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Persistent circadian rhythm of bone turnover in bone metastatic breast cancer suggests greater skeletal retention of bisphosphonates if administered in the night. We assessed differential effects of night vs morning administration of zoledronic acid (ZA) on bone turnover. Forty-four breast cancer patients with bone metastases were randomised to receive intravenous ZA (4 mg) at 1100 or 2300 hours every 28 days for four times. Urinary concentration N-telopeptide of type-I collagen (NTX) and deoxypyridinolines, and serum C-telopeptide of type-I collagen (CTX), bone alkaline phosphatase (ALP), osteocalcin and Parathyroid hormone (PTH) was measured in the morning at baseline and after 4, 7, 14, 28, 56 and 84 days. Urinary ZA concentration was also measured. Zoledronic acid caused significant decreases of NTX and CTX (P

The variability of serum osteoprotegerin (OPG) and soluble RANKL (sRANKL) along the 24-h cycle was assessed in 20 healthy women. No rhythmic variations of serum OPG, sRANKL or sRANKL/OPG ratio were detected as a group phenomenon. Timing of sampling is unlikely to influence the results of measurements of circulating OPG and sRANKL. INTRODUCTION: Physiological bone turnover shows diurnal variations. The aim of the study was to assess variability of OPG and sRANKL serum levels along the 24-h cycle. METHODS: Blood was collected from 20 healthy women (median age 31 years, range 25-65 years) at 4-h intervals between 08:00 and 24:00 and at 2-h intervals between 24:00 and 08:00. Serum albumin, cortisol, osteocalcin (OC), C-terminal telopeptide of type I collagen (CTX), OPG and total sRANKL were measured. Temporal variations were assessed by the COSINOR model. RESULTS: Circadian rhythms of cortisol and albumin documented a normal synchronization within the circadian structure. Serum OC and CTX showed rhythmic variations, peaking at night-time. Rhythmic variations of serum OPG, sRANKL and sRANKL/OPG ratio were not detected as a group phenomenon. On an individual basis, rhythmic changes were detected in ten patients for OPG and eight patients for sRANKL, with very small amplitudes and heterogeneous acrophases. CONCLUSIONS: The absence of consistent rhythmic variations of circulating OPG and sRANKL levels may reflect the absence of rhythmic variations of their expression in the bone microenvironment. Were this the case, the nocturnal rise of bone resorption should be accounted for by different, not RANKL/OPG-mediated factors. Since circulating OPG and sRANKL may derive from sources other than bone, rhythmicity could be masked by non-rhythmic or non-synchronized rhythmic expression in these sources. Timing of sampling is unlikely to influence the results of measurements of circulating OPG and sRANKL

Background Increased osteolysis usually accompanies sclerotic bone metastases from prostate cancer. This provides a rationale for the use of bisphosphonates to treat bone pain and prevent skeletal complications. Methods The fasting urinary levels of calcium, hydroxyproline (OHPRO), pyridinolines (PYD), deoxypyridinolines (DPYD), collagen cross-linked N-telopeptide (NTX) and the serum values of calcium, total alkaline phosphatase and relevant bone isoenzyme, bone gla protein (BGP), carboxy-telopeptide of type I collagen (ICTP) and parathyroid hormone (PTH) were determined at baseline and on the 15th, 30th, 60th and 90th days after single-dose (90 mg) pamidronate administration in 35 consecutive prostate cancer patients with bone metastases. These biochemical indices and serum interleukin 6 (IL-6) were also measured after four days in the last consecutive 17 cases. Results PYD, DPYD and NTX showed a significant decrease lasting four weeks (pConclusions Pamidronate is able to induce a decrease in bone resorption without significantly influencing bone formation. The maximum decrease in bone resorption occurs very early. NTX is the most sensitive bone resorption marker in bisphosphonate therapy monitoring. Changes in IL-6 but not bone resorption markers may be useful in the prediction of symptomatic response.

We assessed the activity and toxicity of the XELBEVOCT regimen in patients with metastatic well-to-moderately differentiated neuroendocrine neoplasms (WMD-NEN). Ancillary studies evaluated hypertension, proteinuria, and vascular endothelial growth factor (VEGF) polymorphisms in predicting progression-free survival (PFS) and the predictive role of serum vitamin D in progression-free survival and proteinuria onset. This prospective phase 2 study included 45 patients with WMD-NEN arising from various primary sites. The treatment regimen was octreotide long-acting release (LAR), 20 mg monthly, metronomic capecitabine, 2000 mg/daily, and intravenous bevacizumab, 5 mg/kg every 2 weeks, without interruption for 9 months. Bevacizumab was continued until disease progression. Partial response was obtained in 8 patients (17.8%, 95% confidence interval [CI], 6.4%-28.2%); tumor response was more frequent in pancreatic than in non-pancreatic malignancies. The median PFS was 14.9 months; median overall survival was not attained. Biochemical and symptomatic responses were observed in 52.9% and 82.3% of cases, respectively. The treatment was well tolerated. Grade 3 toxicities included hand and foot syndrome (11.1%), proteinuria (4.4%), and renal toxicity (2.2%). Proteinuria (all grades) was correlated with longer PFS (p = 0.017). There was an inverse relationship between proteinuria and vitamin D levels. VEGF polymorphisms were not associated with patient outcome. The XELBEVOCT regimen is active and well tolerated in patients with metastatic WMD-NEN. Proteinuria correlated with hypovitaminosis D status and was the best predictive factor of treatment efficacy. Trial registration number NCT01203306 .

We evaluated the incidence of skeletal complications in patients with bone metastatic prostate cancer and hormone refractory disease. We also assessed the predictive role of bone turnover markers determined at baseline.A total of 112 patients were consecutively enrolled in our study from July 1990 to July 1998 and followed until death or the last followup. Bone pain, disease extent in bone, serum prostate specific antigen, hemoglobin, and a panel of bone formation and resorption markers were assessed at baseline before any second line treatment.Skeletal complications in 34 patients (30.3%, estimated yearly incidence 12.3%) involved vertebral deformity or collapse requiring spinal orthosis in 20 (17.9%), spinal cord compression in 7 (6.2%), pathological bone fracture in 10 (8.9%), symptomatic hypercalcemia in 1 (0.9%) and symptomatic hypocalcemia in 1 (0.9%). Median time to the evidence of the initial skeletal complication was 9.5 months. These adverse events did not influence overall survival. At baseline patients with eventual skeletal complications had greater bone pain (p = 0.02), a heavier tumor load in bone (p = 0.005), lower performance status (p = 0.05), and higher serum alkaline phosphatase (p0.02) and urinary deoxypyridoline (p0.05) than their counterparts. Multivariate analysis revealed that only urinary deoxypyridinoline was independently associated with the onset of these events (p0.02). The scatterplot of urinary deoxypyridinoline values in patients with and without skeletal complications enabled us to detect a cutoff of 38 pM./mM. for predicting 51% of skeletal events with only an 8% false-positive rate.Skeletal complications are common in patients with prostate cancer and hormone refractory disease. Bone loss is the major cause of onset. Baseline deoxypyridinoline at the cutoff point noted had moderate sensitivity but high specificity for predicting these adverse skeletal events.

Serum bone alkaline phosphatase (BALP), serum carboxy-terminal propeptide of type I procollagen (PICP) and serum bone gla protein (BGP) as markers of bone formation, serum carboxy-terminal telopeptide of type I collagen (ICTP) as a marker of collagen resorption and fasting molar ratio of urinary calcium to creatinine (CaCr) and serum parathyroid hormone (PTH) were determined in two groups of cancer patients: 48 with advanced or metastatic disease with negative bone scan and 174 with bone metastases categorised as having lytic, mixed or blastic lesions and with more or fewer than or equal to three sites involved. In patients without apparent bone involvement, bone formation markers were rarely elevated. Conversely, serum ICTP was frequently found to be supranormal, showing it to be a non-specific marker for early detection of bone metastases. As expected, values of bone formation markers progressively increased in patients with lytic, mixed and blastic lesions, but ICTP levels did not show any differences according to the types of bone appearances, confirming previous reports of elevated osteoclast activity also in patients with apparent blastic lesions. Serum PTH increased significantly in patients with lytic compared with patients with mixed and blastic appearances, paralleling the bone formation markers, but CaCr showed the opposite pattern. These data are compatible with calcium entrapment in the bone in patients with increased osteoblast activity. This so called 'bone hunger syndrome' is further confirmed by the finding that in the subgroup of blastic appearances CaCr diminished whereas both ICTP and PTH increased according to the extent of tumour load in the bone.

We evaluated the 24-h profile of serum melatonin (MT) in 16 patients with Cushing's syndrome (11 pituitary- and 5 adrenal-dependent) aged 38.1 +/- 13.8 years and in 19 patients with acromegaly due to GH-secreting pituitary adenoma aged 52 +/- 11.9 years. Thirty-three healthy subjects aged 30 +/- 7 years and 26 healthy subjects aged 59 +/- 12 years served as age-matched controls for the two groups of patients, respectively. All the patients had active disease and were off treatment; none had overt hypopituitarism. Blood drawings were collected at 08:00, 12:00, 16:00, 20:00, 22:00, 24:00, 02:00, 04:00 and 06:00 h in controls and patients. Serum MT, cortisol, GH and IGF-I were measured by specific RIA kits and ACTH by IRMA. Cosinor rhythmometry was employed for chronobiological analysis. The acromegalic patients displayed, as a whole group, higher 24-h mean MT levels with respect to age-matched controls (M +/- SD: 52.9 +/- 14.6 vs 32.9 +/- 8.0 pg/ml, p < 0.01) with normally synchronized MT profiles in 11/19 patients. No correlation was apparent between MT and either GH (24-h mean) or IGF-I. In the patients with Cushing's syndrome the circadian profile of serum MT was maintained in 10/16 cases; serum MT levels were comparable to those of controls (48.8 +/- 14.0 vs 50.0 +/- 16.0 pg/ml, NS) without any difference between pituitary- and adrenal-dependent forms. No correlation was found between MT and either ACTH or cortisol levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Serial monitoring of serum and urinary markers of bone regeneration may provide an indirect assessment of the activity of bone metastases. We measured serum levels of carboxyterminal propeptide of type I procollagen (PICP), a new marker of bone formation, in 236 control subjects, stratified according to age and sex, and in 122 cancer patients: 80 with breast cancer (BC) (42 with osteolytic bone metastases) and 42 with prostate cancer (PC) (31 with osteoblastic bone metastases). An inverse correlation between age and serum PICP levels was found in healthy men (r = -0.30), but not in women. PICP levels were significantly higher in patients with osteoblastic metastases compared to age-matched control subjects and patients with advanced disease not involving the skeleton. The sensitivity of PICP was 64% for detecting osteoblastic metastases, and 12% for detecting osteolytic metastases. Serum PICP determination may have a diagnostic role for osteoblastic activity.

Bone metabolic disruption that occurs in bone metastatic prostate cancer could lead to disturbances of calcium metabolism. The prognostic role of either hypocalcemia or hypercalcemia was assessed in a consecutive series of hormone-refractory bone metastatic prostate cancer patients. Serum calcium was measured in 192 patients. The presence of hypocalcemia and hypercalcemia was related with baseline biochemical and clinical characteristics and the role of these two calcium disturbances in predicting prognosis and adverse skeletal-related events (SREs) was assessed. As compared to normocalcemic patients, hypocalcemic patients (n=51) had higher tumor load in bone (P=0.005), higher plasma chromogranin A (CgA, P=0.01), serum alkaline phosphatase (P=0.01), urinary N-telopeptide (NTX, P=0.002) and lower hemoglobin values (P=0.01), while hypercalcemic patients (n=16) had higher plasma CgA (P=0.001) and serum lactate dehydrogenase values (P=0.001), higher bone pain (P=0.003) and a lower frequency of pure osteoblastic lesions (P=0.001). Hypercalcemia was significantly associated with poor prognosis: hazard ratio (HR), 1.9 (95% confidence Interval (CI) 1.2-3.3) and higher risk to develop SREs HR, 2.5 (95% CI 1.2-5.2, P=0.01), while hypocalcemia was not associated with poor prognosis. The prognostic role of hypercalcemia was maintained in multivariate analysis after adjusting for validated prognostic parameters: HR, 2.72 (95% CI 1.1-6.8, P=0.03). These data suggest that serum calcium levels should be taken into account in the clinical decision-making process of bone metastatic prostate cancer patients. Patients with asymptomatic hypercalcemia could benefit of a strict follow-up and an immediate bisphosphonate treatment. Further prospective clinical trials are needed to confirm this finding.

The aim of the present study was to analyze the effects of exogenous melatonin (MT) upon pituitary and adrenal responsiveness to releasing hormones in different phases of the menstrual cycle. We evaluated the response of FSH and LH to 100 micrograms gonadotropin releasing hormone, of TSH and prolactin (PRL) to 200 micrograms thyrotropin releasing hormone (TRH), and of cortisol to 10 micrograms ACTH 1-17. We studied eight young women with normal ovulatory cycles in the early follicular (days 5-7) and luteal (days 22-24) phases. Stimulation tests were performed at 18.00 in baseline conditions as well as 1 h after oral intake of exogenous MT (2 mg as a gelatine capsule). We did not observe any significant change in FSH, LH, TSH and cortisol responses to their respective releasing hormones in either phase of the cycle. PRL response to TRH was higher after MT in the follicular phase, when evaluated in terms of net increment and integrated area of response (p less than 0.02 versus baseline conditions for both variables). In the luteal phase, we recorded larger interindividual variability and higher responses after MT were observed in five out of eight subjects. These results suggest that MT may play a facilitatory role in the TRH-induced PRL release in women of reproductive age.

A number of studies performed in vitro and on experimental animals supported the view that pineal gland inhibits neoplastic growth. Data in humans are scanty and controversial. In the present study we measured serum melatonin (MT), prolactin (PRL) and growth hormone (GH) concentrations, at 08.00 and 24.00, in 132 cancer patients and in 58 healthy control subjects. The patients were stratified according to histology and stage of disease as follows: 30 stage I–II and 45 stage III–IV breast cancer (BC); 39 stage III–IV lung cancer; 18 advanced gastrointestinal (GI) cancer. We also measured MT levels, at the same time-points, in 20 women with primary BC before and after radical mastectomy. Finally, we evaluated the circadian rhythm of serum MT in 18 patients with advanced cancer. On the whole, the patients with advanced tumors showed serum MT levels significantly higher than controls, without any correlation with PRL and GH values. When looking at stage III–IV vs stage I–II BC patients, significantly higher MT levels have been found in the former group. The surgical removal of the primary BC was not associated with any changes in MT values at both time points considered. A highly significant rhythm of serum MT was recorded in advanced cancer patients and the rhythmic parameters were substantially superimposable on those of the control subjects.

We performed a combined stimulation test with the simultaneous application of GnRH (100 micrograms), TRH (200 micrograms) and ACTH (10 micrograms) in 10 healthy adult males at two opposite clock timing, i.e. at 09:00 and 21:00 h. Pituitary (gonadotropins, PRL, TSH) and adrenal (cortisol, aldosterone, progesterone) hormones showed a common trend of enhanced responsiveness to the evening challenge. Differences reached statistical significance in the case of cortisol, aldosterone, PRL and FSH. These findings suggest that the responsiveness of some pituitary and adrenocortical hormones to specific stimuli is physiologically different in humans as a function of the clock timing, being higher in the evening than in the morning. From the clinical standpoint, however, differences in the magnitude of responses were not enough to recommend provocative testing at a particular clock time, at least for routine diagnostic purposes.

We assessed chromogranin A as a tissue biomarker in prostate needle biopsies or as a plasma biomarker, a risk factor for hormone refractory prostate cancer.A total of 211 patients with newly diagnosed prostate cancer treated with luteinizing hormone releasing hormone analogues constituted the study cohort. Univariate and multivariate Cox regression analyses were used to assess the predictive role of tissue and plasma chromogranin A expression.Chromogranin A expression in less than 30% or in 30% or more tumor cells was significantly associated with a shorter time to hormone refractory disease on univariate analysis (HR 2.0, 95% CI 1.3-3.1 and HR 6.0, 95% CI 2.7-12.9), or on multivariate analysis after adjusting for Gleason score, serum prostate specific antigen and disease stage (HR 1.7, 95% CI 1.0-2.8 and HR 3.9, 95% CI 1.7-9.0), respectively. Plasma chromogranin A measured at baseline (HR 3.0, 95% CI 1.8-5.2), and after 1 year (HR 5.8, 95% CI 3.1-10.1) and 2 years (HR 3.5, 95% CI 1.6-7.6), was predictive of hormone refractory risk confirming the tissue results. Plasma as well as tissue chromogranin A expression negatively correlated with overall survival.Chromogranin A expression in prostate cancer biopsies is an independent predictive factor of hormone refractory disease in patients with newly diagnosed prostate cancer on early androgen deprivation therapy. Plasma chromogranin A is also a reliable predictive marker and the predictive significance is maintained over time. These results deserve validation in another data set.

677 Background: Z is an effective bisphosphonate in preventing skeletal related events (SREs) in bone metastatic patients. Decrease in bone resorption markers during Z therapy is a potential surrogate of drug efficacy. Bone resorption markers maintain a circadian rhythmicity in metastatic BC patients (Generali et al ASCO 2005), suggesting that Z may be more active if administered in a chronomodulated way. Raised PTH after Z could impair the drug efficacy (Berruti et al ASCO 2006). Methods: Forty-four BC patients with bone metastases were randomised to receive Z, 4 mg i.v. at 11.00 p.m or 11 a.m every 28 days for 4 times. Serum cross laps (CTX), urinary n-telopeptide (NTX) and serum parathyroid hormone (PTH) levels were measured at baseline and after 4, 7, 14, 28, 56 and 84 days, respectively. Results: Z administration in the night resulted in a greater decrease of either serum CTX or urinary NTX and lower increase in serum PTH ( Table ) than morning administration. Conclusions: Z administration in the night might be more efficacious than morning administration. [Table: see text] No significant financial relationships to disclose.

The presence of neuroendocrine (NE) differentiation in the context of predominantly exocrine prostate cancer may play a key role in androgen-independent tumor growth. The prognostic significance of plasma chromogranin A (CgA) was assessed in a series of consecutive prostate cancer patients with hormone-refractory disease. One hundred and eight patients with newly diagnosed hormone-refractory prostate cancer entered the study. Plasma CgA levels and other biochemical parameters, such as serum prostate specific antigen, serum alkaline phosphatase, serum lactate dehydrogenase, serum albumin and hemoglobin concentration, were measured at baseline (i.e. when hormone refractoriness occurred) and their prognostic role was evaluated together with patient performance status, Gleason score (at diagnosis of prostate cancer) and the presence of visceral metastases. Furthermore, plasma CgA was prospectively evaluated in 50 patients undergoing chemotherapy. At baseline, 45 patients (43.3%) showed elevated CgA values. Plasma CgA negatively correlated with survival, either in univariate analysis (P=0.008) or in multivariate analysis, after adjusting for previously mentioned prognostic parameters (PPP Elevated plasma CgA levels are frequently observed in prostate cancer patients with hormone-refractory disease and correlate with poor prognosis. NE differentiation in hormone-refractory patients is a time-dependent phenomenon and is not influenced by conventional antineoplastic treatments.

Serum parathyroid hormone (PTH), plasma parathyroid hormone related protein (PTHrP) and markers of bone turn-over in serum and urine were monitored monthly, from the start of pamidronate administration, in a breast cancer patient who developed tamoxifen-induced hypercalcaemia (TamIH). At the onset of the hypercalcaemic episode the PTHrP level was three times normal and showed a subsequent decrease until values were below the dection limit by 2 months. A transient rise in bone formation markers appeared during the first month, predicting the subsequent radiological response of bone metastases. Serum and urinary calcium levels significantly decreased but, unexpectedly, an increase in bone collagen resorption indices was observed. Also, serum PTH levels rose considerably, persisting at supranormal levels even after 3 months. This case report indicates that PTHrP could be involved in TamIH. The pamidronate mediated hypocalcaemia is responsible for the PTH rise during the first month, but the elevated PTH level at 3 months was probably due to increased calcium demand as a consequence of calcification occuring in bone. Pamidronate administration is not recommended in the treatment of TamIH; it is short lived and easily controlled by supportive measures.

Summary Background: Chromogranin A (CgA) is a secretory protein present in dense-core vesicles of neuroendocrine (NE) cells. Its ubiquitous presence in NE tissues makes it a suitable circulating marker of neoplasms of NE origin. Patients and Methods: Plasma CgA was determined in 178 patients with NE tumors and in 36 patients with non-endocrine malignancies. Circulating CgA was also serially evaluated in 39 NE cancer patients with advanced disease submitted to systemic therapy and in 14 patients with no evidence of disease (NED). Results: Supranormal CgA values were found in 81% of patients with advanced NE tumors and in only 91% of NED cases. Plasma CgA in patients with well differentiated NE tumors, such as carcinoids, carcinoma of gastrointestinal tract, pheocromocytoma, pancreatic NE carcinoma (either functioning or not functioning), medullary thyroid carcinoma and NE tumors from various primary sites, was higher and more frequently elevated than in patients with small-cell lung cancer (P < 0.001). Plasma CgA did not discriminate patients with NE from those with non NE neoplasms since it was found elevated in 44% of the latter cases. Plasma CgA pattern correlated with the disease response in patients submitted to cytotoxic treatment and with changes in clinical symptomathology in patients receiving somatostatin analogs. Conclusions: Our data confirm that CgA is the best circulating neuroendocrine marker available up to now available for the management of differentiate d neuroendocrine malignancies irrespective of tumor location and functional status. CgA plasma levels could also identify the coexistence of neuroendocrine differentiation in the context of non-endocrine malignancies. Circulating CgA seems to be less useful in undifferentiated tumors such as small-cell lung cancer.

Background: Neuroendocrine (NE) differentiation of prostate adenocarcinoma has received increasing attention in recent years as a result of possible implications for prognosis and therapy. The presence of NE tumor subpopulation can be gauged non invasively by measuring circulating levels of secretory products, primarily chromogranin A (CgA). Methods: This article provides a review on published papers evaluating circulating CgA in prostate cancer patients. Results: Circulating CgA levels were found to be higher in prostate cancer patients than in patients with benign or premalignant prostatic diseases. In patients with malignancy, they correlated either to the stage of disease or to the condition of hormone refractoriness. CgA levels did not correlate with serum prostate specific antigen (PSA) and were supranormal in the majority of advanced patients with PSA within normality. In hormone refractory cases, elevated CgA was a significant predictor of poor prognosis, independently from serum PSA. CgA values were not substantially affected by either endocrine therapy or chemotherapy. They were found to increase during androgen deprivation in some cases and this trend preceded that of PSA. The administration of somatostatin analog in hormone refractory cases was able to reduce plasma CgA values consistently. Conclusions: Present data suggest a potential role of circulating CgA in the management of prostate cancer patients. CgA determination may be useful diagnostically and prognostically and could offer complementary information with respect to PSA. Serial evaluation of circulating CgA could provide information on changes in the NE phenotype expression as a consequence of tumor progression and/or treatment administration.

BACKGROUND. Circulating neuroendocrine markers were measured in patients with prostate carcinoma (PC), prostatic intraepithelial neoplasia (PIN), and benign prostatic hypertrophy (BPH) with the goal to: 1) evaluate the differences in the expression of these markers in patients with benign, premalignant, and primary or metastatic PC; 2) evaluate their prognostic significance; 3) compare values in patients with hormone-naive and hormone-refractory disease; and 4) assess changes after androgen deprivation or chemotherapy. METHODS. Serum neuron specific enolase (NSE) (immunoradiometric assay) and plasma chromogranin A (CgA) (enzyme-linked immunoadsorbent assay) were evaluated in 141 patients with BPH, 54 patients with PIN, and 159 patients with PC; 119 patients were bearing hormone-naive disease and 40 were bearing hormonerefractory disease. CgA was monitored in 31 patients submitted to androgen deprivation and in 24 patients receiving chemotherapy. RESULTS. Supranormal CgA was observed more frequently in patients with American Urologic Association (AUA) Stage D2 disease (45.5%) compared with those with Stage D1 disease (33.3%), Stage C disease (16.7%), Stage A/B disease (18.8%), PIN (25.9%), and BPH (17.0%) (P , 0.02). Supranormal NSE did not change in any of the patient subgroups. Elevated CgA was observed in 36.0% of patients with metastases who had hormone-naive disease and in 45.0% of patients with hormone-refractory disease (P value not significant). Supranormal NSE and CgA values were predictors for poor prognosis in patients with hormone-refractory disease. Elevated baseline CgA values decreased . 50% in 1 of 12 patients who received luteinizing hormone-releasing hormone analogs and in 2 of 12 patients who underwent chemotherapy. CONCLUSIONS. CgA appears to reflect the neuroendocrine activity of PC better than NSE. Elevated CgA values correlate with poor prognosis and are scarcely influenced by either endocrine therapy or chemotherapy. Cancer 2000;88:2590 ‐7. © 2000 American Cancer Society.

To provide preliminary data on whether the diagnostic role of serum prostate specific antigen (PSA) in assessing the response to treatment is improved by concomitant free PSA evaluation both markers were evaluated in 42 patients with advanced prostate cancer who received hormonal therapy and 57 with hormone refractory disease who received chemotherapy.PSA was assessed at baseline and every 3 months during treatment. Free PSA was assessed in stored serum samples obtained at baseline and at maximum PSA decrease. Free PSA was not measurable in 17 patients who received androgen deprivation (40.5%) and 2 who received chemotherapy (3.5%) because it was less than 1.5 ng./ml.Of the 21 patients with greater than 50% PSA decrease after hormonal therapy free-to-total PSA increased in 12 (57.2%) and decreased in 9 (42.9%). Of the 20 patients with PSA response after chemotherapy free-to-total PSA increased in 18 (90.0%) and decreased in 2 (10.0%). Free-to-total PSA increased in 12 of the 20 patients (60.0%) with PSA stabilization after chemotherapy. Patients with an increase in free-to-total PSA after chemotherapy had greater survival compared to those with a decrease or no change (19.8 versus 15.5 months, respectively, p0.03).These data suggest that an effective cytotoxic regimen mainly affects the protein bound PSA fraction. The absence of a clear predominant pattern of free-to-total PSA in patients with PSA response to hormonal therapy and the high percentage of hormone sensitive patients in whom free PSA was not assessable at maximum PSA decrease suggest that free PSA evaluation is less useful in prostate cancer patients undergoing androgen deprivation.

We evaluated the 24-hour profiles of serum melatonin (MT) and cortisol in 12 patients with Cushing’s syndrome and those of serum MT and growth hormone (GH) in 11 patients with acromegaly. The respective age-matched control groups consisted of 22 and 14 healthy subjects. In all cases blood samples were collected at 4-h intervals for a 24-h cycle. The presence of a circadian rhythmicity was assessed by the cosinor analysis, while the diurnal and nocturnal amounts of MT secretion were evaluated in terms of area under the curve (AUC). Spearman’s coefficients of correlation were also calculated. Gross alterations of MT rhythm were not apparent in the Cushing’s patients; pairwise correlations between MT and cortisol and between MT and ACTH (in cases with an ACTH-secreting pituitary adenoma) were not significant. In the acromegalics, two different MT patterns were observed: elevated MT levels with mantained nycthemeral changes in 5 out of 11 patients and normal MT levels with blunted nycthemeral changes in the others. As a group, the acromegalics showed a 24-h MT profile compatible with enhanced secretion, particularly during the day-time period. No significant correlation was found between MT and GH.

Objective: To assess correlation between type of breast cyst and risk of breast cancer in women with gross cystic disease of the breast. Design: Cohort study of women with breast cysts aspirated between 1983 and 1993 who were followed up until December 1994 for occurrence of breast cancer. Setting: Major cancer prevention centre. Subjects: 802 women with aspirated breast cysts. Main outcome measures: Type of breast cyst based on cationic content of cyst fluid: type I (potassium:sodium ratio >1.5), type II (potassium:sodium ratio Results: After median follow up of six years (range 2-12 years) 15 cases of invasive breast cancer and two ductal carcinomas in situ were diagnosed in the cohort: 12 invasive cancers (and two carcinomas in situ) among the 417 women with type I cysts, two cancers among the 325 women with type II cysts, and one among the 60 women with mixed cysts. The incidence of breast cancer in women with type I cysts was significantly higher than that in women with type II cysts (relative risk 4.62 (95% confidence interval 1.26 to 29.7)). These results were confirmed after adjustment for several risk factors for breast cancer (relative risk 4.24 (1.12 to 27.5)). Conclusions: The increased risk of breast cancer of women with breast cysts seems to be concentrated among women with type I breast cysts. Key messages Several studies have shown that women with palpable cysts in their breasts are at increased risk of breast cancer Two types of breast cyst can be identified–type I cysts, with low concentrations of sodium and high concentrations of potassium ions, and type II cysts, with opposite characteristics We investigated the correlation between cyst type and risk of breast cancer in 802 women with aspirated breast cysts After median follow up of six years, the women had a relative risk of breast cancer of 1.69 compared with the general population, and those with type I cysts had a risk four times higher than those with type II cysts The excess risk of breast cancer of women with breast cysts seems to be concentrated among women with type I cysts, but the size and duration of this increased risk are still to be assessed

In order to study the relationship between circulating levels of CA 15-3 and the disease extent in predicting survival, we prospectively followed 312 breast cancer (BC) patients, from October 1988 to March 1995, from the time of first relapse. CA 15-3 values were assessed before treatment onset. Disease extent was defined as the percentage of liver or lung involvement and the number of bone segments positive at scintigraphy. The covariates were primary tumour characteristics (T, N and hormone receptor status) and patient characteristics at recurrence (menopause, performance status and age). Higher CA 15-3 serum levels were found in patients with visceral metastases or with pleural effusion. A logistic regression model selected disease extent in liver, lung and bone as independent variables for the determination of abnormal CA 15-3 values. Univariate survival analysis confirmed the positive prognostic influence of low CA 15-3 serum levels, absence of visceral metastases and the presence of only one metastatic site. Multivariate Cox's survival analysis selected disease extent in liver, lung, bone and soft tissue but not level of CA 15-3 as prognostic factors. In conclusion, CA 15-3 is not an independent variable in determining survival, its prognostic role being linked to the disease extent. This association suggests that CA 15-3 may be useful in assessing disease extent when this is not easily assessable.

BACKGROUND The so-called Bone Hunger Syndrome is a metabolic derangement that sometimes complicates the natural history of prostate cancer patients with osteoblastic bone metastases. An excessive bone formation leads to calcium entrapment in bone and the subsequent increase of parathyroid hormone (PTH) levels, in response to calcium demand. PTH elevation stimulates the osteoclasts in sites distant from those involving the tumor, leading to osteomalacia. METHODS PTH and markers of bone turnover were monitored every 3 weeks, from the start of pamidronate treatment in a prostate cancer patient with progressive disease, to luteinizing hormone releasing hormone analog (LHRH-A) administration, developing hyperparathyroidism, hypophosphatemia, and albumin corrected serum calcium close to the lower limit of normality. Serum bone alkaline phosphatase (BALP), assessed by two different methods: electrophoretic and immunoradiometric, and urinary levels of markers of bone collagen breakdown were also remarkably elevated. RESULTS As a consequence of pamidronate infusion (60 mg e.v. every 3 weeks for a total of four times), BALP and PTH decreased consistently, serum calcium and phosphorus returned within the normal range, while markers of collagen resorption showed a significant decrease at the 9th week, preceded by a transient rise. CONCLUSIONS This case report indicates that bisphosphonates could inhibit both osteoclast and osteoblast activity. The anti-osteoblastic effect is mainly responsible for the improvement of the pretreatment calcium imbalance of our patient towards hypocalcemia and the consequent hyperparathyroidism. Prostate 33:252–255, 1997. © 1997 Wiley-Liss, Inc.

The evaluation of response of osseous metastases to systemic treatments is often low as a consequence of the different radiologic appearances that make objective assessment not only difficult but sometimes impossible. Radiographic evidence of recalcification, the UICC criterion of response, is often evident for 6 months and sometimes may be delayed even more. This accounts for lower response rates in bone with respect to other metastatic sites in clinical trials. A transient rise in bone formation indices may provide an early indication of bone healing and, along with measurement of symptomatic changes, could ameliorate the response evaluation. Among the biochemical markers of bone formation, total alkaline phosphatase (TALP) is widely employed, but it lacks specificity. Estimation of bone isoenzyme (E-BALP) by electrophoretic techniques is time consuming and semiquantitative. The immunoradiometric assay (I-BALP) seems to overcome these limitations. In this study, we compared the two methods of bone isoenzyme estimation with each other and with the levels of bone gla protein (BGP) and carboxyterminal propeptide of type I procollagen (PICP) in a group of 136 cancer patients with bone metastases stratified as having lytic or mixed and blastic lesions at X-ray, and in 62 cancer patients without apparent bone involvement. The same indices were also evaluated prospectively in a patient subset submitted to chemotherapy associated with pamidronate. The aims of the study were to evaluate whether I-BALP is superior to E-BALP and whether both methods of bone isoenzyme estimation are more advantageous than TALP, BGP, and PICP in the assessment of osteoblast activity either in baseline conditions or in response to treatment. In bone metastatic patients with lytic appearances, values above the cut-off limit were observed in 32.1%, 23.3%, 48.9%, 32.9%, and 14% for, TALP, E-BALP, I-BALP, PICP, and BGP, while the corresponding percentages in those with blastic/mixed appearances were 74.0%, 84.8%, 76.9%, 51.9%, and 43.8%, respectively. In the patients without bone involvement, values within the normal range were 90.2%, 98.2%, 89.6%, 71.7%, and 90.2%, respectively. Levels of TALP, E-BALP, and I-BALP were reciprocally correlated in the three groups examined. In bone metastatic patients, however, the degree of correlation of the enzymes with PICP and BGP was weak. Liver isoenzyme of alkaline phosphatase (LALP) was found to correlate with E-BALP, but not with I-BALP, in patients with mixed/blastic lesions. Thirty-eight patients were submitted to pamidronate therapy (60 mg every 3 weeks, administered 4 times) in association with cytotoxic treatment. Osteoblastic markers were determined before any administration. Serum TALP, E-BALP, and I-BALP showed a transient rise in 9 cases, a progressive reduction in 12, no change in 2, and a progressive increase in 6. Changes in E-BALP and I-BALP from baseline were greater than those of TALP. A divergent pattern between TALP and both I-BALP and E-BALP was found in 9 patients, whereas a divergent temporal profile between the two methods of bone isoenzyme estimation was recorded in only 3 patients. Eight out of 38 cases obtained a partial recalcification of lytic and mixed lesions. Seven of them showed the concomitant early increase in TALP, E-BALP, and I-BALP followed by a gradual decline (osteoblastic flare), whereas 1 patient demonstrated the flare of E-BALP and I-BALP but not of TALP. No relationship was found between response and temporal changes in in BGP and PICP serum levels. We conclude that I-BALP is a useful marker for detecting excess osteoblastic activity in patients who have at imaging ``pure'' lytic bone metastases. In the longitudinal evaluation of patients receiving multiple pamidronate infusions plus chemotherapy, TALP, E-BALP, and I-BALP, but not BGP and PICP, appeared to be useful to identify responders in bone. A slight advantage of measurements of serum bone isoenzyme (by both techniques) over TALP is apparent, but this study fails to demonstrate a clear superiority of I-BALP over E-BALP.

The epidemiologic determinants of the mammographic pattern were studied in 710 patients with aspirated gross (> 1 ml) cysts of the breast. The prevalence of the mammograms classified as P2-DY, that are considered to be associated with an increased breast cancer risk, was 636/710 or 89.6%. No relationship between mammographic patterns and characteristics of breast cyst fluid such as K+/Na+ ratio, apocrine changes and dehydroepiandrosterone sulfate concentration was observed. A significant decrease in the proportion of these patterns with increasing age (p = 0.006), Quetelet Index (p < 0.001), parity (p = 0.001), and in postmenopausal women (p = 0.026) was found. Conversely, P2-DY patterns were significantly associated with a later age at menarche (p = 0.023) and alcohol consumption (p = 0.001). In multivariate analysis, an independent association with age was not observed whereas the associations with age at menarche, parity, and relative weight were confirmed. In conclusion, the epidemiologic determinants of mammographic patterns are the same in Gross Cystic Disease patients as in unaffected women, and the lack of correlation between mammographic pattern and cyst type suggests that the latter may represent an independent predictor of breast cancer risk.

The role of circulating tumour markers in providing prognostic information has been scarcely studied. We evaluated the prognostic significance of two mucinous markers: CA 15-3 and CA 125 in 115 breast cancer patients at first recurrence of disease. At diagnosis of advanced disease bone involvement was found in 64 patients, lung in 57, skin lymph nodes in 21, liver in 20, and brain in 5. Patients were recruited and treated in the same institution with conventional chemo- or endocrine therapy. The follow-up ranged from 3 to 54+ months (median 35). Serum samples were drawn at first recurrence of disease before the start of any endocrine and/or chemotherapy. Patients with CA 15-330 U/ml survived significantly longer than those with CA 15-330 U/ml (median 50+ versus 26 months, P0.02). Similarly, overall survival of patients with CA 12535 U/ml was significantly higher in comparison with patients with CA 12535 U/ml (median 34.5 versus 18.5 months, P0.001). CA 125, but not CA 15-3, maintained its prognostic value in the subgroup of patients with visceral metastases. Both markers were found to be independent prognostic variables in multivariate analysis according to Cox's model. CA 15-3 and CA 125 appeared to be powerful prognostic indicators, in addition to visceral metastases, in patients with advanced breast cancer.

Patients bearing macrocysts of the breast are at higher risk of later developing cancer. The fluid filling the cysts (breast cysts fluid, BCF) contains unusual amounts of steroid conjugates, first androgen and estrogen sulfates. Measuring BCF cations (K+, Na+) allows categorization of cysts into two major subsets (type I and type II) that are associated with a different degree and/or turnover of apocrine metaplastic cells in the lining epithelium. Type I cysts (high K+/ Na+ ratio) accumulate huge amounts of dehydroepiandrosterone sulfate, estrone sulfate, androstane-3α, 17β-diol glucuronide, androsterone glucuronide and contain more testosterone and dihydrotestosterone than type II. Conversely, type II cysts (low K+/Na+ ratio) contain more progesterone and pregnenolone. A cohort study was started in 1983 at the Cancer Prevention Center, Ravenna, Italy, with the aim of evaluating the relationships between the biochemistry of BCF and the incidence of breast cancer in women with gross cystic disease (GCD) of the breast. The bimodal distribution of the cationic pattern has been confirmed from data obtained in 798 patients aspirated. The risk of cyst relapse was significantly higher among women with type I cysts or with multiple cysts at presentation. Twelve incident cases of breast cancer have been diagnosed among women whose BCF was categorized. Eleven out of 12 cases had type I or multiple cysts. The cumulative incidence of breast cancer among patients bearing type I cysts was 2.5%. We conclude that women with GCD bearing type I cysts have an increased breast cancer risk when compared with the counterpart bearing type II cysts or the general population.

We measured serum concentrations of bone Gla-protein (osteocalcin, BGP) and carboxyterminal propeptide of type I procollagen (PICP) in 14 patients with active acromegaly. Blood was collected at 0800 for measurement of bone Gla-protein (BGP), carboxyterminal propeptide of type I procollagen (PICP), and insulin-like growth factor I (IGF-I); growth hormone (GH) was then determined at 15-minute intervals for 3 hours and the integrated mean was calculated. The same protocol was repeated at regular intervals during treatment with the long-acting somatostatin analog, octreotide, 150-450 micrograms/day for 6-33 months (median 15). In a case-control analysis, serum BGP concentrations recorded in the acromegalic patients were significantly elevated (14.2 +/- 4.2 micrograms/liter versus 8.0 +/- 3.3 micrograms/liter, P < 0.001). Octreotide treatment induced a roughly parallel reduction in serum GH, IGF-I, and BGP. We found a significant positive correlation between BGP levels recorded before and during therapy and the logarithm of corresponding mean GH levels (r = 0.67, P < 0.001). Also IGF-I concentrations were positively correlated with BGP (r = 0.66, P < 0.001). On the other hand, PICP levels recorded in the acromegalics did not differ from control subjects (146 +/- 46 micrograms/liter versus 127 +/- 44 micrograms/liter, NS) and no correlation was found between either GH and PICP or IGF-I and PICP. To conclude, the present data are compatible with the view that GH and IGF-I play an important role in the control of BGP but not PICP production. It could be that BGP and PICP are submitted to different hormonal modulation.

In all mammals tested so far melatonin (MT), the main pineal hormone, is secreted according to a large-amplitude circadian rhythm. Its profile of secretion is characterized by a brisk surge during the period of darkness with the peak in the first hours after midnight and decline before dawn; during the day-light hours serum MT concentrations are persistently low (Preslock,1984).

Six healthy adult male volunteers underwent serial blood drawings at 4-hour intervals over 24 hours for the definition of melatonin (MT), prolactin (PRL), cortisol, and testosterone circadian patterns. Serum levels of triiodotironine (T3) and thyroxine (T4) were determined at 0800. Systolic and diastolic blood pressure and heart rate were automatically recorded every 30 minutes for 24 hours. The responses of luteinizing hormone (LH), follicle stimulating hormone (FSH), PRL, thyroid stimulating hormone (TSH), cortisol, and aldosterone to a stimulation test with gonadotrophin-releasing hormone (Gn-RH), thyrotrophin-releasing hormone (TRH), adrenocorticotrophin (ACTH), and testosterone to human chorionic gonadotrophin (HCG) were also evaluated. The same protocol was repeated after a two-month course of treatment with MT, 2 mg per os daily at 1800. After treatment, we recorded a marked elevation of mean serum MT levels with a significant phase-advance of its circadian rhythm. The 24-hour patterns of cortisol and testosterone displayed an anticipation of the morning acrophase of about 1.5 hour (not significant) for cortisol and three hours (P less than 0.05) for testosterone. PRL pattern was unchanged as well as serum levels of thyroid hormones. The circadian organization of the cardiovascular variables did not show any changes after MT supplementation; the pituitary, adrenal, and testicular responses to specific stimuli were comparable before and after treatment. These results are compatible with the view that the MT signal may provide temporal cues to the neuroendocrine network for the organization of testicular circadian periodicity.

14091 The primary objective of this study was to validate the clinical significance of blood evaluation of CgA in NET patients at the diagnosis (PHASE I) and during 2 years of follow-up (PHASE II). From May 2003 to October 2004, 276 patients entered the study from 40 Italian centers: 270 were evaluable. All basal and every 3 months collected CgA blood samples were centrally measured in two reference laboratories (Orbassano-Turin and Venice) where ELISA (DAKO,Denmark) or IRMA (CIS-Schering, France) were performed to look at the correlation between the two methods and their sensitivity and specificity. Lab results at the baseline have been recently published (Leon et al., Intern. J. Biol. Markers, 2005). We are now collecting all the correlations between CgA and type and place of NETs; tumor bulk; metastatization; presence or not of specific syndrome;proliferation activity (Ki67); octreoscan; tumor specific markers. 223 patients (83%) had gastroenteropancreatic tumors, whereas 24 (9%) had medullary tyroid cancer, 16 (6%) Merkel cell carcinoma and 6 parathyroid NETs, pheochromocytoma, paraganglioma. Only 26% of GEP tumors presented with specific symptoms. At the entry in the study 58% of patients had a new diagnosis, 23% were in stable disease, whereas 18% had metastatic disease. According to the recent W.H.O. histologic classification (Solcia et al, 2000), 36% specific symptomatic patients had NE tumor, 57% well differentiated cancer and only 3% poor differentiated cancer, whereas 31% not symptomatic patients had NE tumor, 48% well differentiated and 16% poor differentiated cancer. This is the largest study worldwide performed on this topic and all the data about the correlation among all patient variables and CgA blood values will be ready in April 2006. Follow-up data will be evaluable next year. [Table: see text]

737 Background: Circadian rhythmicity is an essential feature of bone metabolism. The assessment of diurnal variation in bone resorption and formation markers in cancer patients with bone metastase...

The prognostic value of CA 15-3 in BC patients at first relapse of disease may be ascribed to the correlation of marker levels with DE or with the tumor differentiation and aggressiveness ( Eur J Cancer, 1994 ). From Oct. 1988 to Sept. 1994 we recorded data from 293 BC patients. CA 15-3 serum levels and DE were separately evaluated before treatment onset. The DE of visceral metastases (mts) was assessed considering the percentage of organ involvement (no mts, 50%). Bone tumor load was defined as the number of skeletal segments positive at scintigraphy. Pleural effusion and lymphangitis were separately considered. Serum CA 15-3 was higher in patients with visceral involvement and with pleural effusion than in those with bone or soft tissue mts. CA 15-3 significantly correlated with liver, pulmonary and bone DE but not with soft tissue DE. Only liver and bone DE demonstrated to be independent variables at multiple regression analysis. Elevated CA 15-3 levels predicted poor survival either in overall patients or in those with visceral involvement. Performance status, menopause, DE in liver and in bone were found to be independence prognostic factors when Cox regression analysis was performed, while CA 15-3 failed to enter the model. To conclude, all these data suggest that the prognostic role of CA 15-3 is linked to the extent of disease.

Ketoconazole is an inhibitor of adrenal steroidogenesis used in the treatment of Cushing's disease. Previous data obtained with single blood sampling were controversial as to increased ACTH levels compensatory to the cortisol fall. We have evaluated by chronobiological procedures the circadian profiles of plasma ACTH and cortisol in three patients with Cushing's disease before and after a six-month course of therapy with ketoconazole (600 mg daily). None of the patients complained of any adverse subjective reaction; in particular no sign or symptom of hypoadrenalism and/or hepatotoxicity was recorded. Ketoconazole treatment markedly improved the clinical setting and caused a highly significant (p less than 0.0001) reduction of mean 24-h cortisol values (ciradian MESOR). The expected rise of ACTH did not take place; rather, we detected a slight decrease of the mean circadian MESOR (p less than 0.05). Our data, althought obtained in a very small number of patients, suggest that ketoconazole may have an additional action at central level, at least in some cases of Cushing's disease.

The morphology of apocrine cells exfoliated in breast cyst fluid (BCF) was studied in 78 BCF samples obtained from 39 premenopausal patients with gross cystic disease who were bearing two simultaneously aspirated cysts. 57/78 samples showed cell clusters suitable for computer-assisted cytometry. This was performed on 5820 cells using a Leitz Texture Analysis System (TAS). We measured the surface areas of cytoplasm, nucleus and nucleolus; we also calculated the nuclear/cytoplasmic (N/C), nuclear/nucleolar (N/n) ratios and the nuclear roundness factor (RF). Cysts were divided according to the cationic pattern of BCF: Type I, K+/Na+ greater than 1.5; Type II, K+/Na+ less than 0.66. The cytometric analysis was made on 47 samples of Type I and 10 samples of Type II. At the light microscope, no difference was apparent between the apocrine cells coming from Type I or Type II cysts. Cytometric measurements showed significant differences for the apocrine cells aspirated from Type I vs. Type II cysts for the mean cytoplasmic area (97.13 +/- 24.28 S.D. mu2 vs. 59.66 +/- 14.90 S.D. mu2, respectively) and the mean nucleolar area (4.35 +/- 0.99 S.D. mu2 vs. 2.75 +/- 0.71 S.D. mu2, respectively). Our data do not allow the inference of apocrine changes in the epithelium lining the cysts simply from the cationic pattern of BCF. The significantly wider cytoplasm and nucleoli of the apocrine cells aspirated from Type I cysts could reflect different functional stages of these particular cells.

Cations (K+ and Na+) content was evaluated in 444 breast cyst fluid (BCF) specimens, aspirated from 391 patients with gross cystic disease of the breast (GCD), a benign form admittedly at major risk of cancer. In 306/444 BCF, dehydroepiandrosterone-sulfate (DHA-S) content was also evaluated. A positive correlation (P less than 0.001) was observed between log K+ vs. log DHA-S whereas a negative correlation was found between log Na+ and log DHA-S (P less than 0.001). Cysts were subdivided in three types according to their cationic concentration: most were of type I (K+/Na+ greater than 1.5) and type II (K+/Na+ less than 0.66) whereas only 10% was of the type III (intermediate). No statistical difference in subtype distribution was apparent when considering patients aspirated in the follicular vs. luteal phase of the menstrual cycle; on the contrary, a significant difference (P less than 0.001) was found between menstruating vs. menopausal patients (type I = 54.8% vs. 32.2%; type II = 34.5% vs. 58.1%, respectively). Ninety-four BCF samples were aspirated simultaneously in 41 patients bearing multiple cysts: the same cationic subtype was present in 29/41 patients. Our data confirm and extend previous observations, and provide conclusive evidence that breast macrocysts can be divided on the basis of their electrolyte composition into different types. Accordingly, the composition of BCF should always be assessed for prospective studies on GCD and breast cancer risk.

We evaluated the effect of a single oral administration of 100 mg melatonin (MT) vs placebo (PL) on the pituitary release of LH, FSH, TSH and prolactin (PRL) after GnRH + TRH and on the adrenocortical release of cortisol, aldosterone and progesterone after ACTH in healthy adult males. We carried out a double blind study in 6 volunteers in winter, at two opposite stages of the circadian cycle: 08:00 and 20:00 h. Injection of GnRH (100 micrograms), TRH (200 micrograms) and ACTH (10 micrograms of the synthetic ACTH 1-17 analogue, Alsactide) was performed one h after MT or PL ingestion. Plasma MT levels were 200-4,000-fold higher after MT than PL thus confirming the effective gastrointestinal absorption of the pineal hormone. The hormonal patterns were superimposable after MT and PL. A higher response of PRL, FSH and cortisol was observed in the evening vs morning protocols independently of previous MT or PL. Our data demonstrate that the acute oral administration of a pharmacological dose of MT at two opposite circadian stages is ineffective to change a variety of pituitary and adrenocortical responses in human male subjects. The circadian chronosusceptibility of pituitary and adrenocortical cells to specific stimuli deserves interest to future investigation.

Gross cystic disease of the breast (GCD) is rarely seen after the menopause. Recent work has shown that by measuring electrolytes in the breast cyst fluid (BCF) it is possible to identify two principal classes of cyst, designated Type 1 (K+/Na+ greater than 1.5) and Type 2 (K+/Na+ less than 0.66). A smaller, intermediate class (Type 3) also appears to exist. We measured K+, Na+ and dehydroepiandrosterone sulphate (DHA-S) in 38 BCF samples aspirated from 33 women with GCD who had undergone spontaneous menopause at least 1 yr previously. Statistically significant correlations were found between DHA-S and cations (positive in relation to K+, P less than 0.001; negative in relation to Na+, P less than 0.001). The distribution of cyst types was shifted with respect to that characteristic of cases that occur at an earlier age: whereas Type 1 cysts predominate in menstruating women, Type 2 cysts proved more numerous in the post-menopausal subjects. The difference was statistically validated (P less than 0.001). The results seem to indicate a sharp reduction in high K+, high DHA-S cysts after the menopause, which may be paralleled by a decrease in the associated apocrine metaplasia. In view of the major biochemical differences between Type 1 and Type 2 cysts and of the suggested differences as regards cancer risk, classification of post-menopausal patients with GCD by cyst type is critical prior to any clinical trial or follow-up.

Several methods for analyzing CgA using either monoclonal or polyclonal antibodies have been developed, which differ in their diagnostic performance. The present paper describes the results of a prospective multicenter study aimed at comparing the clinical value of the two most widely used commercially available CgA assay kits in patients affected by neuroendocrine tumors (NETs). Two hundred sixty-one patients from 40 different centers and 99 healthy subjects were evaluated. CgA levels were measured with two different methods, a two-step immunoradiometric assay (IRMA) and an enzyme-linked immunosorbent assay (ELISA). CgA was measured centrally by two reference laboratories, one of which used IRMA and the other ELISA, and it was measured by the participating institutions with the method routinely used by each of them. The major findings of the present study were: (i) the two assays for the determination of CgA present good diagnostic performance; (ii) both assays are robust and guarantee comparable results when applied in different settings (central vs local laboratory); (iii) the negative/positive cutoff points (87 ng/mL for IRMA and 21.3 U/L for ELISA) were established according to standardized criteria; (iv) the results obtained with the two assays in basal clinical samples of patients affected by NETs show an apparently satisfactory correlation (rs=0.843, p