Dorit Zharhary

Potential Biomarker for Human Uterine Leiomyosarcoma

Sarcomas are a rare form of malignant tumor, with less than 15,000 new cases being diagnosed each year in the United States. In spite of their rarity, sarcomas are highly debilitating malignancies that are often associated with significant morbidity and mortality. They are also biologically very heterogeneous because they originate from many different tissues and cell types. Sarcomas have classically been defined by their tissue of origin and are additionally stratified according to their histopathology or the age of the patient at diagnosis. Uterine mesenchymal tumors that develop in the myometrium have traditionally been divided into benign uterine usual LMA, cellular LMA and malignant Ut-LMS based on cytological atypia, mitotic activity and other criteria. Ut-LMS is relatively rare, having an estimated annual incidence of 0.64 per 100,000 women [1], and is resistant to chemotherapy and radiotherapy; therefore, surgical interventions are virtually the only means of treatment [2, 3]. The prognosis of patients with Ut-LMS is poor, and the 5-year survival rate is approximately 35%. Uterine LMA may occur in 70-80% of women by the age of 50 years [4]. Difficulties have been reported in distinguishing Ut-LMS from other uterine mesenchymal tumors including uterine LMA, and a diagnosis generally requires surgery and cytoscopy. Diagnostic categories for uterine mesenchymal tumors and morphological criteria are used to assign cases. The non-standard subtypes of uterine mesenchymal tumors such as the epithelioid and myxoid types are classified in a different manner using these features; therefore, a diagnostic method needs to be established that can identify non-standard smooth muscle differentiation [5, 6]. High estrogen levels have been shown to significantly influence the development of tumors in the uterine body [7]. However, the molecular mechanisms underlying the transformation of uterine LMA and development of Ut-LMS remain unknown. Tumors that have developed and grown in the myometrium increase in size due to the influence of the female hormone, estrogen, which leads to the generation of more tumors. However, a relationship has not yet been reported between the development of Ut-LMS and hormonal conditions, and no obvious risk factors have been identified. The identification of risk factors associated with the development of human Ut-LMS will contribute significantly to the development of preventive and therapeutic treatments. Cytoplasmic proteins are mostly degraded by a protease complex referred to as the 20S proteasome, which has many substrates that consist of twenty-eight 20 to 30 kDa subunits [8, 9]. Proteasomal degradation is essential for many cellular processes, including the cell cycle, regulation of gene expression and immunological function [10]. A previous study reported that an interferon (IFN)-γ treatment induced the expression of large numbers of responsive genes, the β-ring subunits of proteasomes, i.e., low-molecular mass polypeptide (LMP)2/β1i, LMP7/β5i and LMP10/multicatalytic endopeptidase complex-like (MECL)-1/β2i [11]. Ut-LMS was detected in female LMP2/β1ideficient mice at 6 months or older, and its incidence at 14 months was approximately 40% [12]. Histopathological studies of LMP2/ β1i-lacking uterine tumors have revealed the characteristic abnormalities of Ut-LMS [12]. The non-standard subtypes of uterine mesenchymal tumors such as the epithelioid and myxoid types have been classified in a Manuscript accepted for publication May 15, 2014

Development of unique antibodies directed against each of the six different phosphotyrosine residues within the T cell receptor CD3ζ chain.

Signal transduction from the T cell antigen receptor (TCR)/CD3 complex involves six different immunoreceptor tyrosine-based activation motifs (ITAM) located within the cytoplasmic tails of the CD3 chains. Each ITAM possesses two conserved tyrosine residues that can undergo phosphorylation upon TCR/CD3 crosslinking and become a docking site for SH2-containing effector molecules. Specificity of the SH2 domains is determined by their ability to bind a phosphorylated tyrosine in the context of a longer peptide motif within the target protein. As a result, phosphorylation of different tyrosines within the CD3 cytoplasmic tails creates docking sites for distinct SH2-containing signaling proteins that differentially impact on the quality of the T cell response. In the present study, we prepared antibodies specific for each of the six different phosphotyrosines of the mouse CD3ζ chain. The antibodies were characterized with respect to their cross-reactivity, ability to recognize the phosphorylated versus non-phosphorylated forms of tyrosine-containing motifs, and cross-reactivity with the homologous phospho-motifs on the human CD3ζ protein. The antibodies were found to be specific and selective for phospho-CD3ζ. They can serve as useful tools for distinguishing between the six potential tyrosine phosphorylation sites on the CD3ζ chain, and for correlating the phosphorylation of specific CD3ζ tyrosine residues with activation of signaling pathways that dictate T cell differentiation into responding, anergic, or apoptotic cells.

Pathobiology of Human Uterine Leiomyosarcoma for Development of Novel Diagnosis and Clinical Therapy

Uterine sarcomas comprise a group of rare tumours with differing tumour pathobiology, natural history and response to clinical treatment. Diagnosis is often made following surgical treatment for presumed malignant mesenchymal tumours and benign tumours. Currently pre-operative diagnosis does not reliably distinguish between malignant mesenchymal tumours, Uterine Leiomyosarcoma (U-LMS) and benign tumours including Leiomyomas (LMA). U-LMS is the most common sarcoma but other subtypes include endometrial stromal sarcoma (low grade and high grade), undifferentiated uterine sarcoma and adeno sarcoma. Clinical trials have shown no definite survival benefit for adjuvant radiotherapy or chemotherapy, and have been hampered by the rarity and heterogeneity of these tumour types. There is a role of adjuvant treatment in carefully selected cases following multidisciplinary discussion at U-LMS reference centres. In patients with metastatic LMS then systemic chemotherapy can be considered. Accordingly, it is necessary to analyse risk factors associated with human U-LMS, in order to establish a treatment method. Proteasome β-subunit 9 (PSMB9)/β1i-deficient mice spontaneously develop U-LMS, with a disease prevalence of ~37% by 12 months of age. We found PSMB9/β1i expression to be absent in human U-LMS, but present in human LMA. Therefore, defective PSMB9/β1i expression may be one of the risk factors for human U-LMS. PSMB9/β1i is a potential diagnostic-biomarker for human U-LMS, and may be targeted-molecule for a new therapeutic approach.

A novel diagnostic biomarker for human uterine leiomyosarcoma: PSMB9/β1i

Takuma Hayashi (Shinshu University School of Medicine) discussed a novel biomarker for detecting human uterine leiomyosarcoma (LMS).

Biological Significance of the Proteasome Subunit LMP2/1i as a Tumor Suppressor in Human Uterine Leiomyosarcoma

Uterine leiomyosarcoma (Ut-LMS) develops more often in the muscle tissue layer of the uterine body than in the uterine cervix. The development of gynecologic tumors is often correlated with female hormone secretion; however, the development of Ut-LMS is not substantially correlated with hormonal conditions, and the risk factors are not yet known. Importantly, a diagnostic-biomarker which distinguishes malignant Ut-LMS from other uterine mesenchymal tumors including leiomyoma (LMA) is yet to be established. Accordingly, it is necessary to analyze risk factors associated with Ut-LMS, to establish a clinical treatment method. Proteasome subunit, low-molecular mass polypeptide(LMP2)/b1i-deficient mice spontaneously develop Ut-LMS, with a disease prevalence of ~40% by 14 months of age. Recent experiments with human and mouse uterine tissues revealed defective LMP2/b1i expression in human Ut-LMS that was traced to the interferon (IFN)-g pathway and a specific effect of Janus kinase (JAK)-1 somatic mutations on LMP2/ b1i transcriptional activation. Furthermore, analysis of a human Ut-LMS cell line clarified the biological significance of LMP2/b1i in malignant myometrium transformation and the cell cycle, thus implicating LMP2/b1i as an anti-tumorigenic candidate. Therefore, defective-LMP2/b1i expression may be a risk factor for human Ut-LMS. LMP2/b1i is a potential diagnostic-biomarker for Ut-LMS, and may be a targeted-molecule for a new clinical therapeutic approach.