Investigation of Clinically Significant Molecular Aberrations in Patients with Prostate Cancer: Implications for Personalized Treatment, Prognosis and Genetic Testing.

The data on tumor molecular profiling of European patients with prostate cancer is limited. Our aim was to evaluate the prevalence and prognostic and predictive values of gene alterations in unselected patients with prostate cancer. The presence of gene alterations was assessed in patients with histologically confirmed prostate cancer using the ForeSENTIA® Prostate panel (Medicover Genetics), targeting 36 clinically relevant genes and microsatellite instability testing. The primary endpoint was the prevalence of gene alterations in homologous recombination repair (HRR) genes. Overall, 196 patients with prostate cancer were evaluated (median age 72.2 years, metastatic disease in 141 (71.9%) patients). Gene alterations were identified in 120 (61%) patients, while alteration in HRR genes were identified in 34 (17.3%) patients. The most commonly mutated HRR genes were ATM (17, 8.7%), BRCA2 (9, 4.6%) and BRCA1 (4, 2%). The presence of HRR gene alterations was not associated with advanced stage (p = 0.21), age at diagnosis (p = 0.28), Gleason score (p = 0.17) or overall survival (HR 0.72; 95% CI: 0.41-1.26; p = 0.251). We identified clinically relevant somatic gene alterations in European patients with prostate cancer. These molecular alterations have prognostic significance and therapeutic implications and/or may trigger genetic testing in selected patients. In the era of precision medicine, prospective research on the predictive role of these alterations for innovative treatments or their combinations is warranted.

Genetic Predisposition to Male Breast Cancer: A Case Series.

BACKGROUND/AIM: Male breast cancer (MBC) is a very rare disorder affecting approximately 1 in 833 men. Genetic predisposition is one of the most important risk factors of MBC with BRCA2 being the most commonly mutated gene in males diagnosed with breast cancer. However, a large part of MBC heritability is still unexplained. This study sought to add to the data already available on the genetics of MBC. MATERIALS AND METHODS: Our study initially involved comprehensive analysis of BRCA1 and BRCA2, followed by analysis of 43 genes implicated in cancer predisposition in a series of 100 Greek patients diagnosed with MBC between 1995-2015. RESULTS: Pathogenic variants were identified in 13 patients, with BRCA2 being the most commonly affected gene, followed by BRCA1, RAD50, RAD51B, and MSH3. CONCLUSION: In agreement with previous reports, BRCA2 is the most important genetic factor of MBC predisposition, while the remaining known cancer predisposition genes are each very rarely involved, rendering conclusions as to their cumulative effect difficult to draw.

Development and Analytical Validation of a 6-Plex Reverse Transcription Droplet Digital PCR Assay for the Absolute Quantification of Prostate Cancer Biomarkers in Circulating Tumor Cells of Patients with Metastatic Castration-Resistant Prostate Cancer.

BACKGROUND: Gene expression in circulating tumor cells (CTCs) can be used as a predictive liquid biopsy test in metastatic castration-resistant prostate cancer (mCRPC). We developed a novel 6-plex reverse transcription droplet digital PCR (RT-ddPCR) assay for the absolute quantification of 4 prostate cancer biomarkers, a reference gene, and a synthetic DNA external control (DNA-EC) in CTCs isolated from mCRPC patients. METHODS: A novel 6-plex RT-ddPCR assay was developed for the simultaneous absolute quantification of AR-FL, AR-V7, PSA, and PSMA, HPRT (used as a reference gene), and a synthetic DNA-EC that was included for quality control. The assay was optimized and analytically validated using DNA synthetic standards for each transcript as positive controls. Epithelial cellular adhesion molecule (EpCAM)-positive CTC fractions isolated from 90 mCRPC patients and 11 healthy male donors were analyzed, and results were directly compared with reverse transcription quantitative PCR (RT-qPCR) for all markers in all samples. RESULTS: Linear dynamic range, limit of detection, limit of quantification, intra- and interassay precision, and analytical specificity were determined for each marker. Application of the assay in EpCAM-positive CTC showed positivity for AR-FL (71/90; 78.9%), AR-V7 (28/90; 31.1%), PSA (41/90; 45.6%), PSMA (38/90; 42.2%), and HPRT (90/90; 100%); DNA-EC concentration was constant across all samples. Direct comparison with RT-qPCR for the same markers in the same samples revealed RT-ddPCR to have superior diagnostic sensitivity. CONCLUSIONS: Our 6-plex RT-ddPCR assay was highly sensitive, specific, and reproducible, and enabled simultaneous and absolute quantification of 5 gene transcripts in minute amounts of CTC-derived cDNA. Application of this assay in clinical samples gave diagnostic sensitivity and specificity comparable to, or better than, RT-qPCR.

USP44 Promoter Methylation in Plasma Cell-Free DNA in Prostate Cancer.

Liquid biopsy provides real-time monitoring of tumor evolution and response to therapy through analysis of circulating tumor cells (CTCs) and plasma-circulating tumor DNA (ctDNA). USP44 is a critical gene which plays an important role in cell proliferation; however, its accurate role in other cellular networks is under research. USP44 promoter methylation has been so far reported in colorectal neoplasia and metastatic breast cancer. In this study, we examined for the first time USP44 promoter methylation in plasma cell-free DNA (cfDNA) of patients with prostate cancer (early stage n = 32, metastatic n = 39) and 10 healthy donors (HD). USP44 promoter methylation was detected in plasma cell-free DNA by a newly developed highly specific and sensitive real-time MSP method. Our findings indicate that USP44 promoter is methylated in plasma cell-free DNA of metastatic prostate cancer patients and that detection of USP44 promoter methylation is significantly associated with overall survival (OS) (p = 0.008). We report for the first time that detection of USP44 promoter methylation in plasma cell free DNA provides significant prognostic information in metastatic prostate cancer.

Ipatasertib plus abiraterone and prednisolone in metastatic castration-resistant prostate cancer (IPATential150): a multicentre, randomised, double-blind, phase 3 trial.

BACKGROUND: The PI3K/AKT and androgen-receptor pathways are dysregulated in metastatic castration-resistant prostate cancers (mCRPCs); tumours with functional PTEN-loss status have hyperactivated AKT signalling. Dual pathway inhibition with AKT inhibitor ipatasertib plus abiraterone might have greater benefit than abiraterone alone. We aimed to compare ipatasertib plus abiraterone with placebo plus abiraterone in patients with previously untreated mCRPC with or without tumour PTEN loss. METHODS: We did a randomised, double-blind, phase 3 trial at 200 sites across 26 countries or regions. Patients aged 18 years or older with previously untreated asymptomatic or mildly symptomatic mCRPC who had progressive disease and Eastern Collaborative Oncology Group performance status of 0 or 1 were randomly assigned (1:1; permuted block method) to receive ipatasertib (400 mg once daily orally) plus abiraterone (1000 mg once daily orally) and prednisolone (5 mg twice a day orally) or placebo plus abiraterone and prednisolone (with the same dosing schedule). Patients received study treatment until disease progression, intolerable toxicity, withdrawal from the study, or study completion. Stratification factors were previous taxane-based therapy for hormone-sensitive prostate cancer, type of progression, presence of visceral metastasis, and tumour PTEN-loss status by immunohistochemistry. Patients, investigators, and the study sponsor were masked to the treatment allocation. The coprimary endpoints were investigator-assessed radiographical progression-free survival in the PTEN-loss-by-immunohistochemistry population and in the intention-to-treat population. This study is ongoing and is registered with ClinicalTrials.gov, NCT03072238. FINDINGS: Between June 30, 2017, and Jan 17, 2019, 1611 patients were screened for eligibility and 1101 (68%) were enrolled; 554 (50%) were assigned to the placebo-abiraterone group and 547 (50%) to the ipatasertib-abiraterone group. At data cutoff (March 16, 2020), median follow-up duration was 19 months (range 0-33). In the 521 (47%) patients who had tumours with PTEN loss by immunohistochemistry (261 in the placebo-abiraterone group and 260 in the ipatasertib-abiraterone group), median radiographical progression-free survival was 16·5 months (95% CI 13·9-17·0) in the placebo-abiraterone group and 18·5 months (16·3-22·1) in the ipatasertib-abiraterone group (hazard ratio [HR] 0·77 [95% CI 0·61-0·98]; p=0·034; significant at α=0·04). In the intention-to-treat population, median progression-free survival was 16·6 months (95% CI 15·6-19·1) in the placebo-abiraterone group and 19·2 months (16·5-22·3) in the ipatasertib-abiraterone group (HR 0·84 [95% CI 0·71-0·99]; p=0·043; not significant at α=0·01). Grade 3 or higher adverse events occurred in 213 (39%) of 546 patients in the placebo-abiraterone group and in 386 (70%) of 551 patients in the ipatasertib-abiraterone group; adverse events leading to discontinuation of placebo or ipatasertib occurred in 28 (5%) in the placebo-abiraterone group and 116 (21%) in the ipatasertib-abiraterone group. Deaths due to adverse events deemed related to treatment occurred in two patients (<1%; acute myocardial infarction [n=1] and lower respiratory tract infection [n=1]) in the placebo-abiraterone group and in two patients (<1%; hyperglycaemia [n=1] and chemical pneumonitis [n=1]) in the ipastasertb-abiraterone group. INTERPRETATION: Ipatasertib plus abiraterone significantly improved radiographical progression-free survival compared with placebo plus abiraterone among patients with mCRPC with PTEN-loss tumours, but there was no significant difference between the groups in the intention-to-treat population. Adverse events were consistent with the known safety profiles of each agent. These data suggest that combined AKT and androgen-receptor signalling pathway inhibition with ipatasertib and abiraterone is a potential treatment for men with PTEN-loss mCRPC, a population with a poor prognosis. FUNDING: F Hoffmann-La Roche and Genentech.

Prognostic Significance of Gene Expression and DNA Methylation Markers in Circulating Tumor Cells and Paired Plasma Derived Exosomes in Metastatic Castration Resistant Prostate Cancer.

Liquid biopsy, based on the analysis of circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA), provides non-invasive real-time monitoring of tumor evolution and therapeutic efficacy. We performed for the first time a direct comparison study on gene expression and DNA methylation markers in CTCs and paired plasma-derived exosomes and evaluated their prognostic significance in metastatic castration resistant prostate cancer. This prospective liquid biopsy (LB) study was based on a group of 62 metastatic castration resistant prostate cancer (mCRPC) patients and 10 healthy donors (HD) as controls. Identical blood draws were used to: (a) enumerate CTC and tumor-derived extracellular vesicles (tdEVs) using CellSearch (CS) and (b) analyze CTCs and paired plasma-derived exosomes at the gene expression and DNA methylation level. CTCs were enumerated using CellSearch in 57/62 patients, with values ranging from 5 to 854 cells/7.5 mL PB. Our results revealed for the first time a significantly higher positivity of gene expression markers (CK-8, CK-18, TWIST1, PSMA, AR-FL, AR-V7, AR-567 and PD-L1 mRNA) in EpCAM-positive CTCs compared to plasma-derived exosomes. GSTP1, RASSF1A and SCHLAFEN were methylated both in CTC and exosomes. In CTCs, Kaplan-Meier analysis revealed that CK-19 (p = 0.009), PSMA (p = 0.001), TWIST1 (p = 0.001) expression and GSTP1 (p = 0.001) methylation were correlated with OS, while in exosomes GSTP1 (p = 0.007) and RASSF1A (p = 0.001) methylation was correlated with OS. Our direct comparison study of CTCs and exosomes at gene expression and DNA methylation level, revealed for the first time a significantly higher positivity in EpCAM-positive CTCs compared to plasma-derived exosomes. Future perspective of this study should be the evaluation of clinical utility of molecular biomarkers in CTCs and exosomes on independent multicentric cohorts with mCRPC patients.

Real-World Data on Thromboprophylaxis in Active Cancer Patients: Where Are We? Are We Getting There?

Background: Cancer patients are at high risk for cancer-associated thrombosis (CAT). CAT is the second leading cause of death in these patients but it can be preventable with thromboprophylaxis. Patients and Methods: An observational, prospective, multicenter study aiming to record CAT management in clinical practice was conducted by the Hellenic Society of Medical Oncology (HeSMO). Results: A total of 426 active cancer patients (mean age 65.3 years, mean BMI: 26.1 kg/m2) who received thromboprophylaxis, were included from 18 oncology units. Tumor types were lung 25.1%, pancreas 13.9%, breast 8.7%, stomach 8.5%, ovarian 7.8%, and others 36%, while 69% had metastases. A total of 71% had a Khorana score ≤2 and 61% received High Thrombotic Risk Chemotherapy Agents (HTRCAs, e.g., platinum). For thromboprophylaxis patients received mainly Low Molecular Weight Heparins (LMWHs), on higher than prophylactic doses in 50% of cases. Overall, 16 (3.8%) thrombotic events and 6 (1.4%) bleeding events were recorded. Notably, patients on higher doses of LMWHs compared to patients who received standard prophylactic doses had 70% lower odds to develop thrombotic events (OR: 0.3, 95% CI: 0.10-1.0, p = 0.04). Conclusion: CAT is an important issue in oncology. Along with the Khorana score, factors as metastasis and use of HTRCAs should also be taken into consideration. Thromboprophylaxis for active cancer patients with LMWHs, even on higher doses is safe and efficient.

PIM-1 Is Overexpressed at a High Frequency in Circulating Tumor Cells from Metastatic Castration-Resistant Prostate Cancer Patients.

PIM-1 is an oncogene involved in cell cycle progression, cell growth, cell survival and therapy resistance, activated in many types of cancer, and is now considered as a very promising target for cancer therapy. We report for the first time that PIM-1 is overexpressed in circulating tumor cells (CTCs) from metastatic castration-resistant prostate cancer patients (mCRPC). We first developed and validated a highly sensitive RT-qPCR assay for quantification of PIM-1 transcripts. We further applied this assay to study PIM-1 expression in EpCAM(+) CTC fraction isolated from 64 peripheral blood samples of 50 mCRPC patients. CTC enumeration in all samples was performed using the FDA-cleared CellSearch® system. PIM-1 overexpression was detected in 24/64 (37.5%) cases, while in 20/24 (83.3%) cases that were positive for PIM-1 expression, at least one CTC/7.5 mL PB was detected in the CellSearch®. Our data indicate that PIM-1 overexpression is observed at high frequency in CTCs from mCRPC patients and this finding, in combination with androgen receptor splice variant 7 (AR-V7) expression in CTCs, suggest its potential role as a very promising target for cancer therapy. We strongly believe that PIM-1 overexpression in EpCAM(+) CTC fraction merits to be further evaluated and validated as a non-invasive circulating tumor biomarker in a large and well-defined patient cohort with mCRPC.

Clinical activity of a htert (vx-001) cancer vaccine as post-chemotherapy maintenance immunotherapy in patients with stage IV non-small cell lung cancer: final results of a randomised phase 2 clinical trial.

BACKGROUND: The cancer vaccine Vx-001, which targets the universal tumour antigen TElomerase Reverse Transcriptase (TERT), can mount specific Vx-001/TERT572 CD8 + cytotoxic T cells; this immune response is associated with improved overall survival (OS) in patients with advanced/metastatic non-small cell lung cancer (NSCLC). METHODS: A randomised, double blind, phase 2b trial, in HLA-A*201-positive patients with metastatic, TERT-expressing NSCLC, who did not progress after first-line platinum-based chemotherapy were randomised to receive either Vx-001 or placebo. The primary endpoint of the trial was OS. RESULTS: Two hundred and twenty-one patients were randomised and 190 (101 and 89 patients in the placebo and the Vx-001 arm, respectively) were analysed for efficacy. There was not treatment-related toxicity >grade 2. The study did not meet its primary endpoint (median OS 11.3 and 14.3 months for the placebo and the Vx-001, respectively; p = 0.86) whereas the median Time to Treatment Failure (TTF) was 3.5 and 3.6 months, respectively. Disease control for >6months was observed in 30 (33.7%) and 26 (25.7%) patients treated with Vx-001 and placebo, respectively. There was no documented objective CR or PR. Long lasting TERT-specific immune response was observed in 29.2% of vaccinated patients who experienced a significantly longer OS compared to non-responders (21.3 and 13.4 months, respectively; p = 0.004). CONCLUSION: Vx-001 could induce specific CD8+ immune response but failed to meet its primary endpoint. Subsequent studies have to be focused on the identification and treatment of subgroups of patients able to mount an effective immunological response to Vx-001. CLINICAL TRIAL REGISTRATION: NCT01935154.

Management of Cancer-associated Thrombosis (CAT): Symptomatic or Incidental.

BACKGROUND: Cancer-associated thrombosis (CAT), the second leading cause of death in patients with cancer can be treated with low molecular weight heparin (LMWH) according to guidelines. PATIENTS AND METHODS: A multicenter prospective observational study was carried out to record anti-thrombotic treatment practice, assess thrombosis recurrence and bleeding, and identify potential risk factors. Adult patients from 18 Oncology Departments throughout Greece were followed-up for 12 months. RESULTS: A total of 120 patients with CAT receiving anticoagulant treatment were enrolled (35% incidental); 85% were treated for more than 6 months, 95.8% were treated with tinzaparin and smaller percentages with other agents. Thrombosis recurred in three patients and there was minor bleeding in four patients. Bleeding was associated with high body mass index (>35 kg/m2), trauma history, renal insufficiency and bevacizumab use. CONCLUSION: Incidental thrombosis contributes significantly to CAT burden. Long-term use of LMWH seems to be effective and safe. Several risk factors associated with bleeding should be considered during anti-coagulation therapy planning.

Survival Outcomes From a Cumulative Analysis of Worldwide Observational Studies on Sequential Use of New Agents in Metastatic Castration-Resistant Prostate Cancer.

INTRODUCTION: The sequential use of a number of new agents (NAs) have improved the overall survival (OS) of patients with metastatic castration-resistant prostate cancer whose disease progresses after docetaxel (DOC) treatment. The aim of this study was to assess the cumulative survival outcomes of different sequencing strategies by evaluating the individual data from published studies of patients treated with a post-DOC treatment sequence of 2 NAs. PATIENTS AND METHODS: The patients' individual data were analyzed to investigate whether different sequencing strategies lead to differences in OS. RESULTS: We analyzed the data of 1099 evaluable patients. Among the patients treated with a second-line new hormone agent (NHA), median OS from the start of third-line treatment was significantly longer in the patients treated with cabazitaxel (CABA) than in those treated with abiraterone acetate or enzalutamide. Median cumulative OS (cumOS) from the start of second-line treatment was 21.1 months in the patients who received NHA then NHA, 22.1 months in those who received NHA then CABA, and 21.0 months in those who received CABA then NHA. Among the patients with a second-line progression-free survival of ≥6 months, median cumOS was significantly longer in patients who received CABA-including sequences than in those treated with NHA then NHA sequences (29.5 vs. 24.8 months; P = .03). CONCLUSION: Our findings suggest that the sequential use of NAs with different mechanisms of action improves cumOS regardless of the order in which they are administered, thus supporting the hypothesis of cross-resistance between the 2 NHAs.

Expression pattern of androgen receptors, AR-V7 and AR-567es, in circulating tumor cells and paired plasma-derived extracellular vesicles in metastatic castration resistant prostate cancer.

Androgen-receptor splice variant 7 (AR-V7) is a highly promising liquid biopsy predictive biomarker showing primary or acquired resistance to novel androgen receptor signaling inhibitors in metastatic castration resistant prostate cancer (mCRPC). We present for the first time the expression pattern of AR-FL, AR-V7, and AR-567es at a quantitative level in circulating tumor cells (CTCs) and paired plasma-derived extracellular vesicles in mCRPC. We first developed and analytically validated a novel multiplex RT-qPCR assay for AR full length (AR-FL), AR-V7, AR-567es and AR-total. We then quantified the expression levels of AR-splice variants, CK-19 (epithelial marker) and B2M (reference gene) in EpCAM+ CTCs, and paired plasma-derived extracellular vesicles isolated from peripheral blood (20 mL) of 62 mCRPC patients and 10 healthy donors. CTCs were enumerated using the FDA-cleared CellSearch® system. In CTCs AR-FL was detected in 64/69 (92.3%), AR-V7 in 34/69 (49.3%), AR-567es in 16/69 (23.2%) and AR-total in 62/69 (89.9%). In 52 out of 69 samples, paired plasma-derived extracellular vesicles were analyzed. AR-FL was detected in 40/52 (76.9%), AR-V7 in 4/52 (7.7%), AR-567 in 2/52 (3.8%) and AR total in 39/52 (75.0%). In all cases AR splice variants were expressed in higher levels in CTCs than in paired extracellular vesicles, while AR-V7 was detected in higher percentages than in AR-567es. Using CellSearch®, CTCs were detected in 52/69 (75.4%) mCRPC patient samples; 27/52 (51.9%) of these samples were CTC+/AR-V7+ and 14/52 (26.9%) were CTC+/AR-567es+, while 7/17 (41.2%) were CTC-/AR-V7+ and 2/17 (11.8%) were CTC-/AR-567es+. Our results reveal for the first time a remarkable heterogeneity in the expression levels of AR-FL, AR-V7 and AR-567es in EpCAM+ CTCs and paired extracellular vesicles between individual mCRPC patients. The clinical significance of this finding will be further investigated in a large patient cohort with respect to therapy response.

Prognostic and predictive factors in patients with metastatic or recurrent cervical cancer treated with platinum-based chemotherapy.

BACKGROUND: Recognizing resistance or susceptibility to the current standard cisplatin and paclitaxel treatment could improve therapeutic outcomes of metastatic or recurrent cervical cancer. METHODS: Forty-five tissue samples from patients participating in a phase II trial of cisplatin and ifosfamide, with or without paclitaxel were collected for retrograde analysis. Immunohistochemistry and genotyping was performed to test ERCC1, III ß-tubulin, COX-2, CD4, CD8 and ERCC1 (C8092A and N118 N) and MDR1 (C3435T and G2677 T) gene polymorphisms, as possible predictive and prognostic markers. Results were statistically analyzed and correlated with patient characteristics and outcomes. RESULTS: Patients with higher levels of ERCC1 expression had shorter PFS and OS than patients with low ERCC1 expression (mPFS:5.1 vs 10.2 months, p = 0.027; mOS:10.5 vs. 21.4 months, p = 0.006). Patients with TT in the site of ERCC1 N118 N and GT in the site of MDR1 G2677 T polymorphisms had significantly longer PFS (p = 0.006 and p = 0.027 respectively). ERCC1 expression and the ERCC1 N118 N polymorphism remained independent predictors of PFS. Interestingly, high III beta tubulin expression was associated with chemotherapy resistance and fewer responses [5/20 (25%)] compared to lower III ß-tubulin expression [15/23 (65.2%)] (p = 0.008). Finally, ΙΙΙ ß-tubulin levels and chemotherapy regimen were independent predictors of response to treatment. CONCLUSIONS: ERCC1 expression proved to be a significant prognostic factor for survival in our metastatic or recurrent cervical cancer population treated with cisplatin based chemotherapy. ERCC1 N118 N and MDR1 G2677 T polymorphism also proved of prognostic significance for disease progression, while overexpression of III ß-tubulin was positively correlated with chemotherapy resistance.

Immune response gene expression in colorectal cancer carries distinct prognostic implications according to tissue, stage and site: a prospective retrospective translational study in the context of a hellenic cooperative oncology group randomised trial.

BACKGROUND: Although host immune response is an emerging prognostic factor for colorectal cancer, there is no consensus on the optimal methodology, surrogate markers or tissue for study. PATIENTS AND METHODS: Tumour blocks were prospectively collected from 344 patients with stage II/III colorectal cancer (CRC) treated with adjuvant chemotherapy. Whole section lymphocytic infiltration was studied along with mRNA expression of CD3Z, CD8, CD4, CXCL9, CXCL13, IGHM, FOXP3, SNAI2 and ESR1 by qRT-qPCR in tissue microarray (TMA) cores from the centre of tumour, invasive margin and adjacent normal mucosa. RESULTS: Lymphocytic infiltration, deficient MMR (10.9%), KRAS (40.7%) and BRAF (4.9%) mutations or single mRNA gene expression were not prognostic. Tumour ESR1 gene expression (Hazard Ratio [HR] for relapse 2.33, 95% CI 1.35-4.02; HR for death 1.74, 95% CI 1.02-2.97) and absence of necrosis (HR for relapse 1.71, 95% CI 1.05-2.71; HR for death 1.98, 95% CI 1.14-3.43) were adverse prognostic features. We used CD3Z and CD8 expression in order to devise the mRNA-based Immune Score (mIS) and proceeded to partitioning analysis in 267 patients, with age, stage, tumour site (Right vs Left CRC), KRAS mutation and tumour mIS as input factors. Only in patients with stage III right-sided colon cancer, a low immune response was associated with inferior disease-free survival (mIS-low, HR for relapse 2.28, 95% CI 1.05-8.02). No prognostic significance was seen for tumour mIS in any other stage or site of CRC, or for a similar mIS score derived from adjacent normal mucosa. Independent adverse prognostic significance was retained in multivariable analysis for absence of necrosis, tumour ESR1 expression in all patients and low tumour mIS in stage III right-sided CRC. CONCLUSIONS: In localised CRC, mRNA-based CD3Z/CD8 profiling of tumour immune response may have stage, site and tissue-specific prognostic significance, along with ESR1 expression. TRIAL REGISTRATION: ANZCTR.org.au ACTRN12610000509066.

Do elderly patients with non-small cell lung cancer get the best out of recent advances in first-line treatment? A comparative study in two tertiary cancer centers in Greece.

BACKGROUND: Elderly patients with advanced non-small cell lung cancer (NSCLC) are thought to receive suboptimal treatment mainly due to concerns for poor compliance and/or excessive toxicity. PATIENTS AND METHODS: Using the age of 70 years as the pre-defined cut-off, we compared elderly patients with advanced NSCLC suitable for first line chemotherapy with their younger counterparts in terms of: i) diagnosis and disease characteristics ii) adherence to treatment schedule, including dose intensity (DI), and relative dose intensity (RDI), iii) toxicity, tolerance, and efficacy outcomes. RESULTS: Among 292 eligible patients, data were available for 245, of whom 107 (43.7%) belonged to the elderly group. This group was more likely to present with co-morbidities, non-smoking current status and diagnosis based on cytology alone. As compared to the non-elderly, elderly patients were more likely to receive single-agent therapy (8.0% vs. 29.2% respectively, p < 0.001) and less likely to receive platinum-based chemotherapy (80.3% vs. 57.9%, p < 0.001). Elderly patients also received docetaxel (24.3% vs. 40.4%), and bevacizumab (7.5% vs. 21.3%) significantly less often and received oral vinorelbine (24.3% vs. 11.8%) more frequently. Non-elderly patients were more likely to receive any of the cytotoxic drugs with RDI > 0.8 (49.6% vs. 33.0%, p = 0.012) and RDI > 0.9 (29.6% vs. 16%, p = 0.015). Substantial toxicity, as well as median overall survival did not differ significantly between the two groups. CONCLUSIONS: Only one third of the elderly patients received at least 80% of the scheduled treatment intensity. Nearly half received diagnosis based on cytology alone, which may deprive them from new, histology-driven, therapeutic approaches.

The clinical and pathological significance of RCAS1 expression as a prognostic biomarker in non-small cell lung cancer.

BACKGROUND: Lung cancer remains a major health problem due to its incidence and mortality. Receptor-binding cancer antigen expressed on SiSo cells (RCAS1) is a protein that can be expressed in cancer cells and is involved in tumor cell escape from immune system surveillance. AIM: The aim of this study was to evaluate the clinical significance of immunohistochemical staining for RCAS1 in non-small cell lung cancer (NSCLC). MATERIALS AND METHODS: Tissue microarrays of tumor specimens from 112 consecutive patients with newly diagnosed primary NSCLC were constructed. RCAS1 and Ki-67 immunohistochemistry were studied through computerized image analysis. Associations between RCAS1 and Ki-67 expression and clinico-pathological variables and survival were analyzed. RESULTS: RCAS1 expression was higher in grade III tumors (p=0.009), regardless of the histological type, and in adenocarcinomas with lymphovascular invasion (p=0.014). A positive correlation between RCAS1 and Ki-67 levels was observed (p=0.002). Moreover, there was an inverse correlation of overall survival with RCAS1 (hazard ratio=0.99, p<0.001) and Ki-67 (hazard ratio=1.05, p=0.003) levels. Particularly, patients with higher expression of RCAS1 or Ki-67 had a significantly shorter survival than those with lower expression. CONCLUSION: RCAS1 could be a useful immunohistochemical biomarker, indicating not only tumor aggressiveness but also a poorer prognosis for patients with NSCLC.

Risk factors, pathological and phenotypic features of male breast cancer in Greece.

BACKGROUND: Breast cancer (BC) in males is a rare disease and comprises 0.5-1% of all BC cases. Due to its rarity, there are limited data regarding risk factors, biology and relevant treatment. AIM: A prospective observational study of demographic, clinical and histological characteristics of serially-admitted men with breast cancer was carried out from 1999 to 2009. PATIENTS AND METHODS: Data were recorded and analyzed from a database including 1,315 cases of BC. Registered data concerned age, initial presentation, family and lifestyle history (risk factors), histological features, phenotypic subtypes and TNM staging. RESULTS: Twenty two men with BC were identified, with a median age of 63 years. The most common initial presentation was a palpable lump in 12 patients, nipple contraction in three and ulceration in three. According to their medical history, nine men were overweight, 10 suffered from hypertension and 12 were smokers. The most prevalent phenotype was luminal-A followed by triple-negative type. BC in none of the cases was HER 2-amplified. The majority of cases were grade II or III and stage II or III. CONCLUSION: In the present small study, we confirm that BC in males is rare. It is a disease of middle-age and presents at advanced stages. Most of patients had 1-3 risk factors for BC. Expression of hormonal receptors occurs in the majority of BC tumors in males and with rarity in HER 2 amplification.

Submerged goiter proven to be metastatic infiltration of a neuro-endocrine Merkel cell carcinoma.

BACKGROUND: Merkel cell carcinoma (MCC) is an uncommon neuroendocrine cutaneous carcinoma. Metastases to the thyroid gland are rare and may present diagnostic difficulties. CASE PRESENTATION: A 73-year-old woman presented with a hard mass in the adipose tissue of the right inguinal area. This mass was surgically excised and the histology examination showed the existence of a MCC. CT scans revealed a sizable lesion with imaging features of a submerged goiter, invasive to the upper mediastinum. The patient received chemotherapy following by locoregional radiotherapy at the bed of the excised lesion. During the next 10 months the patient was asymptomatic, serum markers values were normal and CT scans findings were stable. However, afterwards NSE and chromogranin values raised and CT scans revealed an enlargement of the submerged goiter. The patient became symptomatic, mainly experiencing respiratory inconvenience. Surgical excision of the right lobe of the thyroid gland was decided and performed without any complications. The histopathology examination showed infiltration of the thyroid gland by a neuroendocrine carcinoma with characteristics compatible with MCC. CONCLUSIONS: The rare case of metastatic infiltration of the thyroid gland by a MCC based on histological and immunohistochemical findings was described. This case report is of clinical significance indicating that by any abnormal finding in the thyroid gland in patients with a malignant disease, the diagnostic approach should always contain consideration of metastasis from the primary tumor.

Ephrin (Eph) receptor A1, A4, A5 and A7 expression in human non-small cell lung carcinoma: associations with clinicopathological parameters, tumor proliferative capacity and patients' survival.

BACKGROUND: Ephrin (Eph) receptors are frequently overexpressed in a wide variety of human malignant tumors, being associated with tumor growth, invasion, metastasis and angiogenesis. The present study aimed to evaluate the clinical significance of EphA1, A4, A5 and A7 protein expression in non-small cell lung carcinoma (NSCLC). METHODS: EphA1, A4, A5 and A7 protein expression was assessed immunohistochemically in tissue microarrays of 88 surgically resected NSCLC and was analyzed in relation with clinicopathological characteristics and patients' survival. RESULTS: Elevated EphA4 expression was significantly associated with low histopathological stage and presence of inflammation (p = 0.047 and p = 0.026, respectively). Elevated EphA7 expression was significantly associated with older patients' age, presence of fibrosis and smaller tumor size (p = 0.036, p = 0.029 and p = 0.018, respectively). EphA1, A5 and A7 expression were positively associated with tumor proliferative capacity (p = 0.047, p = 0.002 and p = 0.046, respectively). Elevated EphA4, A5 and A7 expression were identified as predictors of favourable patients' survival at both univariate (Log-rank test, 0 = 0.019, p = 0.006 and p = 0.012, respectively) and multivariate levels (Cox-regression analysis, p = 0.029, p = 0.068 and p = 0.044, respectively). CONCLUSIONS: The present study supported evidence that Ephs may be involved in lung cancer progression, reinforcing their utility as clinical biomarkers for patients' management and prognosis, as also as potential targets for future therapeutic interventions.

Prevalence of BRCA1 mutations among 403 women with triple-negative breast cancer: implications for genetic screening selection criteria: a Hellenic Cooperative Oncology Group Study.

In spite the close association of the triple-negative breast cancer immunophenotype with hereditary breast cancers and the BRCA1 pathway, there is a lack of population studies that determine the frequency of BRCA1 mutations among triple-negative breast cancer patients. To address this, we have screened a large sample of 403 women diagnosed with triple-negative invasive breast cancer, independently of their age or family history, for germline BRCA1 mutations. Median age at diagnosis was 50 years (range 20-83). The overall prevalence of triple-negative cases among the initial patient group with invasive breast cancer was 8%. BRCA1 was screened by direct DNA sequencing in all patients, including all exons where a mutation was previously found in the Greek population (exons 5, 11, 12, 16, 20, 21, 22, 23, 24-77% of the BRCA1 coding region), including diagnostic PCRs to detect the three Greek founder large genomic rearrangements. Sixty-five deleterious BRCA1 mutations were identified among the 403 triple-negative breast cancer patients (16%). Median age of onset for mutation carriers was 39 years. Among a total of 106 women with early-onset triple-negative breast cancer (<40 years), 38 (36%) had a BRCA1 mutation, while 27% of women with triple-negative breast cancer diagnosed before 50 years (56/208) had a BRCA1 mutation. A mutation was found in 48% (50/105) of the triple-negative breast cancer patients with family history of breast or ovarian cancer. It is noteworthy, however, that of the 65 carriers, 15 (23%) had no reported family history of related cancers. All but one of the carriers had grade III tumors (98%). These results indicate that women with early-onset triple-negative breast cancer, and ideally all triple-negative breast cancer patients, are candidates for BRCA1 genetic testing even in the absence of a family history of breast or ovarian cancer.