To detect and identify adenovirus (Ad), we used a combination of PCR and restriction fragment length polymorphism (RFLP) analysis. Nested PCR with two primer sets that hybridize to the conserved region for hexon proteins of 14 prototypes of Ad, Ad serotype 1 (Ad1) to Ad8, -11, -14, -19, -37, -40, and -41, amplified a 956-bp DNA fragment. The amplified fragments from the 14 prototypes were completely differentiated with a combination of three restriction endonucleases, EcoT14I, HaeIII, and HinfI. We applied this new method for 127 samples of conjunctival scrapings from patients with conjunctivitis and compared the results with those obtained with the combination of culture isolation and a neutralization test (NT). PCR gave a positive result in 69 of 127 cases (54.3%), while only 61 of the 127 samples (48.0%) tested positive by culture isolation. Compared with isolation, the PCR method had a sensitivity of 100% (61 of 61). Positive PCR samples were further classified as Ad37 (59.5%), -3 (31.9%), -11 (4.3%), -8 (2.9%), and -4 (1.4%) by PCR-RFLP analysis. Of eight samples that were PCR positive and culture isolation negative, six were Ad37 and two were Ad8 by PCR-RFLP analysis. These differentiations of isolation-positive samples were identical to the results obtained by the NT. It took only 3 days to detect and identify Ad by PCR-RFLP analysis, whereas it took at least 3 weeks by culture isolation and NT. Our newly developed method of detecting and typing human Ad by PCR-RFLP analysis is more sensitive, accurate, and rapid than the conventional method of culture isolation and an NT.

Purpose: Molecular targeted therapy using BRAF and/or MEK inhibitors has been applied to BRAFV600E mutant high-grade gliomas (HGGs); however, the therapeutic effect is limited by the emergence of drug resistance. Experimental Design: We established multiple paired BRAFV600E mutant HGG patient-derived xenograft (PDX) models based on tissues collected prior to and at relapse after molecular targeted therapy. Using these models, we dissected treatment resistant mechanisms for molecular targeted therapy and explored therapeutic targets to overcome resistance in BRAFV600E HGG models in vitro and in vivo. Results: We found that, despite causing no major genetic and epigenetic changes, BRAF and/or MEK inhibitor treatment deregulated multiple negative feedback mechanisms, which led to the re-activation of the MAPK pathway through c-Raf and AKT signaling. This altered oncogenic signaling primarily mediated resistance to molecular targeted therapy in BRAFV600E mutant HGG. To overcome this resistance mechanism, we performed a high-throughput drug screening to identify therapeutic agents that potently induce additive cytotoxicity with BRAF and MEK inhibitors. We discovered that HSP90 inhibition combined with BRAF/MEK inhibition coordinately deactivated the MAPK and AKT/mTOR pathways, and subsequently induced apoptosis via dephosphorylation of GSK3β (Ser9) and inhibition of Bcl-2 family proteins. This mediated potent cytotoxicity in vitro and in vivo in refractory models with acquired resistance to molecular-targeted therapy. Conclusions: The combination of an HSP90 inhibitor with BRAF or MEK inhibitors can overcome the limitations of the current therapeutic strategies for BRAFV600E mutant HGG.

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Background: The biology and clinical course of intracranial germinomas differ as per their location of occurrence. Germinoma of the primary midbrain is particularly rare, and its clinical features, treatment strategies, and long-term prognosis remain uncertain. Case Description: A 39-year-old man who had been diagnosed with midbrain germinoma by open biopsy through the occipital transtentorial approach had undergone chemoradiotherapy and achieved 5 years with no recurrence. Conclusion: Germinomas should be considered as a differential diagnosis for adolescents and young adult men with mesencephalic tumors, and reliable sampling followed by chemoradiotherapy must be performed.