Afatinib or Bevacizumab in combination with Osimertinib efficiently control tumor development in orthotopic murine models of non-small lung cancer.

Lung cancer is one of the most common and deadliest cancers. Preclinical models are essential to study new therapies and combinations taking tumor genetics into account. We have established cell lines expressing the luciferase gene from lines with varied genetic backgrounds, commonly encountered in patients with pulmonary adenocarcinoma. We have characterized these lines by testing their response to multiple drugs. Thus, we have developed orthotopic preclinical mouse models of NSCLC with very high engraftment efficiency. These models allow the easy monitoring of tumor growth, particularly in response to treatment, and of tumor cells dissemination in the body. We show that concomitant treatment with osimertinib (3rd generation tyrosine kinase inhibitor targeting mutated EGFR) and bevacizumab (anti-angiogenic targeting VEGF) can have a beneficial therapeutic effect on EGFR-mutated tumors. We also show that the addition of afatinib to osimertinib-treated tumors in escape leads to tumor growth inhibition. No such effect is observed with selumetinib or simvastatin. These preclinical mouse models therefore make it possible to test innovative therapeutic combinations and are also a tool of choice for studying resistance mechanisms.

Orthotopic model of lung cancer: isolation of bone micro-metastases after tumor escape from Osimertinib treatment.

BACKGROUND: Osimertinib is a third generation tyrosine kinase inhibitor (TKI) that targets the epidermal growth factor receptor (EGFR) in lung cancer. However, although this molecule is not subject to some of the resistance mechanisms observed in response to first generation TKIs, ultimately, patients relapse because of unknown resistance mechanisms. New relevant non-small cell lung cancer (NSCLC) mice models are therefore required to allow the analysis of these resistance mechanisms and to evaluate the efficacy of new therapeutic strategies. METHODS: Briefly, PC-9 cells, previously modified for luciferase expression, were injected into the tail vein of mice. Tumor implantation and longitudinal growth, almost exclusively localized in the lung, were evaluated by bioluminescence. Once established, the tumor was treated with osimertinib until tumor escape and development of bone metastases. RESULTS: Micro-metastases were detected by bioluminescence and collected for further analysis. CONCLUSION: We describe an orthotopic model of NSCLC protocol that led to lung primary tumor nesting and, after osimertinib treatment, by metastases dissemination, and that allow the isolation of these small osimertinib-resistant micro-metastases. This model provides new biological tools to study tumor progression from the establishment of a lung tumor to the generation of drug-resistant micro-metastases, mimicking the natural course of the disease in human NSCLC patients.

ING2 tumor suppressive protein translocates into mitochondria and is involved in cellular metabolism homeostasis.

ING2 (Inhibitor of Growth 2) is a tumor suppressor gene that has been implicated in critical biological functions (cell-cycle regulation, replicative senescence, DNA repair and DNA replication), most of which are recognized hallmarks of tumorigenesis occurring in the cell nucleus. As its close homolog ING1 has been recently observed in the mitochondrial compartment, we hypothesized that ING2 could also translocate into the mitochondria and be involved in new biological functions. In the present study, we demonstrate that ING2 is imported in the inner mitochondrial fraction in a redox-sensitive manner in human cells and that this mechanism is modulated by 14-3-3η protein expression. Remarkably, ING2 is necessary to maintain mitochondrial ultrastructure integrity without interfering with mitochondrial networks or polarization. We observed an interaction between ING2 and mtDNA under basal conditions. This interaction appears to be mediated by TFAM, a critical regulator of mtDNA integrity. The loss of mitochondrial ING2 does not impair mtDNA repair, replication or transcription but leads to a decrease in mitochondrial ROS production, suggesting a detrimental impact on OXPHOS activity. We finally show using multiple models that ING2 is involved in mitochondrial respiration and that its loss confers a protection against mitochondrial respiratory chain inhibition in vitro. Consequently, we propose a new tumor suppressor role for ING2 protein in the mitochondria as a metabolic shift gatekeeper during tumorigenesis.

ING3 is required for ATM signaling and DNA repair in response to DNA double strand breaks.

Inhibitor of Growth 3 (ING3) is a candidate tumor suppressor gene whose expression is lost in tumors such as hepatocellular carcinoma, head and neck squamous cell carcinoma and melanoma. In the present study, we show that ING3-depleted human cells and yeast cells deleted for its ortholog YNG2 are sensitive to DNA damage suggesting a conserved role in response to such stress. In human cells, ING3 is recruited to DNA double strand breaks and is required for ATM activation. Remarkably, in response to doxorubicin, ATM activation is dependent on ING3 but not on TIP60, whose recruitment to DNA breaks also depends on ING3. These events lead to ATM-mediated phosphorylation of NBS1 and the subsequent recruitment of RNF8, RNF168, 53BP1, and BRCA1, which are major mediators of the DNA damage response. Accordingly, upon genotoxic stress, DNA repair by non-homologous end joining (NHEJ) or homologous recombination (HR) were impaired in absence of ING3. Finally, immunoglobulin class switch recombination (CSR), a physiological mechanism requiring NHEJ repair, was impaired in the absence of ING3. Since deregulation of DNA double strand break repair is associated with genomic instability, we propose a novel function of ING3 as a caretaker tumor suppressor involved in the DNA damage signaling and repair.

tert-Butylphenolic Derivatives from Paenibacillus odorifer-A Case of Bioconversion.

Two compounds (1) and (2) containing tert-butylphenol groups were, for the first time, produced during the culture of Paenibacillus odorifer, a bacterial strain associated with the crustose lichen, Rhizocarpon geographicum. Their entire structures were identified by one-dimensional (1D) and two-dimensional (2D) NMR and high-resolution electrospray ionisation mass spectrometry (HRESIMS) spectroscopic analyses. Among them, Compound 1 exhibited significant cytotoxicity against B16 murine melanoma and HaCaT human keratinocyte cell lines with micromolar half maximal inhibitory concentration (IC50) values. Furthermore, after supplementation studies, a putative biosynthesis pathway was proposed for Compound 1 throughout a bioconversion by this bacterial strain of butylated hydroxyanisole (BHA), an antioxidant polymer additive.