Association of occult metastases in sentinel lymph nodes and bone marrow with survival among women with early-stage invasive breast cancer.

CONTEXT: Immunochemical staining of sentinel lymph nodes (SLNs) and bone marrow identifies breast cancer metastases not seen with routine pathological or clinical examination. OBJECTIVE: To determine the association between survival and metastases detected by immunochemical staining of SLNs and bone marrow specimens from patients with early-stage breast cancer. DESIGN, SETTING, AND PATIENTS: From May 1999 to May 2003, 126 sites in the American College of Surgeons Oncology Group Z0010 trial enrolled women with clinical T1 to T2N0M0 invasive breast carcinoma in a prospective observational study. INTERVENTIONS: All 5210 patients underwent breast-conserving surgery and SLN dissection. Bone marrow aspiration at the time of operation was initially optional and subsequently mandatory (March 2001). Sentinel lymph node specimens (hematoxylin-eosin negative) and bone marrow specimens were sent to a central laboratory for immunochemical staining; treating clinicians were blinded to results. MAIN OUTCOME MEASURES: Overall survival (primary end point) and disease-free survival (a secondary end point). RESULTS: Of 5119 SLN specimens (98.3%), 3904 (76.3%) were tumor-negative by hematoxylin-eosin staining. Of 3326 SLN specimens examined by immunohistochemistry, 349 (10.5%) were positive for tumor. Of 3413 bone marrow specimens examined by immunocytochemistry, 104 (3.0%) were positive for tumors. At a median follow-up of 6.3 years (through April 2010), 435 patients had died and 376 had disease recurrence. Immunohistochemical evidence of SLN metastases was not significantly associated with overall survival (5-year rates: 95.7%; 95% confidence interval [CI], 95.0%-96.5% for immunohistochemical negative and 95.1%; 95% CI, 92.7%-97.5% for immunohistochemical positive disease; P = .64; unadjusted hazard ratio [HR], 0.90; 95% CI, 0.59-1.39; P = .64). Bone marrow metastases were associated with decreased overall survival (unadjusted HR for mortality, 1.94; 95% CI, 1.02-3.67; P = .04), but neither immunohistochemical evidence of tumor in SLNs (adjusted HR, 0.88; 95% CI, 0.45-1.71; P = .70) nor immunocytochemical evidence of tumor in bone marrow (adjusted HR, 1.83; 95% CI, 0.79-4.26; P = .15) was statistically significant on multivariable analysis. CONCLUSION: Among women receiving breast-conserving therapy and SLN dissection, immunohistochemical evidence of SLN metastasis was not associated with overall survival over a median of 6.3 years, whereas occult bone marrow metastasis, although rare, was associated with decreased survival. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00003854.

Ductal lavage in women from BRCA1/2 families: is there a future for ductal lavage in women at increased genetic risk of breast cancer?

PURPOSE: Ductal lavage has been used for risk stratification and biomarker development and to identify intermediate endpoints for risk-reducing intervention trials. Little is known about patient characteristics associated with obtaining nipple aspirate fluid (NAF) and adequate cell counts (> or =10 cells) in ductal lavage specimens from BRCA mutation carriers. METHODS: We evaluated patient characteristics associated with obtaining NAF and adequate cell counts in ductal lavage specimens from the largest cohort of women from BRCA families yet studied (BRCA1/2 = 146, mutation-negative = 23, untested = 2). Fisher's exact test was used to evaluate categorical variables; Wilcoxon nonparametric test was used to evaluate continuous variables associated with NAF or ductal lavage cell count adequacy. Logistic regression was used to identify independent correlates of NAF and ductal lavage cell count adequacy. RESULTS: From 171 women, 45 (26%) women had NAF and 70 (41%) women had ductal lavage samples with > or =10 cells. Postmenopausal women with intact ovaries compared with premenopausal women [odds ratio (OR), 4.8; P = 0.03] and women without a prior breast cancer history (OR, 5.2; P = 0.04) had an increased likelihood of yielding NAF. Having breast-fed (OR, 3.4; P = 0.001), the presence of NAF before ductal lavage (OR, 3.2; P = 0.003), and being premenopausal (OR, 3.0; P = 0.003) increased the likelihood of ductal lavage cell count adequacy. In known BRCA1/2 mutation carriers, only breast-feeding (OR, 2.5; P = 0.01) and the presence of NAF (OR, 3.0; P = 0.01) were independent correlates of ductal lavage cell count adequacy. CONCLUSIONS: Ductal lavage is unlikely to be useful in breast cancer screening among BRCA1/2 mutation carriers because the procedure fails to yield adequate specimens sufficient for reliable cytologic diagnosis or to support translational research activities.

Evaluation of prime/boost regimens using recombinant poxvirus/tyrosinase vaccines for the treatment of patients with metastatic melanoma.

PURPOSE: Two clinical trials were conducted to evaluate the clinical efficacy and immunologic impact of vaccination against the tyrosinase protein plus systemic interleukin 2 (IL-2) administration in patients with advanced metastatic melanoma. EXPERIMENTAL DESIGN: Full-length tyrosinase was employed as an immunogen to induce diverse immunologic responses against a commonly expressed melanoma antigen. Heterologous prime/boost vaccination with recombinant vaccinia and fowlpox vectors encoding tyrosinase was first explored in a randomized three-arm phase II trial, in which vaccines were administered alone or concurrently with low-dose or high-dose IL-2. In a subsequent single cohort phase II trial, all patients received the same vaccines and high-dose IL-2 sequentially rather than concurrently. RESULTS: Among a total of 64 patients treated on these trials, 8 objective partial responses (12.5%) were observed, all in patients receiving high-dose IL-2. Additional patients showed evidence of lesional regression (mixed tumor response) or overall regression that did not achieve partial response status (minor response). In vitro evidence of enhanced immunity against tyrosinase following protocol treatments was documented in 3 of 49 (6%) patients tested serologically, 3 of 23 (13%) patients tested for T-cell recognition of individual tyrosinase peptides, and 4 of 16 (25%) patients tested for T-cell recognition of full-length tyrosinase protein with real-time reverse transcription-PCR techniques. CONCLUSIONS: Whereas prime/boost immunization with recombinant vaccinia and fowlpox viruses enhanced antityrosinase immunity in some patients with metastatic melanoma, it was ineffective alone in mediating clinical benefit, and in combination with IL-2 did not mediate clinical benefit significantly different from that expected from treatment with IL-2 alone.

The effects of antibody clone and pretreatment method on the results of HER2 immunostaining in cytologic samples of metastatic breast cancer: A query and a review of the literature.

The standardization and use of heat-induced epitope retrieval (HIER) is particularly important with immunohistochemical markers that direct the course of cancer treatment, such as Herceptin therapy. Increasingly, many laboratories are performing immunohistochemical analysis using various antibodies and methodologies for HER2/neu. We attempted to determine the effects of antibody clone and pretreatment methods on the interpretation of HER-2/neu staining in cytologic samples. Cell block sections from 54 cases of metastatic breast cancer (24 FNAs, 30 effusions) were analyzed for HER2 expression using antibodies to CB-11, TAB250, and A0485. Antibodies were analyzed with and without HIER. One pathologist using the FDA-approved scoring system for the HercepTest reviewed all slides in a blinded fashion. Five of fifty-four cases (9%) using CB-11 showed a significant increase in HER2 immunoreactivity using HIER (i.e. from 0/1+ to 2-3+). However, in twenty-nine of fifty-four cases (54%), the cytoplasmic background was significantly higher after HIER. With the A0485 antibody, two of fifty four cases (4%) showed a significant increase in immunoreactivity using HIER, while seventeen of fifty-four cases (31%) exhibited only more pronounced cytoplasmic staining. HIER pretreatment did not increase HER2 staining in any TAB250 stained sample, rather four of fifty-four cases (7%) showed a significant decrease in staining with HIER. We conclude that HIER may enhance membrane staining with the CB-11 and A0485 antibodies, but also increases cytoplasmic background. Loss of antigenicity is seen when HIER is used with TAB250.

Diagnostic effects of prolonged storage on fresh effusion samples.

The effects on morphology and diagnostic interpretation of delayed processing of refrigerated effusion samples have not been well documented. The potential for cellular degeneration has led many laboratories to reflexively fix samples rather than submit fresh/refrigerated samples for cytologic examination. We sought to determine if effusion specimens are suitable for morphologic, immunocytochemical, and DNA-based molecular studies after prolonged periods of refrigerated storage time. Ten fresh effusion specimens were refrigerated at 4 degrees C; aliquots were processed at specific points in time (days 0, 3, 5, 7, 10, 14). Specimens evaluated included four pleural (3 benign, 1 breast adenocarcinoma) and six peritoneal (2 ovarian adenocarcinomas, 1 malignant melanoma, 2 mesotheliomas, 1 atypical mesothelial) effusions. The morphology of the cytologic preparations from the 10 effusions was preserved and interpretable after 14 days of storage at 4 degrees C. The immunocytochemical profile of the samples (AE1/AE3, EMA, calretinin, and LCA) was consistent from day 0 to day 14. Amplifiable DNA was present in all samples tested on day 14. We conclude that cytopathologic interpretation of effusion samples remains reliable with refrigeration at 4 degrees C even if processing is delayed.

Adoptive cell transfer therapy following non-myeloablative but lymphodepleting chemotherapy for the treatment of patients with refractory metastatic melanoma.

PURPOSE: We investigated the combination of lymphodepleting chemotherapy followed by the adoptive transfer of autologous tumor reactive lymphocytes for the treatment of patients with refractory metastatic melanoma. PATIENTS AND METHODS: Thirty-five patients with metastatic melanoma, all but one with disease refractory to treatment with high-dose interleukin (IL) -2 and many with progressive disease after chemotherapy, underwent lymphodepleting conditioning with two days of cyclophosphamide (60 mg/kg) followed by five days of fludarabine (25 mg/m(2)). On the day following the final dose of fludarabine, all patients received cell infusion with autologous tumor-reactive, rapidly expanded tumor infiltrating lymphocyte cultures and high-dose IL-2 therapy. RESULTS: Eighteen (51%) of 35 treated patients experienced objective clinical responses including three ongoing complete responses and 15 partial responses with a mean duration of 11.5 +/- 2.2 months. Sites of regression included metastases to lung, liver, lymph nodes, brain, and cutaneous and subcutaneous tissues. Toxicities of treatment included the expected hematologic toxicities of chemotherapy including neutropenia, thrombocytopenia, and lymphopenia, the transient toxicities of high-dose IL-2 therapy, two patients who developed Pneumocystis pneumonia and one patient who developed an Epstein-Barr virus-related lymphoproliferation. CONCLUSION: Lymphodepleting chemotherapy followed by the transfer of highly avid antitumor lymphocytes can mediate significant tumor regression in heavily pretreated patients with IL-2 refractory metastatic melanoma.

Localization of the ABCG2 mitoxantrone resistance-associated protein in normal tissues.

Reduced drug accumulation due to overexpression of individual members of the ATP binding cassette (ABC) superfamily of membrane transporters has been investigated as a cause of multidrug resistance and treatment failure in oncology. This study was designed to develop an immunohistochemical assay to determine the expression and localization of the 72kDa ABC half-transporter ABCG2 in normal tissues. Formalin-fixed, paraffin embedded archival tissue from 31 distinct normal tissues with an average of eight separate tissue samples of each were immunostained with rabbit-anti-ABCG2 antibody 405 using a modified avidin-biotin procedure. As a negative control, each sample was also stained with antibody pre-adsorbed with peptide to assess background staining. As a means of verification, selected tissues were also stained with the commercially available monoclonal antibody 5D3. ABCG2 positivity was consistently found in alveolar pneumocytes, sebaceous glands, transitional epithelium of bladder, interstitial cells of testes, prostate epithelium, endocervical cells of uterus, squamous epithelium of cervix, small and large intestinal mucosa/epithelial cells, islet and acinar cells of pancreas, zona reticularis layer of adrenal gland, kidney cortical tubules and hepatocytes. Placental syncytiotrophoblasts showed both cytoplasmic and surface staining. Our results support a hypothesis concluding that ABCG2 plays a role in the protection of organs from cytotoxins. However, many of the cell types expressing ABCG2 have a significant secretory function. These data suggest a dual function for ABCG2 in some tissues: the excretion of toxins and xenobiotics including anti-cancer agents and a potential, as-yet undefined role in the secretion of endogenous substrates.

Granulocytic invasion of the central nervous system after hematopoietic stem cell transplantation for systemic lupus erythematosus.

We report on the likely mechanism of an exacerbation of neurological symptoms developed during immune reconstitution after autologous non-myeloablative hematopoietic stem cell transplantation in a 33-year-old man with systemic lupus erythematosus- associated recurrent transverse myelitis. Cerebrospinal fluid examination revealed prominent neutrophilic pleocytosis and no evidence of infection or of reactivation of lupus. Following a course of corticosteroid treatment the exacerbation resolved completely and the patient's neurological function continued to improve, resulting in net gain above pre-treatment for over 1 year follow-up without maintenance immunosuppression. Granulocytic invasion of the central nervous system represents a novel and possibly preventable cause of neurological complications during haematologic reconstitution.

Cytological identification of metastatic epithelial nephroblastoma in pleural fluid: report of a case and review of literature.

Nephroblastoma (Wilms' tumor) is the most common childhood renal tumor and usually presents with a histology and cytology consisting of blastemal, epithelial, and stromal cells. Effusions are not uncommon and may suggest an unfavorable prognosis when containing anaplastic tumor cells. In the present case, we report the cytological appearance of a Wilms' tumor metastatic to the pleura. The effusion consisted primarily of tumor cells demonstrating epithelial differentiation. The tumor cells mostly presented as three-dimensional aggregates in an inflammatory background. Many cystic and tubular structures were identified. The tumor cells demonstrated strong CD56 and WT1 immunoreactivity. The histology of a subsequent surgical specimen reflected the features seen in cytology.

Fine-needle aspiration of a Xp11.2 translocation/TFE3 fusion renal cell carcinoma metastatic to the lung: report of a case and review of the literature.

A 57-yr-old woman presented to the National Cancer Institute (NCI) with a history of nephrectomy for a clear cell renal cell carcinoma (RCC), Fuhrman grade 3 of 4 diagnosed 1 yr prior to admission to the NCI. A CT scan done upon admission revealed multiple bilateral lung masses. A CT-guided fine-needle aspiration (FNA) of one of the lung masses revealed a cellular specimen composed primarily of follicular structures surrounding dense hyalinized central cores. The cells in the follicular structures displayed bland nuclei and had granular to vacuolated cytoplasm. Papillary structures were also appreciated. Immunocytochemical studies showed tumor cells that were strongly vimentin and TFE3 positive. Focal staining for AE1/AE3 and CD10 was observed, as was negative staining for EMA. A surgical biopsy specimen reflected the FNA findings and demonstrated a similar immunoprofile. These findings correspond to the recently described Xp11.2 translocation/TFE3 fusion renal cell carcinoma. To our knowledge, this is the first report describing the cytologic features of an Xp11.2 translocation/TFE3 fusion RCC.

Use of peptide antibodies to probe for the mitoxantrone resistance-associated protein MXR/BCRP/ABCP/ABCG2.

Recent studies have characterized the ABC half-transporter associated with mitoxantrone resistance in human cancer cell lines. Encoded by the ABCG2 gene, overexpression confers resistance to camptothecins, as well as to mitoxantrone. We developed four polyclonal antibodies against peptides corresponding to four different epitopes on the mitoxantrone resistance-associated protein, ABCG2. Three epitopes localized on the cytoplasmic region of ABCG2 gave rise to high-affinity antibodies, which were demonstrated to be specific for ABCG2. Western blot analysis of cells with high levels of ABCG2 showed a single major band of the expected 72-kDa molecular size of ABCG2 under denaturing conditions. Immunoblot analysis performed under non-reducing conditions and after treatment with cross-linking reagents demonstrated a molecular weight shift from 72 kDa to several bands of 180 kDa and higher molecular weight, suggesting detection of dimerization products of ABCG2. Evidence of N-linked glycosylation was also obtained using tunicamycin and N-glycosidase F. Finally, both by light, fluorescence and electron microscopic immunohistochemical staining, we demonstrate cytoplasmic and predominantly plasma membrane localization of ABCG2 in cell lines with high levels of expression. Plasma membrane staining was observed on the surface of the chorionic villi in placenta. These results support the hypothesis that ABCG2 is an ABC half-transporter that forms dimers in the plasma membrane, functioning as an ATP-dependent outward pump for substrate transport.

Breast fine needle aspiration biopsy: prevailing recommendations and contemporary practices.

In 1996, a National Cancer Institute conference was held in Bethesda,Maryland to define parameters for the practice of breast fine needle aspiration (BFNA). Representatives of the American Society of Cytopathology, Papanicolaou Society of Cytopathology, American College of Radiology, American College of Obstetricians & Gynecologists, Society of Surgical Oncology, American Academy of Family Physicians, College of American Pathologists, National Consortium of Breast Centers, International Academy of Cytology, American Society of Clinical Pathologists, American Cancer Society, American College of Surgeons, and American Society for Cytotechnology developed and reviewed recommendations. These guidelines were referred to as "The Uniform Approach to Breast Fine Needle Aspiration Biopsy." This article reviews these recommendations and the contemporary evolution of the practice of BFNA since their original publication.

Antibody αPEP13h reacts with lymphangioleiomyomatosis cells in lung nodules.

BACKGROUND: Lymphangioleiomyomatosis (LAM) is characterized by the proliferation in the lung, axial lymphatics (eg, lymphangioleiomyomas), and kidney (eg, angiomyolipomas) of abnormal smooth muscle-like LAM cells, which express melanoma antigens such as Pmel17/gp100 and have dysfunctional tumor suppressor tuberous sclerosis complex (TSC) genes TSC2 or TSC1. Histopathologic diagnosis of LAM in lung specimens is based on identification of the Pmel17 protein with the monoclonal antibody HMB-45. METHODS: We compared the sensitivity of HMB-45 to that of antipeptide antibody αPEP13h, which reacts with a C-terminal peptide of Pmel17. LAM lung nodules were laser-capture microdissected to identify proteins by Western blotting. RESULTS: HMB-45 recognized approximately 25% of LAM cells within the LAM lung nodules, whereas αPEP13h identified > 82% of LAM cells within these structures in approximately 90% of patients. Whereas HMB-45 reacted with epithelioid but not with spindle-shaped LAM cells, αPEP13h identified both spindle-shaped and epithelioid LAM cells, providing greater sensitivity for detection of all types of LAM cells. HMB-45 recognized Pmel17 in premelanosomal organelles; αPEP13h recognized proteins in the cytoplasm as well as in premelanosomal organelles. Both antibodies recognized a Pmel17 variant of approximately 50 kDa. CONCLUSIONS: Based on its sensitivity and specificity, αPEP13h may be useful in the diagnosis of LAM and more sensitive than HMB-45.

Indinavir-associated interstitial nephritis and urothelial inflammation: clinical and cytologic findings.

The objective of the present study was to characterize the genitourinary syndromes that accompany indinavir-associated pyuria. Of 23 indinavir-treated patients with persistent pyuria, 4 had isolated interstitial nephritis, 10 had both interstitial nephritis and urothelial inflammation, 7 had isolated urothelial inflammation, and 2 had pyuria with nonspecific urinary tract inflammation. A total of 21 patients had multinucleated histiocytes identified by cytologic testing of urine specimens. Urine abnormalities resolved in all 20 patients who stopped receiving indinavir therapy. Pyuria continued in the 3 patients who continued receiving indinavir. Six patients had elevated serum creatinine levels, which returned to baseline levels when indinavir was discontinued. In conclusion, indinavir-associated pyuria was frequently associated with evidence of interstitial nephritis and/or urothelial inflammation, multinucleated histiocytes were commonly present in urine specimens, and cessation of indinavir therapy was associated with prompt resolution of urine abnormalities.

Urinary cytology associated with human polyomavirus and indinavir therapy in HIV-infected patients.

We retrospectively analyzed 155 urine cytology samples (78 from patients treated with indinavir; 77, no indinavir) from 90 HIV+ patients to evaluate possible association between human polyomavirus and hematuria and to describe indinavir-associated urinary cytologic findings. The CD4 count also was recorded. Variables studied included the presence of cellular viral changes consistent with polyomavirus infection (PVCs), microscopic hematuria, multinucleated cells, indinavir crystals, neutrophils, and eosinophils. Twenty-two samples (15.8%) from patients with CD4 counts of more than 200/microL (>200 x 10(6)/L) showed PVCs. Multinucleated cells, of presumed histiocytic origin based on morphologic features and selective immunocytochemical findings, were present in a higher percentage of samples from indinavir-treated patients. Neutrophils were present in a higher percentage of indinavir-treated patients. Indinavir crystals were identified in 9 samples (12%) from patients receiving indinavir The lower percentage of PVCs in HIV+ patients with high CD4 counts likely represents an indirect antipolyomavirus indinavir effect by boosting immunity. Multinucleated cells (presumably histiocytic) and acute inflammation are associated with indinavir therapy. Indinavir crystals have a characteristic fan or circular lamellate appearance. Because indinavir crystals may be associated with genitourinary disease, recognizing and reporting them is clinically relevant in HIV+ patients.

Proteomic evaluation of archival cytologic material using SELDI affinity mass spectrometry: potential for diagnostic applications.

Proteomic studies of cells via surface-enhanced laser desorption/ionization spectrometry (SELDI) analysis have enabled rapid, reproducible protein profiling directly from crude samples. We applied this technique to archival cytology material to determine whether distinct, reproducible protein fingerprints could be identifiedfor potential diagnostic purposes in blinded specimens. Rapid Romanowsky-stained cytocentrifuged specimens from fine-needle aspirates of metastatic malignant melanoma (with both known cutaneous primary and unknown primary sites), clear cell sarcoma, and renal cell carcinoma and reactive effusions were examined using the SELDI technology. A unique characteristic fingerprint was identified for each disease entity. Fifteen "blinded" unknown samples then were analyzed. When the protein profilefingerprints were plotted against the known fingerprints for the aforementioned diagnoses, the appropriate match or diagnosis was obtained in 13 (87%) of 15 cases. These preliminary findings suggest a substantial potential for SELDI applications to specific pathologic diagnoses.

Comparison of three commonly used cytologic preparations in effusion immunocytochemistry.

Discrepant results in effusion immunocytochemistry are often the result of specimen processing. Smears, cytospins, cell blocks, and monolayer preparations have all been used in various published studies; thus, there is no consistency in the immunostaining process for cytology to compare with the surgical pathology "gold standard" results. We sought to evaluate optimal specimen preparation for the immunostaining of effusion samples. Fourteen reactive and 15 malignant effusion samples (various epithelial/mesothelial neoplasms) were each prepared in three forms: air-dried cytospins (postfixed in ethanol), formalin-fixed, paraffin-embedded cell blocks, and liquid-based thin-layer (ThinPrep, CYTYC, Boxborough, MA) processing. All slides were immunostained with antibodies commonly used in effusion cytology: HBME-1, calretinin, E-cadherin, BerEP4, B72.3, LeuM1, and CA19-9. Cytospin and ThinPrep samples performed in a similar manner: high background staining was encountered in 66% of cases, most evident in three-dimensional clusters of cells. In addition, membrane staining patterns were difficult to interpret. Cell blocks provided the best milieu for morphologic interpretation, with less background staining (only 17% of cases) and results that most closely approximated those reported in the surgical pathology literature. The cost per test for cell block immunocytochemistry was also the most economical for our laboratory.

Melanoma metastatic to the breast: utility of fine needle aspiration and immunohistochemistry.

OBJECTIVE: To describe the morphologic spectrum of metastatic malignant melanoma (MM) cells involving the breast and to explore the diagnostic utility of HMB45, Mart-1, Melan-A and T311 (antityrosinase) antibodies in fine needle aspiration material of MM metastatic to the breast. STUDY DESIGN: Cytologic material from 21 cases (18 women) was reviewed for cytomorphology (epithelioid, spindled, mixed) and immunocytochemical staining attributes for Mart-1, HMB45, T311, Melan-A and cytokeratin based on tissue availability. RESULTS: Seventeen cases (81%) demonstrated epithelioid cell morphology, with 14% exhibiting mixed and 5% spindled morphologies. All 21 cases (100%) were immunoreactive with Mart-1 antibody, with 81% (17/21) immunoreactive for HMB45. In 38% of cases there was a similar percentage of cells immunoreactive for Mart-1 and HMB45, while 48% showed a higher percentage of cells immunoreactive for MART-1 than HMB45. Immunoreactivity with T311 was seen in 8 of 11 cases tested (73%). All six cases tested (100%) were immunoreactive with Melan-A. Staining for cytokeratin was negative in all eight cases tested. CONCLUSION: Because the majority of MM metastatic to the breast shows epithelioid cell morphology, it may mimic primary breast carcinoma. Mart-1 should be part of the immunocytochemical panel utilized to confirm the diagnosis of MM metastatic to the breast.

Human melanoma metastases demonstrate nonstochastic site-specific antigen heterogeneity that correlates with T-cell infiltration.

PURPOSE: Metastasis heterogeneity presents a significant obstacle to the development of targeted cancer therapeutics. In this study, we sought to establish from a large series of human melanoma metastases whether there exists a determined pattern in tumor cellular heterogeneity that may guide the development of future targeted immunotherapies. EXPERIMENTAL DESIGN: From a cohort of 1,514 patients with metastatic melanoma, biopsies were procured over a 17-year period from 3,086 metastatic tumors involving various anatomic sites. To allow specific tumor cell profiling, we used established immunohistochemical methods to perform semiquantitative assessment for a panel of prototypic melanocyte differentiation antigens (MDA), including gp100, MART-1, and tyrosinase. To gain insight into the endogenous host immune response against these tumors, we further characterized tumor cell expression of MHC I and MHC II and, also, the concomitant CD4(+) and CD8(+) T-cell infiltrate. RESULTS: Tumor cell profiling for MDA expression demonstrated an anatomic site-specific pattern of antigen expression that was highest in brain, intermediate in soft tissues/lymph nodes, and lowest in visceral metastases. Hierarchical clustering analysis supported that melanoma metastases have a phylogenetically determined, rather than a stochastic, pattern of antigen expression that varies by anatomic site. Furthermore, tyrosinase expression was more frequently lost in metastatic sites outside of the brain and was uniquely correlated with both endogenous CD8(+) and CD4(+) T-cell infiltrates. CONCLUSION: Site-specific antigen heterogeneity represents a novel attribute for human melanoma metastases that should be considered in future therapy development and when assessing the responsiveness to antigen-specific immunotherapies.