Drug-induced immunophenotypic modulation in childhood ALL: implications for minimal residual disease detection.

Assessment of minimal residual disease (MRD) by flow cytometry is considered to be based on the reproducibility of the leukemic immunophenotype detected at diagnosis. However, we previously noticed modulation of surface antigen expression in acute lymphoblastic leukemia (ALL) during the early treatment. Hence, we investigated this in 30 children with B-cell precursor ALL consecutively enrolled in the AIEOP-BFM ALL 2000 protocol. Quantitative expression of seven antigens useful in MRD monitoring was studied at diagnosis and compared to that measured at different time points of remission induction therapy. Downmodulation in the expression of CD10 and CD34 occurred at follow-up. By contrast, upmodulation of CD19, CD20, CD45RA, and CD11a was observed, while the expression of CD58 remained stable. Despite this, we could unambiguously discriminate leukemic cells from normal residual B cells. This holds true when bone marrow (BM) samples from similarly treated T-ALL patients, but not from healthy donors, were used as reference. Our results indicate that immunophenotypic modulation occurs in ALL during the early phases of BFM-type protocols. However, the accuracy of MRD detection by flow cytometry seems not negatively affected if adequate analysis protocols are employed. Investigators should take this phenomenon into account in order to avoid pitfalls in flow cytometric MRD studies.

Synthesis and biological evaluation of a novel pyroglutamyl-modified TRH analogue.

The TRH analogue 3, incorporating the (S)-isothiazolidine-1,1-dioxide-3-carboxylic acid (1) moiety in place of the native L-pyroglutamic acid (pGlu) residue, has been synthesized and fully characterized by 1H and 13C NMR. The effects of replacing pGlu with its sulphonamido counterpart on biological activity have been investigated. This peptide, which is significantly stabilized towards hydrolysis by pyroglutamyl peptidase type I (PP I, EC 3.4.19.3), has shown to maintain in vitro prolactin-releasing activity.

Effects of cocaine- and amphetamine-regulated transcript peptide, leptin and orexins on hypothalamic serotonin release.

We have studied the effects of cocaine- and amphetamine-regulated transcript (CART) peptide-(55-102), leptin, orexin-A and orexin-B on basal and depolarization (K(+) 15 mM)-induced serotonin (5-hydroxytryptamine, 5-HT) release from rat hypothalamic neuronal endings (synaptosomes) in vitro. We have found that leptin and CART peptide-(55-102) have no effect on 5-HT release, while orexin-A and orexin-B inhibit depolarization-stimulated serotonin release. We can conclude that leptin and CART peptide-(55-102), which play a physiological role as feeding inhibitors, do not acutely affect 5-HT release from hypothalamic synaptosomes; on the other hand, feeding induced by orexin-A and orexin-B could be partially explained by decreased 5-HT release.

Chromatin-derived acidic peptides modulate catecholamine release in the hypothalamus.

We have studied the neuromodulatory effects of three synthetic peptides, structurally related to chromatin-derived acidic peptides (ACPs): ACP-1 (Asp-Asp-Ser-Asp-Glu-Glu-Asn), corresponding to the C-terminal fragment of the largest subunit of eukaryotic RNA polymerase II; a more lipophilic derivative, ACP-2 (Ala-Ile-Ser-Pro-Asp-Asp-Ser-Asp-Glu-Glu-Asn); and its phosphorylated form ACP-3 (Ala-Ile-Ser-Pro-Asp-Asp-Ser(P)-Asp-Glu-Glu-Asn). Rat hypothalamic synaptosomes, loaded with [(3)H]norepinephrine or [(3)H]dopamine, were perfused with the above peptides, both basally and during a depolarizing stimulus. We have found: ACP-1 inhibited both dopamine and norepinephrine release; ACP-2 inhibited dopamine release, without affecting norepinephrine release; ACP-3 was almost ineffective, except for a weak dopamine inhibiting effect only at a higher concentration.

Synthesis and preliminary pharmacological evaluation of 5-hydroxy- and 5,6-dihydroxy-1,2,3,7,12,12a-hexahydrobenzo[5,6]cyclohepta[1,2,3-ij]isoquinoline derivatives as dopamine receptor ligands.

A series of 5-hydroxy- and 5,6-dihydroxy-1,2,3,7,12,12a-hexahydrobenzo[5,6]cyclohepta[1,2,3-ij]isoquinoline derivatives (5a--e and 6a--e) were synthesized as conformationally rigid analogues of 1-benzyltetrahydroisoquinoline and evaluated for their affinity at D(1) and D(2) dopamine receptors. All compounds showed lower D(1) and D(2) affinities than dopamine. The 5-hydroxy-1-methyl-2,3,12,12a-hexahydrobenzo[5,6]cyclohepta[1,2,3-ij]isoquinoline 5a and the 5,6-dihydroxy analogue 6a showed D(2) agonist activity. This was proved by their effects on prolactin release from primary cultures of rat anterior pituitary cells. Molecular modeling studies showed that the geometric parameters (namely the distances from meta and para hydroxyl oxygens to the nitrogen and the height of nitrogen from the hydroxylated phenyl ring plane) of the dopaminergic pharmacophore embedded in our compounds have lower values in comparison with those observed in D(1) and D(2) selective ligands.

Cocaine- and amphetamine-regulated transcript peptide-(55-102) and thyrotropin releasing hormone inhibit hypothalamic dopamine release.

Cocaine- and amphetamine-regulated transcript (CART) peptide-(55-102) and thyrotropin releasing hormone (TRH) play an anorectic role in the hypothalamus. Catecholamines are also involved in appetite control and we have previously found that leptin, an adipocyte-derived anorectic hormone, inhibits hypothalamic norepinephrine and dopamine release. We have studied the effect of CART peptide-(55-102) and TRH on basal and depolarization (K+ 15 mM)-induced norepinephrine and dopamine release from rat hypothalamic neuronal endings (synaptosomes) in vitro. We have found that basal catecholamine release was not modified; both CART peptide-(55-102) and TRH, the former with a higher sensitivity, dose-dependently inhibited depolarization-induced dopamine release, and did not affect the stimulated norepinephrine release. Considering the role played by dopamine in the central mechanisms of reward, these findings suggest that the inhibition of dopamine release could underlie the decreased appetitive behaviour induced by CART peptide-(55-102) and TRH.

Atypical endometriosis revisited: clinical and biochemical evaluation of the different forms of superficial implants.

OBJECTIVE: To investigate the activity of different forms of endometriosis implants by clinical and biochemical evaluation. DESIGN: Prospective, blinded study. SETTING: Tertiary-care university hospital. PATIENT(S): Forty-seven infertile patients with minimal or mild endometriosis diagnosed at laparoscopy were included in the clinical investigation. INTERVENTION(S): Patients were submitted to a preoperative evaluation of endometriosis-associated dysmenorrhea by means of a 10-point linear analog scale and to laparoscopic staging of endometriosis following a current classification system. In the biochemical investigation, tissue samples from different endometriosis lesions and control tissues were assessed for prostaglandin (PG) F(2 alpha) production. MAIN OUTCOME MEASURE(S): Evaluation of the correlation between endometriosis-associated dysmenorrhea and the extent of the disease. Evaluation of the production of PGF(2 alpha) by the different tissues sampled. RESULT(S): No positive correlation was present between any of the implant subgroups and the associated dysmenorrhea. White peritoneal implants were associated with milder pain symptoms than black or red lesions. The biologic activity of red and black superficial implants, expressed as the production of PGF(2 alpha), was similar. CONCLUSION(S): No positive correlation was demonstrated between endometriosis-associated dysmenorrhea and the current classification of endometriosis, which includes characterization of the different morphologic aspects of superficial endometriosis implants.

A possible prolactin-related adverse effect of certain antineoplastic anthracyclines.

Nonhormonal antineoplastic therapy can affect the endocrine system with consequent effects on the growth of hormone-sensitive tumors. Amongst anthracycline antibiotics, we have found daunorubicin and epirubicin able to acutely stimulate prolactin (PRL) secretion both in vivo, in the rat, and in vitro, from rat anterior pituitary cell cultures. Despite a similar structure, doxorubicin showed no such activity. Considering the possible role of PRL in breast cancer cell proliferation, the effects of certain anthracyclines might be viewed as an adverse drug reaction involving the anterior pituitary gland.

Binding and preliminary evaluation of 5-hydroxy- and 10-hydroxy-2,3, 12,12a-tetrahydro-1H-[1]benzoxepino[2,3,4-ij]isoquinolines as dopamine receptor ligands.

The N-methyl, N-ethyl, and N-n-propyl derivatives of 5-hydoxy- and 10-hydroxy-2,3,12,12a-tetrahydro-1H-[1]benzoxepino[2,3, 4-ij]isoquinolines were prepared as monophenolic ligands for the dopamine receptor and evaluated for their affinity at D(1)-like and D(2)-like subtypes. All compounds showed very low D(1) affinities. This could be ascribed to the absence of a catechol nucleus or of the beta-phenyldopamine pharmacophore. Only the N-methyl-5-hydroxy- (5a), N-methyl-10-hydroxy- (6a), and N-methyl-4-bromo-10-methoxy-2,3, 12,12a-tetrahydro-1H-[1]benzoxepino[2,3,4-ij]isoquinolines (26a) bound the D(2) receptors with low affinity, in the same range as dopamine. In compounds 5a and 6a, the 2-(3-hydroxyphenyl)ethylamine moiety does not meet the requirements of the D(2) agonist pharmacophore: namely, the 2-(3-hydroxyphenyl)ethylamine does not reach the trans, fully extended conformation. The three compounds did not interact with recombinant human D(4) receptors, and only 5a showed low affinity for rat recombinant D(3) receptors. Analysis of the influence of Na(+) on [(3)H]spiperone binding showed that 5a displays a potential dopamine D(2) agonist profile, whereas 6a probably has a dopamine D(2) antagonist activity. The D(2) agonist activity of 5a was proved by the effects on prolactin release from primary cultures of rat anterior pituitary cells.

Leptin inhibits norepinephrine and dopamine release from rat hypothalamic neuronal endings.

Noradrenergic and dopaminergic afferents arising from the brain stem to the hypothalamus still play a poorly characterised role in food intake control. We have studied the effect of leptin, an adipocyte-derived hormone which has been implicated in the regulation of feeding behaviour, on [3H]norepinephrine and [3H]dopamine release from perfused hypothalamic neuronal endings (synaptosomes) in vitro. We have found that leptin (0.01-10 nM) does not modify basal, while it inhibits depolarization-induced norepinephrine and dopamine release. We can hypothesize that at least part of the anorectic activity of leptin in the hypothalamus is effected through an inhibition of noradrenergic and dopaminergic firing.

Leptin stimulates prostaglandin E2 and F2alpha, but not nitric oxide production in neonatal rat hypothalamus.

Leptin, an adipocyte-derived 16 kDa polypeptide hormone, has been found to regulate food intake and thermogenesis by modulating stimulatory and inhibitory pathways in the feeding circuitry of the hypothalamus, among which corticotropin releasing hormone (CRH). Nitric oxide (NO) and prostaglandins have been shown to be involved in both CRH neurosecretion and feeding regulation. We have investigated the role of NO, prostaglandin E2 and prostaglandin F2alpha as mediators of the hypothalamic effects of leptin and their possible involvement in leptin-stimulated CRH secretion. Using primary cultures of neonatal (5- to 6-day-old) rat hypothalamic cells, we confirmed that leptin (0.1-10 nM) stimulates CRH secretion. This effect was not blocked by L-N(G)-nitro-methyl-arginine (L-NAME, 100 microM), a NO-synthase competitive inhibitor; and leptin did not stimulate NO production. Cyclooxygenase inhibition by indomethacin (10 microM) did not modify leptin-induced CRH secretion, while leptin stimulated prostaglandin E2, and prostaglandin F2alpha secretion. In conclusion, leptin-induced hypothalamic CRH secretion is not modulated by NO-synthase- or cyclooxygenase-mediated mechanisms; leptin does not stimulate NO production, but it stimulates prostaglandin E2 and F2alpha production, which could add to the growing list of mediators of leptin signaling in the hypothalamus.