Gender oncology: recommendations and consensus of the Italian Association of Medical Oncology (AIOM).

BACKGROUND: Following the development of gender medicine in the past 20 years, more recently in the field of oncology an increasing amount of evidence suggests gender differences in the epidemiology of cancers, as well as in the response and toxicity associated with therapies. In a gender approach, critical issues related to sexual and gender minority (SGM) populations must also be considered. MATERIALS AND METHODS: A working group of opinion leaders approved by the Italian Association of Medical Oncology (AIOM) has been set up with the aim of drafting a shared document on gender oncology. Through the 'consensus conference' method of the RAND/University of California Los Angeles (UCLA) variant, the members of the group evaluated statements partly from the scientific literature and partly produced by the experts themselves [good practice points (GPPs)], on the following topics: (i) Healthcare organisation, (ii) Therapy, (iii) Host factors, (iv) Cancer biology, and (v) Communication and social interventions. Finally, in support of each specific topic, they considered it appropriate to present some successful case studies. RESULTS: A total of 42 articles met the inclusion criteria, from which 50 recommendations were extracted. Panel participants were given the opportunity to propose additional evidence from studies not included in the research results, from which 32 statements were extracted, and to make recommendations not derived from literature such as GPPs, four of which have been developed. After an evaluation of relevance by the panel, it was found that 81 recommendations scored >7, while 3 scored between 4 and 6.9, and 2 scored below 4. CONCLUSIONS: This consensus and the document compiled thereafter represent an attempt to evaluate the available scientific evidence on the theme of gender oncology and to suggest standard criteria both for scientific research and for the care of patients in clinical practice that should take gender into account.

Evaluation of ATG5 polymorphisms in Italian patients with systemic lupus erythematosus: contribution to disease susceptibility and clinical phenotypes.

Systemic lupus erythematosus (SLE) is a common heterogeneous autoimmune disease that is caused by the involvement both of genetic and environmental factors. There is evidence that autophagy is involved in several aspects of SLE pathogenesis. In particular, polymorphisms in the ATG5 gene have been observed to be associated with disease susceptibility. Our aim was to verify if ATG5 polymorphisms are involved in the susceptibility to disease and its clinical phenotypes in an Italian cohort of SLE patients. This study involved 315 SLE patients and 265 healthy controls. Three polymorphisms in the ATG5 gene (rs573775, rs6568431 and rs2245214) were investigated by allelic discrimination assay. A case-control association study, a genotype/phenotype correlation analysis and a haplotype study were performed. Moreover, an expression study was conducted in peripheral blood mononuclear cells from 15 SLE patients to verify a possible effect of the three SNPs on the expression of ATG5. Among the three investigated SNPs, only the rs573775 SNP was significantly associated with disease susceptibility with the variant allele conferring a higher risk of developing SLE (OR = 1.50, p = 0.018 and OR = 1.48, p = 0.007 at the genotypic and allelic level, respectively). The variant allele of rs6568431 SNP was more present in patients with anemia (OR = 1.86, p = 0.009) and renal involvement (OR = 1.63, p = 0.06), while the variant allele of rs2245214 SNP was significantly associated with a higher risk of producing anti-DNA autoantibodies (OR = 1.66, p = 0.04). Carriers of the rs6568431 variant allele showed higher messenger RNA levels compared to the carriers of the wild-type allele, suggesting also a potential variant allele dose-dependent effect on gene expression. In conclusion, our study confirms a role for ATG5 polymorphisms both in disease susceptibility and in the modulation of clinical phenotypes in an Italian SLE cohort. These results further suggest that genetic variations in autophagy genes could play a role in autoimmune diseases susceptibility and are worth further investigation.

Immunology of Alveolar and Cystic Echinococcosis (AE and CE).

Cystic and alveolar echinococcosis are severe chronic helminthic diseases caused by the cystic growth or the intrahepatic tumour-like growth of the metacestode of Echinococcus granulosus or Echinococcus multilocularis, respectively. Both parasites have evolved sophisticated strategies to escape host immune responses, mainly by manipulating and directing this immune response towards anergy and/or tolerance. Recent research studies have revealed a number of respective immunoregulatory mechanisms related to macrophages and dendritic cell as well as T cell activities (regulatory T cells, Tregs). A better understanding of this complex parasite-host relationship, and the elucidation of specific crucial events that lead to disease, represents targets towards the development of novel treatment strategies and options.

Role of alpha-synuclein in autophagy modulation of primary human T lymphocytes.

It has been demonstrated that α-synuclein can aggregate and contribute to the pathogenesis of some neurodegenerative diseases and it is capable of hindering autophagy in neuronal cells. Here, we investigated the implication of α-synuclein in the autophagy process in primary human T lymphocytes. We provide evidence that: (i) knocking down of the α-synuclein gene resulted in increased autophagy, (ii) autophagy induction by energy deprivation was associated with a significant decrease of α-synuclein levels, (iii) autophagy inhibition by 3-methyladenine or by ATG5 knocking down led to a significant increase of α-synuclein levels, and (iv) autophagy impairment, constitutive in T lymphocytes from patients with systemic lupus erythematosus, was associated with abnormal accumulation of α-synuclein aggregates. These results suggest that α-synuclein could be considered as an autophagy-related marker of peripheral blood lymphocytes, potentially suitable for use in the clinical practice.

Streptococcal-vimentin cross-reactive antibodies induce microvascular cardiac endothelial proinflammatory phenotype in rheumatic heart disease.

Rheumatic heart disease (RHD) is characterized by the presence of anti-streptococcal group A antibodies and anti-endothelial cell antibodies (AECA). Molecular mimicry between streptococcal antigens and self proteins is a hallmark of the pathogenesis of rheumatic fever. We aimed to identify, in RHD patients, autoantibodies specific to endothelial autoantigens cross-reactive with streptococcal proteins and to evaluate their role in inducing endothelial damage. We used an immunoproteomic approach with endothelial cell-surface membrane proteins in order to identify autoantigens recognized by AECA of 140 RHD patients. Cross-reactivity of purified antibodies with streptococcal proteins was analysed. Homologous peptides recognized by serum cross-reactive antibodies were found through comparing the amino acid sequence of streptococcal antigens with human antigens. To investigate interleukin (IL)-1R-associated kinase (IRAK1) and nuclear factor-κB (NF-κB) activation, we performed a Western blot analysis of whole extracts proteins from unstimulated or stimulated human microvascular cardiac endothelial cells (HMVEC-C). Adhesion molecule expression and release of proinflammatory cytokines and growth factors were studied by multiplex bead based immunoassay kits. We observed anti-vimentin antibodies in sera from 49% RHD AECA-positive patients. Cross-reactivity of purified anti-vimentin antibodies with heat shock protein (HSP)70 and streptopain streptococcal proteins was shown. Comparing the amino acid sequence of streptococcal HSP70 and streptopain with human vimentin, we found two homologous peptides recognized by serum cross-reactive antibodies. These antibodies were able to stimulate HMVEC-C inducing IRAK and NF-κB activation, adhesion molecule expression and release of proinflammatory cytokines and growth factors. In conclusion, streptococcal-vimentin cross-reactive antibodies were able to activate microvascular cardiac endothelium by amplifying the inflammatory response in RHD.

Skewed T-cell receptor variable ß repertoire and massive T-cell activation in idiopathic orofacial granulomatosis.

Orofacial granulomatosis (OFG) is a clinicopathologic entity describing oral lesions with noncaseating granulomas including a spectrum of diseases such as the Melkersson-Rosenthal syndrome. The involvement of abnormal T-cell responses has been suggested in the pathogenesis of OFG although few and contrasting data are currently available on this issue. In a patient with OFG, we observed virtually complete CD4 and CD8 T-cell receptor (TCR) ß-chain variable region (BV) repertoires at the lesion level and in circulation. However, oligoclonal profiles were found in CD4 and, to a greater extent, in CD8 subsets. These findings were seen in association with a massive peripheral T-cell activation, decreased naive T cells, reduced thymic output, altered cytokine production, and increased apoptosis. Our data, pointing to a random influx of T cells at the site of inflammation, argue against the hypothesis of a main allergen acting at the level of oral mucosa. The profound dysregulation of the peripheral T-cell compartment suggests that OFG should be regarded as a systemic disorder with localized manifestations.

Olanzapine-induced neutropenia in a patient with systemiclupus erythematosus: a role of FcγRIIIb polymorphism?

In this study, we report the case of a Chinese patient with systemic lupus erythematosus (SLE) who developed neutropenia after treatment by olanzapine for the SLE-related psychiatric symptoms. The relationship between agranulocytosis, SLE and olanzapine is still unknown. Fcγ receptor IIIb (FcγRIIIb) is a low-affinity receptor, constitutively expressed only by neutrophils; NA1 and NA2 have been identified as representing polymorphisms of FcγRIIIb. NA1 is associated with the incidence of autoimmune neutropenia and is particularly frequent in Asiatic ethnic groups. The Chinese patient resulted to be homozygous for NA1. We suggest that the presence of NA1 allele may be a predisposing factor to olanzapine-induced agranulocytosis in patients with SLE. Hence, the analysis of FcγRIIIb polymorphism should be investigated in other cases of antipsychotic-induced agranulocytosis.

Autoantibodies to the adenosine triphosphate synthase play a pathogenetic role in Alzheimer's disease.

It has become evident that an autoimmune component could play a role in Alzheimer's disease (AD) onset and/or progression. The aim of this study was to identify neuronal antigenic targets specifically recognized by serum autoantibodies and to investigate their cellular effects and their possible pathogenetic role. We identified, by an immunoproteomic approach using mouse brain proteins, the adenosine triphosphate (ATP) synthase ß subunit as a new autoantigen in AD. Using an ELISA assay we found that serum anti-ATP synthase autoantibodies were present in 38% of patients with AD, but in no age-matched healthy subjects or in patients with Parkinson's disease or atherosclerosis. Analytical cytology studies, using SH-SY5Y neuroblastoma cell line, showed that ATP synthase autoantibodies were capable of inducing the inhibition of ATP synthesis, alterations of mitochondrial homeostasis and cell death by apoptosis. These findings suggest that autoantibodies specific to ATP synthase can exert a pathogenetic role via a mechanism that brings into play the impairment of the extracellular ATP homeostasis and the alteration of mitochondrial function triggering cell death by apoptosis.

Favourable prognostic value of antibodies anti-HSP20 in patients with cystic echinococcosis: a differential immunoproteomic approach.

Seeking biomarkers reflecting disease development in cystic echinococcosis (CE), we used a proteomic approach linked to immunological characterisation for the identification of respective antigens. Two-dimensional gel electrophoresis (2-DE) of sheep hydatid fluid, followed by immunoblot analysis (IB) with sera from patients with distinct phases of disease, enabled us to identify by mass spectrometry heat shock protein 20 (HSP20) as a potential marker of active CE. Using IB, antibodies specific to the 34 kDa band of HSP20 were detected in sera from 61/95 (64%) patients with CE, but not in sera from healthy subjects. IB revealed anti-HSP20 antibodies in a higher percentage of sera from patients with active disease than in sera from patients with inactive disease (81 vs. 24%; P = 10(-4)). These primary results were confirmed in a long-term follow-up study after pharmacological and surgical treatment. Herewith anti-HSP20 antibody levels significantly decreased over the course of treatment in sera from patients with cured disease, relative to sera from patients with progressive disease (P = 0·017). Thus, during CE, a comprehensive strategy of proteomic identification combined with immunological validation represents a promising approach for the identification of biomarkers useful for the prognostic assessment of treatment of CE patients.

Association of intracellular pro- and anti-inflammatory cytokines in peripheral blood with the clinical or ultrasound indications for carotid endarterectomy in patients with carotid atherosclerosis.

Early non-invasive diagnostic information would be useful in identifying patients at risk of progressive carotid atherosclerosis, despite an apparently harmless plaque on ultrasound imaging. In this study, we assessed the possible association of intracellular cytokines in peripheral blood with the ultrasound (stenosis > or = 70%) and clinical indications (transient ischaemic attack, amaurosis fugax or stroke) for carotid endarterectomy (CEA) in patients. Intracellular cytokine expression was determined in 106 patients (67 undergoing and 39 not undergoing CEA). Cells primed for the proinflammatory cytokines tumour necrosis factor (TNF)-alpha, interferon (IFN)-gamma, interleukin (IL)-1beta, IL-6, IL-8 and the anti-inflammatory cytokines IL-4 and IL-10 were found in significantly higher percentages in patients undergoing CEA than in patients who were not (P < 0.05). Intracellular cytokine expression was significantly higher in patients undergoing CEA who had stenosis > or = 70% (TNF-alpha, IFN-gamma, IL-1beta, IL-6, IL-4 and IL-10), with previous stroke (IFN-gamma, IL-1beta, IL-6, IL-8, IL-4 and IL-10) and with amaurosis fugax (IFN-gamma, IL-6, IL-4 and IL-10) than in patients not undergoing CEA. Increased intracellular cytokines in patients' peripheral blood might be a warning signal indicating progressive atherosclerosis. If so, intracellular cytokine monitoring could help in selecting patients at high risk of future clinical cardiovascular events and therefore most likely to benefit from CEA or adjustment of pharmacological therapy.

Screening of an Echinococcus granulosus cDNA library with IgG4 from patients with cystic echinococcosis identifies a new tegumental protein involved in the immune escape.

The worldwide problem of chronic Echinococcus granulosus disease calls for new parasite-derived immunomodulatory molecules. By screening an E. granulosus cDNA library with IgG4 from patients with active cystic echinococcosis, we identified a cDNA that encodes a predicted partial protein that immunofluorescence studies localized in the protoscolex tegument and on the germinal layer of cyst wall. We named this protein EgTeg because the 105 amino acid sequence scored highest against a family of Schistosoma tegumental proteins. Evaluating the role of EgTeg in the human early inflammatory response we found that EgTeg significantly inhibited polymorphonuclear cell (PMN) chemotaxis. Cytometric analysis of intracellular cytokines disclosed a significantly higher percentage of cells producing IL-4 than IFN-gamma (P = 0.001, Student's t-test) in T lymphocytes from patients with cystic echinococcosis stimulated with EgTeg. EgTeg induced weak Th1-dependent proliferation in 42% of patients' peripheral blood mononuclear cells. In immunoblotting (IB) analysis of total IgG and IgG subclass responses to EgTeg in patients with cystic echinococcosis, patients with other parasitoses, patients with cystic lesions and healthy controls, total IgG specific to EgTeg yielded high sensitivity (73%) but low specificity (44%) precluding its use in immunodiagnosis. Conversely, IgG4 specific to EgTeg gave acceptable sensitivity (65%) and high specificity (89%) suggesting its use in immunodiagnosis to confirm ultrasound documented cysts suggestive of E. granulosus. Because the new tegumental antigen EgTeg inhibits chemotaxis, induces IL-4-positive T lymphocytes and noncomplement fixing antibodies (IgG4) it is an immunomodulatory molecule associated with chronic infection.

Screening of a HUAEC cDNA library identifies actin as a candidate autoantigen associated with carotid atherosclerosis.

The humoral immune response to endothelium has a pivotal role in the development of atherosclerosis. Using a molecular method, we sought to identify endothelial autoantigens in carotid atherosclerosis. Immunoscreening of a HUAEC expression library with IgG from a pool of two sera from patients with carotid atherosclerosis identified a clone specific to actin. We evaluated actin-specific IgG reactivity in patients with carotid atherosclerosis and compared responses with those in patients with systemic lupus erythematosus and type 1 diabetes mellitus and in healthy subjects. Enzyme-linked immunoassay detected actin-specific IgG in a significantly higher percentage of sera from patients with atherosclerosis and systemic lupus erythematosus than from healthy subjects (16/61, 26% and 13/33, 39%versus 2/41, 5%, P = 0.012 and P < 10(-4), by chi2 test). Mean optical density values were significantly higher in patients than in healthy subjects (P < 10(-4) by Student's t-test). Patients with atherosclerosis and uncomplicated plaques had significantly higher serum anti-actin IgG reactivity than those with complicated plaques (P = 0.048 by Student's t-test). Our findings suggest that actin is an autoantigenic molecule of potential clinical interest in carotid atherosclerosis.

Echinococcus granulosus-specific T-cell lines derived from patients at various clinical stages of cystic echinococcosis.

To investigate the role of T lymphocytes in the immune response to Echinococcus granulosus, using sheep hydatid fluid (SHF) and antigen B (AgB), we generated T-cell lines from patients with active, transitional and inactive hydatid cysts. We established 16 T-cell lines, eight specific to SHF and eight specific to AgB. At surface phenotyping 88-98% of cells displayed the helper/inducer CD4 antigen. In all patients, at all clinical stages of hydatid cyst disease, T-cell stimulation with SHF and AgB invariably amplified a large number of almost identical Vbeta subfamily fragments. Irrespective of antigen-specificity, the two cell lines from the patient with an inactive cyst had a Th1 profile, because they exclusively expressed and produced IFN-gamma. Conversely, the T-cell lines derived from the seven patients with active and transitional hydatid cysts had mixed Th1/Th2 and Th0 clones. The functional characteristics of the 16 T-cell lines differed markedly in the various clinical stages of cystic echinococcosis, thus providing new in vitro evidence that Th1 lymphocytes contribute decisively to the inactive stage of hydatid disease, Th2 lymphocytes in the active and transitional stages. The parasite-specific T-cell lines, especially the two Th1 lines from the patient with an inactive cyst, may help identify Th1 protective epitopes on SHF and AgB.

Critical points in the immunodiagnosis of cystic echinococcosis in humans.

This study discusses the immunodiagnosis of cystic echinococcosis (ce, caused by Echinococcus granulosus). The detection by immunoblotting of antibodies specific for the 8 kDa subunit of antigen B and in particular the IgG4 subclass expression, seems the most promising serodiagnostic tool. Despite the development of molecular methods, nowadays there is no standard, highly sensitive, and specific test available for antibody detection in CE. Furthermore, because serological tests can give only a limited support to clinical findings there is a clear need for new advances in immunodiagnosis of E. granulosus infection.

Molecular and immunological characterization of the C-terminal region of a new Echinococcus granulosus Heat Shock Protein 70.

By screening an expression library of Echinococcus granulosus with IgE from sera of patients with cystic echinococcosis (CE) and allergic reactions, we isolated the C-terminal region of a new heat shock protein (HSP)70 of E. granulosus. The protein, named Eg2HSP70, has close homology with the C-terminal region of Dermatophagoides farinae and human HSP70. We investigated the humoral and cell-mediated immune responses to this antigen in patients with CE grouped according to the presence of allergic reactions. Immunoblotting detected total IgG, IgG4 and IgE specific to Eg2HSP70 (83% of sera contained IgG, 31% IgG4 and 57% IgE). No significant difference was found in immunoglobulin percentages according to the presence of allergic reactions. Immunoblotting inhibition showed that no IgG or IgE specific to Eg2HSP70 cross-reacted with D. farinae or human HSP70. Eg2HSP70-stimulated PBMC from 26 patients produced significantly greater amounts of TNF-alpha, IFN-gamma, and IL-10 than unstimulated cultures in all patients, irrespective of the presence of allergic reactions (P < 0.05). They also produced significantly greater amounts of IL-4 than unstimulated cultures exclusively in patients with allergic reactions (P < 0.05). These findings show that Eg2HSP70 is a new antigenic molecule inducing both B and T cell responses.

Acute generalized exanthematous pustulosis in cystic echinococcosis: immunological characterization.

We report the first case, to the best of our knowledge, of a woman suffering from cystic echinococcosis of the liver, who consequently developed urticaria and acute generalized exanthematous pustolosis (AGEP). Serum immunoglobulin (Ig)E and IgG4 specific to Echinococcus granulosus antigens were detected by immunoblotting. Furthermore, the intracellular cytokine analysis revealed a prevalent T-helper 2 polarization. It can be reasoned that, while the presence of IgE specific to various E. granulosus allergens may be responsible for the chronic urticarial manifestations, the detection of IgG4 specific for E. granulosus antigens, forming immunocomplexes, may be related to the development of the AGEP.