A small-scale experimental study of breast FNA consultation on the internet using Panoptiq.

INTRODUCTION: To test the potential for cytopathology consultation using Panoptiq (ViewsIQ, Richmond, BC, Canada; this is a new type of whole-slide image that is made manually and incorporates video content), we investigated its application in the cytopathological diagnosis of cases that were difficult to diagnose by breast fine-needle aspiration (FNA). MATERIALS AND METHODS: Panoptiq files were created from liquid-based cytology slides prepared by the BD CytoRich Red (BD, Franklin Lakes, NJ) method. The slides were prepared from 23 consecutive samples of breast FNA that had been diagnosed as atypical or suspicious by the Hokkaido Cancer Center, Hokkaido, Japan. Nine volunteer reviewers, who were provided with the URL of the Panoptiq file, the original cytopathological diagnosis, and the clinical information, were asked to classify the cytopathological diagnosis of each case into 4 diagnostic categories (benign, atypical, suspicious, or malignant). We examined the consultation benefit (CB)-how much closer the reviewer's cytopathology diagnosis came to the final histopathological diagnosis than the original cytodiagnosis. The CB scoring system was decided in advance. RESULTS: All 9 reviewers showed a positive total CB score and 2 reviewers showed a significantly higher CB score (Wilcoxon's signed rank test). The representative diagnosis (ie, the most frequently rendered diagnosis in each case) also showed a significant CB. CONCLUSIONS: Our small-scale experimental study, in which Panoptiq was used in the diagnosis of cases that were difficult to diagnose definitively by breast FNA, revealed a positive CB score by every reviewer and the representative diagnosis showed a significant CB. The study suggests that Panoptiq could be used for cytopathology consultation.

Postoperative radiotherapy for T1/2N0M0 mucoepidermoid carcinoma positive for CRTC1/3-MAML2 fusions.

BACKGROUND: The National Comprehensive Cancer Network (NCCN) guidelines recommend considering postoperative radiotherapy (PORT) for completely resected T1/2N0M0 salivary mucoepidermoid carcinomas when they show tumor spillage, perineural invasion, or intermediate/high-grade histology. CRTC1/3-MAML2 fusions have been associated with a favorable clinical outcome. METHODS: Forty-seven T1/2N0M0 mucoepidermoid carcinoma cases positive for CRTC1/3-MAML2 fusions were completely resected and were not treated with PORT. RESULTS: Pathologically, none of the cases showed tumor spillage or perineural invasion. Cases with intermediate/high-grade histology numbered 9 (19%) to 26 (55%) with the currently used 3 different grading systems. During the follow-up (median 60 months), locoregional tumor recurrence occurred in 4 cases, which were treated with surgery alone. At the last follow-up (median 60 months; 7-160), all patients were alive with no evidence of disease. CONCLUSION: An excellent prognosis may be achieved without PORT in T1/2N0M0 mucoepidermoid carcinoma patients positive for CRTC1/3-MAML2 fusions when the tumors are completely resected without tumor spillage.

Mutation analysis of the EGFR pathway genes, EGFR, RAS, PIK3CA, BRAF, and AKT1, in salivary gland adenoid cystic carcinoma.

Adenoid cystic carcinoma (AdCC), one of the most common salivary gland carcinomas, usually has a fatal outcome. Epidermal growth factor receptor (EGFR) pathway gene mutations are important in predicting a patient's prognosis and estimating the efficacy of molecular therapy targeting the EGFR pathway. In this study of salivary gland AdCC (SAdCC), we looked for gene mutations in EGFR, RAS family (KRAS, HRAS, and NRAS), PIK3CA, BRAF, and AKT1, using a highly sensitive single-base extension multiplex assay, SNaPshot. Out of 70 cases, EGFR pathway missense mutations were found in 13 (18.6%): RAS mutations in 10 (14.3%), EGFR in one (1.4%), and PIK3CA in 5 (7.1%). None of the cases showed an EGFR deletion by direct sequencing. Concurrent gene mutations were found in three cases (4.3%). EGFR pathway mutations were significantly associated with a shorter disease-free (p = 0.011) and overall survival (p = 0.049) and RAS mutations were as well; (p = 0.010) and (p = 0.024), respectively. The gene fusion status as determined by a FISH assay had no significant association with mutations of the genes involved in the EGFR pathway. In conclusion, EGFR pathway mutations, especially RAS mutations, may be frequent in SAdCC, and associated with a poor prognosis for the patient.

Improved clonality detection in Hodgkin lymphoma using a semi-nested modification of the BIOMED-2 PCR assay for IGH and IGK rearrangements: A paraffin-embedded tissue study.

The BIOMED-2 PCR protocols targeting IGH and IGK genes may be useful for detecting clonality in Hodgkin lymphoma (HL). The clonality detection rates, however, have not been very high with these methods using paraffin-embedded tumor sections. We previously described the usefulness of the semi-nested BIOMED-2 IGH assay in B-cell malignancies. In this study, we devised a novel semi-nested BIOMED-2 IGK assay. Employing 58 cases of classical HL, we carried out the standard BIOMED-2, BIOMED-2 followed by BIOMED-2 re-amplification, and BIOMED-2 followed by semi-nested BIOMED-2, all targeting IGH and IGK, using paraffin-embedded tissues. In both IGH and IGK assays, semi-nested assays yielded significantly higher clonality detection rates than the standard assays and re-amplification assays. Clonality was detected in 13/58 (22.4%) classical HL cases using the standard IGH/IGK assays while it was detected in 38/58 (65.5%) cases using semi-nested IGH/IGK assays. The detection rates were not associated with the HL subtypes, CD30-positive cell density, CD20-positive cell density, or Epstein-Barr virus (EBV) positivity. In conclusion, tumor clonality was detected in nearly two-thirds of classical HL cases using semi-nested BIOMED-2 IGH/IGK assays using paraffin tumor sections. These semi-nested assays may be useful when the standard IGH/IGK assays fail to detect clonality in histopathologically suspected HLs.

A comparative study of PD-L1 immunohistochemical assays with four reliable antibodies in thymic carcinoma.

Currently, four immunohistochemical assays are registered with the US Food and Drug Administration to detect the expression of PD-L1. We investigated the PD-L1 expression in thymic carcinomas using these four diagnostic assays. The cases of 53 patients were reviewed and their specimens were subjected to four PD-L1 assays with different antibodies (SP142, SP263, 22C3, and 28-8). The PD-L1 expression in tumor cells (TCs) and immune cells (ICs) was evaluated. In TCs, the four assays showed similar scores in each case. Histopathologically, high TC scores were observed in squamous cell carcinomas (SqCCs). Meanwhile, there were no significant relationships among the IC scores in the four assays. In SqCCs, the high expression of PD-L1 (defined as ≥50% TC score) in TCs tended to be associated with early stage cancer. The patients with high expression levels of PD-L1 tended to show longer overall survival in the 22C3 assays (p=0.0200). In thymic carcinomas, the staining pattern showed high concordance among the four assays when TCs - rather than ICs - were stained. High PD-L1 positivity in TCs, especially in SqCCs, indicated that PD-1/PD-L1 targeted therapy may be a promising therapeutic approach.

Epididymal phospholipidosis is a possible mechanism for spermatotoxicity induced by the organophosphorus insecticide fenitrothion in rats.

Fenitrothion (FNT) is used worldwide in agricultural and public health settings. Spermatogenesis is a toxicological target of FNT under high-dose exposure. Although anti-androgenic action is postulated to be the mechanism associated with this toxicity, few studies have examined histopathology of androgen-dependent male accessory sex organs. The present study aimed to reveal the effects of FNT on the accessory organs of rats exhibiting spermatotoxicity in the absence of testicular histopathological changes. Furthermore, a possible novel molecular target was clarified. Male Wistar rats were orally administered 5 or 10 mg/kg FNT or its major metabolite 3-methyl-4-nitrophenol (MNP), or vehicle only, 4 days per week for 9 weeks. Then the epididymis, prostate, and seminal vesicles were collected. FNT and MNP did not show anti-androgenic effects but FNT induced cytoplasmic vacuolation in the epithelial cells of epididymal ducts and hyperplasia of mucosal folds/epithelial papillomatosis in seminal vesicles. FNT and MNP induced epididymal phospholipidosis, which was presumably caused by inhibition of epididymal secreted phospholipase A2 (sPLA2). Percentages of morphologically normal sperm and immature sperm were significantly predicted from both epididymal sPLA2 and phospholipid levels and from epididymal sPLA2, respectively. These results suggest that epididymal phospholipidosis plays an important role in FNT-induced spermatotoxicity. Anti-androgenic actions were not observed.

Clinical significance of tryptophan catabolism in Hodgkin lymphoma.

Indoleamine 2,3-dioxygenase 1 (IDO) is an enzyme catabolizing tryptophan (Trp) into the kynurenine (Kyn) pathway. The purpose of the present study was to determine the clinical significance of Trp catabolism in newly diagnosed Hodgkin lymphoma (HL) patients. We quantified serum Trp and Kyn in 52 HL patients, and analyzed their associations with different clinical parameters including serum soluble CD30 concentration. The IDO expression was evaluated in the patients' affected lymph nodes. The cohort comprised 22 male and 30 female patients (age range, 15-81 years; median, 45 years), with a 5-year overall survival (OS) of 88.6%. The OS was significantly shorter for patients with a high Kyn/Trp ratio (OS at 5 years, 60.0% vs 92.2%), for those with stage IV disease, and for those with lymphocytopenia (<600/mm3 and/or <8% white blood cell count). The latter two parameters are components of the international prognostic score for advanced HL. In contrast, there were no significant differences in OS according to age, serum albumin, hemoglobin, sex, white blood cell count, or serum soluble CD30 (≥ or <285.6 ng/mL). Multivariate analysis using the three variables stage, lymphocytopenia, and serum Kyn/Trp ratio showed that only the latter significantly affected OS. Indoleamine 2,3-dioxygenase 1 was produced by macrophages/dendritic cells, but not by HL tumor cells, and IDO levels determined by immunohistochemistry had a significant positive correlation with the serum Kyn/Trp ratio. In conclusion, quantification of serum Kyn and Trp is useful for predicting prognosis of individual HL patients.

Achromobacter Infection Is Rare in Japanese Patients with Pulmonary B-cell Lymphoma.

Objective Achromobacter xylosoxidans (A. xylosoxidans) has been recently reported to have an association with the development of pulmonary mucosa-associated lymphoid tissue (MALT) lymphoma in patients from European countries. However, the prevalence rates for A. xylosoxidans may vary significantly from country to country. To assess this association, the prevalence of A. xylosoxidans was analyzed in Japanese patients with pulmonary B-cell lymphoma. Methods DNA samples were obtained from formalin-fixed, paraffin-embedded sections of pulmonary MALT lymphomas (n=52), diffuse large B-cell lymphomas (DLBCLs, n=18), and benign pulmonary lesions (n=19). All samples were histopathologically reviewed by experienced hematopathologists, and the clonality of all MALT lymphoma cases was confirmed by a polymerase chain reaction (PCR)-based IGH rearrangement clonality assay. They were also tested for the API2-MALT1 fusion transcript. The presence of bacterial DNA was assessed with a nested PCR, and DNA sequencing was performed to confirm the PCR specificity. Results A. xylosoxidans DNA was detected in 1/52 cases of pulmonary MALT lymphoma, 2/18 cases of DLBCL, and 0/19 cases of benign pulmonary lesions. The prevalence of A. xylosoxidans in pulmonary lymphoma was not significantly higher than in benign lesions. Conclusion The present study shows that A. xylosoxidans infection may not be associated with pulmonary B-cell lymphoma in a Japanese case series. Large-scale international studies are needed to clarify the role of A. xylosoxidans in pulmonary lymphoma.

Improved clonality detection in B-cell lymphoma using a semi-nested modification of the BIOMED-2 PCR assay for IGH rearrangement: A paraffin-embedded tissue study.

The BIOMED-2 PCR protocol for targeting the IGH gene is widely employed for detecting clonality in B-cell malignancies. Unfortunately, the detection of clonality with this method is not very sensitive when paraffin sections are used as a DNA source. To increase the sensitivity, we devised a semi-nested modification of a JH consensus primer. The clonality detection rates of three assays were compared: the standard BIOMED-2, BIOMED-2 assay followed by BIOMED-2 re-amplification, and BIOMED-2 assay followed by semi-nested BIOMED-2. We tested more than 100 cases using paraffin-embedded tissues of various B-cell lymphomas, and found that the clonality detection rates with the above three assays were 63.9%, 79.6%, and 88.0%, respectively. While BIOMED-2 re-amplification was significantly more sensitive than the standard BIOMED-2, the semi-nested BIOMED-2 was significantly more sensitive than both the standard BIOMED-2 and BIOMED-2 re-amplification. An increase in sensitivity was observed in all lymphoma subtypes examined. In conclusion, tumor clonality may be detected in nearly 90% of B-cell lymphoma cases with semi-nested BIOMED-2. This ancillary assay may be useful when the standard BIOMED-2 fails to detect clonality in histopathologically suspected B-cell lymphomas.

MYB, MYBL1, MYBL2 and NFIB gene alterations and MYC overexpression in salivary gland adenoid cystic carcinoma.

AIMS: Adenoid cystic carcinoma (AdCC) is one of the most common salivary gland malignancies and the long-term prognosis is poor. In this study, we examined alterations of AdCC-associated genes, MYB, MYBL1, MYBL2 and NFIB, and their target molecules, including MYC. The results were correlated to clinicopathological profile of the patients. METHODS AND RESULTS: Using paraffin tumour sections from 33 cases of salivary gland AdCC, we performed a detailed fluorescence in-situ hybridization (FISH) analysis for gene splits and fusions of MYB, MYBL1, MYBL2 and NFIB. We found that 29 of 33 (88%) AdCC cases showed gene splits in either MYB, MYBL1 or NFIB. None of the cases showed an MYBL2 gene alteration. AdCCs were divided genetically into six gene groups, MYB-NFIB (n = 16), MYB-X (n = 4), MYBL1-NFIB (n = 2), MYBL1-X (n = 1), NFIB-X (n = 6) and gene-split-negative (n = 4). AdCC patients showing the MYB or MYBL1 gene splits were associated with microscopically positive surgical margins (P = 0.0148) and overexpression of MYC (P = 0.0164). MYC expression was detected in both ductal and myoepithelial tumour cells, and MYC overexpression was associated with shorter disease-free survival of the patients (P = 0.0268). CONCLUSIONS: The present study suggests that (1) nearly 90% of AdCCs may have gene alterations of either MYB, MYBL1 or NFIB, suggesting the diagnostic utility of the FISH assay, (2) MYB or MYBL1 gene splits may be associated with local aggressiveness of the tumours and overexpression of MYC, which is one of the oncogenic MYB/MYBL1 targets and (3) MYC overexpression may be a risk factor for disease-free survival in AdCC.

Cyclin-dependent kinase 9 is a novel specific molecular target in adult T-cell leukemia/lymphoma.

Cyclin-dependent kinase 9 (CDK9), a subunit of the positive transcription elongation factor b (P-TEFb) complex, regulates gene transcription elongation by phosphorylating the C-terminal domain (CTD) of RNA polymerase II (RNAPII). The deregulation of CDK9/P-TEFb has important implications for many cancer types. BAY 1143572 is a novel and highly selective CDK9/P-TEFb inhibitor currently being investigated in phase 1 studies. We evaluated the therapeutic potential of BAY 1143572 in adult T-cell leukemia/lymphoma (ATL). As a result of CDK9 inhibition and subsequent inhibition of phosphorylation at serine 2 of the RNAPII CTD, BAY 1143572 decreased c-Myc and Mcl-1 levels in ATL-derived or human T-cell lymphotropic virus type-1 (HTLV-1)-transformed lines and primary ATL cells tested, leading to their growth inhibition and apoptosis. Median inhibitory concentrations for BAY 1143572 in ATL-derived or HTLV-1-transformed lines (n = 8), primary ATL cells (n = 11), and CD4+ cells from healthy volunteers (n = 5) were 0.535, 0.30, and 0.36 µM, respectively. Next, NOG mice were used as recipients of tumor cells from an ATL patient. BAY 1143572-treated ATL-bearing mice (once daily 12.5 mg/kg oral application) demonstrated significantly decreased ATL cell infiltration of the liver and bone marrow, as well as decreased human soluble interleukin-2 receptor levels in serum (reflecting the ATL tumor burden), compared with untreated mice (n = 8 for both). BAY 1143572-treated ATL-bearing mice demonstrated significantly prolonged survival compared with untreated ATL-bearing mice (n = 7 for both). Collectively, this study indicates that BAY 1143572 showed strong potential as a novel treatment of ATL.

Striated duct adenoma presenting with intra-tumoral hematoma and papillary thyroid carcinoma-like histology.

Striated duct adenoma of the salivary gland is a rare benign tumor characterized by unilayered duct epithelium and striations of the tumor cell membranes. To the best of our knowledge, only six cases have been reported in the literature. Here we report an additional case, which was complicated by an intra-tumoral hematoma on clinical presentation and by papillary thyroid carcinoma-like histology on intra-operative frozen section diagnosis. An asymptomatic 78-year-old male presented with a two-year-history of a painless tumor of the left parotid. An intra-tumoral hematoma, which is unusual for a salivary gland tumor, was suspected from results of pre-operative radiology. The patient then underwent a left parotidectomy. The intra-operative frozen section diagnosis indicated a benign tumor, although ectopic papillary thyroid carcinoma was raised as a differential diagnosis since the eosinophilic tumor cells occasionally possessed nuclear grooves and nuclear pseudo-inclusions. By precise histopathological examination using paraffin sections, the tumor was finally diagnosed as striated duct adenoma. This type of tumor has unique features of hypervascular stroma and papillary thyroid carcinoma-like nuclei. In our case, the former feature was associated with the intra-tumoral hematoma and the latter feature, with difficulty in frozen section tumor diagnosis.

Expression of cancer/testis antigens in salivary gland carcinomas with reference to MAGE-A and NY-ESO-1 expression in adenoid cystic carcinoma.

AIMS: Cancer/testis antigens (CTAs) are detected in cancer cells but not in healthy normal tissues, with the exception of gametogenic tissues. CTAs are highly immunogenic proteins, and thus represent ideal targets for cytotoxic T-lymphocyte-mediated specific immune therapy. The aim of this study was to screen CTA expression in various types of salivary gland carcinoma and to clarify clinicopathological significance of MAGE-A and NY-ESO-1 expression in adenoid cystic carcinomas (AdCCs) of the salivary gland, which is one of the most common salivary gland carcinomas, and usually has a fatal outcome. METHODS AND RESULTS: We used immunohistochemistry to examine the expression of four CTAs (MAGE-A, NY-ESO-1, CT7, and GAGE7) in various types of salivary gland carcinoma (n = 95). When carcinoma cases were divided into low-grade and intermediate/high-grade types, NY-ESO-1 and CT7 were expressed more frequently in intermediate/high-grade carcinomas. We then focused on MAGE-A and NY-ESO-1 expression in a large cohort of adenoid cystic carcinomas (AdCCs) (n = 46). MAGE-A and NY-ESO-1 were frequently expressed in AdCC; specifically, MAGE-A was expressed in >60% of the AdCC cases. MAGE-A expression and tumour site (minor salivary gland) were identified as independent risk factors for locoregional tumour recurrence. CONCLUSIONS: These findings suggest that CTAs may be expressed in a variety of salivary gland carcinomas, especially in those with higher histological grades. In addition, MAGE-A, which is frequently expressed in AdCC cases, may be a useful prognostic factor for poorer locoregional recurrence-free survival.

Quantitative PCR for HTLV-1 provirus in adult T-cell leukemia/lymphoma using paraffin tumor sections.

Detection of HTLV-1 provirus using paraffin tumor sections may assist the diagnosis of adult T-cell leukemia/lymphoma (ATLL). For the detection, non-quantitative PCR assay has been reported, but its usefulness and limitations remain unclear. To our knowledge, quantitative PCR assay using paraffin tumor sections has not been reported. Using paraffin sections from ATLLs and non-ATLL T-cell lymphomas, we first performed non-quantitative PCR for HTLV-1 provirus. Next, we determined tumor ratios and carried out quantitative PCR to obtain provirus copy numbers. The results were analyzed with a simple regression model and a novel criterion, cut-off using 95 % rejection limits. Our quantitative PCR assay showed an excellent association between tumor ratios and the copy numbers (r = 0.89, P < 0.0001). The 95 % rejection limits provided a statistical basis for the range for the determination of HTLV-1 involvement. Its application suggested that results of non-quantitative PCR assay should be interpreted very carefully and that our quantitative PCR assay is useful to estimate the status of HTLV-1 involvement in the tumor cases. In conclusion, our quantitative PCR assay using paraffin tumor sections may be useful for the screening of ATLL cases, especially in HTLV-1 non-endemic areas where easy access to serological testing for HTLV-1 infection is limited.

Cellular-level characterization of B cells infiltrating pulmonary MALT lymphoma tissues.

Mucosa-associated lymphoid tissue (MALT) lymphoma mainly consists of three types of tumor B cells, small (centrocyte-like), scattered large transformed, and intraepithelial. However, it is difficult to differentiate tumor B cells from reactive B cells at the cellular level. We examined five cases of API2-MALT1 fusion-positive MALT lymphoma of the lung. A single paraffin section for each case was subjected to sequential retrieval of whole-slide imaging (WSI) data of hematoxylin and eosin (HE) staining, immunofluorescence staining for CD79a, and fluorescence in situ hybridization (FISH) for the MALT1 split. We counted the number of MALT1 split-positive or MALT1 split-negative cells among CD79a-positive cells. The MALT1 split was detected in 59, 46, and 76 % of small, large, and intraepithelial B cells, respectively. A review of the HE-WSI data showed that cytomorphological distinction between the MALT1 split-positive and MALT1 split-negative B cells was virtually impossible. None of CD79a-negative lymphoid cells, epithelial cells, and microvascular endothelial cells was positive for MALT1 splits. As API2-MALT1 fusion is an early and critical event in the lymphomatogenesis, our findings are best interpreted as that a considerable number of B cells, either small, large, or intraepithelial, are reactive cells and that it is difficult to distinguish cytomorphologically between tumor B cells and reactive B cells. These findings suggest that the tumor architecture may be the central factor for making a correct histopathological diagnosis of MALT lymphoma. The sequential WSI of HE staining, immunofluorescence staining, and FISH as described here is a useful tool for pathological analysis at the cellular level.

Labial salivary gland biopsy for diagnosing immunoglobulin light chain amyloidosis: a retrospective analysis.

Our goal was to evaluate the usefulness of labial salivary gland (LSG) biopsy for diagnosing immunoglobulin light chain (AL) amyloidosis, by comparing bone marrow and skin biopsies in the same patient population. This retrospective study included 34 consecutive patients who showed evidence of monoclonal proteins and symptoms considered to be due to amyloidosis, and who underwent a tissue biopsy from LSG between January 2005 and December 2012 at Nagoya City University Hospital. All samples of superficial tissues, including LSG, bone marrow, and skin, were independently evaluated as having amyloid deposits by a central review, which was blind to clinical information. An AL amyloidosis diagnosis was based on evidence of amyloid deposition in any biopsied tissue. Eighteen patients were diagnosed with AL amyloidosis. The sensitivity for detecting amyloid deposition was highest in biopsies of LSG at 89 %, followed by 77 % for bone marrow, and 72 % for skin. Amyloid deposition was detected in at least one superficial tissue of all the 18 patients. An LSG biopsy may be appropriate as a first-choice procedure to diagnose AL amyloidosis. Multiple biopsies of superficial tissues, including LSG, bone marrow, and skin, are recommended to increase the sensitivity for diagnosing AL amyloidosis.

[Prognostic significance of tryptophan catabolism in adult T-cell leukemia/lymphoma].

PURPOSE: Indoleamine 2,3-dioxygenase 1 (IDO1: IDO) is an enzyme catabolizing tryptophan (Trp) into the kynurenine (Kyn) pathway, and is an important micro-environmental factor suppressing antitumor immune responses. We investigated the prognostic significance of Trp catabolism in adult T-cell leukemia/lymphoma (ATL). EXPERIMENTAL DESIGN: We quantified serum Trp and Kyn in 96 ATL patients, 38 human T-cell lymphotropic virus type-1 asymptomatic carriers (HTLV-1 ACs), and 40 healthy adult volunteers. The relationships between various clinical parameters were analyzed. IDO expression was evaluated in the affected lymph nodes of ATL patients. RESULTS: Serum Kyn concentrations and Kyn/Trp ratios were significantly higher in HTLV-1 ACs than in healthy controls. Both increased significantly with progression from HTLV-1 AC to ATL. There were no significant differences in serum Trp concentrations between ATL patients, HTLV-1 ACs and controls. IDO was possibly produced by ATL and/or cells in the microenvironment. Multivariate analyses demonstrated a high serum Kyn/Trp ratio and high Kyn level, but not a high Trp level, to be significant independent detrimental prognostic factors in ATL and aggressive variant ATL. CONCLUSIONS: Quantification of serum Kyn and Trp is prognostically useful for individual ATL patients. Furthermore, ATL is an appropriate disease for testing novel cancer immunotherapies targeting IDO.

Warthin-like Mucoepidermoid Carcinoma: A Combined Study of Fluorescence In Situ Hybridization and Whole-slide Imaging.

There has been some debate as to whether a subset of metaplastic Warthin tumors (mWTs) harbor the mucoepidermoid carcinoma (MEC)-associated CRTC1-MAML2 fusion. We analyzed 15 tumors originally diagnosed as mWT (mWT-like tumors), 2 of which had concurrent MECs. We looked for the CRTC1/3-MAML2 fusion transcripts and performed immunohistochemistry for p63 and fluorescence in situ hybridization (FISH) for the MAML2 split. To localize MAML2 split-positive cells at the cellular level, whole tumor tissue sections were digitalized (whole-slide imaging [WSI]). The CRTC1-MAML2, but not CRTC3-MAML2 was detected in 5/15 mWT-like tumors. FISH-WSI results showed that all epithelial cells harbored the MAML2 split in fusion-positive mWT-like tumors and were totally negative in fusion-negative mWT-like tumors. A review of the hematoxylin and eosin-stained slides showed that morphology of the "metaplastic" epithelium was virtually indistinguishable between fusion-positive and fusion-negative tumors. However, oncocytic bilayered tumor epithelium, characteristic to typical WT, was always found somewhere in the fusion-negative tumors but not in the fusion-positive tumors. This distinguishing histologic finding enabled 5 pathologists to easily differentiate the 2 tumor groups with 100% accuracy. The age and sex distribution of fusion-positive mWT-like tumor cases was similar to that of fusion-positive MEC cases and significantly different from those of fusion-negative mWT-like tumor and typical WT cases. In addition, only fusion-positive mWT-like tumors possessed concurrent low-grade MECs. In conclusion, a subset of mWT-like tumors were positive for the CRTC1-MAML2 fusion and had many features that are more in accord with MEC than with WT. The term Warthin-like MEC should be considered for fusion-positive mWT-like tumors.

Prognostic Significance of Tryptophan Catabolism in Adult T-cell Leukemia/Lymphoma.

PURPOSE: Indoleamine 2,3-dioxygenase 1 (IDO1: IDO), an enzyme catabolizing tryptophan (Trp) into the kynurenine (Kyn) pathway, is increasingly being recognized as an important microenvironmental factor suppressing antitumor immune responses. The purpose of the present study was to determine the prognostic significance of Trp catabolism in adult T-cell leukemia/lymphoma (ATL). EXPERIMENTAL DESIGN: We quantified serum Trp and Kyn in 96 ATL patients, 38 human T-cell lymphotropic virus type-1 asymptomatic carriers (HTLV-1 ACs), and 40 healthy adult volunteer controls. The relationships between various clinical parameters including overall survival were analyzed. IDO expression was evaluated in the affected lymph nodes of ATL patients. RESULTS: Serum Kyn concentrations and Kyn/Trp ratios were significantly higher in HTLV-1 ACs than healthy controls. Both increased significantly with progression from HTLV-1 AC to ATL. However, there were no significant differences in the serum Trp concentrations between ATL patients, HTLV-1 ACs, and controls. IDO was possibly produced by ATL and/or cells of the microenvironment. Multivariate analyses demonstrated that a high serum Kyn/Trp ratio and high Kyn level, but not a high Trp level, were significantly independent detrimental prognostic factors in ATL, as well as in that subset of patients with aggressive variant ATL. CONCLUSIONS: Quantification of serum Kyn and Trp is useful for predicting prognosis of an individual ATL patient. Furthermore, ATL, especially in patients with a high serum Kyn/Trp ratio, is an appropriate disease for testing novel cancer immunotherapies targeting IDO.

Antitumor effects of bevacizumab in a microenvironment-dependent human adult T-cell leukemia/lymphoma mouse model.

OBJECTIVE: The objective of this study was to evaluate the therapeutic potential of bevacizumab with or without systemic chemotherapy for adult T-cell leukemia/lymphoma (ATL) and clarify the significance of angiogenesis for ATL pathogenesis. METHODS: NOD/Shi-scid, IL-2Rγ(null) (NOG) mice were used as recipients of tumor cells from a patient with ATL, which engraft and proliferate in a microenvironment-dependent manner. The ATL cells could be serially transplanted in NOG mice, but could not be maintained in in vitro cultures. RESULTS: Injection of bevacizumab alone significantly increased necrosis and decreased vascularization in the tumor tissue. Levels of human soluble interleukin two receptor in the serum (reflecting the ATL tumor burden) of bevacizumab-treated mice were significantly lower than in untreated mice. Although bevacizumab monotherapy showed these clear anti-angiogenesis effects, it did not prolong survival. In contrast, injection of bevacizumab together with cyclophosphamide, doxorubicin, vincristine, prednisolone (CHOP) led to a significant prolongation of survival of the ATL mice relative to CHOP alone. CONCLUSIONS: This is the first report to evaluate the efficacy of bevacizumab for ATL in a tumor microenvironment-dependent model. Bevacizumab therapy combined with chemotherapy could be a valuable treatment strategy for that subgroup of ATL probably depending to a large extent on angiogenesis via vascular endothelial growth factor.