apoptosis after 1 day, compared with vehicle. Birinapant induced apoptosis after day 3, but had no effect on tumour necrosis.. Tumour ADC changes after irinotecan treatment were associated with the induction of a mixture of necrotic and apoptotic cell death, whereas induction of apoptosis alone with birinapant was not sufficient to induce changes in tissue microstructure that were detectable with DW-MRI. ADC is a useful non-invasive biomarker for early detection of response to cytotoxic drugs, but false negatives may arise while detecting apoptotic response to birinapant.

Topics: Adenocarcinoma; Animals; Antineoplastic Agents, Phytogenic; Apoptosis; Blotting, Western; Camptothecin; Cell Proliferation; Colorectal Neoplasms; Diffusion Magnetic Resonance Imaging; Dipeptides; Female; Humans; Immunoenzyme Techniques; Indoles; Irinotecan; Lymphatic Metastasis; Mice; Mice, Nude; Necrosis; Tumor Cells, Cultured; Xenograft Model Antitumor Assays