Validation of a Multiplexed Gene Signature Assay for Diagnosis of Canine Cancers from Formalin-Fixed Paraffin-Embedded Tissues.

BACKGROUND: Use of molecular-based diagnostics for companion animals is impeded by availability of technology platforms, tissue acquisition requirements, and species-specific reagents. HYPOTHESIS/OBJECTIVES: To validate a quantitative nuclease protection assay (qNPA) to simultaneously measure RNA expression of multiple genes in archived formalin-fixed paraffin-embedded (FFPE) tumors from dogs. ANIMALS: All tumor biopsy samples were collected retrospectively from surgical biopsies and in the care of veterinarians. METHODS: Retrospective case series. A qNPA 96-well ArrayPlate was built using 30 canine-specific genes, 5 housekeeping genes, positive and negative controls with qualified gene-specific oligonucleotides. Pearson's correlation, coefficient of variation (CV), and multivariate analysis were used to determine analytical performance using 40 FFPE dog tumors. Once validated, 70 FFPE dog tumors were analyzed for differences in gene expression using hierarchical clustering and analysis of variance of log transformed data. Immunohistochemistry (IHC) was performed to correlate gene expression and protein expression in a subset of tumors. RESULTS: The assay was linear with decreasing sample input (R2 = 0.978), reproducible within and between 96-well plates (r = 0.988 and 0.95, respectively) and between different laboratories (CV = 0.96). Hierarchical cluster analysis showed grouping of tumors by histogenesis and oncogenes. Significant differences were found between BCl2, E2F transcription factor 1, MDM2, COX-2, MET proto-oncogene receptor kinase, and other biologically relevant gene expression in tumor subtypes. Immunohistochemistry confirmed protein expression. CONCLUSIONS AND CLINICAL IMPLICATIONS: Because this technology works reliably on FFPE specimens, it can help expedite the broad introduction of multiplexed genomic information for improved diagnostics and discovery of new targets for therapies in veterinary oncology.

Alternating Rabacfosadine/Doxorubicin: Efficacy and Tolerability in Naïve Canine Multicentric Lymphoma.

BACKGROUND: Standard of care treatment for multicentric lymphoma in dogs remains doxorubicin (DOX)-based combination chemotherapy, but owners may hesitate to commit the time and financial resources to complete such a protocol, typically requiring 12-16 visits. Rabacfosadine (RAB), a double prodrug of the nucleotide analog 9-(2-phosphonylmethoxyethyl) guanine, has substantial single-agent activity in dogs with lymphoma, and a different mechanism of action than DOX. HYPOTHESIS/OBJECTIVES: Our objective was to evaluate the efficacy and adverse effect (AE) profile of alternating doses of RAB and DOX in dogs with naïve multicentric lymphoma. ANIMALS: Fifty-four dogs with previously untreated lymphoma. METHODS: Open-label, multicenter prospective clinical trial. Dogs received alternating RAB (1.0 mg/kg IV weeks 0, 6, 12) and DOX (30 mg/m2 IV weeks 3, 9, 15). Dogs that achieved complete response (CR) were followed by monthly evaluations. Complete clinicopathological evaluation and assessment of remission and AEs were performed every 21 days. RESULTS: The overall response rate was 84% (68%; CR; 16%; partial response [PR)]. The overall median progression-free interval (PFI) was 194 days (216 for CR and 63 for PR). Most AEs were mild and self-limiting: gastrointestinal and hematologic AEs were most common. Thirteen dogs experienced dermatologic AEs, and 2 dogs developed grade 5 pulmonary fibrosis. CONCLUSIONS AND CLINICAL IMPORTANCE: Alternating RAB/DOX generally was well tolerated and resulted in PFIs comparable to standard DOX-based multi-agent protocols, with fewer treatment visits. Most adverse events were mild or moderate and self-limiting. Further studies are warranted to explore long-term outcome and other RAB chemotherapy combinations.

Geographical differences in survival of dogs with non-Hodgkin lymphoma treated with a CHOP based chemotherapy protocol.

BACKGROUND: In humans geographical differences in the incidence and presentation of various cancers have been reported. However, much of this information has not been collected in veterinary oncology. AIM: The purpose of this study was to determine if a geographic difference in progression free survival exists for dogs with lymphoma treated within the US. MATERIALS AND METHODS: Medical records of 775 cases of canine lymphoma from 3 US regions (west, south and north), treated with CHOP chemotherapy, were retrospectively evaluated. Cases were collected from referral institutions and were required to have received at least one doxorubicin treatment and have follow up information regarding time to progression. RESULTS: Significant differences in sex (p = 0.05), weight (p = 0.049), stage (p < 0.001), immunophenotype (p = <0.001), and number of doxorubicin doses (p = 0.001) were seen between regions. Upon univariate analysis, progression free survival (PFS) differed by region (p = 0.006), stage (p = 0.009), sub-stage (p = 0.0005), and immunophenotype (p = 0.001). A multivariable Cox regression model showed that dogs in the western region had a significantly shorter PFS when compared to the south and east. CONCLUSION: PFS was significantly affected by stage, sub-stage and phenotype.

Clinical presentation, treatment and outcome in 31 dogs with presumed primary colorectal lymphoma (2001-2013).

The objective of this multicentre retrospective study was to describe clinical presentation, treatment and outcome and to determine prognostic factors for dogs with presumed primary colorectal lymphoma (PCRL). A total of 31 dogs were included. The predominant features of PCRL were high grade (n = 18) and immunophenotype B (n = 24). Most dogs were substage b (n = 25) with higher prevalence of haematochezia (n = 20). One dog had surgery only. Thirty dogs received chemotherapy; amongst them 13 had surgery or radiotherapy. Progression free survival (PFS) was 1318 days and disease-related median survival time (MST) was 1845 days. Fourteen dogs were alive at the end of the study with a median follow-up time of 684 days (3-4678 days). Younger dogs had longer PFS (P = 0.031) and disease-related MST (P = 0.01). Presence of haematochezia corresponded with longer PFS (P = 0.02). Addition of local treatment to chemotherapy did not significantly improve the outcome (P = 0.584). Canine PCRL has considerably longer PFS and MST than other forms of non-Hodgkin's lymphoma.

Recurrence rates and clinical outcome for dogs with grade II mast cell tumours with a low AgNOR count and Ki67 index treated with surgery alone.

Grade II mast cell tumours (MCT) are tumours with variable biologic behaviour. Multiple factors have been associated with outcome, including proliferation markers. The purpose of this study was to determine if extent of surgical excision affects recurrence rate in dogs with grade II MCT with low proliferation activity, determined by Ki67 and argyrophilic nucleolar organising regions (AgNOR). Eighty-six dogs with cutaneous MCT were evaluated. All dogs had surgical excision of their MCT with a low Ki67 index and combined AgNORxKi67 (Ag67) values. Twenty-three (27%) dogs developed local or distant recurrence during the median follow-up time. Of these dogs, six (7%) had local recurrence, one had complete and five had incomplete histologic margins. This difference in recurrence rates between dogs with complete and incomplete histologic margins was not significant. On the basis of this study, ancillary therapy may not be necessary for patients with incompletely excised grade II MCT with low proliferation activity.

Use of adjuvant carboplatin for treatment of dogs with oral malignant melanoma following surgical excision.

Melanoma is the most common oral malignancy in dogs. This retrospective study evaluated adjuvant carboplatin chemotherapy (with or without radiation therapy) in 17 dogs with malignant oral melanoma following surgical resection. The median dosage and number of doses of carboplatin administered to the 17 dogs was 300 mg m(-2) (range, 150-300 mg m(-2)) and 4 (range, 2-11), respectively. The overall median progression-free survival for all dogs was 259 days [95% confidence interval (CI95), 119-399 days]. The first progression-free survival event was local recurrence in seven dogs (41%) and metastases in seven dogs (41%). The median overall survival for all dogs was 440 days (CI95, 247-633 days). The tumour was the cause of death in 10 dogs (59%). On the basis of this study, systemic therapy with carboplatin may be an appropriate adjunct to local treatment for canine malignant melanoma, although future prospective controlled studies are needed to compare treatment modalities for this aggressive neoplasia.

Prolonged prothrombin time correlates with serum monoclonal protein concentration in patients with plasma cell dyscrasia.

INTRODUCTION: Abnormal screening coagulation tests are frequently observed in asymptomatic patients with multiple myeloma and other plasma cell neoplasms. METHODS: Prothrombin time (PT), activated partial thromboplastin time (APTT) and fibrinogen activity were correlated with clinical history and disease parameters in patients referred to the Myeloma Institute for Research and Therapy. RESULTS: An isolated prolonged PT was the most common abnormal coagulation test (25%). Prolonged PT was more frequently observed in patients with multiple myeloma (n = 157) compared to MGUS patients (n = 34) or other diagnostic categories of plasma cell dyscrasia. There were no differences in age, gender, previous chemotherapy, or immunoglobulin isotype in patients with isolated prolonged PT (n = 62) compared to those with normal screening coagulation tests (n = 173). Fibrinogen activity was significantly lower in patients with prolonged PT; however, there was no correlation between fibrinogen activity and PT. Serum M protein concentrations were significantly greater in patients with prolonged PT and were positively correlated with PT. CONCLUSION: An association between disease severity and prolonged PT is suggested by our finding that patients with multiple myeloma were more likely to have prolonged PT than patients with other plasma cell neoplasms. Of the factors examined, the monoclonal protein level was significantly higher in patients with isolated prolonged PT and correlated with PT.

Feline exocrine pancreatic carcinoma: a retrospective study of 34 cases.

Thirty-four cases were reviewed in this retrospective study for information on clinical presentation, prognostic indicators, survival time and response to various therapies. The most common presenting clinical signs were weight loss, decreased appetite, vomiting, palpable abdominal mass and diarrhoea. Metastatic disease was confirmed in 11 cats. The overall median survival was 97 days. The median survival times for patients who received chemotherapy or had their masses surgically removed was 165 days. Those patients who had an abdominal effusion present at the time of diagnosis survived a median of 30 days. Cats that received non-steroidal anti-inflammatory drug therapy had a median survival of 26 days. This study confirms that exocrine pancreatic carcinoma in cats is an aggressive tumour with a high metastatic rate and poor prognosis, although three patients survived over 1 year. Fifteen percent of the patients were diabetic, which raises the question as to what the link between diabetes and pancreatic cancer in people and cats may be.

A randomized trial investigating the efficacy and safety of water soluble micellar paclitaxel (Paccal Vet) for treatment of nonresectable grade 2 or 3 mast cell tumors in dogs.

BACKGROUND: Effective treatments for dogs with advanced stage mast cell tumors (MCT) remain a pressing need. A micellar formulation of paclitaxel (paclitaxel [micellar]) has shown promise in early-phase studies. HYPOTHESIS/OBJECTIVES: The objective was to demonstrate greater activity for paclitaxel (micellar) compared with lomustine. The null hypothesis was µ(p) = µ(L) (ie, proportion of responders for the paclitaxel [micellar] and lomustine groups, respectively). ANIMALS: Two hundred and fifty-two dogs with advanced stage nonresectable grade 2 or 3 MCT. METHODS: Prospective multicenter randomized double-blind positive-controlled clinical trial. The primary endpoint was confirmed overall response rate (CORR) at 14 weeks. A secondary endpoint, biologic observed response rate (BORR), also was calculated. Safety was assessed by the characterization and grading of adverse events (AE). RESULTS: Overall CORR (7% versus 1%; P = .048) and BORR (23% versus 10%; P = .012) were greater for paclitaxel (micellar) compared with lomustine. Paclitaxel (micellar)-treated dogs were 6.5 times more likely to have a confirmed response and 3.1 times more likely to experience a biologic observed response. The majority of AE with paclitaxel (micellar) were transient and clinically manageable. Twenty-seven dogs (33%) receiving lomustine were discontinued because of hepatopathy compared with 3 dogs (2%) receiving paclitaxel (micellar) (P < .0001; odds ratio 26.7). CONCLUSIONS AND CLINICAL IMPORTANCE: Paclitaxel (micellar)'s activity and safety profile are superior to lomustine. The addition of an active and novel taxane to the veterinary armamentarium could fill a substantial need and, as its mechanism of action and AE profile do not overlap with currently available TKI, its availability could lead to effective combination protocols.

Single-agent gemcitabine chemotherapy in dogs with hepatocellular carcinomas.

The goal of this study was to determine the efficacy and tolerability of gemcitabine in dogs diagnosed with hepatocellular carcinoma (HCC). Eighteen dogs were examined retrospectively (4 massive HCC, 10 nodular HCC and 4 diffuse HCC). All dogs received gemcitabine at 350-400 mg m(-2) weekly for 5 weeks. Toxicity was graded using VCOG-CTCAE guidelines and response was monitored with serial abdominal ultrasounds. Fifteen dogs completed all five cycles. Toxicity was minimal and consisted of grade I/II vomiting, anorexia and diarrhoea and two episodes of grade III neutropenia. Median survival time for all dogs was 983 days. Median progression free interval was 971 days. Based on the results of this study, surgery remains the best treatment for HCC, despite incomplete resection. There was no improvement in the survival of those diagnosed with nonresectable HCC treated with gemcitabine chemotherapy.

Evaluation of a multidrug chemotherapy protocol with mitoxantrone based maintenance (CHOP-MA) for the treatment of canine lymphoma.

The purpose of this study was to evaluate the efficacy of adding mitoxantrone to a cyclophosphamide, doxorubicin, vincristine, L-asparaginase and prednisone containing protocol. Sixty-five dogs with multicentric lymphoma were evaluated for overall remission and survival times. Remission and survival time versus stage, substage, pretreatment hypercalcaemia and pretreatment steroid administration were also evaluated. Overall median remission for dogs with multicentric lymphoma was 302 days and overall median survival was 622 days. Of the dogs with multicentric lymphoma, 23 (35%) received all scheduled mitoxantrone doses. Only median survival versus substage was found to be significant (substage a median survival was 679 days and substage b median survival was 302 days, P = 0.025). Increasing the total combined dose of doxorubicin and mitoxantrone may improve remission times when compared with historical controls, and further studies are needed to determine how best to utilize mitoxantrone in multidrug chemotherapy protocols for canine multicentric lymphoma.

Asparaginase and MOPP treatment of dogs with lymphoma.

BACKGROUND: Dogs with multicentric lymphoma are treated with various cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP)-based chemotherapy protocols with variable success. OBJECTIVES: To describe the progression-free survival (PFS) time and overall survival time (OST) of dogs with T-cell lymphoma or hypercalcemic lymphoma treated with L-asparaginase and mechlorethamine, vincristine, prednisone, procarbazine (MOPP). ANIMALS: Fifty dogs with T-cell lymphoma, hypercalcemic lymphoma, or both treated at 3 referral veterinary hospitals. METHODS: Retrospective study. Case were selected based on histologic or cytologic diagnosis of lymphoma; presence of the T-cell phenotype, presence of hypercalcemia or both; and absence of previous chemotherapy. The T-cell phenotype was determined by flow cytometry, immunocytochemistry, immunohistochemistry, or polymerase chain reaction of antigen receptor rearrangement. RESULTS: The overall response rate was 98% (78% complete response, 20% partial response). The median PFS for the entire study population was 189 days with 25% PFS at 939 days. The median OST for the entire study population was 270 days with 25% surviving 939 days. Twenty percent of the dogs required hospitalization for treatment related complications. CONCLUSIONS AND CLINICAL IMPORTANCE: L-Asp/MOPP chemotherapy might result in longer PFS and OST for dogs with multicentric T-cell lymphoma, dogs with hypercalcemic lymphoma or both, than achieved with CHOP.

Effects of health information in youth on adult physical activity: 20-year study results from the Amsterdam growth and health longitudinal study.

In the Amsterdam Growth and Health Longitudinal Study (AGAHLS), a group of apparently healthy males and females (n = 200) were interviewed about their physical activities on eight separate occasions over a period of 20 years between 13 and 33 years of age (multi-measured group: MM). Information about their health was given based on their personally measured lifestyle (activity, diet, smoking) and biological risk characteristics for chronic diseases (medical check-ups). A comparable group of boys and girls (n = 200) was only measured on two occasions (bi-measured group: BM): at 13 and 33 years. Physical activity was estimated with a structured interview. Total physical activity and sports activity were estimated in three intensity levels (light, moderate, and heavy). It was hypothesized that the eight repeated medical check-ups with health information in the MM group would result in a healthier lifestyle with respect to the determinants and levels of habitual physical activity compared to the BM group. Contrary to the hypothesis, males and females in the BM group showed a significantly higher increase or a lower decrease in physical activities compared to the MM group. This negative effect on the physical activity pattern at 33 years in the MM group may have been caused by more underreporting of physical activities than in the BM group. In conclusion, there does not appear to be a significant effect of long-term (multi-measured) health information with medical check-ups during adolescence and young adulthood on level of physical activity in males and females at 33 years of age.

Clinicopathologic and electron microscopic study of cutaneous neuroendocrine (Merkel cell) carcinoma in a cat with comparisons to human and canine tumors.

Malignant neuroendocrine carcinoma of the skin (Merkel cell tumor) was diagnosed in an 18-year-old spayed female Maine Coon Cat. The diagnosis was made on the basis of morphologic and electron microscopic findings. The cat was euthanatized 321 days after surgical excision of the tumor. The tumor's malignancy contrasted with the benign nature of Merkel cell tumors reported in dogs and was consistent with the malignancy of Merkel cell tumors reported in humans.

A 15-year physical activity pattern is positively related to aerobic fitness in young males and females (13-27 years).

The purpose of this paper is to test the hypothesis that daily physical activity over a period of 15 years has been beneficial to aerobic fitness in young male and female participants (13-27 years) in the Amsterdam Growth and Health Longitudinal Study. Only subjects with the maximal data of six sets of measurements were included (83 male and 98 female participants). Daily physical activity was assessed using a standardized interview on activity and expressed as a weighted activity score. Aerobic fitness was assessed using a maximal running test on a treadmill and measuring the maximal oxygen uptake (VO2max) and the maximal slope of the track (Smax). To assess the longitudinal relationship between daily physical activity and aerobic fitness a real longitudinal analysis was carried out with generalized estimating equations, adjusting for differences in initial aerobic fitness at age 13, and for other lifestyle (dietary intake, smoking and alcohol consumption) and biological parameters (biological age, body fat, blood pressure and concentration of serum cholesterol). A significant relationship (P < 0.01) was observed between daily physical activity and both VO2max and Smax. It can be concluded that the development of aerobic fitness between the age of 13 and 27 years is independently and positively related to daily physical activity in this group of male and female participants in the study. The functional implications, however, are small: a relatively high increase in the weighted physical activity score of 30% over a period of 15 years results in a 2%-5% increase in aerobic fitness.

Clustering of risk factors for coronary heart disease. the longitudinal relationship with lifestyle.

PURPOSE: The purpose of this study was to investigate whether or not clustering of biological coronary heart disease (CHD) risk factors exists and to investigate the longitudinal relationship between lifestyle parameters (dietary intake, daily physical activity, smoking behaviour, alcohol consumption) and a biological CHD risk factor clustering score. This was defined as belonging to one or more gender specific 'high risk' quartiles for the following CHD risk factors: ratio between total serum cholesterol and high density lipoprotein cholesterol (TC:HDL), mean arterial blood pressure (MABP), body fatness [sum of skinfolds (SSF)], and cardiopulmonary fitness (VO2-max). METHODS: The data were derived from the Amsterdam Growth and Health Study, an observational longitudinal study in which six repeated measurements were carried out over a period of 15 years covering adolescence and young adulthood. The longitudinal relationships were analysed with generalized estimating equations. RESULTS: The results showed significant clustering for the TC:HDL ratio, SSF, and VO(2)-max. MABP was not significantly associated with the other CHD risk factors. Daily physical activity and alcohol consumption (only for males) were both inversely related to the clustering score. None of the other lifestyle parameters showed significant relationships with the clustering score. CONCLUSIONS: Based on this small longitudinal study, it can be stated that during adolescence and young adulthood both daily physical activity and alcohol consumption were related to a healthy CHD risk profile.

Longitudinal relationships between lifestyle and cardiovascular and bone health status indicators in males and females between 13 and 27 years of age; a review of findings from the Amsterdam Growth and Health Longitudinal Study.

UNLABELLED: The Amsterdam Growth and Health Longitudinal Study is longitudinal co-hort study on 181 males and females initially aged 13 years, with follow-up measurements at ages 14, 15, 16, 21 and 27 years. METHODS: Anthropometrical, biological and lifestyle parameters, and age 27 also bone mineral density (BMD) of the lumbar spine (L2-L4), were measured repeatedly. Adolescent dietary intake and physical activity was related to adult cardiovascular and bone health status indicators by MLR; longitudinal relationships between physical activity and dietary intake, and cardiovascular health status indicators were assessed hy GEE-analysis. RESULTS: Adolescent physical activity was not related to most of the indicators of adult cardiovascular health status, with the exception of a positive relationship with the waist-to-hip in females; in males 'energetic' adolescent physical activity contributed significantly to adult BMD; both in males and females, when taking the entire longitudinal period into account, peak strain physical activity was a relatively more important predictor of adult BMD than 'energetic' physical activity; calcium intake during adolescence was not a significant predictor of bone health status measured at age 27, both in males and females; a consistent significant positive longitudinal relationship was found between physical activity and serum HDL-cholesterol and significant negative longitudinal relationships were found with the subscapular skinfold and with body fat mass; a positive longitudinal relationship was found between serum total cholesterol (TC) and cholesterol intake, saturated fat (SFA) intake and the Keys-score; a negative longitudinal relationship was found between TC and polyunsaturated fit and total energy intake; for HDL a positive longitudinal relationship was found with SFA intake; low tracking was found for physical (in-) activity and dietary intake variables.

Nephrogenic diabetes insipidus in a dog with intestinal leiomyosarcoma.

Nephrogenic diabetes insipidus was diagnosed in a dog with an intestinal leiomyosarcoma. The diagnosis of nephrogenic diabetes insipidus was made on the basis of results of serum biochemical tests, urinalyses, and a water-deprivation test, along with a lack of response to exogenous administration of vasopressin following the water-deprivation test. The temporal association between resection of the intestinal mass and resolution of clinical signs of diabetes insipidus (i.e., polyuria and polydipsia) and between recurrence of clinical signs and detection of metastatic disease suggests that there may have been a causal relationship, and nephrogenic diabetes insipidus may have developed as a paraneoplastic syndrome in this dog.

Body fatness: longitudinal relationship of body mass index and the sum of skinfolds with other risk factors for coronary heart disease.

OBJECTIVE: To analyse the longitudinal relationships between body mass index (BMI)/sum of skinfolds (SSF) and biological and lifestyle risk factors for coronary heart disease (CHD). DESIGN: An observational longitudinal study; that is, the Amsterdam Growth and Health Study. SUBJECTS: 181 males and females, initially aged 13 y. Over a period of 15 y, six repeated measurements were carried out. MEASUREMENTS: BMI and SSF, biological CHD risk factors; that is, total cholesterol (TC), high density lipoprotein (HDL), TC:HDL ratio, systolic/diastolic blood pressure (SBP/DBP) and cardiopulmonary fitness (VO2-max) and lifestyle CHD risk factors (that is, daily physical activity, dietary parameters, smoking, and alcohol consumption). The longitudinal relationships were analysed by an autoregressive model, in which the value of the outcome variable at time-point t is not only related to the value of the predictor variable at t, but also to the value of the outcome variable at t-1. RESULTS: Both BMI and SSF were positively related to TC and the TC:HDL ratio. Only BMI was positively related to SBP and only SSF was negatively related to VO2-max. Physical activity was negatively related to SSF. None of the other lifestyle parameters were related to SSF and/or BMI. CONCLUSIONS: Both BMI and SSF were related to a high risk profile regarding CHD. Different relationships for SSF and BMI are found, because BMI not only reflects body fatness, but also lean body mass. Analyses with BMI as an indicator for body fatness should therefore be interpreted cautiously.

Behavioral variables and development of a central pattern of body fat from adolescence into adulthood in normal-weight whites: the Amsterdam Growth and Health Study.

Associations were investigated between the amount of physical activity, energy and macronutrient intake, smoking behavior, alcohol intake, and a central pattern of body fat (subscapular skinfold thickness and waist circumference) measured six times between the mean ages of 13 and 27 y in a healthy white population. Subjects (84 males, 98 females) were participants in the longitudinal Amsterdam Growth and Health Study. In longitudinal analyses, alcohol intake was positively associated with the subscapular skinfold thickness (beta = 0.09, 95% CI: 0.01, 0.16) in males. In females, the subscapular skinfold thickness was negatively associated with physical activity (beta = -0.10. 95% CI: -0.15, -0.05) and, unexpectedly, energy intake (beta = -0.25, 95% CI: -0.31,-0.19), whereas a positive association was found with carbohydrate intake (beta = 0.09. 95% CI: 0.02, 0.16). In both sexes, the mean value of behavioral variables, obtained from the mean value in adolescence and the values obtained at 21 and 27 y of age was not significantly associated with the subscapular skinfold thickness or waist circumference at the mean age of 27 y, except for a small positive association between physical activity and the subscapular skinfold thickness in males (R2 = 2.3%).