Ignacio Gil-Bazo

Reassessment of Id1 Protein Expression in Human Mammary, Prostate, and Bladder Cancers Using a Monospecific Rabbit Monoclonal Anti-Id1 Antibody

Id proteins are a class of dominant-negative antagonists of helix-loop-helix transcription factors and have been shown to control differentiation of a variety of cell types in diverse organisms. Although the importance of Id1 in tumor endothelial cells is well established, the expression and role of the Id1 protein in human cancer cells is controversial. To explore this issue, we developed and characterized a highly specific rabbit monoclonal antibody against Id1 to assess its expression in human breast, prostate, and bladder malignancies. Our results show that in usual types of human mammary carcinomas, the Id1 protein is expressed exclusively in the endothelium. Interestingly, we detected nuclear expression of the Id1 protein in the tumor cells in 10 of 45 cases of poorly differentiated and highly aggressive carcinoma with metaplastic morphology. Similarly, only 1 of 30 prostate cancer samples showed Id1-positive tumor cells, whereas in almost all, endothelial cells showed high Id1 expression. Intriguingly, whereas normal prostate glands do not show any Id1 protein expression, basal layer cells of benign prostate glands in proximity to tumors expressed high levels of the Id1 protein. In contrast to the lack of Id1 expression in the usual types of mammary and prostate cancers, the majority of transitional cell bladder tumors showed Id1 protein expression in both tumor and endothelial cells. These results suggest that further refinement of Id1 expression patterns in a variety of tumor types will be necessary to identify and study the functional roles played by Id1 in human neoplastic processes.

Identification of TNF-α and MMP-9 as potential baseline predictive serum markers of sunitinib activity in patients with renal cell carcinoma using a human cytokine array

Background:Several drugs are available to treat metastatic renal-cell carcinoma (MRCC), and predictive markers to identify the most adequate treatment for each patient are needed. Our objective was to identify potential predictive markers of sunitinib activity in MRCC.Methods:We collected sequential serum samples from 31 patients treated with sunitinib. Sera of six patients with extreme phenotypes of either marked responses or clear progressions were analysed with a Human Cytokine Array which evaluates 174 cytokines before and after treatment. Variations in cytokine signal intensity were compared between both groups and the most relevant cytokines were assessed by ELISA in all the patients.Results:Twenty-seven of the 174 cytokines varied significantly between both groups. Five of them (TNF-α, MMP-9, ICAM-1, BDNF and SDF-1) were assessed by ELISA in 21 evaluable patients. TNF-α and MMP-9 baseline levels were significantly increased in non-responders and significantly associated with reduced overall survival and time-to-progression, respectively. The area under the ROC curves for TNF-α and MMP-9 as predictive markers of sunitinib activity were 0.83 and 0.77.Conclusion:Baseline levels of TNF-α and MMP-9 warrant further study as predictive markers of sunitinib activity in MRCC. Selection of patients with extreme phenotypes seems a valid method to identify potential predictive factors of response.

TGFBI expression is associated with a better response to chemotherapy in NSCLC

BackgroundLung cancer is one of the most prevalent neoplasias in developed countries. Advances in patient survival have been limited and the identification of prognostic molecules is needed. Resistance to treatment is strongly related to tumor cell adhesion to the extracellular matrix and alterations in the quantity and nature of molecules constituting the tumor cell niche. Recently, transforming growth factor beta-induced protein (TGFBI), an extracellular matrix adaptor protein, has been reported to be differentially expressed in transformed tissues. Loss of TGFBI expression has been described in several cancers including lung carcinoma, and it has been suggested to act as a tumor suppressor gene.ResultsTo address the importance of TGFBI expression in cancer progression, we determined its expression in NSCLC clinical samples using immunohistochemistry. We identified a strong association between elevated TGFBI expression and the response to chemotherapy. Furthermore, we transiently over-expressed and silenced TGFBI in human NSCLC cell lines. Cells over-expressing TGFBI displayed increased sensitivity to etoposide, paclitaxel, cisplatin and gemcitabine. We observed that TGFBI-mediated induction of apoptosis occurred through its binding to αvβ3 integrin. We also determined that full-length TGFBI did not induce caspase 3/7 activation but its proteolytic fragments that were < 3 kDa in size, were able to activate caspase 3, 7 and 8. This pro-apoptotic effect was blocked by anti-αvβ3 integrin antibodies.ConclusionsThe results shown here indicate that TGFBI is a predictive factor of the response to chemotherapy, and suggest the use of TGFBI-derived peptides as possible therapeutic adjuvants for the enhancement of responses to chemotherapy.

Novel therapeutic strategies for patients with NSCLC that do not respond to treatment with EGFR inhibitors

INTRODUCTION Treatment with epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) yields tumour responses in non-small cell lung cancer (NSCLC) patients harbouring activating EGFR mutations. However, even in long-lasting responses, resistance to EGFR TKIs invariably occurs. AREAS COVERED This review examines resistance mechanisms to EGFR TKI treatment, which mainly arise from secondary EGFR mutations. Other resistance-inducing processes include mesenchymal-epithelial transition factor (MET) amplification, epithelial-mesenchymal transformation, phenotypic change from NSCLC to small-cell lung carcinoma, and modifications in parallel signalling pathways. Current therapeutic strategies to overcome these EGFR TKI resistance mechanisms focus on the inhibition or blocking of multiple members of the ErbB family. Several molecules which target multiple ErbB receptors are being investigated in NSCLC and other indications including afatinib, an ErbB Family Blocker, as well as dacomitinib and lapatinib. Novel, non-quinazoline, EGFR inhibitors, that also target EGFR activating and resistance (T790M) mutations, are currently under clinical development. Other therapeutic strategies include inhibition of parallel and downstream pathways, using agents which target heat shock protein (HSP)90 or poly (ADP-ribose) polymerase in addition to mammalian target of rapamycin (mTOR), monoclonal antibodies against the insulin-like growth factor-1 receptor, and fulvestrant-mediated oestrogen receptor regulation. CONCLUSION Improved understanding of mechanisms underlying resistance to EGFR TKIs emphasises the importance of a genotype-guided approach to therapy. Elucidation of resistance mechanisms is indeed crucial to target innovative therapeutic approaches and to improve the efficacy of anticancer regimes in NSCLC.

External-beam radiation therapy and high-dose rate brachytherapy combined with long-term androgen deprivation therapy in high and very high prostate cancer: preliminary data on clinical outcome.

PURPOSE To determine the feasibility of combined long-term androgen deprivation therapy (ADT) and dose escalation with high-dose-rate (HDR) brachytherapy. METHODS AND MATERIALS Between 2001 and 2007, 200 patients with high-risk prostate cancer (32.5%) or very high-risk prostate cancer (67.5%) were prospectively enrolled in this Phase II trial. Tumor characteristics included a median pretreatment prostate-specific antigen of 15.2 ng/mL, a clinical stage of T2c, and a Gleason score of 7. Treatment consisted of 54 Gy of external irradiation (three-dimensional conformal radiotherapy [3DCRT]) followed by 19 Gy of HDR brachytherapy in four twice-daily treatments. ADT started 0-3 months before 3DCRT and continued for 2 years. RESULTS One hundred and ninety patients (95%) received 2 years of ADT. After a median follow-up of 3.7 years (range, 2-9), late Grade ≥2 urinary toxicity was observed in 18% of the patients and Grade ≥3 was observed in 5%. Prior transurethral resection of the prostate (p = 0.013) and bladder D(50) ≥1.19 Gy (p = 0.014) were associated with increased Grade ≥2 urinary complications; age ≥70 (p = 0.05) was associated with Grade ≥3 urinary complications. Late Grade ≥2 gastrointestinal toxicity was observed in 9% of the patients and Grade ≥3 in 1.5%. CTV size ≥35.8 cc (p = 0.007) and D(100) ≥3.05 Gy (p = 0.01) were significant for increased Grade ≥2 complications. The 5-year and 9-year biochemical relapse-free survival (nadir + 2) rates were 85.1% and 75.7%, respectively. Patients with Gleason score of 7-10 had a decreased biochemical relapse-free survival (p = 0.007). CONCLUSIONS Intermediate-term results at the 5-year time point indicate a favorable outcome without an increase in the rate of late complications.

Targeting hypoxia and angiogenesis through HIF-1alpha inhibition.

Hypoxia is an important phenomenon in the tumor microenvironment. Hypoxic tumors are more aggressive and resistant to anti-neoplastic treatments. HIF-1? plays a major role in the response of tumors to hypoxia, and it is mainly responsible for the “angiogenic switch”. HIF-1? contributes to tumor aggressiveness, invasiveness, and resistance to radiotherapy and chemotherapy. Targeting HIF-1? is an attractive strategy, with the potential for disrupting multiple pathways crucial for tumor growth.. We review recent findings on the potential efficacy of small molecules to down-regulate HIF-1?. These promising drugs inhibit HIF-1? synthesis or transcriptional activity by blocking a variety of steps in several different signaling pathways. Blocking HIF-1? activity should not only down-regulate tumor angiogenesis, but also interfere with glycolytic metabolism and tumor cell growth. This strategy could also improve the efficiency of established tumor therapies.

Exosomes isolation and characterization in serum is feasible in non-small cell lung cancer patients: critical analysis of evidence and potential role in clinical practice

Exosomes are nano-sized vesicles of endolysosomal origin, released by several cytotypes in physiological and pathological conditions. Tumor derived exosomes, interacting with other cells of the tumor microenvironment, modulate tumor progression, angiogenic switch, metastasis, and immune escape. Recently, extracellular vesicles were proposed as excellent biomarkers for disease monitoring and prognosis in cancer patients. Non-small cell lung cancer (NSCLC) has a poor 5-year survival rate due to the delay in the detection of the disease. The majority of patients are diagnosed in an advanced disease stage. Exosomes might be promising beneficial tools as biomarker candidates in the scenario of NSCLC, since they contain both, proteins and miRNAs. The clinical case reported in this manuscript is a proof of concept revealing that NSCLC exosomes and sorted miRNAs might constitute, in a near future, novel biomarkers. This review summarizes the role of exosomes in NSCLC, focusing on the importance of exosomal microRNAs in lung cancer diagnosis and prognosis.

Inhibitor of Differentiation-1 as a Novel Prognostic Factor in NSCLC Patients with Adenocarcinoma Histology and Its Potential Contribution to Therapy Resistance

Purpose: High inhibitor of differentiation-1 (Id1) levels have been found in some tumor types. We aimed to study Id1 levels and their prognostic impact in a large series of stages I to IV non-small cell lung cancer (NSCLC) patients. Experiments in cell lines and cells derived from malignant pleural effusions (MPE) were also carried out. Experimental Design: A total of 346 NSCLC samples (three different cohorts), including 65 matched nonmalignant tissues, were evaluated for Id1 expression by using immunohistochemistry. Additional data from a fourth cohort including 111 patients were obtained for Id1 mRNA expression analysis by using publicly available microarrays. In vitro proliferation assays were conducted to characterize the impact of Id1 on growth and treatment sensitivity. Results: Significantly higher Id1 protein levels were found in tumors compared with normal tissues (P < 0.001) and in squamous carcinomas compared with adenocarcinomas (P < 0.001). In radically treated stages I to III patients and stage IV patients treated with chemotherapy, higher Id1 levels were associated with a shorter disease-free survival and overall survival in adenocarcinoma patients in a log-rank test. A Cox model confirmed the independent prognostic value of Id1 levels for both stages I to III and stage IV patients. In silico analysis confirmed a correlation between higher Id1 mRNA levels and poor prognosis for adenocarcinoma subjects. In vitro Id1 silencing in radio/chemotherapy-resistant adenocarcinoma cells from MPEs restored sensitivity to both therapies. Conclusions: In our series, Id1 levels showed an independent prognostic value in patients with adenocarcinoma, regardless of the stage. Id1 silencing may sensitize adenocarcinoma cells to radiotherapy and chemotherapy. Clin Cancer Res; 17(12); 4155–66. ©2011 AACR.

Entrectinib: a potent new TRK, ROS1, and ALK inhibitor

Introduction: Receptor tyrosine kinases (RTKs) and their signaling pathways, control normal cellular processes; however, their deregulation play important roles in malignant transformation. In advanced non-small cell lung cancer (NSCLC), the recognition of oncogenic activation of specific RTKs, has led to the development of molecularly targeted agents that only benefit roughly 20% of patients. Entrectinib is a pan-TRK, ROS1 and ALK inhibitor that has shown potent anti-neoplastic activity and tolerability in various neoplastic conditions, particularly NSCLC. Areas covered: This review outlines the pharmacokinetics, pharmacodynamics, mechanism of action, safety, tolerability, pre-clinical studies and clinical trials of entrectinib, a promising novel agent for the treatment of advanced solid tumors with molecular alterations of Trk-A, B and C, ROS1 or ALK. Expert opinion: Among the several experimental drugs under clinical development, entrectinib is emerging as an innovative and promising targeted agent. The encouraging antitumor activity reported in the Phase 1 studies, together with the acceptable toxicity profile, suggest that entrectinib, thanks to its peculiar mechanism of action, could play an important role in the treatment-strategies of multiple TRK-A, B, C, ROS1, and ALK- dependent solid tumors, including NSCLC and colorectal cancer. That being said, further evidence for its clinical use is still needed.

Improving disease control in advanced colorectal cancer: Panitumumab and cetuximab

Colorectal cancer remains a major public health concern in Europe and North America. It is responsible for one million new cases and half a million deaths per year worldwide. During the past few years new effective treatments have evolved improving the outcome of patients with this disease. Several alternatives are currently available for advanced colorectal cancer patients including different chemotherapeutic regimens (fluoropyrimidines, irinotecan and oxaliplatin) and targeted therapies such as bevacizumab and cetuximab. Different combinations achieve a median survival of over 2 years. Intense efforts focus on identifying agents targeting growth factor receptors, signal transduction pathways or angiogenesis mediators. One of the last available drugs for the management of advanced colorectal cancer is panitumumab, a well-tolerated and effective anti-EGFR monoclonal antibody approved as a single agent in chemotherapy refractory patients. We discuss the current evidence supporting panitumumab for metastatic colorectal cancer treatment, potential predictive biomarkers and ongoing clinical trials with different combinations including panitumumab.