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Objectives: Autologous hematopoietic stem cell transplantation (HSCT) is a treatment option which may be considered for severe diffuse cutaneous systemic sclerosis (dcSSc) patients not responding to cyclophophamide (CY). We present two cases of dcSSc not responding to CY >10 g who were successfully treated with HSCT. Patients and methods: Two dcSSc patients were unresponsive to monthly i.v. pulse of CYC (0.75 g m2). Both patients had significant reduction of DLCO and mild-moderate pulmonary hypertension and HSCT was considered due to the rapid progression of the disease. Following informed consent and ethics committee approval, HSCT was performed. Mobilisation was performed with CY 4g/m2 and recombinant human granulocyte colony stimulating factor (rHu GCSF) followed by a successful apheresis (CD34+ cells, >7X106). Conditioning regimens were: CY 100mg/kg body weight plus thiotepa 10 mg/ kg in the first patient and CY 200 mg/kg in the second. Both graft products were CD34 selected. No arrythmias occurred during the procedure and no other severe side effects were observed during hospitalisation. Results: Follow up: Patients underwent a monthly follow up with physical examination, pulmonary function tests and echocardiography every 3 months. Chest CT has been performed 6 months post transplantation. The following was observed: skin score (from 40 to 10 for the first patient and from 38 to 12 for the second one), LVEF and pulmonary function remained stable, PAP decreased from 45 mmHg to 35 mmHg and from 40 to 32 mmHg. No late complications or cardiac toxicity was observed. Conclusion: These two dcSSc cases demonstrate that HSCT may be successfully performed without serious side effects in cases in whom despite a cumulative CY dose was ineffective. This suggests an “immunological threshold” effect which may be exploited in other severe, therapy refractory autoimmune cases.

Objective: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) identified in China, in December 2019 determines COronaVIrus Disease 19 (COVID-19). Whether or not the virus was present in Italy earlier the first autochthonous COVID-19 case was diagnosed is still uncertain. We aimed to identify anti-SARS-CoV-2 antibodies in sera collected from 4th November 2019 to 9th March 2020, in order to assess the possible spread of the virus in Italy earlier than the first official national diagnosis., Patients and Methods: Anti-SARS-CoV-2 antibodies were evaluated in retrospective serum samples from 234 patients with liver diseases (Hep-patients) and from 56 blood donors (BDs). We used two rapid serologic tests which were confirmed by a validated chemoluminescence assay., Results: Via rapid tests, we found 10/234 (4.3%) IgG-positive and 1/234 (0.4%) IgM-positive cases in the Hep-patient group. Two/56 (3.6%) IgG-positive and 2/56 (3.6%) IgM-positive cases were detected in BD group. Chemoluminescence confirmed IgG-positivity in 3 Hep-patients and 1 BD and IgM-positivity in 1 Hep-patient. RNAemia was not detected in any of the subjects, rendering the risk of transfusion transmission negligible., Conclusions: Our results suggest an early circulation of SARS-CoV-2 in Italy, before the first COVID-19 cases were described in China. Rapid tests have multiple benefits; however, a confirmation assay is required to avoid false positive results.

Objective: Terlipressin has been successfully used as rescue treatment in hypotensive adults and children with septic shock, but only exceptionally in neonates. The aim of this study is to describe original clinical scenarios in which terlipressin, in newborns and infants, resolved the catecholamine-refractory hypotension., Design: Retrospective study., Setting: Neonatal intensive care unit., Patients: All newborns with hypotension unresponsive to volume replacement and catecholamines, and treated with terlipressin, from January 2008 to December 2009. In this study, also an infant (11 months old) born extremely preterm was included., Interventions: None., Measurements and Main Results: Four hypotensive patients received as rescue therapy terlipressin, which produced a dramatic increase in mean arterial pressure, diuresis, and reduction of lactate levels. In three newborns, hypotension, associated with pulmonary hypertension, was resolved with terlipressin. Two of them (one with systemic inflammatory response syndrome, the other with congenital diaphragmatic hernia) died in the following days for causes unrelated to hypotension; the third (on mild hypothermia for hypoxic-ischemic encephalopathy) recovered. We report furthermore an infant with septic shock and on treatment with β-blockers in whom terlipressin normalized blood pressure. In two patients, cranial Doppler ultrasonography showed the recovery of diastolic cerebral flow in the anterior cerebral artery and the normalization of resistance index within 30 mins from the first dose of terlipressin. In two infants, hyponatremia was detected., Conclusion: Although the number of reported infants is little, terlipressin appears to be an effective rescue treatment in different scenarios of refractory neonatal hypotension. Further controlled studies are required to confirm its efficacy and safety.

The chemolithoautotrophic bacterium Acidithiobacillus ferrooxidans is of great importance in biomining operations. During the bioleaching of ores, microorganisms are subjected to a variety of environmental stresses and to the limitations of some nutrients, such as inorganic phosphate (P(i)), which is an essential component for all living cells. Although the primary source of phosphorus for microorganisms is P(i), some bacteria are also able to metabolize P(i) esters (with a C-O-P bond) and phosphonates (with a very inert C-P bond). By using bioinformatic analysis of genomic sequences of the type strain of A. ferrooxidans (ATCC 23270), we found that as part of a Pho regulon, this bacterium has a complete gene cluster encoding C-P lyase, which is the main bacterial enzyme involved in phosphonate (Pn) degradation in other microorganisms. A. ferrooxidans was able to grow in the presence of methyl-Pn or ethyl-Pn as an alternative phosphorus source. Under these growth conditions, a great reduction in inorganic polyphosphate (polyP) levels was seen compared with the level for cells grown in the presence of P(i). By means of reverse transcription-PCR (RT-PCR), DNA macroarrays, and real-time RT-PCR experiments, it was found that A. ferrooxidans phn genes were cotranscribed and their expression was induced when the microorganism was grown in methyl-Pn as the only phosphorus source. This is the first report of phosphonate utilization in a chemolithoautotrophic microorganism. The existence of a functional C-P lyase system is a clear advantage for the survival under P(i) limitation, a condition that may greatly affect the bioleaching of ores.

Background: Controlled-rate freezing (CRF) followed by storage in liquid nitrogen is employed by most centers as the standard procedure for peripheral blood progenitor cell (PBPC) cryopreservation. Uncontrolled-rate freezing (URF) at -80 degrees C is more simple, time-saving, less expensive, and, possibly, as effective as CRF. The aim of this retrospective analysis was to compare CRF and URF in childhood transplantation., Study Design and Methods: A total of 54 PBPC transplants performed in 39 children aged 3 to 16 years (median, 9.5 years) were analyzed: 23 transplants in 16 children with CRF versus 31 transplants performed in 23 children with -80 degrees C URF. All grafts contained at least 2 x 10(6) per kg unselected CD34+ cells, enumerated before freezing. Nucleated cells infused ranged from 1.32 x 10(8) to 4.3 x 10(8) per mL with a median of 3.1 x 10(8) per mL. Cryoprotectant solution consisted of a final dimethyl sulfoxide (DMSO) concentration of 10 percent DMSO with autologous plasma., Results: The two study groups did not differ in terms of timing of neutrophil and platelet recovery or transfusion requirements. Adverse events related to graft infusion, severe complications, and transplant-related mortality were not significantly different between CRF and URF groups. In both groups only mild adverse events were observed during graft administration. URF procedures, however, were simpler and less expensive. At a median follow-up of 72 months, no secondary myelodysplasia was observed in either group., Conclusion: Our analysis suggests that URF is safe and effective in the pediatric population.

Steganacin and podophyllotoxin are two naturally occurring lignans first isolated from plant sources, which share the capability to disrupt tubulin assembly. Although not strictly essential for its activity, the lactone ring on both structures represents Achilles' heel, as it is a potential site of metabolic degradation and epimerization on its C2 carbon brings about a significant loss in potency. In the present manuscript, we have used the ruthenium-catalyzed [3+2] azide-alkyne cycloaddition, a click-chemistry reaction, to replace the lactone ring with a 1,5-disubstituted triazole in few synthetic steps. The compounds were cytotoxic, although to a lesser degree compared to podophyllotoxin, while retaining antitubulin activity. The present structures might therefore represent a good platform for the fast generation of metabolically stable compounds with few stereogenic centers that might be of value from a medicinal chemistry point of view.

Autologous hematopoietic stem cell transplantation (AHSCT) has been proposed as a rescue treatment in multiple sclerosis (MS) patients not responding to first- or second-line therapies. To date, most of the treated cases had a secondary progressive disease course. However, patients with high inflammatory activity, but no secondary progression of the disease, could be candidates to take greater advantage of AHSCT. In this paper, we report two cases with very active, relapsing-remitting (RR) MS, who underwent AHSCT, and obtained a dramatic resolution to disease activity.

Over the last decade, hematopoietic stem cells transplantation (HSCT) has been increasingly used in the treatment of severe progressive autoimmune diseases. We report a retrospective survey of 183 multiple sclerosis (MS) patients, recorded in the database of the European Blood and Marrow Transplantation Group (EBMT). Transplant data were available from 178 patients who received an autologous graft. Overall, transplant related mortality (TRM) was 5.3% and was restricted to the period 1995-2000, with no further TRM reported since then. Busulphan-based regimens were significantly associated with TRM. Clinical status at the time of transplant and transplant techniques showed some correlations with toxicity. No toxic deaths were reported among the 53 patients treated with the BEAM (carmustine, etoposide, cytosine-arabinoside, melphalan)/antithymocyte globulin (ATG) regimen without graft manipulation, irrespective of their clinical condition at the time of the transplant. Improvement or stabilization of neurological conditions occurred in 63% of patients at a median follow-up of 41.7 months, and was not associated with the intensity of the conditioning regimen. In this large series, HSCT was shown as a promising procedure to slow down progression in a subset of patients affected by severe, progressive MS; the safety and feasibility of the procedure can be significantly improved by appropriate patient selection and choice of transplant regimen.

We have synthesized rigid analogues of combretastatin bearing a furan ring in place of the olefinic bridge. These compounds are cytotoxic at nanomolar concentrations in neuroblastoma cells, display a similar structure-activity relationship compared to combretastatin A4, and inhibit tubulin polymerization. We also show that the furan ring can be further functionalized. Thus, it is possible that combretafurans could act as scaffolds for the development of dual-action antitumoral agents.

Resveratrol is a phytoalexin able to display an array of biological activities. We decided to replace the double bond with a triazole ring using the archetypical click reaction: the Huisgen [3 + 2] cycloaddition. Seventy-two triazole derivatives were synthesized via a parallel combinatorial approach. Preliminary data suggest that this procedure can lead to the synthesis of compounds that display some, but not all, of resveratrol's actions with increased potency.

Objectives: Autologous hematopoietic stem cell transplantation (HSCT) is a treatment option which may be considered for severe diffuse cutaneous systemic sclerosis (dcSSc) patients not responding to cyclophophamide (CY). We present two cases of dcSSc not responding to CY >10 g who were successfully treated with HSCT., Patients and Methods: Two dcSSc patients were unresponsive to monthly i.v. pulse of CYC (0.75 g m²). Both patients had significant reduction of DLCO and mild-moderate pulmonary hypertension and HSCT was considered due to the rapid progression of the disease. Following informed consent and ethics committee approval, HSCT was performed. Mobilisation was performed with CY 4 g/m² and recombinant human granulocyte colony stimulating factor (rHu GCSF) followed by a successful apheresis (CD34+ cells, >7X106). Conditioning regimens were: CY 100mg/kg body weight plus thiotepa 10 mg/ kg in the first patient and CY 200 mg/kg in the second. Both graft products were CD34 selected. No arrhythmias occurred during the procedure and no other severe side effects were observed during hospitalisation., Results: Follow up: Patients underwent a monthly follow up with physical examination, pulmonary function tests and echocardiography every 3 months. Chest CT has been performed 6 months post transplantation. The following was observed: skin score (from 40 to 10 for the first patient and from 38 to 12 for the second one), LVEF and pulmonary function remained stable, PAP decreased from 45 mmHg to 35 mmHg and from 40 to 32 mmHg. No late complications or cardiac toxicity was observed., Conclusion: These two dcSSc cases demonstrate that HSCT may be successfully performed without serious side effects in cases in whom despite a cumulative CY dose was ineffective. This suggests an "immunological threshold" effect which may be exploited in other severe, therapy refractory autoimmune cases.

Aggressive forms of multiple sclerosis (MS) represent a limited group of demyelinating diseases that rapidly progress to severe disability. Currently available therapies are poorly effective against these clinical entities. Recently, it has been demonstrated that intense immunosuppression followed by autologous haematopoietic stem cell transplantation (AHSCT) can affect the clinical course of individuals with severe MS and completely abrogate the inflammatory activity detected by MRI. We report the result of the Italian phase 2 GITMO study, a multicentre study in which 21 MS patients, who were rapidly deteriorating and not responding to the usual therapeutic strategies, were treated with this procedure. The clinical effect of the treatment is long lasting, with a striking abrogation of inflammation detected by MRI findings. These results support a role for intense immunosuppression followed by ASCT as treatment in rapidly evolving MS cases unresponsive to conventional therapies.

Background: Growth hormone (GH) treatment in patients with GH deficiency (GHD) can determine changes in the thyroid function. The clinical significance of these changes remains controversial, and all studies have so far covered rather a short period--usually no longer than one year., Objective: To determine the effect of long-term recombinant hGH treatment in children with idiopathic GHD on the thyroid function., Patients and Methods: Nineteen prepubertal children (12 boys and 7 girls, mean age 9.2 +/- 3.1 years) with idiopathic GHD were studied and followed for twenty-four months. None of the patients showed multiple pituitary hormone deficiencies. Nineteen healthy children matched for age and sex acted as controls., Results: Patients with GHD showed a significant increase in TT (3) at twelve months and in FT (3) at six and twelve months after starting GH treatment, with a significant decrease at eighteen and twenty-four months. TT (4) level decreased significantly at twelve months and increased significantly at eighteen and twenty-four months. FT (4) also decreased, but only slightly, after twelve months of hGH treatment, and then increased significantly at twenty-four months. TSH levels did not vary significantly during the course of therapy. TT (3)/TT (4) and FT (3)/FT (4) ratios increased significantly after six and twelve months of therapy and significantly decreased later, approaching pre-therapy values. The SDS of Growth Velocity (SDS-GV) increased remarkably during the first year of therapy and then decreased significantly during the second year, although it remained significantly higher than the pre-therapy values. TT (3) and TT (3)/TT (4) ratio displayed a significant correlation with SDS-GV at twelve months of therapy. In a multiple regression analysis with age, bone age, parental height, GH dose, TT (3,) TT (3)/TT (4), and the SDS of IGF-I, only the TT (3)/TT (4) ratio at twelve months of therapy (p < 0.001) was identified as a significant predictor of SDS-GV., Conclusion: Our data confirm that changes in thyroid function are present in GHD children during long-term hGH therapy. These changes probably resulted from the effect of hGH on the peripheral metabolism of thyroid hormones and appear to be transitory, disappearing during the second year of hGH treatment. We speculate on the functional significance of these changes, and in particular, on their role in catch-up growth after hGH therapy.

A series of chimeric compounds bearing the combretastatin and the nitrogen mustard cores were synthesized. All the compounds were cytotoxic and inhibited tubulin polymerization. When combretastatin was joined to chlorambucil via an ester linkage, the resultant compound proved to be significantly more potent than the two compounds put together. When combretastatin was joined to nitrogen mustard via an ether linkage or when a true hybrid was synthesized, loss of potency was observed. Nonetheless, these latter compounds appeared to be more efficacious and surprisingly were able to inhibit tubulin depolymerization at high concentrations.

Combretastatin A-4 is an antitumoral and antitubulin agent that is active only in its cis configuration. In the present manuscript, we have synthesized cis-locked combretastatins embodying a furazan ring (combretafurazans). To achieve this, we have developed a new strategy that exploits the dehydration of vicinal dioximes using the Mitsunobu reaction. Among the advantages of following such a strategy are the mild conditions used for the construction of the diarylfurazan derivatives, allowing for the presence of highly functionalized substrates and deactivated aromatic rings. Combretafurazans are more potent in vitro cytotoxic compounds compared to combretastatins in neuroblastoma cells, yet maintaining similar structure-activity relationship and pharmacodynamic profiles.

Hematopoietic stem cell transplantation (HSCT) has been proposed for the treatment of severe multiple sclerosis (MS). In a phase 2 multicenter study we selected 19 non-primary progressive MS patients showing high disease activity on the basis of both brain magnetic resonance imaging (MRI) and sustained clinical deterioration despite conventional treatments. After stem cell mobilization with cyclophosphamide (CY) and filgrastim, patients were conditioned with BCNU (1,3-bis(2-chloroethyl)-1-nitrosourea), cytosine arabinoside, etoposide, and melphalan (BEAM) followed by antithymocyte globulin (ATG). Unmanipulated peripheral blood stem cells (PBSCs) were then infused. No maintenance treatment was administered with a median follow-up of 36 months (range, 12 to 72 months). All patients showed clinical stabilization or improvement; 3 subsequently deteriorated, 1 beyond the baseline. No MRI active lesions were detected after the HSCT except in 1 patient who showed a new lesion at 4.5 years. Infections were limited and restricted to 3 months after HSCT. Health-related quality of life was assessed through the 54-item MS quality of life (MSQOL-54) questionnaire, showing a statistically significant improvement in both composite scores and in most of the individual domains. HSCT is able to induce a prolonged clinical stabilization in severe progressive MS patients, resulting in both sustained treatment-free periods and quality of life improvement.

Autologous haematopoietic stem cell transplantation is now a feasible and effective treatment for selected patients with severe autoimmune diseases. Worldwide, over 650 patients have been transplanted in the context of phase I and II clinical trials. The results are encouraging enough to begin randomised phase III trials. However, as predicted, significant transplant-related morbidity and mortality have been observed. This is primarily due to complications related to either the stage of the disease at transplant or due to infections. The number of deaths related to cardiac toxicity is low. However, caution is required when cyclophosphamide or anthracyclines such as mitoxantrone are used in patients with a possible underlying heart damage, for example, systemic sclerosis patients. In November 2002, a meeting was held in Florence, bringing together a number of experts in various fields, including rheumatology, cardiology, neurology, pharmacology and transplantation medicine. The object of the meeting was to analyse existing data, both published or available, in the European Group for Blood and Marrow Transplantation autoimmune disease database, and to propose a safe approach to such patients. A full cardiological assessment before and during the transplant emerged as the major recommendation.

We report the case of a 31-year-old man with anaplastic large-cell lymphoma successfully treated with chemotherapy who showed mediastinal widening 5 months after autologous stem-cell transplantation. CT scan and PET evaluations were consistent with the diagnosis of benign thymic hyperplasia. Because of the rapid and aggressive course of this type of lymphoma, and the progressive widening of the mass at CT scan, we performed a mediastinal biopsy that confirmed these findings, showing normal thymic tissue. This is the first case of benign thymic hyperplasia defined with FDG-PET and confirmed by histologic evaluation.

Background: Autologous hematopoietic stem cell transplantation (ASCT) has been recently utilized with encouraging results in patients with poorly controlled MS., Objective: To determine in severe cases of MS the effect of ASCT on gadolinium (Gd)-enhanced MRI and to obtain information on clinical course and safety., Methods: In a cooperative study, 10 patients with rapidly evolving secondary progressive MS were transplanted, after BEAM conditioning regimen (carmustine, etoposide, cytosine-arabinoside, and melphalan), with unmanipulated autologous peripheral blood SC mobilized with high-dose cyclophosphamide (CY; 4 g/m2) and granulocyte-colony-stimulating factor. Triple-dose Gd-enhanced scans were performed monthly for a pretreatment period of 3 months and compared with serial monthly Gd-enhanced MRI for the following 6 months and then once every 3 months., Results: The median follow-up is now 15 months (range 4 to 30 months). The number of Gd-enhancing lesions decreased immediately after mobilization with CY and finally dropped to zero in all cases after the conditioning regimen. The number of new T2-weighted positive lesions paralleled data obtained for Gd-enhanced MRI. Clinically, patients improved slightly or remained stable., Conclusion: These results demonstrate that the therapeutic sequence CY-BEAM-ASCT has the capacity to completely suppress MR-enhancing activity, an effect that is sustained with time. The final impact of this procedure on disease course remains to be established.

In a series of 20 patients presenting a threat of premature delivery, the vascular resistance index of Pourcelot, in the umbilical artery, was measured before, and then 24 hours after starting a continuous Salbutamol perfusion. A moderate but significant decreased placental vascular resistances is noted under betamimetic treatment. Relaxation of the myometrium obtained with a tocolytic treatment could be implicated in this alteration of placental resistances.

The diagnosis of a thanatophorous nanism is evoked in the presence of an abnormality of the size of the long bones. This study records the symptoms which, on sonograms, permit to recognize this fatal form of nanism among other forms of metaphysis chondrodysplasia, in order to establish a diagnosis before deciding to interrupt the pregnancy.