Denise L. Johnson

Characterization of circulating T cells specific for tumor- associated antigens in melanoma patients

We identified circulating CD8+ T-cell populations specific for the tumor-associated antigens (TAAs) MART-1 (27-35) or tyrosinase (368-376) in six of eleven patients with metastatic melanoma using peptide/HLA-A*0201 tetramers. These TAA-specific populations were of two phenotypically distinct types: one, typical for memory/effector T cells; the other, a previously undescribed phenotype expressing both naive and effector cell markers. This latter type represented more than 2% of the total CD8+ T cells in one patient, permitting detailed phenotypic and functional analysis. Although these cells have many of the hallmarks of effector T cells, they were functionally unresponsive, unable to directly lyse melanoma target cells or produce cytokines in response to mitogens. In contrast, CD8+ T cells from the same patient were able to lyse EBV-pulsed target cells and showed robust allogeneic responses. Thus, the clonally expanded TAA-specific population seems to have been selectively rendered anergic in vivo. Peptide stimulation of the TAA-specific T-cell populations in other patients failed to induce substantial upregulation of CD69 expression, indicating that these cells may also have functional defects, leading to blunted activation responses. These data demonstrate that systemic TAA-specific T-cell responses can develop de novo in cancer patients, but that antigen-specific unresponsiveness may explain why such cells are unable to control tumor growth.

Human melanoma-initiating cells express neural crest nerve growth factor receptor CD271

The question of whether tumorigenic cancer stem cells exist in human melanomas has arisen in the last few years. Here we show that in melanomas, tumour stem cells (MTSCs, for melanoma tumour stem cells) can be isolated prospectively as a highly enriched CD271+ MTSC population using a process that maximizes viable cell transplantation. The tumours sampled in this study were taken from a broad spectrum of sites and stages. High-viability cells isolated by fluorescence-activated cell sorting and re-suspended in a matrigel vehicle were implanted into T-, B- and natural-killer-deficient Rag2−/−γc−/− mice. The CD271+ subset of cells was the tumour-initiating population in 90% (nine out of ten) of melanomas tested. Transplantation of isolated CD271+ melanoma cells into engrafted human skin or bone in Rag2−/−γc−/− mice resulted in melanoma; however, melanoma did not develop after transplantation of isolated CD271− cells. We also show that in mice, tumours derived from transplanted human CD271+ melanoma cells were capable of metastatsis in vivo. CD271+ melanoma cells lacked expression of TYR, MART1 and MAGE in 86%, 69% and 68% of melanoma patients, respectively, which helps to explain why T-cell therapies directed at these antigens usually result in only temporary tumour shrinkage.

Prognostic value of tumor "budding" in patients with colorectal cancer.

From 1970 to 1985, 663 patients underwent curative resection of colon and rectal adenocarcinomas. All surgical specimens were examined for tumor “budding,” defined as small clusters of undifferentiated cancer cells ahead of the invasive front of the lesion. Patients were divided into two groups according to degree of budding: none or mild (BD-1) and moderate or severe (BD-2). BD-1 occurred in 493 patients (74.4 percent), and BD-2 was found in 170 patients (25.6 percent). More severe budding was associated with worse outcome: 71.1 percent of BD-2 patients had recurrence, compared with 20.0 percent of BD-1 patients (P <0.005). The five-year survival rate was worse in BD-2 than in BD-1 (22.2 percentvs.70.7 percent;P <0.001). The 10-year survival rate was also worse in BD-2 than in BD-1 (13.8 percentvs.50.6 percent;P <0.001). The incidence of BD-2 rose with the Dukes stage. However, the five-year survival rate of Dukes B patients with BD-2 lesions was worse than that of Dukes C patients with BD-1 cancers (29.1 percentvs.66.2 percent;P <0.001). Moreover, there was no difference in five-year survival among BD-1 patients with either Dukes B or C lesions (68.3 percentvs.66.2 percent). The presence of more severe budding appears to indicate a vigorous biologic activity of colorectal cancer. Thus, meticulous follow-up—and possibly adjuvant chemotherapy—may be beneficial for patients with marked budding, regardless of their Dukes stage.

New models and online calculator for predicting non-sentinel lymph node status in sentinel lymph node positive breast cancer patients

BackgroundCurrent practice is to perform a completion axillary lymph node dissection (ALND) for breast cancer patients with tumor-involved sentinel lymph nodes (SLNs), although fewer than half will have non-sentinel node (NSLN) metastasis. Our goal was to develop new models to quantify the risk of NSLN metastasis in SLN-positive patients and to compare predictive capabilities to another widely used model.MethodsWe constructed three models to predict NSLN status: recursive partitioning with receiver operating characteristic curves (RP-ROC), boosted Classification and Regression Trees (CART), and multivariate logistic regression (MLR) informed by CART. Data were compiled from a multicenter Northern California and Oregon database of 784 patients who prospectively underwent SLN biopsy and completion ALND. We compared the predictive abilities of our best model and the Memorial Sloan-Kettering Breast Cancer Nomogram (Nomogram) in our dataset and an independent dataset from Northwestern University.Results285 patients had positive SLNs, of which 213 had known angiolymphatic invasion status and 171 had complete pathologic data including hormone receptor status. 264 (93%) patients had limited SLN disease (micrometastasis, 70%, or isolated tumor cells, 23%). 101 (35%) of all SLN-positive patients had tumor-involved NSLNs. Three variables (tumor size, angiolymphatic invasion, and SLN metastasis size) predicted risk in all our models. RP-ROC and boosted CART stratified patients into four risk levels. MLR informed by CART was most accurate. Using two composite predictors calculated from three variables, MLR informed by CART was more accurate than the Nomogram computed using eight predictors. In our dataset, area under ROC curve (AUC) was 0.83/0.85 for MLR (n = 213/n = 171) and 0.77 for Nomogram (n = 171). When applied to an independent dataset (n = 77), AUC was 0.74 for our model and 0.62 for Nomogram. The composite predictors in our model were the product of angiolymphatic invasion and size of SLN metastasis, and the product of tumor size and square of SLN metastasis size.ConclusionWe present a new model developed from a community-based SLN database that uses only three rather than eight variables to achieve higher accuracy than the Nomogram for predicting NSLN status in two different datasets.

Profile of immune cells in axillary lymph nodes predicts disease-free survival in breast cancer.

Background While lymph node metastasis is among the strongest predictors of disease-free and overall survival for patients with breast cancer, the immunological nature of tumor-draining lymph nodes is often ignored, and may provide additional prognostic information on clinical outcome. Methods and Findings We performed immunohistochemical analysis of 47 sentinel and 104 axillary (nonsentinel) nodes from 77 breast cancer patients with 5 y of follow-up to determine if alterations in CD4, CD8, and CD1a cell populations predict nodal metastasis or disease-free survival. Sentinel and axillary node CD4 and CD8 T cells were decreased in breast cancer patients compared to control nodes. CD1a dendritic cells were also diminished in sentinel and tumor-involved axillary nodes, but increased in tumor-free axillary nodes. Axillary node, but not sentinel node, CD4 T cell and dendritic cell populations were highly correlated with disease-free survival, independent of axillary metastasis. Immune profiling of ALN from a test set of 48 patients, applying CD4 T cell and CD1a dendritic cell population thresholds of CD4 ≥ 7.0% and CD1a ≥ 0.6%, determined from analysis of a learning set of 29 patients, provided significant risk stratification into favorable and unfavorable prognostic groups superior to clinicopathologic characteristics including tumor size, extent or size of nodal metastasis (CD4, p < 0.001 and CD1a, p < 0.001). Moreover, axillary node CD4 T cell and CD1a dendritic cell populations allowed more significant stratification of disease-free survival of patients with T1 (primary tumor size 2 cm or less) and T2 (5 cm or larger) tumors than all other patient characteristics. Finally, sentinel node immune profiles correlated primarily with the presence of infiltrating tumor cells, while axillary node immune profiles appeared largely independent of nodal metastases, raising the possibility that, within axillary lymph nodes, immune profile changes and nodal metastases represent independent processes. Conclusion These findings demonstrate that the immune profile of tumor-draining lymph nodes is of novel biologic and clinical importance for patients with early stage breast cancer.

Impaired interferon signaling is a common immune defect in human cancer

Immune dysfunction develops in patients with many cancer types and may contribute to tumor progression and failure of immunotherapy. Mechanisms underlying cancer-associated immune dysfunction are not fully understood. Efficient IFN signaling is critical to lymphocyte function; animals rendered deficient in IFN signaling develop cancer at higher rates. We hypothesized that altered IFN signaling may be a key mechanism of immune dysfunction common to cancer. To address this, we assessed the functional responses to IFN in peripheral blood lymphocytes from patients with 3 major cancers: breast cancer, melanoma, and gastrointestinal cancer. Type-I IFN (IFN-α)-induced signaling was reduced in T cells and B cells from all 3 cancer-patient groups compared to healthy controls. Type-II IFN (IFN-γ)-induced signaling was reduced in B cells from all 3 cancer patient groups, but not in T cells or natural killer cells. Impaired-IFN signaling was equally evident in stage II, III, and IV breast cancer patients, and downstream functional defects in T cell activation were identified. Taken together, these findings indicate that defects in lymphocyte IFN signaling arise in patients with breast cancer, melanoma, and gastrointestinal cancer, and these defects may represent a common cancer-associated mechanism of immune dysfunction.

Positron emission tomography is superior to computed tomography for metastatic detection in melanoma patients.

BackgroundWhole-body positron emission tomography (PET) provides diagnostic information not currently available with traditional imaging and may improve the accuracy of staging melanoma patients.MethodsA retrospective cohort review was performed of 104 patients with primary or recurrent melanoma who underwent PET to determine sensitivity/specificity for metastatic detection compared with body computed tomography (CT). One hundred fifty-seven PET and 70 CT scans were analyzed, with a median patient follow-up of 24 months. Metastases were confirmed with positive histology (87.5%) or documented disease progression (12.5%). Fifty-three patients prospectively underwent consecutive studies within a mean 3-week interval for direct comparative analysis.ResultsPET demonstrated 84% sensitivity (95% confidence interval [CI], .78 to .89) and 97% specificity (95% CI, .91 to .99), whereas CT showed 58% sensitivity (95% CI, .49 to .66) and 70% specificity (95% CI, .51 to .84). Exclusion of areas not evaluated on CT (head, neck/supraclavicular, extremities) increased CT sensitivity to 69% (95% CI, .59 to .77). Sixty-six consecutive PET and CT scans were performed with 81% and 57% of metastases detected, respectively.ConclusionsPET is more sensitive and specific than CT for detection of melanoma metastasis and should be considered the primary staging study for recurrent disease. PET shows greater ability to detect soft tissue, small-bowel, and lymph node metastasis that do not meet criteria designated as abnormal by CT. PET is superior to CT even when sites not routinely evaluated by CT are excluded from comparative analysis.

Long-term results of curative resection of “minimally invasive” colorectal cancer

PURPOSE: The aim of this study was to determine the long-term outcome after curative resection of colorectal cancers that extend only into the submucosa (“minimally invasive”) and to evaluate potential histologic predictors of lymph node metastases. METHODS: Seventy-nine patients who underwent curative resection of minimally invasive colorectal cancer and were followed for at least five years were studied retrospectively. RESULTS: The series was comprised of 53 men and 26 women, with a mean age of 61 years. The lesion was in the colon in 47 patients and the rectosigmoid or rectum in 32 patients. Open surgery followed attempted endoscopic tumor removal in 25 patients. Lymph node metastasis, found in 11/79 patients (13.9 percent), was associated with worse outcome: 36.4 percent of node(+) patients developed recurrence,vs.only 5.9 percent of node(−) patients (P<0.005). The cumulative survival rate was also worse in node(+)vs.node(−) patients: 72.7 percentvs.91.1 percent at five years (P<0.05) and 45.5 percentvs.65.3 percent at ten years (P<0.05). Five histopathologic characteristics were identified as risk factors for lymph node metastasis: 1) small clusters of undifferentiated cancer cells ahead of the invasive front of the lesion (“tumor budding”); 2) a poorly demarcated invasive front; 3) moderately or poorly differentiated cancer cells in the invasive front; 4) extension of the tumor to the middle or deep submucosal layer; 5) cancer cells in lymphatics. Whereas patients with three or fewer risk factors had no nodal spread, the rate of lymph node involvement with four or more risk factors was 33.3 percent and 66.7 percent, respectively. CONCLUSIONS: Metastasis is not infrequent in “minimally invasive” colorectal cancer. Appropriate bowel resection with lymph node dissection is indicated if such a lesion exhibits more than three histologie risk factors for metastasis.

The effect of fluorine-18 fluorodeoxyglucose positron emission tomography on the management of cutaneous malignant melanoma.

PURPOSE To assess the effect of positron emission tomography (PET) with fluorine-18 fluorodeoxyglucose (FDG) on the treatment of patients with cutaneous malignant melanoma. MATERIALS AND METHODS We retrospectively reviewed the medical records of 38 patients with newly diagnosed (n = 25) and recurrent (n = 13) cutaneous melanoma who were referred for evaluation with FDG PET imaging at our institution. We compared the PET findings with computed tomography (CT), which was available in 21 (55%) patients, and tabulated the changes in the diagnostic evaluation and therapeutic management that were prompted by PET during a follow-up period of 10 to 36 months. RESULTS Compared with PET, the extent of disease was underestimated by CT in 5 (13%) patients. Planned surgical resection of metastases was canceled in two of these patients. In another patient, surveillance PET detected an unsuspected hypermetabolic abdominal mass that was noted on a subsequent CT to arise from the small bowel. The mass was found to be jejunal metastatic melanoma at the time of resection. Overall, PET influenced surgical management in 3 (8%) patients, but it did not affect the wait-and-watch strategy or decision to initiate immunotherapy in the others. CONCLUSIONS FDG PET contributes important information not provided by CT and has a substantial effect on the clinical management of patients with malignant melanoma.

The Role of a Positron- and High-energy Gamma Photon Probe in Intraoperative Localization of Recurrent Melanoma

Purpose: This preliminary study retrospectively evaluated the ability of intraoperative localization of recurrent melanoma using F-18 fluorodeoxyglucose (FDG) and a probe sensitive to both high-energy gamma rays and positrons to enable complete tumor resection and improved patient outcome. Materials and Methods: Three hours before surgery for resection of recurrent melanoma, 5 patients (mean age, 52 ± 22 years) with a history of local surgery, radiation therapy, and/or large habitus received 14.6 ± 3.2 mCi of F-18 FDG. Intraoperative tumor localization was performed with a radiation probe (PET-Probe; IntraMedical Imaging LLC, Los Angeles, CA). Intraoperative tumor tissue activities, background tissue activities, pathology results, and patient follow up (clinical/imaging) were recorded. Results: Eight of the 19 surgical specimens were identified by the probe as having increased FDG uptake when compared with the surrounding tissues before resection. All 8 specimens contained melanoma. Of the 11 specimens that were not identified using the probe, one contained melanoma, yielding a sensitivity of 89% (8 of 9) and a specificity of 100% (10 of 10). In 3 of the 5 cases, the probe allowed the identification of nonvisualized and nonpalpable tumor foci that were later confirmed pathologic. At an average follow up of 210 days (range, 30–515 days), 2 of 5 patients had no evidence of recurrent melanoma by clinical or radiographic evaluations. Conclusion: In the setting of recurrent melanoma, there appear to be potential benefits to intraoperative detection with FDG and a positron-detecting probe, particularly in cases with challenging or altered anatomy.