CD30 expression in an emerging group of mesenchymal spindle cell neoplasms with ALK fusion detected by flow cytometry and immunohistochemistry.

An emerging group of spindle cell neoplasms harboring fusions involving NTRK or non-NTRK kinase genes often share characteristic S100 and/or CD34 expression; however, the diagnostic utility of immunohistochemical stains is not well established in this family owing to their lack of specificity. Recently, CD30 expression in spindle cell neoplasms with kinase gene fusions, such as NTRK, BRAF, RAF1, and RET, has been increasingly identified. We herein report a 10-year-old girl with high-grade spindle cell sarcoma of the neck. Prior to histopathological evaluation, flow cytometry (FCM) analysis and touch smear cytology of the tumor tissue revealed CD34+ and dimCD30+ spindle cell populations. Histopathologically, the case was characterized by monomorphic spindle-shaped cytomorphology with CD30, S100, and CD34 positivity and harbored close similarities with spindle cell neoplasms with NTRK or non-NTRK gene fusions. Subsequently, a comprehensive next-generation sequencing sarcoma panel identified a rare PLEKHH2::ALK fusion, and a diagnosis of ALK-rearranged spindle cell neoplasm was made. The patient showed significant tumor response to single-agent treatment with alectinib, an ALK-tyrosine kinase inhibitor. This case supports that CD30 is expressed in an ALK-rearranged mesenchymal neoplasm. The benefit of the early detection of CD30 expression by FCM for a prompt diagnosis and treatment is highlighted in the context of an aggressive clinical course. This case represents a learning experience regarding the need to the check the status of CD30 expression in these tumors and suggests the potential clinical benefits of CD30-targeted therapy.

Undiagnosed Cardiac Sarcoidosis Causing Refractory Heart Failure After Acute Myocardial Infarction due to Thromboembolism.

A 61-year-old woman suffered chest pain and was admitted to a nearby hospital emergency department. She was diagnosed with acute myocardial infarction probably due to thromboembolism in the left anterior descending coronary artery and aspiration thrombectomy was performed. Afterwards, she developed refractory heart failure with severe global left ventricular dysfunction and was transferred to our hospital. An 18F-FDG-PET/CT scan revealed abnormal 18F-FDG uptake in non-infarcted regions of the left ventricle. Non-caseating granulomas were detected by biopsy from a skin eruption. She was diagnosed with cardiac sarcoidosis. In cases of refractory heart failure which cannot be explained only by myocardial infarction, evaluation of other undiagnosed cardiomyopathies is important for optimal management.

Acid-Tolerant Reversibly Switchable Green Fluorescent Protein for Super-resolution Imaging under Acidic Conditions.

Reversibly switchable fluorescent proteins (RSFPs) are crucial tags for super-resolution observation of protein localization and dynamics inside living cells. However, due to the high fluorescence pKa (∼5-6) of most RSFPs, their usage in acidic conditions (pH 4.5-6.0) has been limited. Here, we investigated a new photochromic mechanism in Gamillus, a recently developed green fluorescent protein with acid tolerance. Gamillus exhibits negative switching with especially high contrast in acidic conditions, and its off switching is caused by trans-to-cis isomerization of the chromophore hydroxyphenyl ring that accompanies protonation. Through a combination of rational design and saturation mutagenesis, we developed two variants with enhanced switching contrasts and off-switching speeds, designated rsGamillus-S and rsGamillus-F, respectively. The fluorescence intensity, off-switching speed, and switching contrast of the rsGamillus variants are only slightly affected by changes in pH between 4.5 and 7.5. Exploiting these properties, we succeeded in high-contrast super-resolution imaging of cellular architectures in acidic conditions.

Crystallographic Analysis of Drug and Inhibitor-Binding Structure of RND-Type Multidrug Exporter AcrB in Physiologically Relevant Asymmetric Crystals.

Xenobiotic extruding pumps have recently been known to be widely distributed in living organisms from mammalian to bacteria as a host-defense mechanism in cellular level. These pumps not only confer multidrug resistance of cancer cells and pathogenic bacteria but also cause hereditary diseases through the mutation. Our purposes are to elucidate the molecular structures and mechanisms of these xenobiotic exporters.We had succeeded to determine the crystal structure of bacterial major multidrug exporter AcrB at 3.5 Å resolution (Murakami et al., Nature 419:587-593, 2002) and elucidated the structural bases of substrate recognition that the pump recognize the places and thus act as a "membrane vacuum cleaner." After that we also determined the crystal structure of the drug-binding form of AcrB in space group C2 in which asymmetric unit contains structurally asymmetric homo-trimer of AcrB (Murakami et al., Nature 443:173-179, 2006; Nakashima et al., Nature 480:565-569, 2011; Nakashima et al., Nature 500:120-126, 2013). Analyses revealed the existence of a specific mechanism to recognize numerous substrates that the multisite binding is the base of multidrug recognition rather than induced-fit, and functional-rotation mechanism in which three monomers undergo a strictly coordinated sequential conformational change cycle of access, binding, and extrusion. Determination of physiological asymmetric AcrB structure was crucially important to understand these transport mechanisms.