Prevalence of Helicobacter pylori infection among patients with esophageal carcinoma.

BACKGROUND: Helicobacter pylori (H. pylori) is a widespread microorganism related to gastric adenocarcinoma (AC). In contrast, it has been reported that an inverse association exists between H. pylori infection and esophageal carcinoma. The mechanisms underlying this supposedly protective effect remain controversial. AIM: To determine the prevalence of H. pylori infection in esophageal carcinoma patients, we performed a retrospective observational study of esophageal tumors diagnosed in our hospital. METHODS: We retrospectively reviewed the prevalence of H. pylori infection in a cohort of patients diagnosed with esophageal carcinoma. Concomitant or previous proton pump inhibitor (PPI) usage was also recorded. RESULTS: A total of 89 patients with esophageal carcinoma (69 males, 77.5%), with a mean age of 66 years (range, 26-93 years) were included. AC was the most frequent pathological variant (n = 47, 52.8%), followed by squamous cell carcinoma (n = 37, 41.6%). Fourteen ACs (29.8%) originated in the gastroesophageal junction and 33 (70.2%) in the esophageal body. Overall, 54 patients (60.7%) presented at stages III and IV. Previous H. pylori infection occurred only in 4 patients (4.5%), 3 with AC (6.3% of all ACs) and 1 with squamous cell carcinoma (2.7% of all squamous cell tumors). All patients with previous H. pylori infection had stage III-IV. Only one patient had received prior H. pylori eradication therapy, whereas 86 (96.6%) had received previous or concomitant PPI treatment. CONCLUSION: In our cohort of patients, and after histologic evaluation of paraffin-embedded primary tumors, we found a very low prevalence of previous H. pylori infection. We also reviewed the medical history of the patients, concluding that the majority had received or were on PPI treatment. The minimal prevalence of H. pylori infection found in this cohort of patients with esophageal carcinoma suggests a protective role.

Clinical features and lipid metabolism genes as potential biomarkers in advanced lung cancer.

BACKGROUND: Lung cancer is one of the most lethal tumors with a poor survival rate even in those patients receiving new therapies. Metabolism is considered one of the hallmarks in carcinogenesis and lipid metabolism is emerging as a significant contributor to tumor metabolic reprogramming. We previously described a profile of some lipid metabolism related genes with potential prognostic value in advanced lung cancer. AIM: To analyze clinical and pathological characteristics related to a specific metabolic lipid genomic signature from patients with advanced lung cancer and to define differential outcome. METHODS: Ninety samples from NSCLC (non-small cell lung cancer) and 61 from SCLC (small cell lung cancer) patients were obtained. We performed a survival analysis based on lipid metabolic genes expression and clinical characteristics. The primary end point of the study was the correlation between gene expression, clinical characteristics and survival. RESULTS: Clinical variables associated with overall survival (OS) in NSCLC patients were clinical stage, adenocarcinoma histology, Eastern Cooperative Oncology Group (ECOG), number and site of metastasis, plasma albumin levels and first-line treatment with platinum. As for SCLC patients, clinical variables that impacted OS were ECOG, number of metastasis locations, second-line treatment administration and Diabetes Mellitus (DM). None of them was associated with gene expression, indicating that alterations in lipid metabolism are independent molecular variables providing complementary information of lung cancer patient outcome. CONCLUSIONS: Specific clinical features as well as the expression of lipid metabolism-related genes might be potential biomarkers with differential outcomes.

Brain metastases and lung cancer: molecular biology, natural history, prediction of response and efficacy of immunotherapy.

Brain metastases stemming from lung cancer represent a common and challenging complication that significantly impacts patients' overall health. The migration of these cancerous cells from lung lesions to the central nervous system is facilitated by diverse molecular changes and a specific environment that supports their affinity for neural tissues. The advent of immunotherapy and its varied combinations in non-small cell lung cancer has notably improved patient survival rates, even in cases involving brain metastases. These therapies exhibit enhanced penetration into the central nervous system compared to traditional chemotherapy. This review outlines the molecular mechanisms underlying the development of brain metastases in lung cancer and explores the efficacy of novel immunotherapy approaches and their combinations.

Phenolic diterpenes from Rosemary supercritical extract inhibit non-small cell lung cancer lipid metabolism and synergise with therapeutic drugs in the clinic.

Lung cancer is one of the most deadly and common cancers in the world. The molecular features of patient's tumours dictate the different therapeutic decisions, which combines targeted therapy, chemotherapy, and immunotherapy. Altered cellular metabolism is one of the hallmarks of cancer. Tumour cells reprogram their metabolism to adapt to their novel requirements of growth, proliferation, and survival. Together with the Warburg effect, the role of lipid metabolism alterations in cancer development and prognosis has been highlighted. Several lipid related genes have been shown to promote transformation and progression of cancer cells and have been proposed as biomarkers for prognosis. Nevertheless, the exact mechanisms of the regulation of lipid metabolism and the biological consequences in non-small cell lung cancer (NSCLC) have not been elucidated yet. There is an urgent necessity to develop multidisciplinary and complementary strategies to improve NSCLC patients´ well-being and treatment response. Nutrients can directly affect fundamental cellular processes and some diet-derived ingredients, bioactive natural compounds and natural extracts have been shown to inhibit the tumour growth in preclinical and clinical trials. Previously, we described a supercritical extract of rosemary (SFRE) (12 - 16% composition of phenolic diterpenes carnosic acid and carnosol) as a potential antitumoral agent in colon and breast cancer due to its effects on the inhibition of lipid metabolism and DNA synthesis, and in the reduction of resistance to 5-FluoroUracil (5-FU). Herein, we demonstrate SFRE inhibits NSCLC cell bioenergetics identifying several lipid metabolism implicated targets. Moreover, SFRE synergises with standard therapeutic drugs used in the clinic, such as cisplatin, pemetrexed and pembrolizumab to inhibit of cell viability of NSCLC cells. Importantly, the clinical relevance of SFRE as a complement in the treatment of NSCLC patients is suggested based on the results of a pilot clinical trial where SFRE formulated with bioactive lipids (PCT/ES2017/070263) diminishes metabolic and inflammatory targets in peripheral-blood mononuclear cells (PBMC), such as MAPK (p=0.04), NLRP3 (p=0.044), and SREBF1 (p=0.047), which may augment the immune antitumour function. Based on these results, SFRE merits further investigation as a co-adjuvant in the treatment of NSCLC. Clinical trial registration: ClinicalTrials.gov Identifier NCT05080920.

Ewing Sarcoma Meets Epigenetics, Immunology and Nanomedicine: Moving Forward into Novel Therapeutic Strategies.

Ewing Sarcoma (EWS) is an aggressive bone and soft tissue tumor that mainly affects children, adolescents, and young adults. The standard therapy, including chemotherapy, surgery, and radiotherapy, has substantially improved the survival of EWS patients with localized disease. Unfortunately, this multimodal treatment remains elusive in clinics for those patients with recurrent or metastatic disease who have an unfavorable prognosis. Consistently, there is an urgent need to find new strategies for patients that fail to respond to standard therapies. In this regard, in the last decade, treatments targeting epigenetic dependencies in tumor cells and the immune system have emerged into the clinical scenario. Additionally, recent advances in nanomedicine provide novel delivery drug systems, which may address challenges such as side effects and toxicity. Therefore, therapeutic strategies stemming from epigenetics, immunology, and nanomedicine yield promising alternatives for treating these patients. In this review, we highlight the most relevant EWS preclinical and clinical studies in epigenetics, immunotherapy, and nanotherapy conducted in the last five years.

Multidisciplinary Consensus on the Prevention and Treatment of Osteoporosis and Fragility Fractures in Patients with Prostate Cancer Receiving Androgen-Deprivation Therapy.

Patients with prostate cancer (PCa) on androgen-deprivation therapy (ADT) are at high risk of osteoporosis and fragility fractures. We aimed to provide some practical insights into the delivery of optimal bone health care for PCa patients, particularly those on ADT. An interdisciplinary group of experts, including urologists and rheumatologists developed recommendations based on their expertise, current evidence and guidelines. The multidisciplinary group's main recommendations are: fragility fracture risk should be assessed in all PCa patient, especially, in those under ADT. FRAX® tool may be incorporated into clinical practice to identify patients at high risk of fracture. Bone mineral density (BMD) should be measured routinely by dual energy X-ray absorptiometry in all patients scheduled for or on ADT. Thoracic and lumbar spine X-ray may be performed at the initial evaluation of patients with the diagnosis of osteoporosis and in case of suspected clinical vertebral fracture. Basic laboratory tests are recommended to exclude secondary osteoporosis. Treatment with bisphosphonates or denosumab should be considered in patients on ADT with fragility fracture, osteoporosis (BMD T-score ≤-2.5), or high risk of fracture according to FRAX®. Referral to a bone metabolism specialist should be contemplated in some cases. The recommendations provided in this document, tailored for clinicians treating PCa patients, may be of help to identify and treat patients at high risk of fracture.

A Multicenter Analysis of the Outcome of Cancer Patients with Neutropenia and COVID-19 Optionally Treated with Granulocyte-Colony Stimulating Factor (G-CSF): A Comparative Analysis.

BACKGROUND: Approximately 15% of patients infected by SARS-CoV-2 develop a distress syndrome secondary to a host hyperinflammatory response induced by a cytokine storm. Myelosuppression is associated with a higher risk of infections and mortality. There are data to support methods of management for neutropenia and COVID-19. We present a multicenter experience during the first COVID-19 outbreak in neutropenic cancer patients infected by SARS-CoV-2. METHODS: Clinical retrospective data were collected from neutropenic cancer patients with COVID-19. Comorbidities, tumor type, stage, treatment, neutropenia severity, G-CSF, COVID-19 parameters, and mortality were analyzed. A bivariate analysis of the impact on mortality was carried out. Additionally, we performed a multivariable logistic regression to predict respiratory failure and death. RESULTS: Among the 943 cancer patients screened, 83 patients (11.3%) simultaneously had neutropenia and an infection with COVID-19. The lungs (26%) and breasts (22%) were the primary locations affected, and most patients had advanced disease (67%). In the logistic model, as adjusted covariates, sex, age, treatment (palliative vs. curative), tumor type, and the lowest level of neutrophils were used. A significant effect was obtained for the number of days of G-CSF treatment (OR = 1.4, 95% CI [1,1,03,92], p-value = 0.01). CONCLUSIONS: Our findings suggest that a prolonged G-CSF treatment could be disadvantageous for these cancer patients with infections by COVID-19, with a higher probability of worse outcome.

RAS Mutational Status in Advanced Colorectal Adenocarcinoma Treated With Anti-angiogenics: Preliminary Experience With Liquid Biopsy.

AIM: To determinate molecular changes in the downstream epidermal growth factor receptor signaling pathway using serial liquid biopsies in patients with metastatic colorectal tumors (mCRC) under anti-angiogenic treatment. PATIENTS AND METHODS: Determination of RAS mutation in primary tissue samples from colorectal tumors was performed in the 23 patients included in the study at diagnosis using quantitative-polymerase chain reaction. Sequential mutations were studied in circulating tumor (ct) DNA obtained from plasma samples. RESULTS: Twenty-three patients with RAS-mutated primary tumors were included. In the first ctDNA determination, 17 of these patients were found to have wild-type RAS status. Remarkably, three out of these 17 wild-type cases changed to RAS-mutated in subsequent ctDNA assays. CONCLUSION: Serial liquid biopsies in patients with mCRC might be a useful tool for identifying changes in the RAS mutation status in patients who had undergone previous anti-angiogenic therapy. The understanding of these changes might help to better define the landscape of mCRC and be the path to future randomized studies.

Metabolic Health Together with a Lipid Genetic Risk Score Predicts Survival of Small Cell Lung Cancer Patients.

Small cell lung cancer (SCLC) prognosis is the poorest of all types of lung cancer. Its clinical management remains heterogeneous and therefore, the capability to predict survival would be of great clinical value. Metabolic health (MH) status and lipid metabolism are two relevant factors in cancer prevention and prognosis. Nevertheless, their contributions in SCLC outcome have not yet been analyzed. We analyzed MH status and a transcriptomic panel of lipid metabolism genes in SCLC patients, and we developed a predictive genetic risk score (GRS). MH and two lipid metabolism genes, racemase and perilipin 1, are biomarkers of SCLC survival (HR = 1.99 (CI95%: 1.11-3.61) p = 0.02, HR = 0.36 (CI95%: 0.19-0.67), p = 0.03 and HR = 0.21 (CI95%: 0.09-0.47), respectively). Importantly, a lipid GRS of these genes predict better survival (c-index = 0.691). Finally, in a Cox multivariate regression model, MH, lipid GRS and smoking history are the main predictors of SCLC survival (c-index = 0.702). Our results indicate that the control of MH, lipid gene expression and environmental factors associated with lifestyle is crucial for increased SCLC survival. Here, we propose for the first time, a metabolic precision approach for SCLC patients.

Metabolic enzyme ACSL3 is a prognostic biomarker and correlates with anticancer effectiveness of statins in non-small cell lung cancer.

Lung cancer is one of the most common cancers, still characterized by high mortality rates. As lipid metabolism contributes to cancer metabolic reprogramming, several lipid metabolism genes are considered prognostic biomarkers of cancer. Statins are a class of lipid-lowering compounds used in treatment of cardiovascular disease that are currently studied for their antitumor effects. However, their exact mechanism of action and specific conditions in which they should be administered remains unclear. Here, we found that simvastatin treatment effectively promoted antiproliferative effects and modulated lipid metabolism-related pathways in non-small cell lung cancer (NSCLC) cells and that the antiproliferative effects of statins were potentiated by overexpression of acyl-CoA synthetase long-chain family member 3 (ACSL3). Moreover, ACSL3 overexpression was associated with worse clinical outcome in patients with high-grade NSCLC. Finally, we found that patients with high expression levels of ACSL3 displayed a clinical benefit of statins treatment. Therefore, our study highlights ACSL3 as a prognostic biomarker for NSCLC, useful to select patients who would obtain a clinical benefit from statin administration.

Severe osteomalacia with multiple insufficiency fractures secondary to intravenous iron therapy in a patient with Rendu-Osler-Weber syndrome.

SUMMARY: This case report describes a 65-year-old man with a Rendu-Osler-Weber syndrome with secondary chronic anaemia, who received multiple intravenous (IV) iron infusions and sustained diffuse bone pain secondary to multiple insufficiency fractures. Laboratory study confirmed fibroblast growth factor 23 (FGF-23)-mediated hypophosphatemia as the main cause of a severe osteomalacia induced by ferric carboxymaltose (FCM).After 3 months or oral phosphate replacement and switching to iron sucrose, serum phosphate levels were normalized and patient improved clinically. INTRODUCTION: Some drugs can induce asymptomatic hypophosphatemia, which if sustained, can lead to a severe osteomalacia with multiple skeletal fractures. This complication has also been described with IV iron therapy. METHODS: This case report describes a patient with Rendu-Osler-Weber syndrome with chronic iron deficiency anaemia, recurrently treated with FCM, who developed a severe osteomalacia with multiple skeletal fractures. RESULTS: Laboratory study showed hypophosphatemia, with high ALP and high FGF-23. Images studies confirmed bone mass loss and multiple insufficiency fractures. A Fluorodeoxyglucose Positron Emission Tomography/Computed Tomography (18F-FDG PET/CT) did not show hidden tumor, so a diagnosis of FCM-induced hypophosphatemic osteomalacia was performed. Phosphate replacement improved clinical symptoms of the patient. CONCLUSION: Intravenous iron therapy, mainly FCM form, can cause hypophosphatemia, and in some cases induce a severe osteomalacia with multiple fractures, so it seems advisable to monitor serum phosphate levels in high risk patients, as those who receive repeated dose.

Clinical-pathological and molecular characterization of long-term survivors with advanced non-small cell lung cancer.

Objective: Long-term survivors (LS) of non-small cell lung cancer (NSCLC) without driver alterations, displaying an overall survival (OS) of more than 3 years, comprise around 10% of cases in several series treated with chemotherapy. There are classical prognosis factors for these cases [stage, Eastern Cooperative Oncology Group (ECOG), etc.], but more data are required in the literature. In this multi-center study, we focused on LS of advanced NSCLC with OS above 36 months to perform a clinical-pathological and molecular characterization. Methods: In the first step, we conducted a clinical-pathological characterization of the patients. Afterwards, we carried out a genetic analysis by comparing LS to a sample of short-term survivors (SS) (with an OS less than 9 months). We initially used whole-genome RNA-seq to identify differentiating profiles of LS and SS, and later confirmed these with reverse transcription-polymerase chain reaction (RT-PCR) for the rest of the samples. Results: A total of 94 patients were included, who were mainly men, former smokers, having adenocarcinoma (AC)-type NSCLC with an ECOG of 0-1. We obtained an initial differential transcriptome expression, displaying 5 over- and 33 under-expressed genes involved in different pathways: namely, the secretin receptor, surfactant protein, trefoil factor 1 (TFF1), serpin, Ca-channels, and Toll-like receptor (TLRs) families. Finally, RT-PCR analysis of 40 (20 LS/20 SS) samples confirmed that four genes (surfactant proteins and SFTP) were significantly down-regulated in SS compared to LS by using an analysis of covariance (ANCOVA) model: SFTPA1 (P = 0.023), SFTPA2 (P = 0.027), SFTPB (P = 0.02), and SFTPC (P = 0.047). Conclusions: We present a sequential genetic analysis of a sample of NSCLC LS with no driver alterations, obtaining a differential RNA-seq/RT-PCR profile showing an abnormal expression of SF genes.

Tolerability and Safety of a Nutritional Supplement with Potential as Adjuvant in Colorectal Cancer Therapy: A Randomized Trial in Healthy Volunteers.

Bioactive supplements display relevant therapeutic properties when properly applied according to validated molecular effects. Our previous research efforts established the basis to develop a dietary supplement based on a Rosmarinus officinalis supercritical extract. This was enriched in phenolic diterpenes (RE) with proven properties against signaling pathways involved in colon tumorigenesis, and shark liver oil rich in alkylglycerols (AKG) as a bioactive lipid vehicle to improve RE bioavailability and synergize with the potential therapeutic action of the extract. Herein, we have investigated the tolerability and safety of the supplement and the biological and molecular effects from an immuno-nutritional perspective. Sixty healthy volunteers participated in a six week, double-blind, randomized parallel pilot study with two study arms: RE-AKG capsules (CR) and control capsules (CC). Mean age (±SD) of volunteers was 28.32 (±11.39) and 27.5 (±9.04) for the control and the study groups, respectively. Safety of the CR product consumption was confirmed by analyzing liver profile, vital constants, and oxidation markers (LDLox in blood and isoprostanes and thromboxanes in urine). The following were monitored: (1) the phenotyping of plasmatic leukocytes and the ex vivo response of lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMCs); (2) expression of genes associated with immune-modulation, inflammation, oxidative stress, lipid metabolism, and tumorigenesis; and (3) the correlation of selected genetic variants (SNPs) with the differential responses among individuals. The lack of adverse effects on liver profile and oxidation markers, together with adequate tolerability and safe immunological adaptations, provide high-quality information for the potential use of CR as co-adjuvant of therapeutic strategies against colorectal cancer.

Antinuclear antibodies and cancer: A literature review.

Antinuclear antibodies (ANAs) are a spectrum of autoantibodies targeted to various nuclear and cytoplasmic components of the cells. They are very useful as serological markers for different autoimmune disease, like systemic lupus erythematosus (SLE), Sjögren's syndrome (SS), scleroderma, polymyositis, or mixed connective tissue disease. In these years, an increasing attention has been focussed in the relationship between tumours and autoimmunity. Different authors have demonstrated that ANAs are presented, not only in autoimmune diseases, also in serum of patients with different types of cancers. These data suggested that ANAs could be involved in the pathogenesis of cancer as well as other premalignant disease. In this review, we are going to describe all data reported about the presence of these antibodies in samples from patients with cancer as well as the potential role of some of these proteins in early detection and prognosis.

Lipid metabolism and lung cancer.

Lung cancer is currently one of the most serious health issues in developed and developing countries. There are multiple available treatment options; however survival still remains very poor. Despite metabolism alteration being one of the hallmarks described in human cancer, lipid metabolism disorders are less known. They are recently becoming more important in this setting and therefore achieving a deeper knowledge might be helpful to obtain new strategies to accurate diagnosis, estimate prognosis, and develop therapeutic agents based on bioactive compounds such as cerulenin, SCD1, ACLY inhibitors, statins, polyphenolic compounds, etc. The present paper reviews the basis of lipid metabolism in lung cancer and suggests potential biomarkers. Further investigation is crucial to improve our knowledge in this area.

Genetic polymorphisms of SCN9A are associated with oxaliplatin-induced neuropathy.

BACKGROUND: Oxaliplatin is a chemotherapy agent active against digestive tumors. Peripheral neuropathy is one of the most important dose-limiting toxicity of this drug. It occurs in around 60-80% of the patients, and 15% of them develop severe neuropathy. The pathophysiology of oxaliplatin neurotoxicity remains unclear. SCN9A is a gene codifying for a subtype sodium channel (type IX, subunit α) and mutations in this gene are involved in neuropathic perception. In this study we investigated whether SCN9A genetic variants were associated with risk of neurotoxicity in patients diagnosed of cancer on treatment with oxaliplatin. METHODS: Blood samples from 94 patients diagnosed of digestive cancer that had received oxaliplatin in adjuvant or metastatic setting were obtained from three hospitals in Madrid. These patients were classified into two groups: "cases" developed oxaliplatin-induced grade 3-4 neuropathy (n = 48), and "controls" (n = 46) had no neuropathy or grade 1. The neuropathy was evaluated by an expert neurologist and included a clinical examination and classification according to validated neurological scales: National Cancer Institute Common Toxicity Criteria (NCI-CTC), Oxaliplatin-Specific Neurotoxicity Scale (OSNS) and Total Neuropathy score (TNS). Genotyping was performed for 3 SCN9A missense polymorphisms: rs6746030 (R1150W), rs74401238 (R1110Q) and rs41268673 (P610T), and associations between genotypes and neuropathy were evaluated. RESULTS: We found that SCN9A rs6746030 was associated with protection for severe neuropathy (OR = 0.39, 95% CI = 0.16-0.96; p = 0.041). Multivariate analysis adjusting for diabetes provided similar results (p = 0.036). No significant differences in neuropathy risk were detected for rs74401238 and rs41268673. CONCLUSION: SCN9A rs6746030 was associated with protection for severe oxaliplatin-induced peripheral neuropathy. The validation of this exploratory study is ongoing in an independent series.

Current evidence for cancer stem cells in gastrointestinal tumors and future research perspectives.

Cancer stem cells (CSCs) are a very heterogeneous subpopulation of "stem-like" cancer cells that have been identified in many cancers, including leukemias and solid tumors. It is believed that CSCs drive tumor growth, malignant behavior and are responsible for the initiation of metastatic spread. In addition, CSCs have been implicated in chemotherapy and radiotherapy resistance. Current evidence supports the theory that CSCs share at least two main features of normal stem cells: self-renewal and differentiation, properties that contribute to tumor survival even in the presence of aggressive chemotherapy; however, the mechanism(s) governing the unique biology of CSCs remain unclear. In the field of gastrointestinal cancer, where we face very low survival rates across different tumor types, unraveling the role of CSCs in gastrointestinal tumors should improve our knowledge of cancer biology and chemoresistance, ultimately benefiting patient survival. Towards this end, much effort is being invested in the characterization of CSCs as a means of overcoming drug resistance and controlling metastatic spread. In this review we will cover the concept of CSCs, the current evidence for CSCs in gastrointestinal tumors and future research directions.

Reference intervals for bone turnover markers in Spanish premenopausal women.

BACKGROUND: The aims of this study were to establish robust reference intervals and to investigate the factors influencing bone turnover markers (BTMs) in healthy premenopausal Spanish women. METHODS: A total of 184 women (35-45 years) from 13 centers in Catalonia were analyzed. Blood and second void urine samples were collected between 8 a.m. and 10 a.m. after an overnight fast. Serum procollagen type I amino-terminal propeptide (PINP) and serum cross-linked C-terminal telopeptide of type I collagen (CTX-I) were measured by two automated assays (Roche and IDS), bone alkaline phosphatase (bone ALP) by ELISA, osteocalcin (OC) by IRMA and urinary NTX-I by ELISA. PTH and 25-hydroxyvitamin D (25OHD) levels were measured. All participants completed a questionnaire on lifestyle factors. RESULTS: Reference intervals were: PINP: 22.7-63.1 and 21.8-65.5 µg/L, bone ALP: 6.0-13.6 µg/L, OC: 8.0-23.0 µg/L, CTX-I: 137-484 and 109-544 ng/L and NTX-I: 19.6-68.9 nM/mM. Oral contraceptive pills (OCPs) influenced PINP (p=0.007), and low body mass index (BMI) was associated with higher BTMs except for bone ALP. Women under 40 had higher median values of most BTMs. CTX-I was influenced by calcium intake (p=0.010) and PTH (p=0.007). 25OHD levels did not influence BTMs. Concordance between the two automated assays for PINP and particularly CTX-I was poor. CONCLUSIONS: Robust reference intervals for BTMs in a Southern European country are provided. The effects of OCPs and BMI on their levels are significant, whilst serum 25OHD levels did not influence BTMs. Age, calcium intake, BMI and PTH influenced CTX-I. The two automated assays for measuring PINP and CTX-I are not interchangeable.