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Denosumab is the world's first human monoclonal antibody for the treatment of osteoporosis and shares an active pathway with bisphosphonates, strongly suppressing osteoclast activities. This is the first case report describing a possible relationship between the development of osteonecrosis of external auditory canal and denosumab administration. We herein report an 81-year-old woman diagnosed with left osteonecrosis of external auditory canal who had a history of denosumab administration. She underwent left radical mastoidectomy due to being refractory to conservative treatment. No major complications or recurrence were observed in the left ear after surgery, but bone erosion in the right ear has continued to progress slowly despite the cessation of denosumab administration. Otolaryngologists should be aware of the association between osteonecrosis of external auditory canal and denosumab administration and consider performing long-term observation even after cessation.

Ultrasound-guided, lymph node, fine-needle aspiration cytology is important in diagnosing axillary lymph node metastasis in breast cancer. However, poor needle visibility can render the procedure difficult. We describe a case in which state-of-the-art enhancement techniques using matrix linear probes can provide better needle visibility and improve the certainty and efficiency of the examination.

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Comprehensive genomic profiling (CGP) provides information regarding cancer-related genetic aberrations. However, its clinical utility in recurrent/metastatic head and neck cancer (R/M HNC) remains unknown. Additionally, predictive biomarkers for immune checkpoint inhibitors (ICIs) should be fully elucidated because of their low response rate. Here, we analyzed the clinical utility of CGP and identified predictive biomarkers that respond to ICIs in R/M HNC. We evaluated over 1100 cases of HNC using the nationwide genetic clinical database established by the Center for Cancer Genomics and Advanced Therapeutics (C-CAT) and 54 cases in an institution-based study. The C-CAT database revealed that 23% of the cases were candidates for clinical trials, and 5% received biomarker-matched therapy, including NTRK fusion. Our institution-based study showed that 9% of SCC cases and 25% of salivary gland cancer cases received targeted agents. In SCC cases, the tumor mutational burden (TMB) high (≥10 Mut/Mb) group showed long-term survival (>2 years) in response to ICI therapy, whereas the PD-L1 combined positive score showed no significant difference in progression-free survival. In multivariate analysis, CCND1 amplification was associated with a lower response to ICIs. Our results indicate that CGP may be useful in identifying prognostic biomarkers for immunotherapy in patients with HNC.

Comprehensive screenings to clarify indirect cell–cell interactions, such as those in the tumor microenvironment, especially comprehensive assessments of supporting cells’ effects, are challenging. Therefore, in this study, indirect CRISPR screening for drug resistance with cell–cell interactions was invented. The photoconvertible fluorescent protein Dendra2 was inducted to determine the drug resistance responsible factors of supporting cells with CRISPR screenings. Random mutated supporting cells co-cultured with leukemic cells induced drug resistance with cell– cell interactions. Supporting cells responsible for drug resistance were isolated with green-to-red photoconversion, and 39 candidate genes were identified. Knocking out C9orf89, MAGI2, MLPH, or RHBDD2 in supporting cells reduced the ratio of apoptosis of cancer cells. In addition, the low expression of RHBDD2 in supporting cells, specifically fibroblasts, of clinical pancreatic cancer showed a shortened prognosis, and a negative correlation with CXCL12 was observed. Indirect CRISPR screening was established to isolate the responsible elements of cell–cell interactions. This screening method could reveal new mechanisms in all kinds of cell–cell interactions by revealing live phenotype-inducible cells, and it could be a new platform for discovering new targets of drugs for conventional chemotherapies.

Preoperatively diagnosed ductal carcinoma in situ (DCIS) is sometimes upstaged to invasive cancer by postoperative pathological examination. Various preoperative factors associated with upstaging to invasive cancer have been reported; however, this subject remains to be clarified. DCIS takes various forms on imaging, but many cases show non-mass-type lesions. In non-mass-type DCIS, recognizing the presence of invasion is difficult. To investigate predictors associated with upstaging to invasive cancer more precisely, we examined only non-mass-type DCIS. The present study retrospectively analyzed 101 patients diagnosed with non-mass-type DCIS preoperatively on breast biopsy at our institution between 2007 and 2017. Data were analyzed using Fisher's exact probability test and two-sample t-tests. Multivariate analysis was performed using logistic regression. The results showed that 27 patients (27%) were finally diagnosed with invasive cancer. Univariate analysis revealed abnormal result of palpation on breast examination (P=0.05), comedo necrosis (P=0.05), and HER2 status (P=0.02) as significant predictors. Multivariate analysis revealed an abnormal result of palpation as an independent predictor of invasive cancer underestimation (odds ratio 4.76; confidence interval 1.44-15.7; P=0.01). In conclusion, preoperatively diagnosed non-mass-type DCIS represented an underestimation in approximately 27% of cases. In particular, the presence of a clinically abnormal palpation increases the chance of upstaging to invasive cancer.

We used virtual navigator real-time ultrasound (US) fusion imaging with 18F-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) to identify a lesion that could not be detected on the US alone in a preoperative breast cancer patient. Of the patient’s two lesions of breast cancer, the calcified lesion could not be identified by US alone. By fusing US with 18F-FDG PET/CT, which had been performed in advance, the location of the lesion could be estimated and marked, which benefited planning an appropriate surgery. The fusion of US and 18F-FDG PET/CT was a simple and noninvasive method for identifying the lesions detected by 18F-FDG PET/CT.

CRISPR/Cas has emerged as an excelle nt gene-editing technology and is used worldwide for research. The CRISPR library is an ideal tool for identifying essential genes and synthetic lethality targeted for cancer therapies in human cancers. Synthetic lethality is defined as multiple genetic abnormalities that, when present individually, do not affect function or survival, but when present together, are lethal. Recently, many CRISPR libraries are available, and the latest libraries are more accurate and can be applied to few cells. However, it is easier to efficiently search for cancer targets with their own screenings by effectively using databases of CRISPR screenings, such as Depmap portal, PICKLES (Pooled In-Vitro CRISPR Knockout Library Essentiality Screens), iCSDB, Project Score database, and CRISP-view. This review will suggest recent optimal CRISPR libraries and effective databases for Novel Approaches in the Discovery and Design of Targeted Therapies.

Decorin (DCN), an extracellular matrix proteoglycan found in tumor surrounding tissues, is a natural inhibitor of tumor cell proliferation and invasion. We conducted a cross-sectional observation study to evaluate the association of the pathological stage with the levels of DCN in plasma or tumor surrounding tissue. Among 118 patients who underwent breast surgery, 35 were designated as carcinoma in situ (Stage 0), 39 were Stage I, and 44 were Stage II or III. The stromal expression of DCN was quantified using a semiquantitative digital image analysis after immunohistochemical staining. The concentration of DCN was evaluated with a specific ELISA. As we have previously shown, stromal DCN expression was attenuated in the patients with Stage I, whereas stromal and plasma DCN was elevated paradoxically in those with Stage II/III. The elevated plasma DCN is an independent predictive factor of Stage II/III by the multivariate logistic regression analysis. The plasma level of DCN was negatively correlated with stromal DCN expression only in patients with advanced disease (Stage II/III). The plasma level of DCN could become a useful biomarker for patients in the advanced stages. Extensive studies and further assessments are warranted for evaluating the prognostic significance and tumor characteristics to understand the clinical significances of stromal and systemic DCN.

Pathological phenotypes of myeloid neoplasms are closely related to genetic/chromosomal abnormalities of neoplastic cells whereas the bone marrow microenvironment, including stromal elements and hematopoietic stem cell niche cells, have a great influence on the differentiation/proliferation of both hematopoietic and neoplastic cells. The pathology of myeloid neoplasms might be generated through the interaction of hematopoietic (stem) cells and stromal cells. The present study aims to provide the morphological/functional aspects of the bone marrow environment in myeloid neoplasms. Among the myeloid neoplasms, myelodysplastic syndromes (MDS) exhibit significant and complex interactions between neoplastic cells and stromal cells. Hematopoietic cells in MDS are greatly influenced by macrophages/niche cells via several signaling pathways. As such, the pathological significance of cell proliferation, cell apoptosis, and anti-apoptosis signals in the bone marrow of myeloid neoplasms, especially MDS bone marrow, will be discussed.

Objective The aim of this study was to evaluate the performance of preoperative axillary lymph node assessment in breast cancer using time-of-flight 18F-fluorodeoxyglucose PET/computed tomography (TOF [F-18]FDG-PET/CT). Methods Eighty-two women with breast cancer (mean age, 59.3 years; range, 30-84 years) underwent TOF [F-18]FDG-PET/CT scanning before surgery between January 2016 and June 2018 at our hospital. Visual analysis of FDG uptake and the maximum standardized uptake value (SUVmax) of axillary lymph nodes were compared with the pathological diagnoses. Results There were 77 patients with invasive breast carcinoma (mean invasive long diameter, 18.5 mm; range, 2-90 mm) and five patients with noninvasive carcinoma. Axillary lymph node metastases were histologically confirmed in 13 of 82 patients (15.9%). SUVmax showed an area under a receiver operating characteristic curve of 0.916, and the cut-off value of 1.1 was appropriate. By visual assessment, there were 11 true positives, 15 false positives, 54 true negatives and two false negatives; the sensitivity, specificity, positive predictive value, negative predictive value and accuracy were 85%, 78%, 42%, 96% and 79%, respectively. SUVmax showed values of 69%, 99%, 90%, 94% and 94%, respectively. Conclusions The sensitivity of TOF [F-18]FDG-PET/CT was as high as 85% by visual analysis. SUVmax using TOF [F-18]FDG-PET/CT showed high diagnostic performance for N-staging in breast cancer patients, especially high negative predictive value. The specificity, positive predictive value and accuracy of SUVmax were higher than those of visual analysis.

Lung squamous cell carcinoma (LSCC) exhibits poor response to treatment compared with other lung cancer subtypes, resulting in worse prognosis. Therefore, new therapeutic strategies are required for advanced LSCC. Ferroptosis is a recently discovered nonapoptotic cell death caused by intracellular lipid peroxidation that can bring about effective cell death in cancer cells resistant to apoptosis. Hence, ferroptosis is a potential therapeutic strategy for refractory cancer.In this study, we performed clinicopathological and molecular analyses on tumor specimens from 270 patients with squamous cell lung cancer, focusing on the expression of glutathione peroxidase 4 (GPX4) and ferroptosis suppressor protein 1 (FSP1), which are known to be key regulators of ferroptosis, and the accumulation of 4-hydroxynoneral (4-HNE), a lipid peroxidation marker.Immunohistochemistry revealed that patients with low 4-HNE accumulation and low levels of GPX4 or FSP1 had significantly worse prognoses than other patients (P = 0.001). This stratification was an independent prognostic predictor (P = 0.003). A dramatic cell death synergistic effect was observed on LSCC-derived LK-2 and EBC1 cells treated with GPX4 and FSP1 inhibitors. This effect was completely inhibited by treatment with the ferroptosis inhibitor. Notably, this was not the case in LK-2 cells treated with the apoptosis inhibitor, and in these cells, ferroptosis was induced.Ferroptosis regulators GPX4 and FSP1 are associated with lung squamous cell cancer cancer's prognosis. We present the clinicopathological and molecular basis of novel therapeutic strategies for refractory LSCC.

This retrospective study examined the relationship between the standardized uptake value max (SUVmax) of fluorine-18 fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) and the prognostic stage of breast cancer. We examined 358 breast cancers in 334 patients who underwent 18F-FDG PET/CT for initial staging between January 2016 and December 2019. We extracted data including SUVmax of 18F-FDG PET and pathological biomarkers, including estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), and nuclear grade. Anatomical and prognostic stages were determined per the American Joint Committee on Cancer (eighth edition). We examined whether there were statistical differences in SUVmax between each prognostic stage. The mean SUVmax values for clinical prognostic stages were as follow: stage 0, 2.2 ± 1.4, stage IA, 2.6 ± 2.1, stage IB, 4.2 ± 3.5, stage IIA, 5.2 ± 2.8, stage IIB, 7.7 ± 6.7, and stage III + IV, 7.0 ± 4.5. The SUVmax values for pathological prognostic stages were as follows: stage 0, 2.2 ± 1.4, stage IA, 2.8 ± 2.2, stage IB, 5.4 ± 3.6, stage IIA, 6.3 ± 3.1, stage IIB, 9.2 ± 7.5, and stage III + IV, 6.2 ± 5.2. There were significant differences in mean SUVmax between clinical prognostic stage 0 and ≥II (p &lt, 0.001) and I and ≥II (p &lt, 0.001). There were also significant differences in mean SUVmax between pathological prognostic stage 0 and ≥II (p &lt, 0.001). In conclusion, mean SUVmax increased with all stages up to prognostic stage IIB, and there were significant differences between several stages. The SUVmax of 18F-FDG PET/CT may contribute to prognostic stage stratification, particularly in early cases of breast cancers.

Background and Objectives: It is necessary to properly diagnose and manage axillary lymphadenopathy caused by a variety of diseases. This study aimed to evaluate the utility of ultrasound (US)-guided sampling in patients with axillary lymphadenopathy. Materials and Methods: Patients with axillary lymphadenopathy (excluding patients with newly diagnosed breast cancer) who underwent US-guided fine needle aspiration (FNA) or core needle biopsy (CNB) at a single center between February 2016 and September 2020 were retrospectively examined. The association between US imaging findings and malignancy was investigated and the diagnostic performance of US-guided sampling was assessed. Results: Fifty-five patients (including eight males) were included in the study, of these, 34 patients (61.8%) were finally diagnosed with a malignant lymph node lesion. Twenty-two patients (40.0%) had undergone FNA and 33 (60.0%) had undergone CNB. Larger short and long axis diameters, thicker lymph node cortex, and the absence of fatty hilum on the US were significantly associated with malignancy (p &lt, 0.05). The diagnostic performance of FNA, CNB, and FNA + CNB was excellent (sensitivity, specificity, and accuracy of 0.909, 0.900, and 0.917 for FNA, 0.958, 1.000, and 0.970 for CNB, and 0.941, 0.952, and 0.945 for FNA + CNB, respectively). Conclusions: US-guided FNA and CNB play an important role in the diagnosis and management of patients with axillary lymphadenopathy

Background: Indocyanine green angiography enables real-time visualization of blood vessels at depths of up to 10 mm beneath the body surface, thereby aiding the evaluation of the viability of skin flaps and predicting necrosis in surgical fields requiring good tissue perfusion. Although skin-flap necrosis also occurs in mastectomy without reconstruction, most studies have focused on reconstructive plastic surgery. Several patients undergoing mastectomy are eligible for postoperative adjuvant therapy, but complications can lead to delays in treatment and thus require prevention. However, a lack of a standard protocol for evaluating skin-flap perfusion using indocyanine green necessitates the study of its characteristics to facilitate comparison of the perfusion rate among individuals. Methods: This retrospective study focused on the characteristics of indocyanine green and established a protocol for indocyanine green angiography using laser-assisted imaging (SPY system) to predict postoperative skin-flap necrosis from intraoperative images of 30 patients who underwent mastectomy without reconstruction. Results: Our protocol predicted postoperative skin-flap necrosis as follows. First, the intravenous dose and concentration were set at 2.5 mg/mL and 0.05 mg/kg, respectively. Second, the timing of measurement was set to 100 s after the entry of indocyanine green into the skin (plateau phase); the analysis pattern was set to single frame. Third, comparisons among individuals were made using relative values. Conclusions: We analyzed the area of postoperative flap necrosis using this protocol. We found that the intraoperative images showed decreased perfusion in that area, which was useful in predicting skin-flap necrosis, as reported by previous breast reconstruction studies.

At our institution (Tokyo Medical and Dental University, Medical Hospital), latissimus dorsi flap (LD flap) reconstruction without a prosthetic implant is a popular surgical strategy following total mastectomy in patients with breast cancer. The LD flap, especially the extended LD flap, is usually rotated anteriorly through the axilla. However, if future additional surgery is required for axillary recurrence after LD flap reconstruction, the flap interferes with the visual field, making the surgical procedure more difficult. Because cases of axillary lymph node recurrence alone are rare, to the best of our knowledge, no paper has reported in detail on the technique and course of lymph node recurrence after LD flap reconstruction. The present study describes two cases of successful axillary lymph node dissection without sacrificing the flap for breast cancer recurrence after LD flap reconstruction. A brief analysis of immediate breast reconstruction with an LD flap performed in patients with breast cancer at our institution is also reported.

In haematological malignancies, such as malignant lymphoma, reprogramming of fatty acid metabolism favours tumour cell survival and drug resistance. Hydroxyacyl-CoA dehydrogenase trifunctional multienzyme complex subunit alpha (HADHA), an enzyme involved in fatty acid beta-oxidation (FAO), is overexpressed in high-grade lymphoma and is a predictor of poor prognosis in diffuse large B-cell lymphoma (DLBCL). HADHB forms a heterodimer with HADHA and functions as an FAO enzyme together with HADHA; however, the relevance of its expression in malignant lymphoma is unknown. In this study, we investigated the roles and antitumour effects of HADHB expression in malignant lymphoma. Immunohistochemical analysis showed that HADHB was frequently overexpressed in the high-grade lymphoma subtype. HADHB overexpression was observed in 68% (87/128) of DLBCL cases and was an independent predictor of poor prognosis (p=0.001). In vitro analysis demonstrated that HADHB knockdown suppressed cell proliferation in LCL-K and MD901 cells (p

Background Adenomyoepithelioma (AME) is a rare type of benign breast tumor. Many AMEs have demonstrated benign behavior, but very few cases of malignant AME have been reported. Case presentation A 53-year-old woman visited our hospital with a 16-mm, palpable mass in the right breast. Core needle biopsy was performed. The pathological diagnosis was AME. Partial mastectomy was performed. Two years later, local recurrence developed, and the patient again underwent partial mastectomy. The pathology showed malignant AME, and the margin was negative. Eight months later, local recurrence developed again in the same location, and total mastectomy was performed. The pathological diagnosis was malignant AME. The patient has remained recurrence-free without adjuvant therapy.Conclusions A very rare case of AME with repeated local recurrences and malignant transformation was reported.

The overexpression of glutathione peroxidase 4 (GPX4; an enzyme that suppresses peroxidation of membrane phospholipids) is considered a poor prognostic predictor of diffuse large B-cell lymphoma (DLBCL). However, the mechanisms employed in GPX4 overexpression remain unknown. GPX4 is translated as a complete protein upon the binding of SECISBP2 to the selenocysteine insertion sequence (SECIS) on the 3′UTR of GPX4 mRNA. In this study, we investigated the expression of SECISBP2 and its subsequent regulation of GPX4 and TXNRD1 in DLBCL patients. Moreover, we determined the significance of the expression of these selenoproteins in vitro using MD901 and Raji cells. SECISBP2 was positive in 45.5% (75/165 cases) of DLBCL samples. The SECISBP2-positive group was associated with low overall survival (OS) as compared to the SECISBP2-negative group (P = 0.006). Similarly, the SECISBP2 and GPX4 or TXNRD1 double-positive groups (P

Highly malignant tumors overexpress the minichromosome maintenance 2 (MCM2) protein in the nucleus, which is associated with advanced tumor grade, advanced stage, and poor prognosis. In this study, we showed that MCM2 is highly expressed in clinical samples of ovarian clear cell carcinoma. Although MCM2 expression was mainly localized to the nuclei as in other cancers, a few cases exhibited cytoplasmic localization of MCM2. Surprisingly, tumor samples with cytoplasmic MCM2 demonstrated excellent prognosis, showing 100% survival during the observation period of more than 200 months. However, cases with nuclear expression of MCM2 exhibited approximately 78% 5-year-survival rate. In a previous study, we showed that Friend leukemia virus (FLV) envelope protein gp70 bound to MCM2, impaired its nuclear translocation, and enhanced DNA damage-induced apoptosis in FLV-infected hematopoietic cells with high levels of MCM2. As expected, clear cell carcinoma cells with cytoplasmic expression of MCM2 exhibited significantly higher apoptotic cell ratio than that of cells with nuclear MCM2 expression. In vitro experiments using ovarian cancer cells with cytoplasmic expression of MCM2 demonstrated that transfection of MCM2-ΔN enhanced DNA damage-induced apoptosis. Therefore, cytoplasmic localization of MCM2 significantly correlated with increased apoptosis in clear cell carcinoma cells, resulting in improved prognosis.

The oncological safety of immediate breast reconstruction (IBR) in lymph node-positive patients is unclear. In the present study, the impact of IBR on recurrence based on data of patients with axillary lymph node metastases only was examined. The subjects were 232 patients who underwent breast surgery. The patients were grouped into 2 cohorts: non-IBR patients who underwent mastectomy with axillary lymph node dissection; and IBR patients with tissue expander or flap transfer and axillary lymph node dissection. The Non-IBR group included 165 patients, and the IBR group included 67 patients. For the comparison of oncological outcomes between the 2 groups, propensity score matching was performed. The propensity scores were calculated by logistic regression analysis, including age, tumor staging, human epidermal growth factor receptor 2 status, and estrogen receptor status. There was no difference in locoregional recurrence-free survival (LRRFS) between the non-IBR and IBR groups. The 5-year LRRFS rate was 78.9% in the non-IBR group and 85.1% in the IBR group. There was no difference in recurrence-free survival (RFS) between the non-IBR and IBR groups. The 5-year RFS rate was 75.6% in the non-IBR group and 78.8% in the IBR group. In all patients, the 5-year LRRFS rate was 77.3%, and the RFS rate was 70.5%. Multivariate Cox regression analysis to identify factors affecting RFS in all patients showed that estrogen receptor status and high nuclear grade were significant prognostic factors; IBR was irrelevant. This is the first report of an analysis using propensity score matching limited to node-positive breast cancer patients, and it showed that IBR is relatively safe in such patients.

Ferroptosis, a term first proposed in 2012, is iron-dependent, non-apoptotic regulatory cell death induced by erastin. Ferroptosis was originally discovered during synthetic lethal screening for drugs sensitive to RAS mutant cells, and is closely related to synthetic lethality. Ferroptosis sensitizes cancer stem cells and tumors that undergo epithelial−mesenchymal transition and are resistant to anticancer drugs or targeted therapy. Therefore, ferroptosis-inducing molecules are attractive new research targets. In contrast, synthetic lethal strategies approach mechanisms and genetic abnormalities that cannot be directly targeted by conventional therapeutic strategies, such as RAS mutations, hypoxia, and abnormalities in the metabolic environment. They also target the environment and conditions specific to malignant cells, have a low toxicity to normal cells, and can be used in combination with known drugs to produce new ones. However, the concept of synthetic lethality has not been widely adopted with ferroptosis. In this review, we surveyed the literature on ferroptosis-related factors and synthetic lethality to examine the potential therapeutic targets in ferroptosis-related molecules, focusing on factors related to synthetic lethality, discovery methods, clinical application stages, and issues in drug discovery.

Preoperatively diagnosed ductal carcinoma in situ (DCIS) is sometimes upstaged to invasive cancer by postoperative pathological examination. Various preoperative factors associated with upstaging to invasive cancer have been reported; however, this subject remains to be clarified. DCIS takes various forms on imaging, but many cases show non-mass-type lesions. In non-mass-type DCIS, recognizing the presence of invasion is difficult. To investigate predictors associated with upstaging to invasive cancer more precisely, we examined only non-mass-type DCIS. The present study retrospectively analyzed 101 patients diagnosed with non-mass-type DCIS preoperatively on breast biopsy at our institution between 2007 and 2017. Data were analyzed using Fisher's exact probability test and two-sample t-tests. Multivariate analysis was performed using logistic regression. The results showed that 27 patients (27%) were finally diagnosed with invasive cancer. Univariate analysis revealed abnormal result of palpation on breast examination (P=0.05), comedo necrosis (P=0.05), and HER2 status (P=0.02) as significant predictors. Multivariate analysis revealed an abnormal result of palpation as an independent predictor of invasive cancer underestimation (odds ratio 4.76; confidence interval 1.44-15.7; P=0.01). In conclusion, preoperatively diagnosed non-mass-type DCIS represented an underestimation in approximately 27% of cases. In particular, the presence of a clinically abnormal palpation increases the chance of upstaging to invasive cancer.

Background MYC is one of the proto-oncogenes contributing to tumorigenesis in many human cancers. Although the mechanism of MYC regulation is still not fully understood, learning about the comprehensive mechanism controlling the transcriptional activity of MYC will lead to therapeutic targets. The CRISPR/Cas9 library system is a simple and powerful screening technique. This study aims to identify new transcriptional upstream activators of MYC using the CRISPR activation library with new promoter-reporter systems. Methods and Results The MYC promoter-reporter system was developed with a photoconvertible fluorescent protein, Dendra2, and named “pMYC-promoter-Dendra2.” This MYC promoter-reporter system was designed to harbor a proximal MYC promoter at (3.1 kb). Both the CRISPR activation library and pMYC-promoter-Dendra2 were induced to HEK 293T cells, and Dendra2-positive cells, that are supposed that MYC should be upregulated, were collected individually by a cell sorter. Among the 169 cells collected, 12 clones were successfully established. Then, pMYC-promoter-Dendra2 was transfected again into these 12 clones, and two of 12 clones showed Dendra2 positivity. In this procedure, the cells with non-specific autofluorescence were correctly distinguished by utilizing the photoswitchable character of Dendra2. Using extracted genomic DNA of these two Dendra2 positive clones, polymerase chain reaction (PCR) was performed to amplify the guide RNA (gRNA) containing region, which was introduced by the CRISPR activation library. Eventually, PLEKHO2, MICU, MBTPS1, and M1AP were identified, and these gRNAs were transfected individually into HEK 293T cells again using the CRISPR activation system. Only M1AP gRNA transfected cells showed Dendra2-positive fluorescence. Then, the overexpression vector for M1AP with a doxycycline-inducible vector confirmed that M1AP induced high MYC expression by real-time quantitative PCR and western blot. Furthermore, the dual-luciferase assay showed a significant increase of promoter activity, and MYC mRNA was higher in M1AP- overexpressing cells. M1AP is highly expressed in several cancers, though, a positive correlation between M1AP and MYC was observed only in human acute myeloid leukemia. Conclusion The present study confirmed that the experimental method using the CRISPR library technology functions effectively for the identification of molecules that activate endogenous MYC. This method will help elucidate the regulatory mechanism of MYC expression, as well as supporting further drug research against malignant tumors.

Because fungi usually spread from the paranasal sinuses to the orbital apex in invasive fungal sinusitis (IFS), IFS often presents as an orbital apex syndrome (OAS) characterized by dysfunction of cranial nerves II, III, IV, V1, and VI. We report a case of sphenoid sinus mucormycosis that presented as isolated retrobulbar optic neuropathy. A 94‐year‐old woman presented with acute blindness in the right eye. Examination revealed the absence of light perception and pupillary reflex in the right eye. Head MRI showed a mass in the right sphenoid sinus, which was contiguous with the right optic nerve. She underwent endoscopic surgery, and a histopathological diagnosis of mucormycosis was established. Treatment with intravenous liposomal amphotericin B reduced the size of the mass. She has survived for more than 1 year without recurrence. Clinicians should consider that IFS can present as isolated retrobulbar optic neuropathy.

We encountered an 83-year-old man with 3-repeat dominant grain-like tau deposition. Tau-positive lesions exhibited apparent similarity to argyrophilic grains in terms of their distribution in the ambient gyrus, amygdala, and dorsomedial temporal tip and the characteristic comma-like morphology. The abundant oligodendroglial tau immunoreactivities were 3-repeat dominant. Tuft-shaped astrocytes showed partial 3-repeat tau immnoreactivities. These grain-like structures, as well as tuft-shaped astrocytes and oligodendroglia, exhibited predominant 3-repeat tau immunoreactivity, suggesting that grain-like structures and their characteristic distribution are mutually linked and not unique to 4-repeat tau deposition. pTDP immunoreactivity, extensive macrophage infiltration, and spongiosis were associated with these 3-repeat tau deposits.

Even though the BCR-ABL tyrosine kinase inhibitor imatinib significantly improves the prognosis of chronic myelogenous leukaemia (CML) patients, drug resistance is a major obstacle to better management. We examined the interaction of recently defined bone marrow microenvironment factors CXCL12 and ATP-binding cassette (ABC) transporters in the bone marrow of CML patients in the chronic phase and blast crisis.Expression levels of mRNA extracted from frozen specimens of CML patients were measured by real-time polymerase chain reaction. The expression of the ABC transporters MDR1, ABCC1, ABCG2, and CXCL12 was significantly higher in the bone marrow samples of CML blast crisis than in those of CML chronic phase. Immunohistochemical staining for CXCL12 revealed that the proportion of CXCL12 positive reticular cell areas correlated well with the mRNA levels of CXCL12 in CML bone marrow. Finally, co-culture experiments of K562 CML cells with CXCL12 expressing mesenchymal cells (OP9 cells or human CXCL12 transfected 3T3 cells) revealed enhanced mRNA levels for MDR1 in a CXCL12 rich environment.These results suggest that imatinib treatment restores the bone marrow microenvironment in CML with the presence of CXCL12 expressing reticular cells but in turn induces the overexpression of MDR1 in haematopoietic cells due to up-regulated expression of CXCL12.

Summary Background Mycosis fungoides (MF) classically presents from patch stage to plaque stage over a number of years and finally progresses to tumour stage with nodal or visceral involvement. The mechanism of progression remains incompletely elucidated. Chemokines and their receptors are known to be involved in disease mechanisms, with CXCL12 and CXCR4 playing a critical role in carcinogenesis, invasion and cancer cell migration in various carcinomas. Objectives To investigate the expression of CXCL12 and CXCR4 in different cutaneous stages of MF. Methods Formalin-fixed, paraffin-embedded skin samples from 40 patients with MF (21 patch stage, 10 plaque stage, nine tumour stage) and 30 non-neoplastic control skin samples were analysed. CXCL12 and CXCR4 were assessed by quantitative reverse-transcription polymerase chain reaction and immunohistochemical staining. Results The expression level of mRNA for CXCL12 in plaque-stage MF was significantly higher than in control skin (P = 0·0035), or patch-stage (P = 0·0108) or tumour-stage disease (P = 0·0089). The CXCR4 mRNA expression level in plaque-stage disease was significantly higher than in control skin (P = 0·0090) or patch-stage disease (P = 0·0387). CXCL12- and CXCR4-positive cell rates in patch-stage and plaque-stage MF were significantly higher than those in control skin (P

Aims: Multidrug resistance (MDR) in B-cell lymphomas still constitutes a major obstacle to the effectiveness of chemotherapy even in the anti-CD20 antibody therapy era. The aim of this study was to investigate the expression of MDR-associated molecules in reactive lymphadenopathy (RL), follicular lymphoma (FL), and diffuse large B-cell lymphoma (DLBCL). Methods and results: The expression of mRNA for ABC-transporter family genes was determined by real-time RT-PCR in lymph nodes from RL, FL, and DLBCL cases. MDR1 exhibited significantly stronger expression in RL, FL, and DLBCL than Raji B-cell lymphoma cells. RL and FL showed significantly higher expression than DLBCL. Immunohistochemically, MDR1 positive cells were localized in the germinal centers of RL and center of the nodular lesions of FL showing associations with CD21 positive follicular dendritic cells (FDCs). Raji cells were co-cultured with FDC sarcoma-derived cells and the expression of MDR1 and drug resistance were analyzed. The co-culture of Raji cells with FDCs induced strong expression of MDR1 and introduced resistance to doxorubicin-induced apoptosis. Conclusions: These results suggest that FDCs induce MDR1 expression in reactive as well as neoplastic B-cells. Inhibition of the interaction of FDCs with B-cells may provide a novel strategy for treating the chemotherapy resistant fraction.