Molecular profiling of single circulating tumor cells from lung cancer patients.

Circulating tumor cells (CTCs) are established cancer biomarkers for the "liquid biopsy" of tumors. Molecular analysis of single CTCs, which recapitulate primary and metastatic tumor biology, remains challenging because current platforms have limited throughput, are expensive, and are not easily translatable to the clinic. Here, we report a massively parallel, multigene-profiling nanoplatform to compartmentalize and analyze hundreds of single CTCs. After high-efficiency magnetic collection of CTC from blood, a single-cell nanowell array performs CTC mutation profiling using modular gene panels. Using this approach, we demonstrated multigene expression profiling of individual CTCs from non-small-cell lung cancer (NSCLC) patients with remarkable sensitivity. Thus, we report a high-throughput, multiplexed strategy for single-cell mutation profiling of individual lung cancer CTCs toward minimally invasive cancer therapy prediction and disease monitoring.

Altered local and systemic immune profiles underlie lymph node metastasis in breast cancer patients.

Cancer-mediated immune dysfunction contributes to tumor progression and correlates with patient outcome. Metastasis to tumor draining lymph nodes (TDLNs) is an important step in breast cancer progression and is used to predict patient outcome and survival. Although lymph nodes are important immune organs, the role of immune cells in TDLNs has not been thoroughly investigated. We hypothesized that the host immune response in node negative (NN) patients is more intact and thereby can resist tumor invasion compared to node positive (NP) patients. As such, lymph node metastasis requires breakdown of the host immune response in addition to escape of cancer cells from the tumor. To investigate the immunological differences between NN and NP breast cancer patients, we purified and profiled immune cells from the three major compartments where cancer and immune cells interact: tumor, TDLNs and peripheral blood. Significant down-regulation of genes associated with immune-related pathways and up-regulation of genes associated with tumor-promoting pathways was consistently observed in NP patients' TDLNs compared to NN patients. Importantly, these signatures were seen even in NP patients' tumor-free TDLNs, suggesting that such immune changes are not driven solely by local tumor invasion. Furthermore, similar patterns were also observed in NP patients' tumor and blood immune cells, suggesting that immunological differences between NN and NP patients are systemic. Together, these findings suggest that alterations in overall immune function may underlie risk for LN metastasis in breast cancer patients.

A clinical trial of lovastatin for modification of biomarkers associated with breast cancer risk.

Pre-clinical and epidemiologic studies provide rationale for evaluating lipophilic statins for breast cancer prevention. We conducted a single-arm, biomarker modulation trial of lovastatin among women with increased risk of breast cancer. Eligibility criteria included a deleterious germline mutation in BRCA1, BRCA2, CDH1, or TP53; lifetime breast cancer risk of ≥20 % as estimated by the Claus model; or personal history of estrogen receptor and progesterone receptor-negative breast cancer. Participants received 40 mg of lovastatin orally twice daily for 6 months. We evaluated the following biomarkers before and after lovastatin use: breast duct cytology (primary endpoint), serum lipids, C-reactive protein, insulin-like growth factor-1, IGF binding protein-3, lipid peroxidation, oxidative DNA damage, 3-hydroxy-3-methylglutaryl CoA reductase genotype, and mammographic density. Thirty women were enrolled, and 26 (86.7 %) completed the study. For the primary endpoint of changes in breast duct cytology sampled by random periareolar fine needle aspiration, most participants [57.7 %, 95 % confidence interval (CI) 38.9-74.5 %] showed no change after lovastatin; 19.2 % (CI 8.1-38.3 %) had a favorable change in cytology, 7.7 % (95 % CI 1.0-25.3 %) had an unfavorable change, and 15.4 % (95 % CI 5.5-34.2 %) had equivocal results due to acellular specimens, usually after lovastatin. No significant changes were observed in secondary biomarker endpoints. The study was generally well-tolerated: 4 (13.3 %) participants did not complete the study, and one (3.8 %) required a dose reduction. This trial was technically feasible, but demonstrated no significant biomarker modulation; contributing factors may include insufficient sample size, drug dose and/or duration. The results are inconclusive and do not exclude a favorable effect on breast cancer risk.

Spatial organization of dendritic cells within tumor draining lymph nodes impacts clinical outcome in breast cancer patients.

BACKGROUND: Dendritic cells (DCs) are important mediators of anti-tumor immune responses. We hypothesized that an in-depth analysis of dendritic cells and their spatial relationships to each other as well as to other immune cells within tumor draining lymph nodes (TDLNs) could provide a better understanding of immune function and dysregulation in cancer. METHODS: We analyzed immune cells within TDLNs from 59 breast cancer patients with at least 5 years of clinical follow-up using immunohistochemical staining with a novel quantitative image analysis system. We developed algorithms to analyze spatial distribution patterns of immune cells in cancer versus healthy intra-mammary lymph nodes (HLNs) to derive information about possible mechanisms underlying immune-dysregulation in breast cancer. We used the non-parametric Mann-Whitney test for inter-group comparisons, Wilcoxon Matched-Pairs Signed Ranks test for intra-group comparisons and log-rank (Mantel-Cox) test for Kaplan Maier analyses. RESULTS: Degree of clustering of DCs (in terms of spatial proximity of the cells to each other) was reduced in TDLNs compared to HLNs. While there were more numerous DC clusters in TDLNs compared to HLNs,DC clusters within TDLNs tended to have fewer member DCs and also consisted of fewer cells displaying the DC maturity marker CD83. The average number of T cells within a standardized radius of a clustered DC was increased compared to that of an unclustered DC, suggesting that DC clustering was associated with T cell interaction. Furthermore, the number of T cells within the radius of a clustered DC was reduced in tumor-positive TDLNs compared to HLNs. Importantly, clinical outcome analysis revealed that DC clustering in tumor-positive TDLNs correlated with the duration of disease-free survival in breast cancer patients. CONCLUSIONS: These findings are the first to describe the spatial organization of DCs within TDLNs and their association with survival outcome. In addition, we characterized specific changes in number, size, maturity, and T cell co-localization of such clusters. Strategies to enhance DC function in-vivo, including maturation and clustering, may provide additional tools for developing more efficacious DC cancer vaccines.

A comparative analysis of cutaneous marginal zone lymphoma and cutaneous chronic lymphocytic leukemia.

The morphologic distinction between cutaneous marginal zone lymphoma (CMZL) and secondary cutaneous involvement by B-cell chronic lymphocytic leukemia (B-CLL) can be difficult. Both entities can show very similar architectural patterns of involvement in the skin and not uncommonly, the skin can be the first site of presentation of B-CLL in the elderly. We reviewed biopsies of 13 patients with cutaneous B-CLL and 14 patients with CMZL to compare their histologic and immunohistochemical features. CMZL and cutaneous B-CLL both predominantly exhibited a nodular pattern of skin involvement (9 of 13 B-CLL, 9 of 14 CMZL) with a minority of cases demonstrating a diffuse pattern (4 of 13 B-CLL, 4 of 14 CMZL). Although reactive germinal centers (12 of 14 cases) and plasma cells (10 of 14 cases) were seen more often in CMZL, plasma cells were also observed in cases of B-CLL (4 of 13). The lesional cells of B-CLL expressed CD79, CD5, CD23, and CD43, although CMZL did not express CD5 or CD43. Although we noted light chain restriction in 13 of 14 cases of CMZL cases, we also observed light chain restriction in 4 of 13 cases of B-CLL. Our results indicate that CMZL and B-CLL can be morphologically similar and both may show light chain restriction. Complete immunophenotyping is necessary to ensure that all cases are correctly classified.

Cytotoxic T lymphocyte responses against melanocytes and melanoma.

BACKGROUND: Vitiligo is a common toxicity associated with immunotherapy for melanoma. Cytotoxic T lymphocytes (CTLs) against melanoma commonly target melanoma-associated antigens (MAAs) which are also expressed by melanocytes. To uncouple vitiligo from melanoma destruction, it is important to understand if CTLs can respond against melanoma and melanocytes at different levels. METHODS: To understand the dichotomous role of MAA-specific CTL, we characterized the functional reactivities of established CTL clones directed to MAAs against melanoma and melanocyte cell lines. RESULTS: CTL clones generated from melanoma patients were capable of eliciting MHC-restricted, MAA-specific lysis against melanocyte cell lines as well as melanoma cells. Among the tested HLA-A*0201-restricted CTL clones, melanocytes evoked equal to slightly higher degranulation and cytolytic responses as compared to melanoma cells. Moreover, MAA-specific T cells from vaccinated patients responded directly ex vivo to melanoma and melanocytes. Melanoma cells express slightly higher levels of MART-1 and gp100 than melanocytes as measured by quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) and immunohistochemistry. CONCLUSIONS: Our data suggest that CTLs respond to melanoma and melanocytes equally in vitro and directly ex vivo.

Adenomatoid tumors of the female and male genital tracts: a clinicopathological and immunohistochemical study of 44 cases.

Adenomatoid tumors of the female and male genital tracts are well characterized as mesothelial in origin, but a detailed histological and immunohistochemical analysis comparing both traditional and newer mesothelial markers across gender and site has not been formally conducted. A variety of morphologic features previously described as characteristic of adenomatoid tumors were evaluated in 44 adenomatoid tumors from the male and female genital tracts. Immunohistochemical analysis with pankeratin (AE1/CAM5.2), WT-1, calretinin, CK5/6, D2-40, and caldesmon was also performed. The extent and intensity of staining were scored semiquantitatively on one representative section per case and mean value for each parameter was calculated. All (n=44) the adenomatoid tumors from both the female and male genital tracts demonstrated a distinctive thread-like bridging strand pattern. Lymphoid aggregates were seen in all 12 adenomatoid tumors of male patients, but in only 4 of 32 (13%) tumors in female patients (P<0.0001). The remaining morphologic features were variably present with no clear sex predilection. Pankeratin, calretinin, and D2-40 reactivity were identified in all female (n=32) and male (n=12) genital tract adenomatoid tumors. Adenomatoid tumors expressed WT-1 in 11/12 (92%) male patients and in 31/32 (97%) female patients. In male patients, reactivity for CK5/6 and caldesmon was found in 1/12 (8%) and 0/12 (0%) adenomatoid tumors (respectively), whereas reactivity in female patients was found in 5/32 (16%) and 1/32 (3%); respectively. Female tumors differ from their male counterparts by the frequent absence of lymphoid aggregates and the presence of a circumscribed margin when occurring in the fallopian tube. Of the putative mesothelial markers evaluated, calretinin, D2-40, and WT-1 show a similar immunoprofile and have a higher sensitivity than CK5/6 and caldesmon in genital tract adenomatoid tumors. However, the presence of additional, often strong expression of WT-1 in normal tissues of the female genital tract limits the utility of WT-1 in this setting.

Immunohistochemical characterization of nasal-type extranodal NK/T-cell lymphoma using a tissue microarray: an analysis of 84 cases.

Nasal-type extranodal natural killer (NK)/T-cell lymphoma is an uncommon malignancy. By using a tissue microarray, we characterized 84 cases of extranodal NK/T-cell lymphoma with regard to expression of 18 immunohistochemical markers and the presence of Epstein-Barr virus (EBV) RNA. In our series, CD2 was positive in 69 (93%) of 74 cases, CD3 in 68 (84%) of 81, CD5 in 22 (27%) of 81, CD20 in 0 (0%) of 82, CD29 in 75 (91%) of 82, CD30 in 29 (35%) of 84, CD43 in 81 (96%) of 84, CD54 in 58 (72%) of 81, CD56 in 46 (58%) of 79, CD62L in 23 (28%) of 83, CD183 in 66 (80%) of 83, BCL2 in 33 (39%) of 84, cutaneous lymphocyte antigen in 21 (25%) of 84, granzyme B in 70 (83%) of 84, Ki-67 in 59 (71%) of 83, linker for activation of T cells in 60 (71%) of 84, perforin in 66 (86%) of 77, TIA1 in 76 (90%) of 84, and EBV in 73 (87%) of 84. Hierarchical cluster analysis separated primary cutaneous cases from cases manifesting in other sites based on lower expression of the cell adhesion molecule CD54.

Sentinel lymph node B cells can predict disease-free survival in breast cancer patients.

Tumor invasion into draining lymph nodes, especially sentinel lymph nodes (SLNs), is a key determinant of prognosis and treatment in breast cancer as part of the TNM staging system. Using multicolor histology and quantitative image analysis, we quantified immune cells within SLNs from a discovery cohort of 76 breast cancer patients. We found statistically more in situ CD3+ T cells in tumor negative vs. tumor positive nodes (mean of 8878 vs. 6704, respectively, p = 0.006), but no statistical difference in CD20+ B cells or CD1a+ dendritic cells. In univariate analysis, a reduced hazard was seen with a unit increase in log CD3 with HR 0.49 (95% CI 0.30-0.80) and log CD20 with HR 0.37 (95% CI 0.22-0.62). In multivariate analysis, log CD20 remained significant with HR 0.42 (95% CI 0.25-0.69). When restricted to SLN tumor negative patients, increased log CD20 was still associated with improved DFS (HR = 0.26, 95% CI 0.08-0.90). The CD20 results were validated in a separate cohort of 21 patients (n = 11 good outcome, n = 10 poor outcome) with SLN negative triple-negative breast cancer (TNBC) ("good" mean of 7011 vs. "poor" mean of 4656, p = 0.002). Our study demonstrates that analysis of immune cells within SLNs, regardless of tumor invasion status, may provide additional prognostic information, and highlights B cells within SLNs as important in preventing future recurrence.

Computed Tomography Features associated With the Eighth Edition TNM Stage Classification for Thymic Epithelial Tumors.

PURPOSE: The eighth edition of the TNM classification of malignant tumors for the first time includes an official staging system for thymic epithelial tumors (TETs) recognized by the American Joint Committee on Cancer (AJCC) and the Union for International Cancer Control (UICC). Staging is critical for the management of TETs, and determining stage accurately from imaging has the potential to improve clinical outcomes. We examine preoperative computed tomography (CT) characteristics of TETs associated with AJCC/UICC pathologic TNM stage. MATERIALS AND METHODS: In this retrospective study, patients were included if they met all the following criteria: (1) diagnosis of TET, (2) had primary curative intent surgery performed at Stanford University, and (3) had available preoperative CT imaging for review. Tumor pathology was staged according to the eighth edition TNM classification. Fifteen CT scan features were examined from each patient case according to the International Thymic Malignancy Interest Group standard report terms in a blinded manner. A Lasso-regularized multivariate model was used to produce a weighted scoring system predictive of pathologic TNM stage. RESULTS: Examining the 54 patients included, the following CT characteristics were associated with higher pathologic TNM stage when using the following scoring system: elevated hemidiaphragm (score of 6), vascular endoluminal invasion (score of 6), pleural nodule (score of 2), lobulated contour (score of 2), and heterogeneous internal density (score of 1). Area under the receiver operating characteristic curve was 0.76. CONCLUSIONS: TETs with clearly invasive or metastatic features seen on CT are associated with having higher AJCC/UICC pathologic TNM stage, as expected. However, features of lobulated contour and heterogeneous internal density are also associated with higher stage disease. These findings need to be validated in an independent cohort.

Rare Solitary Fibrous Tumor in the Pediatric Neck: A Case Report and Review of the Literature.

Solitary fibrous tumors (SFT) are a rare type of mesenchymal-derived tumor not commonly found in the pediatric population, especially in the head and neck. Tumors of this nature are most commonly seen in the adult population and are identified with unique immunohistochemical markers, specifically signal transducer and activator of transcription 6 (STAT6) and hematopoietic progenitor cell antigen (CD34). Including SFTs in the differential diagnosis while working up a mass can be difficult considering their relatively non-descript appearance on imaging and the low yield immunohistochemical staining that must be ordered to confirm diagnosis. The current literature identifies only a handful of cases of SFTs occurring in the pediatric population, with a majority arising from the pleura. We present the case of a 13-year-old male who underwent radical excision of a left occipital triangle neck mass after radiological and pathological workup failed to conclusively make a diagnosis. Postoperative pathologic analysis revealed it to be an SFT. Due to the exceptionally rare presentation of SFTs in pediatric patients, the aim of this case report is to discuss diagnostic measures, solitary fibrous tumor etiology, as well as a recent risk stratification system used for the evaluation of postoperative disease progression. Our hope is that clinicians will include SFTs in their differential diagnosis when working up a neck mass in the pediatric population.

Expression of CD163 (hemoglobin scavenger receptor) in normal tissues, lymphomas, carcinomas, and sarcomas is largely restricted to the monocyte/macrophage lineage.

CD163, a hemoglobin scavenger receptor, is expressed in monocytes and macrophages. We tested the expression of the CD163 protein in 1,105 human malignancies and normal tissues using tissue microarrays and conventional paraffin-embedded tissue sections. Besides staining nonneoplastic monocytes and histiocytes (tissue macrophages), membranous/cytoplasmic staining for CD163 was primarily limited to neoplasms with monocytic/histiocytic differentiation. CD163 reactivity was not observed in normal tissues, lymphomas, carcinomas, and in a majority of mesenchymal neoplasms, including follicular dendritic cell tumors (0 of 4), although it stained admixed histiocytes. Staining for CD163 was seen in Rosai-Dorfman disease (5 of 6), histiocytic sarcoma (3 of 4), littoral cell angioma (6 of 6), and Langerhans cell histiocytosis (3 of 5). A subset of atypical fibrous histiocytomas (9 of 16), benign fibrous histiocytomas (6 of 9), and atypical fibroxanthomas (1 of 3) also showed CD163 staining. Our studies also confirm earlier work showing that CD163 is expressed in acute myeloid leukemia with monocytic differentiation (AML, FAB subtype M5) (2 of 6), as well as a majority of giant cell tenosynovial tumors (7 of 8). Its limited range of expression and tissue specificity indicate that CD163 may have significant diagnostic utility in separating specific tumors with monocytic and histiocytic derivation from other entities in their differential diagnosis.

C-Kit (CD117) expression in AIDS-related, classic, and African endemic Kaposi sarcoma.

Kaposi sarcoma (KS) is a multicentric vascular neoplasm characterized histologically by the progressive proliferation of spindle-shaped tumor cells in all epidemiologic (AIDS-related, classic, endemic, and iatrogenic) forms. Human herpesvirus 8 (HHV8) is associated with all epidemiologic forms of KS and has been shown in vitro to induce the tyrosine receptor kinase c-kit in HHV8-infected cells. To date, c-kit immunoreactivity has not been systematically studied in KS lesions. Therefore, the aim of this study was to evaluate c-kit expression by immunohistochemistry in different proliferative stages and epidemiologic forms of KS. Archival cases of formalin-fixed, paraffin-embedded KS lesions, including 9 classic, 11 AIDS-related, and 15 African (endemic) forms at various histologic stages (5 patch, 8 plaque, 22 nodular), biopsied from different sites, were stained using immunohistochemistry with antibodies to HHV8 (LNA-1) and c-kit (CD117). C-kit immunoreactivity of lesional cells was demonstrated in 15 (43%) cases overall. A total of five (56%) classic, five (45%) AIDS-related, and five (33%) endemic KS cases were positive for c-kit. There was no difference in the intensity of c-kit staining between the different epidemiologic groups and histologic stages of KS. HHV8 (LNA-1) immunoreactivity was present in all (100%) classic, 10 (91%) AIDS-related, and 9 (60%) endemic cases. LNA-1 staining was demonstrated in 13 (93%) of the c-kit-positive and 15 (75%) of the c-kit-negative KS lesions. These findings indicate that c-kit expression in lesional cells can be detected by immunohistochemistry in different epidemiologic forms and histologic stages of KS. Furthermore, the expression of c-kit does not correspond with the presence of HHV8 (LNA-1) immunoreactivity in KS lesions.

Follicular dendritic cell immunohistochemical markers in angioimmunoblastic T-cell lymphoma.

Angioimmunoblastic T-cell lymphoma is characterized by a paracortical proliferation of medium to large neoplastic T cells, often with clear cytoplasm, in a background of arborizing high endothelial venules, many surrounded by follicular dendritic cells (FDCs). IHC staining may be applied to highlight these extrafollicular FDCs, traditionally using CD21, or CD23. Several alternative FDC markers have been described, including CNA.42, cystatin A/acid cysteine proteinase inhibitor (ACPI, involved in antigen presentation), and fascin (an actin binding protein). The authors stained a collection of 45 angioimmunoblastic T-cell lymphomas with CD21, CD23, CNA.42, cystatin A, and fascin for direct comparison of FDC staining characteristics in this setting. CD21 highlighted the expected dendritic network of cell processes, within residual follicles and outside of follicles, often adjacent to proliferating vessels. CD23 exhibited similar staining quality but was less sensitive than CD21. CNA.42 showed only diffuse weak labeling of FDCs. Cystatin A stained the cytoplasm of follicular dendritic cells within and outside of follicles; however, staining was often not sharply localized to dendritic cell processes, and scoring was further complicated by reactivity with other cell types in over half of the cases. Likewise, fascin stained a variety of cell types, including strong staining of interdigitating dendritic-like cells, moderate staining of endothelial cells, and only weak staining of follicular dendritic cells within and outside of follicles. Thus, CD21 remains the most reliable marker of follicular dendritic cells in angioimmunoblastic T-cell lymphoma.

The vascular marker CD31 also highlights histiocytes and histiocyte-like cells within cutaneous tumors.

OBJECTIVES: While useful in diagnosing angiosarcomas, CD31 can also highlight histiocytes within soft tissue tumors and lead to errors in diagnosis. We sought to determine how often CD31 highlights cutaneous histiocytomas and histiocytoma mimics. METHODS: We examined eight epithelioid cell histiocytomas (ECHs), 12 xanthogranulomas (XGs), nine cases of Langerhans cell histiocytosis (LCH), eight reticulohistiocytomas, 11 xanthomas, 29 atypical fibroxanthomas, nine granular cell tumors, four cases of angiolymphoid hyperplasia with eosinophilia, nine intradermal Spitz nevi, and nine angiosarcomas with antibodies directed against CD31, CD34, CD163, and factor VIII. RESULTS: CD31 marked cells in three of 12 XGs, four of nine cases of LCH, one of eight reticulohistiocytomas, one of 11 xanthomas, 10 of 29 atypical fibroxanthomas, four of four cases of angiolymphoid hyperplasia with eosinophilia, nine of nine angiosarcomas, zero of nine granular cell tumors, and zero of eight ECHs. CD34 and factor VIII were negative in all nonvascular cases. CONCLUSIONS: Our results indicate that CD31 can mark lesional cells and imitate vascular tumors in cutaneous histiocytomas and histiocytoma mimics, an error that can be avoided by using a panel of antibodies.

Diffuse high intensity PD-L1 staining in thymic epithelial tumors.

INTRODUCTION: Blockade of the immune checkpoint programmed death receptor ligand-1 (PD-L1)/PD-1 pathway has well-established clinical activity across many tumor types. PD-L1 protein expression by immunohistochemistry is emerging as a predictive biomarker of response to these therapies. Here, we examine PD-L1 expression in a thymic epithelial tumor (TET) tissue microarray (TMA). METHODS: The TMA contained 69 TETs and 17 thymic controls, with each case represented by triplicate cores. The TMA was stained with rabbit monoclonal antibody (clone 15; Sino Biological, Beijing, China) to human PD-L1. PD-L1 staining was scored based on intensity as follows: 0 = none, 1 = equivocal/uninterpretable, 2 = weak, and 3 = intermediate-strong. Those cases with all cores scoring three in the epithelial component were categorized as PD-L1 high and the remaining as PD-L1 low. RESULTS: PD-L1 high scores were more frequent in TETs than in controls (68.1% versus 17.6%; p = 0.0036). PD-L1 scores and histology were significantly correlated, with higher intensity staining in World Health Organization (WHO). B2/B3/C TETs. Only 14.8% of TETs had PD-L1 staining of associated lymphocytes. In an adjusted analysis (age/sex), PD-L1 high TETs had a significantly worse overall survival (hazard ratio: 5.40, 95% confidence interval: 1.13-25.89; p = 0.035) and a trend for worse event-free survival (hazard ratio: 2.94, 95% confidence interval: 0.94-9.24; p = 0.064). CONCLUSIONS: PD-L1 expression was present in all cases of TETs within the epithelial component but only in a minority in the lymphocytic component. TETs stained more intensely for PD-L1 than in controls, and PD-L1 high TETs were associated with more aggressive histology and worse prognosis. This study lends rationale to a clinical trial with anti-PD-1/PD-L1 therapy in this rare tumor type.

Human herpesvirus-8 latent nuclear antigen-1 expression in endemic Kaposi sarcoma: an immunohistochemical study of 16 cases.

Human herpesvirus-8 (HHV-8) infection is considered the initiating factor in all forms of Kaposi sarcoma (KS). Latent nuclear antigen (LNA-1) is constitutively expressed in all HHV-8-infected cells. An antibody to LNA-1 has recently become commercially available. The current study addresses the role of immunohistochemistry in the diagnosis of KS, particularly the endemic form. Seven recent cases of KS, 1 atypical vascular lesion in a patient subsequently diagnosed with KS, and 16 endemic cases collected in South Africa in the early 1960s were stained with an antibody to LNA-1. Nine benign vascular lesions and three angiosarcomas were also stained. All 7 recent cases expressed the antigen as did the atypical vascular lesion. Of particular interest was the finding that 10 of the 16 endemic cases were positive. None of the other vascular lesions showed staining. A subset of the endemic lesions was stained for CD31, an antigen universally expressed in KS. CD31 staining was reduced compared with a positive control suggesting that the current study may underestimate the sensitivity of LNA-1 immunohistochemistry in endemic KS because of poor antigen preservation in the archival tissue. Our results confirm the utility of LNA-1 immunohistochemistry as an aid in the diagnosis of KS.

Multiple myeloma recurrence with optic nerve infiltration diagnosed by vitrectomy, immunohistochemistry, and in situ hybridization.

PURPOSE: We present a case of multiple myeloma recurrence diagnosed by optic nerve infiltration. METHODS: Interventional case report with clinical, surgical, immunohistochemical, and fluorescence in situ hybridization correlation. RESULTS: A 51-year-old woman with a history of bilateral invasive ductal breast carcinoma and multiple myeloma, both in remission on maintenance bortezomib, was referred for sudden, painless loss of vision OS. Examination demonstrated anterior vitritis with severe optic disc elevation, with yellow-white thickening, peripapillary hemorrhages, and a retinal detachment inferiorly. Diagnostic vitrectomy showed CD138-positive and BRST2-negative cells. Fluorescence in situ hybridization was positive for del(13q) and p53 deletion and negative for CCND1/IGH. CONCLUSIONS: This is the first report of optic nerve infiltration of multiple myeloma as evidence of recurrence while on maintenance chemotherapy. We demonstrate that diagnostic vitrectomy and immunohistochemistry of vitreous fluid is feasible for the diagnosis of recurrent multiple myeloma.

Isolation and mutational analysis of circulating tumor cells from lung cancer patients with magnetic sifters and biochips.

Detection and characterization of circulating tumor cells (CTCs) may reveal insights into the diagnosis and treatment of malignant disease. Technologies for isolating CTCs developed thus far suffer from one or more limitations, such as low throughput, inability to release captured cells, and reliance on expensive instrumentation for enrichment or subsequent characterization. We report a continuing development of a magnetic separation device, the magnetic sifter, which is a miniature microfluidic chip with a dense array of magnetic pores. It offers high efficiency capture of tumor cells, labeled with magnetic nanoparticles, from whole blood with high throughput and efficient release of captured cells. For subsequent characterization of CTCs, an assay, using a protein chip with giant magnetoresistive nanosensors, has been implemented for mutational analysis of CTCs enriched with the magnetic sifter. The use of these magnetic technologies, which are separate devices, may lead the way to routine preparation and characterization of "liquid biopsies" from cancer patients.

Discordant immunophenotypic profiles of adhesion molecules and cytokines in acute myeloid leukemia involving bone marrow and skin.

We investigated the role of adhesion molecules in skin involvement by acute myeloid leukemia (AML) using immunohistochemical analysis. Ten paired cases of skin and bone marrow biopsy specimens from patients with myeloid leukemia cutis (MLC) and 15 bone marrow biopsy specimens from patients without MLC were studied with antibodies directed against CD29, CD34, CD54, CD62-L, CD183, and cutaneous lymphocyte antigen (CLA). CLA was expressed in all cases of leukemia whereas CD54 was negative within blasts. CD62-L was expressed in 4 of 10 specimens of marrow infiltrates with MLC and 6 of 10 specimens of matching skin infiltrates; in marrows without MLC, only 2 of 15 were positive. CD29 was expressed in 1 of 10 marrow infiltrate specimens with MLC and 4 of 10 matching skin infiltrate specimens; in marrows without MLC, only 1 of 15 were positive. CD183 was expressed in 1 of 10 marrow infiltrate specimens with MLC and 4 of 10 matching skin infiltrate specimens; in marrows without MLC, CD183 was negative. The gain of CD62-L, CD29, and CD183 expression in bone marrow and skin infiltrates in leukemia cutis, relative to bone marrow infiltrates of cases without MLC, suggests a role for these markers in AML homing to skin.