Kanako C. Hatanaka

A pilot study of operational tolerance with a regulatory T‐cell‐based cell therapy in living donor liver transplantation

Potent immunosuppressive drugs have significantly improved early patient survival after liver transplantation (LT). However, long‐term results remain unsatisfactory because of adverse events that are largely associated with lifelong immunosuppression. To solve this problem, different strategies have been undertaken to induce operational tolerance, for example, maintenance of normal graft function and histology without immunosuppressive therapy, but have achieved limited success. In this pilot study, we aimed to induce tolerance using a novel regulatory T‐cell‐based cell therapy in living donor LT. Adoptive transfer of an ex vivo‐generated regulatory T‐cell‐enriched cell product was conducted in 10 consecutive adult patients early post‐LT. Cells were generated using a 2‐week coculture of recipient lymphocytes with irradiated donor cells in the presence of anti‐CD80/86 monoclonal antibodies. Immunosuppressive agents were tapered from 6 months, reduced every 3 months, and completely discontinued by 18 months. After the culture, the generated cells displayed cell‐number‐dependent donor‐specific inhibition in the mixed lymphocyte reaction. Infusion of these cells caused no significant adverse events. Currently, all patients are well with normal graft function and histology. Seven patients have completed successful weaning and cessation of immunosuppressive agents. At present, they have been drug free for 16‐33 months; 4 patients have been drug free for more than 24 months. The other 3 recipients with autoimmune liver diseases developed mild rejection during weaning and then resumed conventional low‐dose immunotherapy. Conclusions: A cell therapy using an ex vivo‐generated regulatory T‐cell‐enriched cell product is safe and effective for drug minimization and operational tolerance induction in living donor liver recipients with nonimmunological liver diseases. (Hepatology 2016;64:632‐643)

p53 accumulation is a strong predictor of recurrence in estrogen receptor-positive breast cancer patients treated with aromatase inhibitors.

Aromatase inhibitors have played a central role in endocrine therapy for estrogen receptor (ER)‐positive breast cancer in postmenopausal women. However, factors predictive of the efficacy of aromatase inhibitors, and prognostic factors, both for early and late recurrence in women treated with adjuvant aromatase inhibitors have not been identified. Whole genome analysis identified that a TP53 gene mutation exists in ER‐positive breast cancers, although the frequency of TP53 gene mutation in luminal tumors is lower compared with basal‐like or human epidermal growth factor receptor type 2 (HER2)‐positive breast cancers. We examined expression of p53, as well as ER, progesterone receptor, HER2 and Ki‐67 using immunohistochemistry in postmenopausal ER‐positive breast cancer patients who were treated with aromatase inhibitors as adjuvant endocrine therapy. There were 53 (21%) tumors that contained 10% or more p53‐positive cells. High p53 expression was positively correlated with tumor grade, HER2 score and Ki‐67 expression. Significant association was observed between disease‐free survival and high p53 expression in multivariate analysis (P < 0.0001). Compared with women without recurrence, women with early recurrence had significantly higher p53 expression (P < 0.0001), as did women with late recurrence (P = 0.037). The present study demonstrates that p53 accumulation is a strong predictor of both early and late recurrence in ER‐positive breast cancer patients treated with aromatase inhibitors as adjuvant endocrine therapy. TP53 gene alteration might be a key biological characteristic of ER‐positive breast cancer.

Nasal manifestations of immunoglobulin G4‐related disease

Immunoglobulin (Ig)G4‐related disease is a systemic syndrome, characterized by sclerosing lesions that mainly affect the exocrine tissue. Although some patients with IgG4‐related disease complain of nasal symptoms, there are few reports concerning the nasal manifestations of this disease. We investigated the clinical and pathological features of the nasal manifestations of IgG4‐related disease.

Heat shock factor 1 accelerates hepatocellular carcinoma development by activating nuclear factor-κB/mitogen-activated protein kinase

Heat shock factor 1 (HSF1), a major transactivator of stress responses, has been implicated in carcinogenesis in various organs. However, little is known about the biological functions of HSF1 in the development of hepatocellular carcinoma (HCC). To clarify the functional role of HSF1 in HCC, we established HSF1-knockdown (HSF1 KD) KYN2 HCC cells by stably expressing either small hairpin RNA (shRNA) against HSF1 (i.e. HSF1 KD) or control shRNA (HSF1 control). Tumorigenicity was significantly reduced in orthotopic mice with HSF1 KD cells compared with those with HSF1 control cells. Reduced tumorigenesis in HSF1 KD cells appeared attributable to increased apoptosis and decreased proliferation. Tumor necrosis factor-α-induced apoptosis was increased in HSF1 KD cells and HSF1(-/-) mouse hepatocytes compared with controls. Decreased expression of IκB kinase γ, a positive regulator of nuclear factor-κB, was also observed in HSF1 KD cells and HSF1(-/-) mouse hepatocytes. Furthermore, expression of bcl-2-associated athanogene domain 3 (BAG3) was dramatically reduced in HSF1 KD cells and HSF1(-/-) mouse hepatocytes. We also found that epidermal growth factor-stimulated mitogen-activated protein kinase signaling was impaired in HSF1 KD cells. Clinicopathological analysis demonstrated frequent overexpression of HSF1 in human HCCs. Significant correlations between HSF1 and BAG3 protein levels and prognosis were also observed. In summary, these results identify a mechanistic link between HSF1 and liver tumorigenesis and may provide as a potential molecular target for the development of anti-HCC therapies.

β1-integrin via NF-κB signaling is essential for acquisition of invasiveness in a model of radiation treated in situ breast cancer

IntroductionDuctal carcinoma in situ (DCIS) is characterized by non-invasive cancerous cell growth within the breast ducts. Although radiotherapy is commonly used in the treatment of DCIS, the effect and molecular mechanism of ionizing radiation (IR) on DCIS are not well understood, and invasive recurrence following radiotherapy remains a significant clinical problem. This study investigated the effects of IR on a clinically relevant model of Akt-driven DCIS and identified possible molecular mechanisms underlying invasive progression in surviving cells.MethodsWe measured the level of phosphorylated-Akt (p-Akt) in a cohort of human DCIS specimens by immunohistochemistry (IHC) and correlated it with recurrence risk. To model human DCIS, we used Akt overexpressing human mammary epithelial cells (MCF10A-Akt) which, in three-dimensional laminin-rich extracellular matrix (lrECM) and in vivo, form organotypic DCIS-like lesions with lumina expanded by pleiomorphic cells contained within an intact basement membrane. In a population of cells that survived significant IR doses in three-dimensional lrECM, a malignant phenotype emerged creating a model for invasive recurrence.ResultsP-Akt was up-regulated in clinical DCIS specimens and was associated with recurrent disease. MCF10A-Akt cells that formed DCIS-like structures in three-dimensional lrECM showed significant apoptosis after IR, preferentially in the luminal compartment. Strikingly, when cells that survived IR were repropagated in three-dimensional lrECM, a malignant phenotype emerged, characterized by invasive activity, up-regulation of fibronectin, α5β1-integrin, matrix metalloproteinase-9 (MMP-9) and loss of E-cadherin. In addition, IR induced nuclear translocation and binding of nuclear factor-kappa B (NF-κB) to the β1-integrin promoter region, associated with up-regulation of α5β1-integrins. Inhibition of NF-κB or β1-integrin signaling abrogated emergence of the invasive activity.ConclusionsP-Akt is up-regulated in some human DCIS lesions and is possibly associated with recurrence. MCF10A-Akt cells form organotypic DCIS-like lesions in three-dimensional lrECM and in vivo, and are a plausible model for some forms of human DCIS. A population of Akt-driven DCIS-like spheroids that survive IR progresses to an invasive phenotype in three-dimensional lrECM mediated by β1-integrin and NF-κB signaling.

Clinicopathologic Study of Pineal Parenchymal Tumors of Intermediate Differentiation

OBJECTIVE Pineal parenchymal tumors of intermediate differentiation (PPTID) are extremely rare tumor entities, and only limited data are available regarding their pathologic features and biologic behaviors. Because grading criteria of pineal parenchymal tumors (PPTs) have yet to be established, the treatment strategy and prognosis of PPTIDs remain controversial. We describe the clinicopathologic study of six patients with PPTID and compare responses for the treatment and prognosis with cases of pineocytoma (PC) and pineoblastoma (PB). From this analysis, we attempt to clarify the treatment strategy for PPTIDs. METHODS This study included 15 patients with PPTs, consisting of 6 PCs, 6 PPTIDs, and 3 PBs. We focused on the 6 patients with PPTIDs. All PPTID cases were treated surgically, and radiotherapy and chemotherapy were administered as adjuvant therapies in some cases. We have earlier reported the histopathologic study (Neuropathology 32:647-653, 2012). Briefly, we examined mitotic figures and necrosis by hematoxylin-eosin staining and immunohistochemical markers such as neuronal markers (synaptophysin, neurofilament (NF), and neuronal nuclear antigen), and an MIB-1 labeling index was determined. RESULTS In the PPTID cases, the extent of resection was variable and the recurrence rates among patients varied according to stage and treatment. All PC patients underwent total resection with no recurrence. All PB patients underwent resection and adjuvant therapy with radiotherapy and chemotherapy. There were no recurrences in patients with PC or PB. The results of histopathologic findings have been already reported as mentioned above. Briefly, the results indicated no mitotic figure or necrosis in any of the six cases of PPTID, but those features were observed in PB cases. All cases even including PC and PB were immunopositive for neuronal markers. The MIB-1 labeling index of PPTID was 3.5%, whereas it was 0% in PC and 10.5% in PB. CONCLUSIONS Good radiosensitivity of PPTIDs was observed in our series. Because there are cases with discrepancies between images and pathologic findings, it is very difficult to determine the proper treatment strategy for PPTIDs. Proliferative potential was correlated with World Health Organization grade, although the immunoreactivity of neuronal markers did not correlate with the histologic grade.

Chromophobe renal cell carcinoma, oncocytic variant: a proposal of a new variant giving a critical diagnostic pitfall in diagnosing renal oncocytic tumors

In chromophobe renal cell carcinoma (RCC), two forms of typical and eosinophilic variants have been reported to date. We have previously reported a new variant of chromophobe RCC, namely an oncocytic variant. However, little is known on the histological features of this variant. In this article, we report such five cases. Macroscopically, the tumor was well demarcated, but unencapsulated. The cut surface of the tumor showed brown in color, but neither hemorrhage nor necrosis was seen. Microscopically, the tumor consisted of predominant tubular configuration with or without various proportion of solid-sheet pattern. In one tumor, tumor cells microscopically invaded branches of renal vein. In addition, the constituting cells were characterized by the oncocytic cytoplasm, trivial to minimal variation in tumor size, indistinct to slightly distinct cell border, centrally located round nuclei and the absence of perinuclear halo. These characteristics entirely resembled renal oncocytoma. However, neoplastic cells immunohistochemically showed the diffuse and strong labeling for cytokeratin 7 and mitochondrial antigen in all cases. In addition, in fluorescence in situ hybridization (FISH) study the loss of more than four chromosomes among chromosomes 7, 10, 13, 17 and 21 was confirmed in all tumors and the diagnosis of chromophobe RCC was rendered. In conclusion, we propose a new variant, namely an oncocytic variant, of chromophobe RCC morphologically resembling renal oncocytoma and biologically showing characteristics of chromophobe RCC, and this recognition is practically crucial in the differential diagnosis from renal oncocytoma.

High level expression of AMAP1 protein correlates with poor prognosis and survival after surgery of head and neck squamous cell carcinoma patients

BackgroundDespite recent advances in cancer therapeutics in general, the survival of patients with head and neck squamous cell carcinomas (HNSCCs) has not improved substantially over the past few decades. HNSCC cells often exhibit invasive and metastatic phenotypes, and expression of epidermal growth factor receptor (EGFR) and cortactin has been highly implicated in the development of malignancy in HNSCCs. We have shown previously that an Arf6 pathway, in which Arf6 is activated by GEP100 and employs AMAP1 (also called DDEF1 or ASAP1) as its downstream effector, is pivotal for the invasion and metastasis of different breast cancer cells. This pathway is activated by receptor tyrosine kinases, including EGFR; and moreover, AMAP1 physically associates with cortactin, in which inhibition of this binding effectively blocks invasion and metastasis. We here investigated whether the expression of Arf6 pathway components correlates with the poor prognosis of HNSCC patients. We have shown previously that AMAP1 protein levels are not correlated with its mRNA levels, and hence we here employed immunohistochemical staining of HNSCC clinical specimens to investigate AMAP1 protein levels.ResultsWe found that high levels of AMAP1 protein expression on its own, as well as its co-overexpression with EGFR statistically correlates with poor disease-free survival and poor overall survival, while high levels of cortactin expression or its co-expression with EGFR did not.ConclusionOur identification of predictive biomarkers, together with our previous findings on the coherent signaling pathway that these biomarkers ultimately generate should be powerful information for the further development of HNSCC therapeutics.

Imaging characteristics of cardiac dominant diffuse large B-cell lymphoma demonstrated with MDCT and PET/CT

PurposeTo investigate the specific imaging findings of multidetector row CT (MDCT) and PET/CT with18F-FDG in cardiac dominant diffuse large B-cell lymphoma (DLBCL) in comparison with other cardiac tumours.MethodsFive patients with DLBCL and 12 patients with other cardiac tumours including pericardial tumours were retrospectively reviewed. Among the patients with other cardiac tumours, seven had metastatic tumours, three had benign tumours, and two had other malignant cardiac tumours. The location of the cardiac mass, the encasement of the coronary artery surrounded by the mass, and pericardial effusion were evaluated using MDCT. The disease activity of the cardiac tumour was also evaluated by PET/CT.ResultsFour of the five DLBCL patients had primarily right-sided cardiac lesions, which was seen significantly more frequently in DLBCL than in other cardiac tumours (p = 0.028). All cardiac DLBCL lesions were located around the atrioventricular groove and encased the coronary arteries. ECG-gated cardiac MDCT showed that there was no apparent stenosis of the coronary arteries. Large amounts of pericardial effusion were seen in all DLBCL patients. PET/CT revealed significantly higher FDG uptake in DLBCL than in other cardiac malignant tumours, with no overlap (p = 0.0007).ConclusionThe combination of a right-sided cardiac mass with a large pericardial effusion and no apparent stenosis of the encased coronary artery revealed by MDCT and a high maximum standard uptake value were the specific findings in cardiac dominant DLBCL.

Differences in morphological features and minimum apparent diffusion coefficient values among breast cancer subtypes using 3-tesla MRI

PURPOSE To compare the morphology and minimum apparent diffusion coefficient (ADC) values among breast cancer subtypes. METHODS Ninety-three patients, who underwent breast MRI and collectively had 98 pathologically proven invasive carcinomas, were enrolled. Morphology was evaluated according to BIRADS-MRI. Minimum ADC was measured. Morphology and minimum ADC were compared among subtypes. Multivariate logistic regression analyses were used to identify the characteristics associated with different subtypes. RESULTS Oval/round shape was significantly associated with triple-negative (TN) cancer (TN vs. non-TN: 90.9% vs. 45.2%; p=0.0123). Rim enhancement was significantly less frequent in Luminal A (Luminal A vs. non-Luminal A: 34.2% vs. 76.1%; p=0.0003). The minimum ADC of Luminal A was significantly higher than that of Luminal B (HER2-negative) (834 vs. 748×10(-6)mm(2)/s; p<0.025). The minimum ADC of the TN-special type was significantly higher than that of TN-ductal (997 vs. 702×10(-6)mm(2)/s; p<0.025). On the multivariate analysis comparing the characteristics associated with Luminal A vs. Luminal B (HER2-negative), the internal enhancement characteristics of the mass and minimum ADC were significant factors. CONCLUSION Morphology and minimum ADC would be useful in distinguishing breast cancer subtypes.