Chemotherapy toxicity and activity in patients with pancreatic ductal adenocarcinoma and germline BRCA1-2 pathogenic variants (gBRCA1-2pv): a multicenter survey.

BACKGROUND: Germline BRCA1-2 pathogenic variants (gBRCA1-2pv)-related pancreatic ductal adenocarcinoma (PDAC) showed increased sensitivity to DNA cross-linking agents. This study aimed at exploring safety profile, dose intensity, and activity of different chemotherapy regimens in this setting. PATIENTS AND METHODS: gBRCA1-2pv PDAC patients of any age and clinical tumor stage who completed a first course of chemotherapy were eligible. A descriptive analysis of chemotherapy toxicity, dose intensity, response, and survival outcomes was performed. RESULTS: A total of 85 gBRCA1-2pv PDAC patients treated in 21 Italian centers between December 2008 and March 2021were enrolled. Seventy-four patients were assessable for toxicity and dose intensity, 83 for outcome. Dose intensity was as follows: nab-paclitaxel 72%, gemcitabine 76% (AG); cisplatin 75%, nab-paclitaxel 73%, capecitabine 73%, and gemcitabine 65% (PAXG); fluorouracil 35%, irinotecan 58%, and oxaliplatin 64% (FOLFIRINOX). When compared with the literature, grade 3-4 neutropenia, thrombocytopenia, and diarrhea were increased with PAXG, and unmodified with AG and FOLFIRINOX. RECIST responses were numerically higher with the three- (81%) or four-drug (73%) platinum-containing regimens that outperformed AG (41%) and oxaliplatin-based doublets (56%). Carbohydrate antigen 19.9 (CA19.9) reduction >89% at nadir was reported in two-third of metastatic patients treated with triplets and quadruplets, as opposed to 33% and 45% of patients receiving oxaliplatin-based doublets or AG, respectively. All patients receiving AG experienced disease progression, with a median progression-free survival (mPFS) of 6.4 months, while patients treated with platinum-containing triplets or quadruplets had an mPFS >10.8 months. Albeit still immature, data on overall survival seemed to parallel those on PFS. CONCLUSIONS: Our data, as opposed to figures expected from the literature, highlighted that platinum-based regimens provoked an increased toxicity on proliferating cells, when dose intensity was maintained, or an as-expected toxicity, when dose intensity was reduced, while no change in toxicity and dose intensity was evident with AG. Furthermore, an apparently improved outcome of platinum-based triplets or quadruplets over other regimens was observed.

Germinal BRCA1-2 pathogenic variants (gBRCA1-2pv) and pancreatic cancer: epidemiology of an Italian patient cohort.

OBJECTIVE: Germline BRCA1-2 pathogenic variants (gBRCApv) increase the risk of pancreatic cancer and predict for response to platinating agents and poly(ADP-ribose) polymerase inhibitors. Data on worldwide gBRCApv incidence among pancreatic ductal adenocarcinoma (PDAC) patients are sparse and describe a remarkable geographic heterogeneity. The aim of this study is to analyze the epidemiology of gBRCApv in Italian patients. MATERIALS AND METHODS: Patients of any age with pancreatic adenocarcinoma, screened within 3 months from diagnosis for gBRCApv in Italian oncologic centers systematically performing tests without any selection. For the purposes of our analysis, breast, ovarian, pancreas, and prostate cancer in a patient's family history was considered as potentially BRCA-associated. Patients or disease characteristics were examined using the χ2 test or Fisher's exact test for qualitative variables and the Student's t-test or Mann-Whitney test for continuous variables, as appropriate. RESULTS: Between June 2015 and May 2020, 939 patients were tested by 14 Italian centers; 492 (52%) males, median age 62 years (range 28-87), 569 (61%) metastatic, 273 (29%) with a family history of potentially BRCA-associated cancers. gBRCA1-2pv were found in 76 patients (8.1%; 9.1% in metastatic; 6.4% in non-metastatic). The gBRCA2/gBRCA1 ratio was 5.4 : 1. Patients with gBRCApv were younger compared with wild-type (59 versus 62 years, P = 0.01). The gBRCApv rate was 17.1% among patients <40 years old, 10.4% among patients 41-50 years old, 9.2% among patients 51-60 years old, 6.7% among patients aged 61-70 years, and 6.2% among patients >70 years old (none out of 94 patients >73 years old). gBRCApv frequency in 845 patients <74 years old was 9%. Patients with/without a family history of potentially BRCA-associated tumors had 14%/6% mutations. CONCLUSION: Based on our findings of a gBRCApv incidence higher than expected in a real-life series of Italian patients with incident PDAC, we recommend screening all PDAC patients <74 years old, regardless of family history and stage, due to the therapeutic implications and cancer risk prevention in patients' relatives.

Intrahepatic cholangiocarcinoma development in a patient with a novel BAP1 germline mutation and low exposure to asbestos.

BRCA1 associated protein-1 (BAP1) germline mutations define a novel hereditary cancer syndrome, namely BAP1 tumor predisposition syndrome (BAP1-TPDS), characterized by an increased susceptibility to develop different cancer types, including mesothelioma, uveal and cutaneous melanoma, renal cell carcinoma, and basal cell and squamous cell carcinoma. Currently, the role of BAP1 germline mutations in intrahepatic cholangiocarcinoma (iCCA) pathogenesis is less known. Here we report the first clinical case of a female patient who developed an iCCA when she was 47-years-old and was found to carry a novel germline mutation at a splicing site of exon 4 in BAP1 gene (NM_004656.4: c.255_255+6del). An accurate anamnesis revealed the absence of risk factors linked to iCCA development, except for a low occupational exposure to asbestos. In tumor tissue, BAP1 sequencing, multiplex ligation-dependent probe amplification and immunoistochemistry showed the loss of heterozygosity and lack of nuclear expression, suggesting that BAP1 wild-type allele and functional protein were lost in cancer cells, in line with the classical two-hit model of tumor suppressor genes. Further studies are needed to confirm whether iCCA may be included into BAP1-TPDS cancer phenotypes and whether minimal asbestos exposure may facilitate the development of this malignancy in individuals carrying BAP1 germline mutations.

Membrane human equilibrative nucleoside transporter 1 is associated with a high proliferation rate and worse survival in resected intrahepatic cholangiocarcinoma patients not receiving adjuvant treatments.

Human equilibrative nucleoside transporter 1 (hENT-1) is a membrane nucleoside transporter mediating the intracellular uptake of nucleosides and their analogues. hENT-1 was recently reported to have a predictive role in intrahepatic cholangiocarcinoma (iCC) patients receiving adjuvant gemcitabine-based chemotherapy, but its biological and clinical significance in iCC remains unsettled. This study investigated the role of hENT-1 in regulating tumour growth and predicting the survival of 40 resected iCC patients not receiving adjuvant treatments. hENT-1 expression was found to be significantly higher in iCC than in the matched non-tumoural liver. Patients harbouring hENT-1 localised on the tumour cell membrane had a worse overall survival than membrane hENT-1-negative patients (median 21.2 months vs 30.3 months, p = 0.031), with an adjusted hazard ratio of 2.8 (95% confidence interval 1.01-7.76). Moreover, membrane hENT-1-positive patients had a higher percentage of Ki67-positive cells in tumour tissue than membrane hENT-1-negative patients (median 23% vs 5%, p < 0.0001). Functional analyses in iCC cell lines revealed that hENT-1 silencing inhibited cell proliferation and induced apoptosis in HUH-28 cells expressing hENT-1 on the cell membrane, but not in SNU-1079 cells expressing the transporter only in the cytoplasm. Overall, these findings suggest that membrane hENT-1 is involved in iCC proliferation and associated with worse survival in resected iCC patients. Further prospective studies on larger cohorts are required to confirm these results and better define the potential prognostic role of membrane hENT-1 in this setting of patients.

An update of treatments of hepatocellular carcinoma in patients refractory to sorafenib.

Hepatocellular carcinoma (HCC) remains among the neoplastic diseases with the most unfavorable prognosis. Historically, systemic treatments for HCC have been scarce. In particular, sorafenib was the only registered drug for the treatment of unresectable HCC until 2017. Last year, regorafenib was approved by the global regulatory agencies as second-line therapy. Since then, further randomized, controlled trials have been successful in this patient population. This paper deals with the most recent data concerning cabozantinib and ramucirumab, the two drugs that have recently demonstrated efficacy in phase III, randomized, controlled trials in HCC patients who failed previous treatment with sorafenib.

Regorafenib for the treatment of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a worldwide problem, with a high prevalence in nonindustrialized countries and a rising incidence in industrialized countries as well. Its close association with chronic liver diseases and liver cirrhosis represents a significant challenge in its treatment. A front-line systemic treatment for unresectable cases of HCC (sorafenib) was identified only in 2007. Following a decade of failed clinical trials with a wide range of drugs for second-line treatment, regorafenib proved its efficacy as a second-line treatment in 2016, when the randomized, placebo-controlled, phase III RESORCE trial demonstrated a meaningful increase in overall survival in the regorafenib treatment arm compared with the placebo arm (10.6 vs. 7.8 months). In this monograph we review the main preclinical and clinical findings in the trials assessing regorafenib for the treatment of HCC patients.

TB and MDR/XDR-TB in European Union and European Economic Area countries: managed or mismanaged?

In spite of the growing awareness of emerging drug-resistant Mycobacterium tuberculosis, the extent of inappropriate tuberculosis (TB) case management may be underestimated, even in Europe. We evaluated TB case management in the European Union/European Economic Area countries, with special focus on multidrug-resistant (MDR) and extensively drug-resistant (XDR)-TB, using a purposely developed, standardised survey tool. National reference centres in five countries representing different geographical, socioeconomic and epidemiological patterns of TB in Europe were surveyed. 40 consecutive, original clinical TB case records (30 MDR/XDR-TB cases) were reviewed in each of the five countries. The findings were recorded and, through the survey tool, compared with previously agreed and identified international standards. Deviations from international standards of TB care were observed in the following areas: surveillance (no information available on patient outcomes); infection control (lack of respiratory isolation rooms/procedures and negative-pressure ventilation rooms); clinical management of TB, MDR-TB and HIV co-infection (inadequate bacteriological diagnosis, regimen selection and treatment duration); laboratory support; and diagnostic/treatment algorithms. Gaps between present international standards of care and the management of MDR/XDR-TB patients were identified. Training, increased awareness, promotion of standards and allocation of appropriate resources are necessary to ensure appropriate care and management as well as to prevent further emergence of drug resistance.

Improving signal/noise resolution in single-molecule experiments using molecular constructs with short handles.

We investigate unfolding/folding force kinetics in DNA hairpins exhibiting two and three states with newly designed short dsDNA handles (29 bp) using optical tweezers. We show how the higher stiffness of the molecular setup moderately enhances the signal/noise ratio (SNR) in hopping experiments as compared to conventional long-handled constructs (≅700 bp). The shorter construct results in a signal of higher SNR and slower folding/unfolding kinetics, thereby facilitating the detection of otherwise fast structural transitions. A novel analysis, as far as we are aware, of the elastic properties of the molecular setup, based on high-bandwidth measurements of force fluctuations along the folded branch, reveals that the highest SNR that can be achieved with short handles is potentially limited by the marked reduction of the effective persistence length and stretch modulus of the short linker complex.

Presynaptic inhibition of spontaneous acetylcholine release mediated by P2Y receptors at the mouse neuromuscular junction.

At the neuromuscular junction, ATP is co-released with the neurotransmitter acetylcholine (ACh) and once in the synaptic space, it is degraded to the presynaptically active metabolite adenosine. Intracellular recordings were performed on diaphragm fibers of CF1 mice to determine the action of extracellular ATP (100 muM) and the slowly hydrolysable ATP analog 5'-adenylylimidodiphosphate lithium (betagamma-imido ATP) (30 muM) on miniature end-plate potential (MEPP) frequency. We found that application of ATP and betagamma-imido ATP decreased spontaneous secretion by 45.3% and 55.9% respectively. 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX), a selective A(1) adenosine receptor antagonist and alpha,beta-methylene ADP sodium salt (alphabeta-MeADP), which is an inhibitor of ecto-5'-nucleotidase, did not prevent the inhibitory effect of ATP, demonstrating that the nucleotide is able to modulate spontaneous ACh release through a mechanism independent of the action of adenosine. Blockade of Ca(2+) channels by both, Cd(2+) or the combined application of nitrendipine and omega-conotoxin GVIA (omega-CgTx) (L-type and N-type Ca(2+) channel antagonists, respectively) prevented the effect of betagamma-imido ATP, indicating that the nucleotide modulates Ca(2+) influx through the voltage-dependent Ca(2+) channels related to spontaneous secretion. betagamma-Imido ATP-induced modulation was antagonized by the non-specific P2 receptor antagonist suramin and the P2Y receptor antagonist 1-amino-4-[[4-[[4-chloro-6-[[3(or4)-sulfophenyl] amino]-1,3,5-triazin-2-yl]amino]-3-sulfophenyl] amino]-9,10-dihydro-9,10-dioxo-2-anthracenesulfonic acid (reactive blue-2), but not by pyridoxal phosphate-6-azo(benzene-2,4-disulfonic acid) tetrasodium salt (PPADS), which has a preferential antagonist effect on P2X receptors. Pertussis toxin and N-ethylmaleimide (NEM), which are blockers of G(i/o) proteins, prevented the action of the nucleotide, suggesting that the effect is mediated by P2Y receptors coupled to G(i/o) proteins. The protein kinase C (PKC) antagonist chelerythrine and the calmodulin antagonist N-(6-aminohexil)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7) occluded the effect of betagamma-imido ATP, while the protein kinase A (PKA) antagonist KT-5720 and the inhibitor of the calcium/calmodulin-dependent protein kinase II (CAMKII) KN-62 failed to do so. betagamma-Imido ATP did not affect 10, 15 and 20 mM K(+)-evoked release and application of reactive blue-2 before incubation in high K(+) induced a higher asynchronous secretion. Thus, our results show that at mammalian neuromuscular junctions, ATP induces presynaptic inhibition of spontaneous ACh release due to the modulation of Ca(2+) channels related to tonic secretion through the activation of P2Y receptors coupled to G(i/o) proteins. We also demonstrated that at increasing degrees of membrane depolarization evoked by K(+), endogenously released ATP induces presynaptic inhibition as a means of preventing excessive neurotransmitter secretion.

Recombinant IFN-alpha2b treatment activates poly (ADPR) polymerase-1 (PARP-1) in KB cancer cells.

In the present paper, we investigated the relationship between the growth inhibitory effects of recombinant interferon-alpha2b (rIFN-alpha2b) and poly (ADPR) polymerase-1 (PARP-1) activity in the human squamous KB cancer cell line. Growth inhibition of the KB cells mediated by 1000 IU/ml of rIFN-alpha2b was accompanied by a transient rise in PARP-1 specific activity 24 h after rIFN-alpha2b treatment, confirmed by both the increase of intracellular poly (ADP-ribose) content and the PARP-1 auto-modification level. At longer times of incubation, the onset of apoptosis accompanied KB cell growth inhibition, as demonstrated by both flow cytometry and western-blotting analysis showing an 89 kDa apoptotic fragment of PARP-1. Moreover, pretreatment of the cells with the PARP-1 inhibitor, 3-aminobenzamide (3-ABA), at non-cytotoxic concentrations (1 mM), reduced the cell-growth inhibition, cell-cycle perturbation and apoptosis caused by rIFN-alpha2b. Taken together, these results strongly suggest that PARP-1 may be directly involved in the effects of rIFN-alpha2b in the KB cancer cell line.

[Cytomegalovirus infection in the adult]. / Infezione da Cytomegalovirus nell'adulto.

Aim of this report is to review the epidemiological and clinical features of HCMV infection in the adult. In all geographical areas high diffusion of HCMV infection involving all socioeconomic groups is observed; significant most elevated seroprevalence in North African and Asian ethnic groups is compared to Western populations is pointed out; besides, HCMV is absolutely the virus most frequently transmitted in the perinatal period. In the immunocompetent host, the HCMV infection is symptomless in the great majority of cases; in the symptomatic cases it shows the clinical features of a self-limited mononucleosis-like syndrome. In the immunocompromised patients, either in subjects infected with HIV or in onco-hematologic patients or recipients of solid organ or bone marrow transplants patients, HCMV infection leads to serious illness. The most frequent clinical pictures are: pneumonia, retinitis, hepatitis, polyradiculopathy, encephalitis, gastrointestinal tract disease, adrenal involvement; cases of myocarditis, pancreatitis, genitourinary localizations are less frequent. The clinical pictures are different in the different clinical groups: retinitis, in subjects with HIV infection and pneumonia in recipients of transplants, are respectively the main clinical manifestations; the mechanisms of such differences are not clearly defined. A to the diagnosis, the serological tests (evidence of IgM activity, IgG avidity) are useful in the immunocompetent host; whereas, in the immunocompromised host cytological detection (demonstration of typical cytological aspects and positive immunohistology for HCMV antigens) and/or virological detection (isolation of virus or evidence of viral antigens or viral DNA) are needed. The most used therapeutical choices are ganciclovir, foscarnet and cidofovir; these three drugs have similar antiviral effectiveness, but they show different outlines of toxicity and praticality of use.

Gemcitabine combined with continuous infusion 5-fluorouracil in advanced and symptomatic pancreatic cancer: a clinical benefit-oriented phase II study.

Gemcitabine and 5-fluorouracil are the only two compounds with reproducible activity against advanced pancreatic cancer (APC). We have evaluated a novel combination of gemcitabine and 5-fluorouracil on the clinical benefit response (CBR) end point. Eleven consecutive patients with symptomatic APC were entered in a two-stage phase II trial. Gemcitabine was administered by intravenous (i.v.) bolus injection at the dose of 1,000 mg m(-2) on days 1, 8, 15 and 5-fluorouracil 500 mg m(-2) was given by continuous i.v. infusion on days 1-5. Treatment was repeated every 28 days. A CBR was achieved in 7/11 patients. The mean time to loss of CBR was 26.5 weeks (range 14-18, median 22). Toxicity was mild and no APC patient experienced WHO grade 3 toxicity. The gemcitabine/5-fluorouracil combination is well tolerated and produces a symptomatic relief in the majority of APC patients.

Antitumor activity of sequential treatment with topotecan and anti-epidermal growth factor receptor monoclonal antibody C225.

Epidermal growth factor (EGF)-related proteins such as transforming growth factor alpha (TGF-alpha) control cancer cell growth through autocrine and paracrine pathways. Overexpression of TGF-alpha and/or its receptor (EGFR) has been associated with a more aggressive disease and a poor prognosis. The blockade of EGFR activation has been proposed as a target for anticancer therapy. Monoclonal antibody (MAb) C225 is an anti-EGFR humanized chimeric mouse MAb that is presently in Phase II clinical trials in cancer patients. Previous studies have suggested the potentiation of the antitumor activity of certain cytotoxic drugs, such as cisplatin and doxorubicin, in human cancer cell lines by treatment with anti-EGFR antibodies. We have evaluated in human ovarian, breast, and colon cancer cell lines, which express functional EGFR, the antiproliferative activity of MAb C225 in combination with topotecan, a cytotoxic drug that specifically inhibits topoisomerase I and that has shown antitumor activity in these malignancies. A dose-dependent supraadditive increase of growth inhibition in vitro was observed when cancer cells were treated with topotecan and MAb C225 in a sequential schedule. In this respect, the cooperativity quotient, defined as the ratio between the actual growth inhibition obtained by treatment with topotecan followed by MAb C225 and the sum of the growth inhibition achieved by each agent, ranged from 1.2 to 3, depending on drug concentration and cancer cell line. Treatment with MAb C225 also markedly enhanced apoptotic cell death induced by topotecan. For example, in GEO colon cancer cells, 5 nM topotecan, followed by 0.5 microg/ml MAb C225, induced apoptosis in 45% cells as compared with untreated cells (6%) or to 5 nM topotecan-treated cells (22%). Treatment of mice bearing established human GEO colon cancer xenografts with topotecan or with MAb C225 determined a transient inhibition of tumor growth because GEO tumors resumed the growth rate of untreated tumors at the end of the treatment period. In contrast, an almost complete tumor regression was observed in all mice treated with the two agents in combination. This determined a prolonged life span of the mice that was significantly different as compared with controls (P < 0.001), to MAb C225-treated group (P < 0.001), or to the topotecan-treated group (P < 0.001). All mice of the topotecan plus MAb C225 group were the only animals alive 14 weeks after tumor cell injection. Furthermore, 20% of mice in this group were still alive after 19 weeks. The combined treatment with MAb C225 and topotecan was well tolerated by mice with no signs of acute or delayed toxicity. These results provide a rationale for the evaluation of the anticancer activity of the combination of topoisomerase I inhibitors and anti-EGFR blocking MAbs in clinical trials.

Tuberculosis and Pelger-Huët anomaly. Case report.

We describe a patient with pulmonary tuberculosis and a rare disturbance of leukocyte segmentation, known as "Pelger-Huët anomaly", which can be observed in various diseases such as malignancies and/or infections. The importance of this association is equivocal: some authors have related to the association the particular severity of tuberculosis or the death they observed; in the case reported we noted no evidence of such a relation, notwithstanding the presence of the homozygous form of the Pelger-Huët anomaly. We suggest therefore that, when Pelger-Huët anomaly is found, an underlying disease should be searched for; the course of this illness, however, might not be affected.

Desferioxamine increases iron depletion and apoptosis induced by ara-C of human myeloid leukaemic cells.

We investigated whether changes in iron metabolism and the transferrin receptor (TRF-R) expression were involved in the antileukaemic effects of arabinoside cytosine (ara-C). Treatment with 100 nM ara-C for 48h reduced thymidine uptake and increased the surface expression of the TRF-R on leukaemic blasts derived from 13/16 (81%) patients and on the HL-60 and U-937 cell lines. Whereas intracellular non-haem iron was strongly depleted 24 h after ara-C addition, TRF-R up-regulation and recovery of intracellular non-haem iron concentration occurred together after a longer exposure of the cultured cells to the drug. Since iron is an essential regulator of cell proliferation we have evaluated the effects of the combination between ara-C and the iron chelator desferioxamine (DSF) on the growth of HL-60 and U-937 cells. We found that desferioxamine strongly potentiated the effects of ara-C on leukaemic cell growth inhibition and apoptosis. This is the first report of a positive interaction between ara-C and an iron chelator in terms of antileukaemic effects.

Use of an amino-cupric-silver technique for the detection of early and semiacute neuronal degeneration caused by neurotoxicants, hypoxia, and physical trauma.

A new amino-cupric silver protocol is described for detection of neuronal degeneration. We describe its selectivity in visualizing both early and semiacute degeneration after intracerebral or systemic administration of a variety of neurotoxicants in rats, and after transient ischemic episodes in gerbils. As early as 5 min after physical trauma, or 15 min following either intrastriatal injections of glutamate analogs or exposure to ischemic episodes, neuronal silver staining was evident at primary sites of trauma (i.g. injection sites) and at hodologically related secondary sites. With intoxication by peripheral injections of trimethyltin (IP) or intracerebral injections of Doxorubicin, reproducible patterns of degeneration are demonstrable after 24 h or after 9-13 days, respectively. The amino-cupric silver method permits simultaneous detection of all neuronal compartments against a clear background. Degeneration in the neuronal cell bodies, dendrites, axons and terminals, as well as the recruitment of new structures in a progressive pathologic process, could be accurately followed. The inclusion of new reagents increased the sensitivity vis-à-vis previous versions of the cupric-silver method. The advantages and disadvantages of the current method in comparison with other means of neurotoxic assessment are discussed in detail, with special emphasis on its unique ability to discriminate irreversible degenerative phenomena and degeneration of axonal components in cases where the cell body remains apparently intact. The amino-cupric silver method is an especially useful tool for surveying neuronal damage in basic neuroscience investigations and in neuropathologic and neurotoxic assessment.

[Diagnostic, prognostic and therapeutic problems of cystic pneumatosis of the colon]. / Problemi diagnostici, prognostici e terapeutici della pneumatosi cistica del colon.

Two cases of cystic pneumatosis of the colon raised clinical problems that were mainly concerned with the diagnostic difficulties posed by this disease. These can only be solved by correct employment of radiology and endoscopy. Reference is made to the proper therapeutic approach, which must be moderate and conservative in all cases. An account is given of the pathogenesis of this form and support is expressed for the most commonly held theory, namely that mechanical factors are responsible for the penetration of air into the wall of the intestine.