RIP140 regulates transcription factor HES1 oscillatory expression and mitogenic activity in colon cancer cells.

The transcription factor receptor-interacting protein 140 (RIP140) regulates intestinal homeostasis and tumorigenesis through Wnt signaling. In this study, we investigated its effect on the Notch/HES1 signaling pathway. In colorectal cancer (CRC) cell lines, RIP140 positively regulated HES1 gene expression at the transcriptional level via a recombining binding protein suppressor of hairless (RBPJ)/neurogenic locus notch homolog protein 1 (NICD)-mediated mechanism. In support of these in vitro data, RIP140 and HES1 expression significantly correlated in mouse intestine and in a cohort of CRC samples, thus supporting the positive regulation of HES1 gene expression by RIP140. Interestingly, when the Notch pathway is fully activated, RIP140 exerted a strong inhibition of HES1 gene transcription controlled by the level of HES1 itself. Moreover, RIP140 directly interacts with HES1 and reversed its mitogenic activity in human CRC cells. In line with this observation, HES1 levels were associated with a better patient survival only when tumors expressed high levels of RIP140. Our data identify RIP140 as a key regulator of the Notch/HES1 signaling pathway, with a dual effect on HES1 gene expression at the transcriptional level and a strong impact on colon cancer cell proliferation.

Generation and Characterization of a New Preclinical Mouse Model of EGFR-Driven Lung Cancer with MET-Induced Osimertinib Resistance.

Despite the introduction of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) to treat advanced lung cancer harboring EGFR-activating mutations, the prognosis remains unfavorable because of intrinsic and/or acquired resistance. We generated a new state-of-the-art mouse strain harboring the human EGFRT790M/L858R oncogene and MET overexpression (EGFR/MET strain) that mimics the MET amplification occurring in one out of five patients with EGFR-mutated lung cancer that relapsed after treatment with osimertinib, a third-generation anti-EGFR TKI. We found that survival was reduced in EGFR/MET mice compared with mice harboring only EGFRT790M/L858R (EGFR strain). Moreover, EGFR/MET-driven lung tumors were resistant to osimertinib, recapitulating the phenotype observed in patients. Conversely, as also observed in patients, the crizotinib (anti-MET TKI) and osimertinib combination improved survival and reduced tumor burden in EGFR/MET mice, further validating the model's value for preclinical studies. We also found that in EGFR/MET mice, MET overexpression negatively regulated EGFR activity through MIG6 induction, a compensatory mechanism that allows the coexistence of the two onco-genic events. Our data suggest that single EGFR or MET inhibition might not be a good therapeutic option for EGFR-mutated lung cancer with MET amplification, and that inhibition of both pathways should be the best clinical choice in these patients.

The evolution of resistance and tolerance as cancer defences.

Although there is a plethora of cancer associated-factors that can ultimately culminate in death (cachexia, organ impairment, metastases, opportunistic infections, etc.), the focal element of every terminal malignancy is the failure of our natural defences to control unlimited cell proliferation. The reasons why our defences apparently lack efficiency is a complex question, potentially indicating that, under Darwinian terms, solutions other than preventing cancer progression are also important contributors. In analogy with host-parasite systems, we propose to call this latter option 'tolerance' to cancer. Here, we argue that the ubiquity of oncogenic processes among metazoans is at least partially attributable to both the limitations of resistance mechanisms and to the evolution of tolerance to cancer. Deciphering the ecological contexts of alternative responses to the cancer burden is not a semantic question, but rather a focal point in understanding the evolutionary ecology of host-tumour relationships, the evolution of our defences, as well as why and when certain cancers are likely to be detrimental for survival.

Incidence of thyroid dysfunctions during treatment with nivolumab for non-small cell lung cancer: Retrospective study of 105 patients.

INTRODUCTION: Immunotherapy is a standard not only in second line but also in first line treatment in patients with non-small cell lung cancer (NSCLC) and other tumors. Thyroid dysfunctions are the most common endocrine toxicities. OBJECTIVE: To determine the incidence of thyroid dysfunctions during treatment with a PD-1 monoclonal antibody (nivolumab) in patients with NSCLC. METHODS: Retrospective study of patients treated with nivolumab for NSCLC between May 2015 and December 2016; euthyroidism within the 3 months preceding immunotherapy; monitoring of thyroid function tests until stopping nivolumab, death or February 2017. Patients treated with levothyroxine, amiodarone or another immunotherapy were excluded. RESULTS: Among 183 patients treated, 105 fullfilled the inclusion criteria (72 males, median age: 61 years [range: 41-80]). Fifteen patients (14.3%) experienced a thyroid dysfunction; among them, compared to the "control" group (n=90), we found more females (53.3% vs. 27.8%; P=0.07), and younger patients (median age: 56 years vs. 62 years; P=0.02). Thirteen patients had thyrotoxicosis (median onset: 8 weeks), and then hypothyroidism was observed in 5 patients. Isolated hypothyroidism was rare (n=2) and late (median: 30 weeks). Three patients had anti-TPO antibodies. Three patients discontinued immunotherapy transiently due to thyroid dysfunctions. After a median follow-up of 9 months [95% CI, 7.5-10.3], one patient (6.7%) in the "thyroid dysfunctions" group and 30 patients (33.3%) in the "control" group died, with a trend toward a higher overall survival in the "thyroid dysfunctions" group (HR: 0.16 [95% CI, 0.02-1.15]; P=0.07). CONCLUSION: Thyroid dysfunctions (isolated thyrotoxicosis, biphasic thyroiditis and hypothyroidism) were common, and required patients with NSCLC to be screened during nivolumab therapy.

An oligoclonal antibody durably overcomes resistance of lung cancer to third-generation EGFR inhibitors.

Epidermal growth factor receptor (EGFR) mutations identify patients with lung cancer who derive benefit from kinase inhibitors. However, most patients eventually develop resistance, primarily due to the T790M second-site mutation. Irreversible inhibitors (e.g., osimertinib/AZD9291) inhibit T790M-EGFR, but several mechanisms, including a third-site mutation, C797S, confer renewed resistance. We previously reported that a triple mixture of monoclonal antibodies, 3×mAbs, simultaneously targeting EGFR, HER2, and HER3, inhibits T790M-expressing tumors. We now report that 3×mAbs, including a triplet containing cetuximab and trastuzumab, inhibits C797S-expressing tumors. Unlike osimertinib, which induces apoptosis, 3×mAbs promotes degradation of the three receptors and induces cellular senescence. Consistent with distinct mechanisms, treatments combining 3×mAbs plus sub-inhibitory doses of osimertinib synergistically and persistently eliminated tumors. Thus, oligoclonal antibodies, either alone or in combination with kinase inhibitors, might preempt repeated cycles of treatment and rapid emergence of resistance.

The RAS-related GTPase RHOB confers resistance to EGFR-tyrosine kinase inhibitors in non-small-cell lung cancer via an AKT-dependent mechanism.

Although lung cancer patients harboring EGFR mutations benefit from treatment with EGFR-tyrosine kinase inhibitors (EGFR-TKI), most of them rapidly relapse. RHOB GTPase is a critical player in both lung carcinogenesis and the EGFR signaling pathway; therefore, we hypothesized that it could play a role in the response to EGFR-TKI In a series of samples from EGFR-mutated patients, we found that low RHOB expression correlated with a good response to EGFR-TKI treatment while a poor response correlated with high RHOB expression (15.3 versus 5.6 months of progression-free survival). Moreover, a better response to EGFR-TKI was associated with low RHOB levels in a panel of lung tumor cell lines and in a lung-specific tetracycline-inducible EGFRL858R transgenic mouse model. High RHOB expression was also found to prevent erlotinib-induced AKT inhibition in vitro and in vivo Furthermore, a combination of the new-generation AKT inhibitor G594 with erlotinib induced tumor cell death in vitro and tumor regression in vivo in RHOB-positive cells. Our results support a role for RHOB/AKT signaling in the resistance to EGFR-TKI and propose RHOB as a potential predictor of patient response to EGFR-TKI treatment.

RhoB determines tumor aggressiveness in a murine EGFRL858R-induced adenocarcinoma model and is a potential prognostic biomarker for Lepidic lung cancer.

PURPOSE: A crucial event in lung adenocarcinoma progression is the switch from an aerogenous spread toward an infiltrating tumor. Loss of RhoB expression has been suggested to be critical for lung cancer invasion. Here, we tested RhoB expression as a prognostic biomarker in non-small cell lung cancer (NSCLC) with a special focus on lepidic pattern. EXPERIMENTAL DESIGN: We analyzed RhoB expression using both IHC and RT-qPCR in two series of operated patients (n = 100 and 48, respectively) and in a series of advanced lepidic adenocarcinoma (n = 31) from different hospitals. Next, we examined the role of RhoB in lung cancer progression in transgenic mice that express inducible EGFR(L858R) crossed with Rhob null mice. RESULTS: We identified that loss of RhoB expression was strongly associated with worse survival (P = 0.0001) and progression-free survival (P < 0.001) in the first series. We then confirmed these results after multivariate analyses of the second series. In the series of adenocarcinoma with lepidic features issued from a clinical trial (IFCT-0401), we showed that loss of RhoB expression was associated with higher aggressiveness of stage IV. Finally, we showed that EGFR(L858R)/Rhob(+/+) mice developed mainly diffuse lung tumors with a lepidic pattern, whereas EGFR(L858R)/Rhob(+/-) and EGFR(L858R)/Rhob(-/-) developed a greater number of tumors, and aggressive adenocarcinomas with invasive properties. CONCLUSIONS: We showed that RhoB is not only a strong prognostic factor in NSCLC but it is also critical for the acquisition of an aggressive phenotype of adenocarcinoma.

Loss of RhoB expression promotes migration and invasion of human bronchial cells via activation of AKT1.

Lung cancer is the leading cause of cancer-related death worldwide, mainly due to its highly metastatic properties. Previously, we reported an inverse correlation between RhoB expression and the progression of the lung cancer, occurring between preinvasive and invasive tumors. Herein, we mimicked the loss of RhoB observed throughout lung oncogenesis with RNA interference in nontumoral bronchial cell lines and analyzed the consequences on both cell transformation and invasion. Down-regulation of RhoB did not modify the cell growth properties but did promote migration and invasiveness. Furthermore, RhoB depletion was accompanied by modifications of actin and cell adhesion. The specific activation of the Akt1 isoform and Rac1 was found to be critical for this RhoB-mediated regulation of migration. Lastly, we showed that RhoB down-regulation consecutive to K-RasV12 cell transformation is critical for cell motility but not for cell proliferation. We propose that RhoB loss during lung cancer progression relates to the acquisition of invasiveness mediated by the phosphatidylinositol 3-kinase (PI3K)/AKT and Rac1 pathways rather than to tumor initiation.

Development of a conditional bioluminescent transplant model for TPM3-ALK-induced tumorigenesis as a tool to validate ALK-dependent cancer targeted therapy.

Overexpression and activation of TPM3-ALK tyrosine kinase fusion protein is a causal oncogenic event in the development of Anaplastic Large Cell Lymphoma and Inflammatory Myofibroblastic ALK-positive tumors. Thus, the development of ALK specific tyrosine kinase inhibitors is a current therapeutic challenge. Animal models are essential to assess, in vivo, the efficiency of ALK-oncogene inhibitors and to identify new and/or additional therapeutic targets in the ALK tumorigenesis pathway. Using the tetracycline system to allow conditional and concomitant TPM3-ALK and luciferase expression, we have developed a unique transplant model for bioluminescent TPM3-ALK-induced fibroblastic tumors in athymic nude mice. The reversible TPM3-ALK expression allowed us to demonstrate that this oncogene is essential for the tumor growth and its maintenance. In addition, we showed that this model could be used to precisely assess tumor growth inhibition upon ALK chemical inactivation. As proof of principle, we used the general tyrosine kinase inhibitor herbimycin A to inhibit ALK oncoprotein activity. As expected, herbimycin A treatment reduced tumor growth as assessed both by tumor volume measurement and bioluminescent imaging. We conclude that this transplant model for TPM3-ALK-induced tumors represents a valuable tool not only to accurately and rapidly evaluate in vivo ALK-targeted therapies but also to gain insight into the mechanism of ALK-positive tumor development.