Endocrinologist (diabetes, hormones), Surgical Specialist
27 years of experience
Video profile
Accepting new patients
University City
Hospital of the University of PA
3400 Spruce St
Fl 4
Philadelphia, PA 19104
215-662-2050
Locations and availability (6)

Education ?

Medical School Score
UMDNJ Robert Wood Johnson (1983)
  • Currently 2 of 4 apples

Awards & Distinctions ?

Awards  
One of America's Leading Experts on:
Breast Cancer (Breast Neoplasm)
Melanoma (Skin cancer)
Castle Connolly America's Top Doctors® (2009, 2013 - 2014)
Castle Connolly America's Top Doctors® for Cancer (2009, 2014)
Castle Connolly's Top Doctors™ (2012 - 2013)
Patients' Choice Award (2010 - 2013)
Appointments
University of Pennsylvania
Associate Professor of Surgery Co-Director, Rena Rowan Breast Center
Associations
American Board of Surgery
American College of Surgeons
American Society of Clinical Oncology

Affiliations ?

Dr. Czerniecki is affiliated with 7 hospitals.

Hospital Affilations

Score

Rankings

  • University of PA Medical Center/Presbyterian
    51 N 39th St, Philadelphia, PA 19104
    • Currently 3 of 4 crosses
    Top 50%
  • Hospital of the University of PA
    Medical Oncology
    3400 Spruce St, Philadelphia, PA 19104
    • Currently 3 of 4 crosses
    Top 50%
  • Pennsylvania Hospital University PA Health System
    Medical Oncology
    800 Spruce St, Philadelphia, PA 19107
    • Currently 3 of 4 crosses
    Top 50%
  • Graduate Hospital
    1800 Lombard St, Philadelphia, PA 19146
    • Currently 1 of 4 crosses
  • Clinical Practices of the University of Pennsylvania
  • University of Penn Med Center-Presb Med Group
  • University Of Pennsylvania Medical Center
  • Publications & Research

    Dr. Czerniecki has contributed to 63 publications.
    Title The Significance of Her-2/neu Receptor Positivity and Immunophenotype in Ductal Carcinoma in Situ with Early Invasive Disease.
    Date October 2011
    Journal Journal of Surgical Oncology
    Excerpt

    Biologic markers that predict development of invasive breast cancer (IBC) in patients diagnosed with ductal carcinoma in situ (DCIS) are needed to improve personalized therapy. In this study, we examined the incidence of early IBC in DCIS subgroups defined by immunophenotype.

    Title Computer Aided Automatic Detection of Malignant Lesions in Diffuse Optical Mammography.
    Date June 2010
    Journal Medical Physics
    Excerpt

    Computer aided detection (CAD) data analysis procedures are introduced and applied to derive composite diffuse optical tomography (DOT) signatures of malignancy in human breast tissue. In contrast to previous optical mammography analysis schemes, the new statistical approach utilizes optical property distributions across multiple subjects and across the many voxels of each subject. The methodology is tested in a population of 35 biopsy-confirmed malignant lesions.

    Title Asymptomatic Changes in Cardiac Function Can Occur in Ductal Carcinoma-in-situ Patients Following Treatment with Her-2/neu-pulsed Dendritic Cell Vaccines.
    Date October 2009
    Journal American Journal of Surgery
    Excerpt

    Targeting HER-2/neu with Trastuzumab has been associated with development of cardiac toxicity.

    Title Differentiation of Benign and Malignant Breast Tumors by In-vivo Three-dimensional Parallel-plate Diffuse Optical Tomography.
    Date July 2009
    Journal Journal of Biomedical Optics
    Excerpt

    We have developed a novel parallel-plate diffuse optical tomography (DOT) system for three-dimensional in vivo imaging of human breast tumor based on large optical data sets. Images of oxy-, deoxy-, and total hemoglobin concentration as well as blood oxygen saturation and tissue scattering were reconstructed. Tumor margins were derived using the optical data with guidance from radiology reports and magnetic resonance imaging. Tumor-to-normal ratios of these endogenous physiological parameters and an optical index were computed for 51 biopsy-proven lesions from 47 subjects. Malignant cancers (N=41) showed statistically significant higher total hemoglobin, oxy-hemoglobin concentration, and scattering compared to normal tissue. Furthermore, malignant lesions exhibited a twofold average increase in optical index. The influence of core biopsy on DOT results was also explored; the difference between the malignant group measured before core biopsy and the group measured more than 1 week after core biopsy was not significant. Benign tumors (N=10) did not exhibit statistical significance in the tumor-to-normal ratios of any parameter. Optical index and tumor-to-normal ratios of total hemoglobin, oxy-hemoglobin concentration, and scattering exhibited high area under the receiver operating characteristic curve values from 0.90 to 0.99, suggesting good discriminatory power. The data demonstrate that benign and malignant lesions can be distinguished by quantitative three-dimensional DOT.

    Title Her-2/neu Overexpression As a Predictor for the Transition from in Situ to Invasive Breast Cancer.
    Date June 2009
    Journal Cancer Epidemiology, Biomarkers & Prevention : a Publication of the American Association for Cancer Research, Cosponsored by the American Society of Preventive Oncology
    Excerpt

    The clinical implications of HER-2/neu (HER2) expression in ductal carcinoma in situ (DCIS) lesions have yet to be clearly elucidated; this despite the more frequent expression of HER2 in high-grade DCIS lesions compared with invasive cancers. We hypothesized that HER2 overexpression in DCIS is associated with more rapid progression to invasive disease. Immunohistochemical staining for estrogen receptor, progesterone receptor, and HER2 was done on DCIS specimens. Univariate analysis and a multivariate logistic regression were done to determine whether estrogen receptor, progesterone receptor, or HER2 status, comedo necrosis, nuclear grade, lesion size, or patient age predicted the presence of associated invasive disease in patients with DCIS. Invasive foci were found in association with HER2 overexpressing DCIS at a higher frequency than with DCIS that did not overexpress HER2. Although high nuclear grade, large lesion size, and HER2 overexpression were all associated with the presence of invasive disease on univariate analysis, HER2 was the only significant predictor for the presence of invasive disease after multivariate adjustment (odds ratio, 6.4; P = 0.01). These data indicate that HER2 overexpression in DCIS lesions predicts the presence of invasive foci in patients with DCIS and suggest that targeting of HER2 in an early disease setting may forestall or prevent disease progression.

    Title Ductal Carcinoma in Situ: State of the Science and Roadmap to Advance the Field.
    Date January 2009
    Journal Journal of Clinical Oncology : Official Journal of the American Society of Clinical Oncology
    Excerpt

    Ductal carcinoma in situ (DCIS) is the fourth leading cancer for women in the United States. Understanding of the biology and clinical behavior of DCIS is imperfect. This article highlights the current knowledge base and the scientific roadmap needed to advance the field.

    Title Stat3- and Stat5-dependent Pathways Competitively Regulate the Pan-differentiation of Cd34pos Cells into Tumor-competent Dendritic Cells.
    Date September 2008
    Journal Blood
    Excerpt

    The clinical outcomes of dendritic cell (DC)-based immunotherapy remain disappointing, with DCs often displaying a tenuous capacity to complete maturation and DC1 polarization in the tumor host. Surprisingly, we observed that the capacity for successful DC1 polarization, including robust IL12p70 production, could be regulated by STAT-dependent events even prior to DC differentiation. Exposure of CD34(pos) cells to single-agent granulocyte-macrophage colony-stimulating factor (GMCSF) induced multilineage, STAT5-dependent differentiation, including DCs that failed to mature in the absence of further exogenous signals. In contrast, Flt3L induced nearly global differentiation of CD34(pos) cells into spontaneously maturing DCs. IL-6 synergized with Flt3L to produce explosive, STAT3-dependent proliferation of phenotypically undifferentiated cells that nevertheless functioned as committed DC1 precursors. Such precursors not only resisted many tumor-associated immunosuppressants, but also responded to tumor contact or TGFbeta with facilitated DC maturation and IL12p70 production, and displayed a superior capacity to reverse tumor-induced T-cell tolerance. GMCSF preempted Flt3L or Flt3L plus IL-6 licensing by blocking STAT3 activation and promoting STAT5-dependent differentiation. Paradoxically, following overt DC differentiation, STAT5 enhanced whereas STAT3 inhibited DC1 polarization. Therefore, nonoverlapping, sequential activation of STAT3 and STAT5, achievable by sequenced exposure to Flt3L plus IL-6, then GMCSF, selects for multilog expansion, programming, and DC1 polarization of tumor-competent DCs from CD34(pos) cells.

    Title Method of Primary Tumor Detection As a Risk Factor for Local and Distant Recurrence After Breast-conservation Treatment for Early-stage Breast Cancer.
    Date September 2008
    Journal Clinical Breast Cancer
    Excerpt

    BACKGROUND: Recent studies have shown that breast cancer detected by screening has a more favorable prognosis than interval breast cancer. To further understand the biologic significance of this finding, we investigated the association of disease recurrence, local and distant, with the method of detection of the primary breast cancer in a cohort of 1686 women treated with breast conservation. PATIENTS AND METHODS: The charts of 1686 women with primarily stage I or II invasive breast cancer treated by breast conservation between 1977 and 2002 were reviewed. The median length of follow-up was 6 years. Univariate and multivariate analyses using binary logistic regression were performed for 2 subgroups: (1) those with local recurrence versus those without; and (2) those with distant metastasis versus those without distant metastasis. RESULTS: Our data confirmed several of the well-known risk factors for local and distant recurrence. In addition, we found that individuals with breast cancer detected on physical examination alone have a significantly higher risk for local recurrence compared with patients with cancer detected on mammogram alone, independent of tumor size (odds ratio [OR], 2.369; 95% CI, 1.235-4.547; P = .01). We also found a similar correlation for risk of distant metastasis in these 2 groups of women (OR, 2.201; 95% CI, 1.211-3.998; P = .01). CONCLUSION: Breast cancers that are palpable might represent an aggressive biologic subtype with an increased risk of local and distant recurrence. Risk stratification might need to include this clinical feature in addition to conventional prognostic factors.

    Title Radiation Therapy and Toll-like Receptor Signaling: Implications for the Treatment of Cancer.
    Date February 2008
    Journal Oncogene
    Excerpt

    The identification of pathogen-associated molecular patterns, conserved microbial structures that act on Toll-like receptors, has led to a novel avenue of investigation aimed at developing a new generation of cancer immunotherapies. Ligation of Toll-like receptors results in the induction of robust immune responses that may be directed against tumor-associated antigens. Recent data suggest that such strategies may result in enhanced antitumor immunity. Nonetheless, as clinically effective immunotherapy for cancer remains a somewhat distant goal, attention has shifted toward multimodality approaches to cancer therapy, sometimes combining novel immune interventions and conventional treatments. The traditional view of radiation therapy as immunosuppressive has now been challenged, prompting a re-evaluation of its potential as an adjunct to immunotherapy. Radiation therapy can enhance the expression of tumor-associated antigens, induce immune-mediated targeting of tumor stroma, and diminish regulatory T cell activity. Recent evidence suggests that radiation therapy may also activate effectors of innate immunity through TLR-dependent mechanisms, thereby augmenting the adaptive immune response to cancer. In this paper, we will review evidence for enhanced tumor-directed immunity resulting from radiation exposure and early promising data suggesting synergistic effects of radiation and TLR-targeted immunotherapies.

    Title Biopsy Findings After Breast Conservation Therapy for Early-stage Invasive Breast Cancer.
    Date November 2007
    Journal International Journal of Radiation Oncology, Biology, Physics
    Excerpt

    PURPOSE: To determine the patterns and factors predictive of positive ipsilateral breast biopsy after conservation therapy for early-stage breast cancer. METHODS AND MATERIALS: We performed a retrospective review of Stage I-II breast cancer patients initially treated with lumpectomy and radiotherapy between 1977 and 1996, who later underwent post-treatment ipsilateral breast biopsies. RESULTS: A total of 223 biopsies were performed in 193 treated breasts: 171 single and 22 multiple biopsies. Of the 223 biopsies, 56% were positive and 44% were negative for recurrence. The positive biopsy rate (PBR) was 59% for the first and 32% for subsequent biopsies. The median time to the first post-treatment biopsy was 49 months. Of the patients with negative initial biopsy findings, 11% later developed local recurrence. The PBR was 40% among patients with physical examination findings only, 65% with mammographic abnormalities only, and 79% with both findings (p = 0.001). Analysis of the procedure type revealed a PBR of 86% for core and 58% for excisional biopsies compared with 28% for aspiration cytology alone (p = 0.025). The PBR varied inversely with age at the original diagnosis: 49% if >or=51 years, 57% if 36-50 years, and 83% if <or=35 years (p = 0.05). The PBR correlated directly with the interval after radiotherapy: 49% if <or=60 months, 59% if 60.1-120 months, 77% if 120.1-180 months, and 100% if >180 months after completing postlumpectomy radiotherapy (p = 0.01). The PBR was not linked with recurrence location, initial pathologic T or N stage, estrogen receptor/progesterone receptor status, or final pathologic margins (all p >or= 0.15). CONCLUSION: After definitive radiotherapy for early-stage breast cancer, a greater PBR was associated with the presence of both mammographic and clinical abnormalities, excisional or core biopsies, younger age at the initial diagnosis, and longer intervals after radiotherapy completion.

    Title Regional Nodal Metastatic Disease is the Strongest Predictor of Survival in Patients with Thin Vertical Growth Phase Melanomas: a Case for Sln Staging Biopsy in These Patients.
    Date October 2007
    Journal Annals of Surgical Oncology
    Excerpt

    BACKGROUND: The benefit of sentinel lymph node (SLN) biopsy for patients with thin (< or =1.0 mm) melanomas, even for prognostic value, is controversial. This may partly result from the relatively small number and short follow-up of SLN-positive patients in this group. Previously, we have shown that clinical regional nodal metastatic disease (RNMD) serves as a good surrogate for SLN positivity. Here, we use RNMD as a validated surrogate for SLN positivity and examine its prognostic value in a large pre-SLN group of patients with thin vertical growth phase (VGP) lesions who would today commonly be offered SLN biopsy in our practice. METHODS: Between 1972 and 1991, 472 patients with thin VGP melanomas with at least 10 years' follow-up were eligible for the study. Kaplan-Meier survival curves were computed for patients with and without RNMD. A multivariate Cox model and classification tree analysis were used to evaluate clinical and histopathologic predictors of survival. RESULTS: Sixty-seven patients (14.2%) developed recurrence, 53.7% of whom developed RNMD. Forty-five patients (9.5%) experienced melanoma-related deaths (MRD). The most statistically significant predictor of MRD was RNMD (hazard ratio [HR] 13.5, P < .0001). Thickness (HR 10.5, P = .004), axial location (HR 4.6, P = .001), and age >60 years (HR 2.7, P = .005) additionally were independently associated with an increased risk of MRD. RNMD patients demonstrated a 44.4% 10-year disease-specific mortality. CONCLUSIONS: RNMD was the most statistically significant factor associated with MRD in patients with thin VGP lesions. This supports the prognostic use of SLN biopsy in this group, recognizing that additional factors, including thickness, axial location, and older age were independently associated with a worse survival outcome.

    Title Development of Vaccines for High-risk Ductal Carcinoma in Situ of the Breast.
    Date August 2007
    Journal Cancer Research
    Excerpt

    Certain ductal carcinoma in situ (DCIS) lesions overexpress the HER-2/neu receptor at this early stage of breast cancer development. Recently, we showed that a HER-2-targeted dendritic cell vaccine could be used to eliminate HER-2-overexpressing cells in patients that harbor these high-risk DCIS lesions. Our findings suggest that vaccinating such patients might diminish the risk of recurrence, protect against the development of invasive breast cancer, and minimize morbidity associated with current treatments. We discuss several implications of this work for developing effective cancer vaccines.

    Title Identification of High-risk Patients Among Those Diagnosed with Thin Cutaneous Melanomas.
    Date April 2007
    Journal Journal of Clinical Oncology : Official Journal of the American Society of Clinical Oncology
    Excerpt

    PURPOSE: Most patients with melanoma have microscopically thin (< or = 1 mm) primary lesions and are cured with excision. However, some develop metastatic disease that is often fatal. We evaluated established prognostic factors to develop classification schemes with better discrimination than current American Joint Committee on Cancer (AJCC) staging. PATIENTS AND METHODS: We studied patients with thin melanomas from the US population-based Surveillance, Epidemiology, and End Results (SEER) cancer registry (1988 to 2001; n = 26,291) and those seen by the University of Pennsylvania's Pigmented Lesion Group (PLG; 1972 to 2001; n = 2,389; Philadelphia, PA). AJCC prognostic factors were thickness, anatomic level, ulceration, site, sex, and age; PLG prognostic factors also included a set of biologically based candidate prognostic factors. Recursive partitioning was used to develop a SEER-based classification tree that was validated using PLG data. Next, a new PLG-based classification tree was developed using the expanded set of prognostic factors. RESULTS: The SEER-based classification tree identified additional criteria to explain survival heterogeneity among patients with thin, nonulcerated lesions; 10-year survival rates ranged from 89.1% to 99%. The new PLG-based tree identified groups using level, tumor cell mitotic rate, and sex. With survival rates from 83.4% to 100%, it had better discrimination. CONCLUSION: Prognostication and related clinical decision making in the majority of patients with melanoma can be improved now using the validated, SEER-based classification. Tumor cell mitotic rate should be incorporated into the next iteration of AJCC staging.

    Title Targeting Her-2/neu in Early Breast Cancer Development Using Dendritic Cells with Staged Interleukin-12 Burst Secretion.
    Date March 2007
    Journal Cancer Research
    Excerpt

    Overexpression of HER-2/neu (c-erbB2) is associated with increased risk of recurrent disease in ductal carcinoma in situ (DCIS) and a poorer prognosis in node-positive breast cancer. We therefore examined the early immunotherapeutic targeting of HER-2/neu in DCIS. Before surgical resection, HER-2/neu(pos) DCIS patients (n = 13) received 4 weekly vaccinations of dendritic cells pulsed with HER-2/neu HLA class I and II peptides. The vaccine dendritic cells were activated in vitro with IFN-gamma and bacterial lipopolysaccharide to become highly polarized DC1-type dendritic cells that secrete high levels of interleukin-12p70 (IL-12p70). Intranodal delivery of dendritic cells supplied both antigenic stimulation and a synchronized preconditioned burst of IL-12p70 production directly to the anatomic site of T-cell sensitization. Before vaccination, many subjects possessed HER-2/neu-HLA-A2 tetramer-staining CD8(pos) T cells that expressed low levels of CD28 and high levels of the inhibitory B7 ligand CTLA-4, but this ratio inverted after vaccination. The vaccinated subjects also showed high rates of peptide-specific sensitization for both IFN-gamma-secreting CD4(pos) (85%) and CD8(pos) (80%) T cells, with recognition of antigenically relevant breast cancer lines, accumulation of T and B lymphocytes in the breast, and induction of complement-dependent, tumor-lytic antibodies. Seven of 11 evaluable patients also showed markedly decreased HER-2/neu expression in surgical tumor specimens, often with measurable decreases in residual DCIS, suggesting an active process of "immunoediting" for HER-2/neu-expressing tumor cells following vaccination. DC1 vaccination strategies may therefore have potential for both the prevention and the treatment of early breast cancer.

    Title Use of Radiotracer for Sentinel Lymph Node Mapping in Breast Cancer Optimizes Staging Independent of Site of Administration.
    Date January 2007
    Journal Clinical Nuclear Medicine
    Excerpt

    PURPOSE: In an effort to optimize sentinel lymph node (SLN) mapping for breast cancer, sites of mapping agent administration and types of mapping agents used continue to be evaluated. This study compares SLN mapping using peritumoral (PT) or subareolar (SA) injection of radiolabeled colloid and examines the relative contributions of radiotracer and blue dye to SLN identification. MATERIALS AND METHODS: A retrospective review was performed of 456 patients with breast cancer and clinically negative axillae who underwent SLN mapping. Sequential groups of patients were injected with filtered Tc-99m SC, 326 peritumorally (group 1) and 130 subareolarly (group 2). All patients had intraoperative SA injection of 1% isosulfan blue dye. RESULTS: The SLN identification and isotope success rates were 97% and 96% in group 1 and 98% and 98% in group 2, respectively. Eighty-one patients (25%) in group 1 and 44 patients (34%) in group 2 had positive SLNs. Of these patients, 15% from group 1 and 14% from group 2 had only positive nodes detected by radiotracer, and 9 of these patients (6 from group 1 and 3 from group 2) had other nodes identified by both radiotracer and blue dye that were negative for metastases. Six percent of patients with positive SLNs were upstaged because of use of radiotracer. CONCLUSIONS: PT and SA injection of radiotracer have comparable success rates for axillary SLN identification. Given that 15% of patients in group 1 and 14% in group 2 had only positive SLNs detected by radiotracer, independent of site of administration, radiotracer remains essential for optimizing breast SLN mapping.

    Title Predicting Sentinel Node Status in Ajcc Stage I/ii Primary Cutaneous Melanoma.
    Date January 2007
    Journal Cancer
    Excerpt

    BACKGROUND: Sentinel lymph node (SLN) status is an important prognostic factor for survival for patients with primary cutaneous melanoma. To address the issue of selecting patients at high and low risk for a positive SLN, prognostic factors were sought that predict SLN involvement by examining characteristics of both the primary tumor and the patient within the context of a biological model of melanoma progression. METHODS: The study included 682 patients with primary vertical growth phase (VGP) melanoma and no clinical evidence of metastatic disease who underwent SLN biopsy (1995-2003). Logistic regression and classification tree analyses were used to investigate the association between SLN positivity and Breslow thickness, Clark level, tumor infiltrating lymphocytes (TIL), ulceration, mitotic rate (MR), lesion site, gender, and age. RESULTS.: In all, 88 of the 682 patients had > or =1 positive SLN (12.9%). In the multivariate analysis, MR, TIL, and thickness were found to be independent prognostic factors for SLN positivity. In the classification tree, four different risk groups were defined, ranging from minimal risk (2.1%) to high risk (40.4%). In lesions < r =2.0 mm, MR was important in risk-stratifying patients, and in lesions >2.0 mm TIL was important. CONCLUSIONS: By incorporating biologically based variables such as VGP, TIL, and MR along with thickness into a prognostic model, both patients at high risk and minimal risk for SLN positivity can be identified. If validated, this model can be used in patient management and trial design to select patients to undergo or be spared SLN biopsy.

    Title High-avidity Antitumor T-cell Generation by Toll Receptor 8-primed, Myeloid- Derived Dendritic Cells is Mediated by Il-12 Production.
    Date October 2006
    Journal Surgery
    Excerpt

    BACKGROUND: High-level production of heterodimeric p70 interleukin (IL)-12 by myeloid-derived dendritic cells (DCs) requires 2 signals: interferon gamma (IFN-gamma) and a maturation signal provided by CD40 ligation (CD40L) or lipopolysaccharide (LPS). METHODS: In the current study we demonstrate that signaling through toll-like receptor (TLR) 8, but not TLR3, TLR2, or TLR4, provides a priming signal to myeloid-derived DC for high IL-12 p70 heterodimer production. RESULTS: All the TLR agonists induced maturation of DC as evidenced by increased expression of CD83, CD80, and CD86. Both IFN-gamma and TLR7/8 agonist R848 increased expression of TLR8 in immature monocyte-derived DCs. The combination of TLR7/8 agonist R848 and maturation signals LPS or CD40L induced high-level expression of IL-12p35 and p40 similar to that induced by IFN-gamma plus LPS. In contrast, receptor agonists specific for TLR7 did not prime for IL-12 production. The p70 IL-12 produced by the TLR8-primed DC polarized CD4+ T for Th1 cytokine production and induced CD8+ T cells, displaying high functional avidity with enhanced tumor cell recognition. CONCLUSIONS: The data suggest that toll 8 receptor agonists are useful for inducing type-1 polarized DCs for vaccine design in treating cancer and infectious disease.

    Title Predictors of Regional Nodal Disease in Patients with Thin Melanomas.
    Date September 2006
    Journal Annals of Surgical Oncology
    Excerpt

    BACKGROUND: Most melanoma patients present with thin (<or=1.0 mm) lesions. Indications for sentinel lymph node (SLN) biopsy are not well defined for this group. Previously, we reported an association between mitotic rate (MR) and SLN positivity in these patients. The study was limited by a relatively small sample size and low statistical power. In this study, we evaluated a large population of patients with thin melanoma from the pre-SLN era to identify predictors of regional nodal disease (RND) that may serve as a surrogate for SLN positivity. METHODS: Eight hundred eighty-two patients evaluated between 1972 and 1991 were included in the study. Univariate and multivariate regression analyses were performed by using clinical and histological data to identify factors associated with RND. A multivariate logistic regression model was developed and applied to the previously reported group of patients with thin melanomas who underwent SLN biopsy between 1996 and 2004 for validation. RESULTS: Thirty-eight patients (4.3%) had evidence of RND. In the multivariate analysis, a MR>0, vertical growth phase (VGP), male sex, and ulceration were statistically significant predictors of RND. Patients at the highest risk according to a classification tree analysis (VGP and MR>0) had an RND rate of 11.9%. The regression model developed predicted well the SLN status in the validation sample. CONCLUSIONS: Investigation of a large pre-SLN population identified MR>0, ulceration, VGP, and male sex as independently predictive of RND in patients with thin melanomas. These factors may help to identify subgroups of these patients that have clinically significant risks of SLN positivity.

    Title Dendritic Cells in Melanoma Immunotherapy.
    Date August 2006
    Journal Current Treatment Options in Oncology
    Excerpt

    No consensus exists regarding optimal systemic treatment for melanoma in the adjuvant or metastatic disease setting. Dendritic cell vaccine therapy, though investigational at present, offers very promising preliminary data that warrant exploration, and patients with existing disease and those seeking adjuvant treatment should evaluate open protocols using this strategy.

    Title Human Leukocyte Antigen-a2-restricted Ctl Responses to Mutated Braf Peptides in Melanoma Patients.
    Date May 2006
    Journal Cancer Research
    Excerpt

    Mutated BRAF (BRAF(V600E)) is a potential immunotherapeutic target for melanoma because of its tumor specificity and expression in the majority of these lesions derived from different patients. BRAF(V600E) is expressed intracellularly and not on the cell surface, therefore providing a target for T cells but not B cells. Demonstration of patients' T cell responses to BRAF(V600E) would suggest the feasibility of active specific immunotherapy targeting the mutation in these patients. In the present study, BRAF(V600E) peptides with putative binding sites for human leukocyte antigen (HLA)-A2 were used to stimulate T lymphocytes of HLA-A2-positive melanoma patients. Four of five patients with BRAF(V600E)-positive lesions showed lymphoproliferative responses to BRAF(V600E) peptide stimulation. These responses were specific for the mutated epitope and HLA-A2 was restricted in three patients. Lymphocytes from these three patients were cytotoxic against HLA-A2-matched BRAF(V600E)-positive melanoma cells. None of the four patients with BRAF(V600E)-negative lesions and none of five healthy donors had lymphoproliferative responses specific for the mutated epitope. The high prevalence (approximately 50%) of HLA-A2 among melanoma patients renders HLA-A2-restricted BRAF(V600E) peptides attractive candidate vaccines for these patients.

    Title Repeat Operative Sentinel Lymph Node Biopsy.
    Date April 2006
    Journal Clinical Breast Cancer
    Excerpt

    Because sentinel lymph node (SLN) biopsy continues to be used for staging in patients with breast cancer, physicians treating these patients will be faced with in-breast recurrences and new primary breast cancers in the treated breast. Repeat operative SLN biopsy might be feasible in this clinical scenario. This report describes the case of a patient with an ipsilateral different-site, recurrent, infiltrating ductal carcinoma 14 months after lumpectomy; negative SLN biopsy result; and radiation therapy, now with a positive SLN biopsy result.

    Title Diffuse Optical Measurement of Blood Flow in Breast Tumors.
    Date January 2006
    Journal Optics Letters
    Excerpt

    Blood flow contrast between tumor and normal tissues in patients with malignant and benign breast cancer was measured by diffuse optical correlation methods. The measurements were carried out with a hand-held optical probe that was manually scanned over the tumor-bearing breast. Increased blood flow was observed in tumor regions relative to healthy tissue, and control subjects did not exhibit significant blood flow heterogeneity. The measurements introduce a new optical contrast for diffuse optical mammography.

    Title Breast Cancer Detection Based on Incremental Biochemical and Physiological Properties of Breast Cancers: a Six-year, Two-site Study.
    Date December 2005
    Journal Academic Radiology
    Excerpt

    RATIONALE AND OBJECTIVES: To demonstrate that near-infrared spectroscopy would achieve sufficient sensitivity and specificity in human breast cancer to reach ROC/AUC values in the 90s and yet to warn of the potential liabilities of introduction of a novel technology in this field. MATERIALS AND METHODS: 116 subjects from two nations (44 were cancer-verified by biopsy and histopathology) were reviewed. NIR spectroscopy of total hemoglobin and its relative oxygenation were monitored in breast cancers and compared to their contralateral breast in a 2D nomogram for diagnostic evaluation. A novel handheld NIR breast cancer detector pad with a 3-wavelength LED and 8 detectors with 4 cm separation between source and detectors was placed on the subject's breast. The method is convenient, rapid, and safe and has achieved high patient compliance with minimal patient apprehension of compression, confinement, or radioactivity. RESULTS: The absorbance increments of the cancerous region are referred to the mirror image location on the contralateral breast. The two metrics are increased hemoglobin concentration due to angiogenesis and decreased hemoglobin saturation due to hypermetabolism of the cancer. The 2D nomogram display of these two metrics shows Zone 1 contains verified cancers and Zone 2 contains noncancers. ROC evaluation of the nomogram gives 95% AUC for the two sites, Philadelphia and Leipzig. CONCLUSION: A simple, economical breast cancer detector has achieved high patient compliance and a high ROC/AUC score for a population which involved a range of tumors down to and including those of 0.8-1 cm in diameter.

    Title Mitotic Rate As a Predictor of Sentinel Lymph Node Positivity in Patients with Thin Melanomas.
    Date August 2005
    Journal Annals of Surgical Oncology
    Excerpt

    BACKGROUND: Lymphatic mapping and sentinel lymphadenectomy (LM/SL) provide important prognostic information for patients with early-stage melanoma. Although the use of this technique in patients with thin melanomas (< or =1.00 mm) is not routine, risk factors that may predict sentinel lymph node (SLN) positivity in this patient population are under investigation. We sought to determine whether mitotic rate (MR) is associated with SLN positivity in thin-melanoma patients and, therefore, whether it may be used to risk-stratify and select patients for LM/SL. METHODS: Clinical and histopathologic variables were reviewed for 181 patients with thin melanomas who underwent LM/SL from January 1996 through January 2004. Univariate and multivariate logistic regression analyses were performed to identify factors associated with SLN positivity. Risk groups were defined on the basis of the development of a classification tree. RESULTS: The overall SLN positivity rate was 5%. All patients with positive SLNs had an MR of >0. By univariate analysis, MR and thickness were significant predictors of SLN positivity. The association between MR and SLN positivity remained significant controlling for each of the other variables evaluated. On the basis of a classification tree, patients with an MR >0 and tumor thickness > or =.76 mm were identified as a higher-risk group, with an SLN positivity rate of 12.3%. CONCLUSIONS: In patients with thin melanomas, MR >0 seems to be a significant predictor of SLN positivity that may be used to risk-stratify and select patients for LM/SL. To confirm these results, the predictive value of MR for SLN positivity needs to be validated in other populations of thin-melanoma patients.

    Title Diffuse Optical Tomography of Breast Cancer During Neoadjuvant Chemotherapy: a Case Study with Comparison to Mri.
    Date August 2005
    Journal Medical Physics
    Excerpt

    We employ diffuse optical tomography (DOT) to track treatment progress in a female subject presenting with locally advanced invasive carcinoma of the breast during neoadjuvant chemotherapy. Three-dimensional images of total hemoglobin concentration and scattering identified the tumor. Our measurements reveal tumor shrinkage during the course of chemotherapy, in reasonable agreement with magnetic resonance images of the same subject. A decrease in total hemoglobin concentration contrast between tumor and normal tissue was also observed over time. The results demonstrate the potential of DOT for measuring physiological parameters of breast lesions during chemotherapy.

    Title Combining Innate Immunity with Radiation Therapy for Cancer Treatment.
    Date June 2005
    Journal Clinical Cancer Research : an Official Journal of the American Association for Cancer Research
    Title Skin-sparing Mastectomy and Immediate Reconstruction is an Acceptable Treatment Option for Patients with High-risk Breast Carcinoma.
    Date March 2005
    Journal Cancer
    Excerpt

    BACKGROUND: Skin-sparing mastectomy (SSM) followed by immediate reconstruction is an effective treatment option for patients with early-stage breast carcinoma, but its use in patients with more advanced disease is controversial. METHODS: A retrospective review was performed that included 38 consecutive patients with high-risk breast carcinoma who underwent SSM and immediate reconstruction (between July 1996 and January 2002). Tumor characteristics, type of reconstruction, margin status, timing of adjuvant therapy, postoperative complications, and incidence of recurrence were evaluated. RESULTS: High-risk patients (Stage IIA [n=4 patients] Stage IIB [n=23 patients] Stage IIIA [n=8 patients] and Stage IIIB [n=3 patients]) underwent immediate reconstruction after SSM with the use of a transverse rectus abdominis myocutaneous flap (n=31 patients), a latissimus dorsi myocutaneous flap plus an implant (n=3 patients), or tissue expanders with subsequent implant placement (n=4 patients). The median follow-up was 52.9 months (range, 27.5-92.0 months), and the median time to recurrence has not yet been reached at the time of last follow-up. The median interval from surgery to the initiation of postoperative adjuvant therapy was 38 days (range, 25-238 days). Local recurrence was seen in 1 patient (2.6%), systemic recurrence in was seen in 10 patients (26.3%), and both local and distant metastases in were seen in 2 other patients (5.3%). CONCLUSIONS: SSM with immediate reconstruction appeared to be an oncologically safe treatment option for high-risk patients with advanced stages of breast carcinoma. In addition to the aesthetic and psychological benefits of performing SSM with immediate reconstruction, local recurrence rates and disease-free survival were favorable when combined with the use of radiation therapy and adjuvant chemotherapy, as indicated.

    Title Cutting Edge: Innate Immune System Discriminates Between Rna Containing Bacterial Versus Eukaryotic Structural Features That Prime for High-level Il-12 Secretion by Dendritic Cells.
    Date July 2004
    Journal Journal of Immunology (baltimore, Md. : 1950)
    Excerpt

    RNA derived from bacterial but not eukaryotic sources, when transfected into human monocyte-derived dendritic cell precursors, induces high-level IL-12 secretion in conjunction with dendritic cell maturation stimuli. In vitro-transcribed mRNA that mimics the structure of bacterial mRNA in the lack of a long 3'-poly(A) tail likewise induces IL-12 secretion, but this property is lost upon efficient enzymatic 3'-polyadenylation. Among other tested RNAs, only polyuridylic acid induced IL-12 p70. This RNA response phenomenon appears biologically distinct from the classically defined response to dsRNA. RNA-transfected APC also polarize T cells in an IL-12-dependent manner toward the IFN-gamma(high)IL-5 (low) Th1 phenotype, suggesting a link between the detection of appropriately structured RNA and the skewing of immune responses toward those best suited for controlling intracellular microbes. RNA structured to emulate bacterial patterns constitutes a novel vaccine strategy to engender polarized Th1-type immune responses.

    Title Outcome After Treatment of Patients with Mammographically Occult, Magnetic Resonance Imaging-detected Breast Cancer Presenting with Axillary Lymphadenopathy.
    Date July 2004
    Journal Clinical Breast Cancer
    Excerpt

    Although magnetic resonance imaging (MRI) for patients with mammographically occult breast cancer with axillary lymphadenopathy has been accepted for staging, treatment outcome data in this patient group is lacking. In this study, 16 patients, median age of 45 years (range, 27-66 years), presented with malignant axillary lymphadenopathy, negative mammograms, negative breast physical examination, and abnormal breast MRI. All 16 patients were found to have >/= 1 suspicious lesions on breast MRI. Ten patients had a solitary enhancing lesion; 1 patient had 2 enhancing lesions; 3 patients had 3 enhancing lesions; 1 patient had a mass lesion and diffuse patchy enhancement in the breast; and 1 patient had regional enhancement but no discrete lesion on MRI. Six patients underwent breast-conservation surgery using MRI-guided wire localization and 10 patients had modified radical mastectomy. Fourteen patients received adjuvant chemotherapy and the remaining 2 patients received neoadjuvant chemotherapy. With a median follow-up of 5 years (range, 1.2-7.6 years), the 5-year actuarial local control was 100%, relapse-free survival was 74%, and overall survival was 87%. Three patients developed distant metastases. Two patients died from distant metastases, and 1 patient is alive with metastatic disease. One patient had a relapse in the contralateral axilla and was treated with paclitaxel and is disease free. Although the patient population is small, the outcome after treatment for this group of patients with a mammographically occult, MRI-detected breast cancer presenting with axillary adenopathy is similar to the expected outcome for patients with breast cancer with positive axillary lymph nodes.

    Title Mri Before Reexcision Surgery in Patients with Breast Cancer.
    Date March 2004
    Journal Ajr. American Journal of Roentgenology
    Excerpt

    OBJECTIVE: The aims of this study were to assess the diagnostic accuracy of MRI in evaluating patients for residual cancer, identify the prevalence of multicentric or multifocal disease, and evaluate the impact of MRI on surgical treatment planning. SUBJECTS AND METHODS. Of 101 potentially eligible patients, 80 candidates for breast conservation therapy who had primary breast cancer in 82 breasts diagnosed by excisional biopsy with close or positive margins were included in the study group. All patients underwent contrast-enhanced MRI before further surgery and subsequently underwent either reexcision lumpectomy or mastectomy with histopathologic correlation. RESULTS: Residual carcinoma, either invasive or in situ, was present in 59.8% of the breasts. The sensitivity and specificity of MRI for detecting residual disease were 61.2% and 69.7%, respectively. Twenty-three additional lesions distant from the biopsy site were identified in 19 breasts, and 18 suspicious lesions underwent biopsy. Histology results indicated that six lesions were malignant, so the overall prevalence was 7.3%. The positive predictive value of identifying an additional suspicious lesion was 33.3%. In 24 breasts, MRI changed which procedure would be performed next from reexcision lumpectomy to mastectomy (n = 9), biopsy of an additional lesion in the ipsilateral (n = 12) or contralateral (n = 2) breast, or neoadjuvant chemotherapy (n = 1). Approximately 25% of the breasts underwent mastectomy as definitive surgical treatment. CONCLUSION: Overlap in the appearances of benign and malignant lesions limits MRI evaluation for residual disease. MRI can show additional suspicious lesions that are likely to be multicentric or multifocal disease. These findings changed the original treatment plan for approximately 30% of breasts.

    Title Rapid High Efficiency Sensitization of Cd8+ T Cells to Tumor Antigens by Dendritic Cells Leads to Enhanced Functional Avidity and Direct Tumor Recognition Through an Il-12-dependent Mechanism.
    Date November 2003
    Journal Journal of Immunology (baltimore, Md. : 1950)
    Excerpt

    Myeloid-origin dendritic cells (DCs) can develop into IL-12-secreting DC1 or non-IL-12-secreting DC2 depending on signals received during maturation. Through rapid culture techniques that prepared either mature, CD83+ DC1 or DC2 from CD14+ monocytes in only 2 days followed by a single 6-7 day DC-T cell coculture, we sensitized normal donor CD8+ T cells to tumor Ags (HER-2/neu, MART-1, and gp100) such that peptide Ag-specific lymphocytes constituted up to 16% of the total CD8+ population. Both DC1 and DC2 could sensitize CD8+ T cells that recognized peptide-pulsed target cells. However, with DC2, a general decoupling was observed between recognition of peptide-pulsed T2 target cells and recognition of Ag-expressing tumor cells, with peptide-sensitized T cells responding to tumor only about 15% of the time. In contrast, direct recognition of tumor by T cells was dramatically increased (to 85%) when DC1 were used for sensitization. Enhanced tumor recognition was accompanied by 10- to 100-fold increases in peptide sensitivity and elevated expression of CD8beta, characteristic of high functional avidity T cells. Both of these properties were IL-12-dependent. These results demonstrate the utility of rapid DC culture methods for high efficiency in vitro T cell sensitization that achieves robust priming and expansion of Ag-specific populations in 6 days. They also demonstrate a novel function of IL-12, which is enhancement of CD8+ T cell functional avidity. A new approach to DC-based vaccines that emphasizes IL-12 secretion to enhance functional avidity and concomitant tumor recognition by CD8+ T cells is indicated.

    Title Intranodal Administration of Peptide-pulsed Mature Dendritic Cell Vaccines Results in Superior Cd8+ T-cell Function in Melanoma Patients.
    Date October 2003
    Journal Journal of Clinical Oncology : Official Journal of the American Society of Clinical Oncology
    Excerpt

    PURPOSE: We evaluated the feasibility, safety, and immunogenicity of mature, peptide-pulsed dendritic cell (DC) vaccines administered by different routes. PATIENTS AND METHODS: We performed a randomized, phase I, dose-escalation study in 27 patients with metastatic melanoma receiving four autologous peptide-pulsed DC vaccinations. Patients were randomly assigned to an intravenous (IV), intranodal (IN), or intradermal (ID) route of administration (ROA). For each route, primary end points were dose-limiting toxicity, maximum-tolerated dose, and T-cell sensitization. Sensitization was evaluated through tetramer staining, in vitro peptide recognition assays, and delayed-type hypersensitivity (DTH) responses. RESULTS: Twenty-two (81.5%) of 27 patients completed all four vaccinations. Vaccinations were well tolerated; a few patients exhibited grade 1 to 2 toxicities including rash, fever, and injection site reaction. All routes of administration induced comparable increases in tetramer-staining CD8+ T cells (five of seven IV, four of seven IN, and four of six ID patients). However, the IN route induced significantly higher rates for de novo development of CD8+ T cells that respond by cytokine secretion to peptide-pulsed targets (six [85.7%] of seven IN patients v two [33%] of six ID patients v none [0%] of six IV patients; P =.005) and de novo DTH (seven [87.5%] of eight IN patients v two [33.3%] of six ID patients v one [14.3%] of seven IV patients; P =.01) compared with other routes. CONCLUSION: Administration of this peptide-pulsed mature DC vaccine by IN, IV, or ID routes is feasible and safe. IN administration seems to result in superior T-cell sensitization as measured by de novo target-cell recognition and DTH priming, indicating that IN may be the preferred ROA for mature DC vaccines.

    Title Changes in the Surgical Management of Patients with Breast Carcinoma Based on Preoperative Magnetic Resonance Imaging.
    Date August 2003
    Journal Cancer
    Excerpt

    BACKGROUND: Breast magnetic resonance imaging (MRI) is a developing technique for the evaluation of patients with primary breast carcinoma. The authors assessed the impact of preoperative breast MRI on surgical management. METHODS: The current study was a retrospective review of 267 patients with primary breast tumors who had MRI studies prior to undergoing definitive surgery. RESULTS: Two hundred sixty-seven patients with invasive breast carcinoma who had preoperative breast MRI studies and had complete clinical, radiologic, and pathologic data available were identified and formed the basis of this analysis. The overall sensitivity of MRI for detecting primary, intact breast tumors was 95%. Planned surgical management was altered in 69 of 267 patients (26%); and, in 49 of those patients (71%), there was pathologic verification of malignancy in the surgical specimen that confirmed the need for wider or separate excision or mastectomy. Forty-four of 267 patients (16.5%) had conversion of planned breast conservation to mastectomy. In a univariate analysis, change in management was associated significantly with histology; management was altered in 11 of 24 lobular tumors (46%) compared with 58 of 243 ductal tumors (24%; P = 0.02). CONCLUSIONS: Breast MRI was very sensitive for the detection of primary, intact, invasive breast carcinoma and improved local staging in almost 20% of patients. Preoperative breast MRI studies may be particularly useful in surgical planning for and management of patients with lobular carcinoma.

    Title Mr Imaging Screening of the Contralateral Breast in Patients with Newly Diagnosed Breast Cancer: Preliminary Results.
    Date April 2003
    Journal Radiology
    Excerpt

    PURPOSE: To investigate the role of screening magnetic resonance (MR) imaging in the detection of synchronous contralateral breast cancer in patients with newly diagnosed breast cancer. MATERIALS AND METHODS: Between January 1999 and July 2001, 182 patients with newly diagnosed breast cancer (after either core or excisional biopsy with positive or close margins of resection) underwent bilateral contrast material-enhanced MR imaging at 1.5 T with a dedicated bilateral breast multicoil array. The contralateral breast was imaged for cancer screening. Family history of breast cancer, index cancer histology, breast density, and age at diagnosis of first breast cancer were assessed as potential risk factors for synchronous contralateral breast cancer. RESULTS: Fifteen patients (8.2%) had a suspicious enhancing lesion depicted in the contralateral breast. Seven patients (3.8%) had malignant results: ductal carcinoma in situ (DCIS) in four, invasive ductal carcinoma with DCIS in two, and invasive ductal carcinoma in one. Eight patients (4.4%) had benign results: fibrocystic changes in four, atypical ductal hyperplasia in two, atypical lobular hyperplasia and focal lobular carcinoma in situ in one, and ductal hyperplasia in one. Six patients with negative MR findings underwent prophylactic mastectomy; no malignancy was found. No significant differences were noted among patients with true-positive (n = 7), false-positive (n = 8), or negative (n = 167) MR findings with regard to family history of breast cancer (P <.27), index cancer histology (P <.19), breast density (P <.34), or age at diagnosis of first breast cancer (P <.10). CONCLUSION: The preliminary results demonstrate the feasibility of using MR imaging of the breast in a screening role, specifically to evaluate the contralateral breast in patients with newly diagnosed breast cancer to detect mammographically and clinically occult synchronous breast cancer.

    Title Magnetic Resonance Imaging-guided Biopsy of Mammographically and Clinically Occult Breast Lesions.
    Date July 2002
    Journal Annals of Surgical Oncology
    Excerpt

    BACKGROUND: Breast magnetic resonance imaging (MRI) is a very sensitive technique for detection of breast cancer. We report on MRI-guided needle localization for biopsy of abnormalities seen only on MRI. METHODS: A retrospective review was performed of 231 patients with invasive breast cancer or ductal carcinoma-in-situ who had MRI as part of their evaluation and treatment at the University of Pennsylvania between 1992 and 1998. Clinical, radiological, and pathologic data were examined. RESULTS: MRI needle localization was performed in 41 (18%) patients. MRI needle localization was required for a finding of a mammographically or clinically occult lesion in 31 patients, better MRI definition of tumor in 5 patients, and surgeon's choice in 5 patients. In all cases, MRI localization and excisional biopsy were successfully completed. Nineteen of 31 patients were found to have additional mammographically and clinically occult tumors. There were 12 (29%) false-positive MRI scans. CONCLUSIONS: MRI has a high sensitivity for detection of breast cancer; additional mammographically and clinically occult sites of tumor are detected in approximately 1 (15%) of 7 breast cancer patients. These otherwise occult sites of disease can be appropriately biopsied with MRI needle-localization techniques.

    Title Mature Cd83(+) Dendritic Cells Infected with Recombinant Gp100 Vaccinia Virus Stimulate Potent Antimelanoma T Cells.
    Date July 2002
    Journal Annals of Surgical Oncology
    Excerpt

    BACKGROUND: Mature dendritic cells (DCs) are potent antigen-presenting cells that activate naive T lymphocytes and initiate cellular immune responses. The ability of CD83(+) mature DCs infected with vaccinia virus encoding the gp100 melanoma transgene (rV-gp100) to stimulate an antimelanoma CD8(+) T-cell response was investigated. METHODS: Monocyte-derived immature or CD83(+) mature DCs were infected with rV-gp100. The activation state of the DCs and the expression of gp100 protein were evaluated by flow cytometry. The reactivity of antimelanoma CD8(+) T cells was confirmed by measuring specific interferon gamma secretion by using enzyme-linked immunosorbent assay in a mixed-tumor lymphocyte culture. RESULTS: Both immature and CD83(+) mature DCs expressed gp100 protein when the DCs were infected with rV-gp100. Calcium-signaling agents were required to induce maturation of both infected and noninfected immature DCs. Only rV-gp100-infected CD83(+) DCs induced CD8(+) T cells, after a single stimulation that recognized both peptide-pulsed target cells to multiple gp100 epitopes and a melanoma cell line that endogenously expressed gp100 antigen. CONCLUSIONS: CD83(+) DCs transduced with rV-gp100 are capable of generating a strong CD8(+) T-cell response against melanoma tumor cells. Expression of melanoma antigens by mature DCs offers the potential advantage of presenting multiple endogenously processed T-cell epitopes and using multiple HLA restriction elements for antimelanoma vaccine therapy.

    Title Subareolar and Peritumoral Injection Identify Similar Sentinel Nodes for Breast Cancer.
    Date April 2002
    Journal Annals of Surgical Oncology
    Excerpt

    BACKGROUND: Sentinel lymph node (SLN) mapping with radioisotope and blue dye is rapidly becoming the standard of care for breast cancer. The optimal location for injection of radioisotope and blue dye is still being investigated. The goal of this study was to determine whether blue dye injection into the subareolar (SA) location localized the same sentinel nodes as the peritumoral (PT) location for patients with breast cancer. METHODS: Three hundred thirty-two patients with biopsy-proven operable breast cancer or ductal carcinoma in situ at two institutions underwent SLN mapping. Eighty-three patients had PT injection of blue dye (group 1), and 249 patients had SA injection of blue dye (group 2). All patients underwent PT injection of (99m)Tc-labeled sulfur colloid. RESULTS: The two groups were similar in age, previous biopsy type, and tumor size, location, and histology. The mean number of SLNs identified was 2.4 (range, 0-9) in group 1 and 2.5 (range, 0-11) in group 2. The SLN identification rate was 95% for group 1 and 97% for group 2. The isotope success rate was 94% for both groups. The blue dye success rate was 84% for group 1 and 90% for group 2. The isotope/blue dye concordance rate was 87% for group 1 and 90% for group 2. At a median follow-up of 28 months (range, 14 to 40), there were no axillary recurrences in any of the 332 patients. CONCLUSIONS: These data suggest that delivery of mapping reagents in the SA and PT locations identifies similar lymph nodes. Because of simplicity and the similarity in node identification between SA and PT injection, further investigation of the SA site for delivery of SLN mapping reagents for breast cancer is warranted.

    Title Diverse Functional Activity of Cd83+ Monocyte-derived Dendritic Cells and the Implications for Cancer Vaccines.
    Date March 2002
    Journal Critical Reviews in Immunology
    Excerpt

    Antigen-loaded dendritic cells (DCs) provide key regulatory signals to T cells during a developing antitumor response. In addition to providing costimulation, mature DC provides cytokine and chemokine signals that can define the T1 vs T2 nature of the antitumor T-cell response as well as whether T cells engage in direct interactions with tumor cells. In serum-free culture conditions that hasten the differentiation of monocytes into mature DCs, certain agents, such as CD40L, accelerate phenotypic maturation (e.g., CD83 and costimulatory molecule expression) without influencing the acquisition of Dc1/Dc2 characteristics. In contrast, exposure to serum-free medium and interferon-gamma (IFN-gamma) rapidly influences CD83+ DCs to secrete high levels of IL-12, IL-6, and MIP-1beta, and promotes Dcl differentiation. In contrast, CD83+ DCs matured in serum-free medium in the absence of IFN-gamma, or in the presence of calcium signaling agents, prostaglandin-E2, or IFN-alpha, produce no IL-12, scant IL-6, and prodigious IL-8, MDC, and TARC, and promote Dc2 differentiation. T cells sensitized via IL-12-secreting, peptide-pulsed DCs secrete cytokines when subsequently exposed to relevant peptide-pulsed antigen-presenting cells (APCs) or to HLA-compatible tumor cells endogenously expressing the peptide. In contrast, T cells sensitized via IL-12 nonsecreting DC were limited to antigenic reactivation through APC contact rather than tumor cell contact. Therefore, the development of antitumor responses can be dramatically influenced not only by costimulation, but also by the cytokine and chemokine production of DCs, which must be considered in the development of cancer vaccines.

    Title T-cell Adoptive Therapy of Tumors: Mechanisms of Improved Therapeutic Performance.
    Date March 2002
    Journal Critical Reviews in Immunology
    Excerpt

    The T cells of many cancer patients are naturally sensitized to tumor-associated antigens (Ag), or they can readily be sensitized with vaccine maneuvers. In melanoma patients, the adoptive transfer of such T cells can often be causally linked to the objective regression of established tumors. So far, few patients have shown sustained clinical benefit from such therapy, but preclinical mouse studies have now clearly delineated the hurdles that must be overcome to render T-cell-based antitumor therapy effective. Contrary to earlier expectations, it is now established that remarkably potent CD4+ and CD8+ pre-effector T cells are naturally sensitized even in mice bearing progressive, weakly immunogenic tumors. However, such T cells often display signal transduction impairments as a consequence of the tumor environment, which limit their acquisition of optimal effector function. Extracorporealization and culture of these tumor-sensitized T cells with appropriate activation stimuli not only restores normal signal transduction, but also confers resolute effector activity that can often sustain tumor rejection upon reinfusion. In mouse studies, the L-selectin(low) fraction of T cells in tumor-draining lymph nodes (TDLN) constitutes the potent pre-effector population and comprises both CD4+ and helper-independent CD8+ T cells. Appropriate in vitro activation confers an apparently unrestricted trafficking capacity to this fraction, and even the ability to proliferate within the tumor bed, leading to unprecedented tumor rejection at anatomic sites (e.g., subcutaneous and intracranial) that were historically refractory to such treatment. Such results underscore the surprising capacity of appropriately activated effector T cells to withstand the immunosuppressive, tolerogenic, and apoptotic influences of the typical tumor environment. Given the increasingly appreciated and critical communications between T cells and host Ag-presenting cells (APC), which cross-present tumor Ag, it is likely that dendritic cell-based vaccine maneuvers that promote sensitization of T1-committed L-selectin(low) antitumor T cells will play an increasingly important role in adoptive therapy strategies.

    Title Calcium Signaling Inhibits Interleukin-12 Production and Activates Cd83(+) Dendritic Cells That Induce Th2 Cell Development.
    Date December 2001
    Journal Blood
    Excerpt

    Mature dendritic cells (DCs), in addition to providing costimulation, can define the Th1, in contrast to the Th2, nature of a T-cell response through the production of cytokines and chemokines. Because calcium signaling alone causes rapid DC maturation of both normal and transformed myeloid cells, it was evaluated whether calcium-mobilized DCs polarize T cells toward a Th1 or a Th2 phenotype. After human monocytes were cultured for 24 hours in serum-free medium and granulocyte-macrophage colony-stimulating factor to produce immature DCs, additional overnight culture with either calcium ionophore (CI) or interferon gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), and soluble CD40L resulted in phenotypically mature DCs that produced interleukin-8 (IL-8) and displayed marked expression of CD80, CD86, CD40, CD54, CD83, DC-LAMP, and RelB. DCs matured by IFN-gamma, TNF-alpha, and soluble CD40L were additionally distinguished by undetectable CD4 expression, marked secretion of IL-12, IL-6, and MIP-1beta, and preferential ability to promote Th1/Tc1 characteristics during T-cell sensitization. In contrast, DCs matured by CI treatment were distinguished by CD4 expression, modest or absent levels of IL-12, IL-6, and MIP-1beta, and preferential ability to promote Th2/Tc2 characteristics. Calcium signaling selectively antagonized IL-12 production by mature DCs activated with IFN-gamma, TNF-alpha, and soluble CD40L. Although the activation of DCs by calcium signals is largely mediated through calcineurin phosphatase, the inhibition of IL-12 production by calcium signaling was independent of this enzyme. Naturally occurring calcium fluxes in immature DCs, therefore, negatively regulate Dc1 differentiation while promoting Dc2 characteristics and Th2/Tc2 polarization. Calcium-mobilized DCs may have clinical usefulness in treating disease states with excessive Th1/Tc1 activity, such as graft-versus-host disease or autoimmunity.

    Title Locally Recurrent Malignant Melanoma Characteristics and Outcomes: a Single-institution Study.
    Date October 2001
    Journal Annals of Plastic Surgery
    Excerpt

    Despite improvements in the identification and treatment of melanoma, local recurrence continues to challenge the success of current melanoma therapy. A retrospective analysis of 1,996 patients presenting from 1990 to 1997 at the Pigmented Lesion Group of the University of Pennsylvania was performed to assess clinical characteristics and outcomes of locally recurrent melanoma. The cases were analyzed by chart and pathological slide review. A control group was identified for statistical comparison. The incidence of locally recurrent melanoma during the study period was 2.2%. Lentigo maligna melanoma (LMM) accounted for 37% of the local recurrences. Increased tumor thickness and microsatellites were associated with "early" local recurrence and decreased survival from time of recurrence. Nineteen percent of the local recurrences occurred more than 5 years after the initial definitive treatment. The preponderance of locally recurrent LMM suggests the need for refinements in the techniques of margin identification and surgical excision of LMM. Tumors with increased thickness and microsatellites should receive particularly close attention. Lastly, with nearly 20% of the local recurrences occurring more than 5 years after the initial date of treatment, the authors suggest extending the follow-up time for all melanoma lesions beyond 5 years.

    Title Revolutionary Impact of Lymphoscintigraphy and Intraoperative Sentinel Node Mapping in the Clinical Practice of Oncology.
    Date September 2001
    Journal Seminars in Nuclear Medicine
    Excerpt

    Intraoperative lymphatic mapping is a rapidly emerging diagnostic approach that is revolutionizing the management of patients who have solid malignant tumors. The procedure is being performed for the most part with radiopharmaceuticals and vital blue dyes. It is widely believed that passive trapping of radioactive particles determines the sentinel lymph node (SLN) for intraoperative delineation of potential draining sites. In this article, we show that dendritic cells within the SLN actively take up and trap radioactive particles and thus define the SLN immunologically. The role of preoperative lymphoscintigraphy and the selection of the site of placement of mapping reagents for intraoperative lymphatic mapping are established for patients with melanoma. For patients with breast cancer, the role of preoperative lymphoscintigraphy is controversial. We have shown that this procedure can be performed with success in identifying SLN as hot spots 87% of the time, with 20% of the cases showing draining nodes to other basins in addition to or independent of the axilla. The use of preoperative lymphoscintigraphy for patients with breast cancer can therefore be justified. The selection of the site for placement of radiotracer and blue dye can vary for patients with breast cancer depending on the primary site of the lesion. However, based on data from our institution and others, the delivery of the mapping reagents (both radioactive tracers and blue dye) to the subareolar space may help to standardize breast cancer SLN mapping.

    Title The Impact of a Multidisciplinary Breast Cancer Center on Recommendations for Patient Management: the University of Pennsylvania Experience.
    Date May 2001
    Journal Cancer
    Excerpt

    BACKGROUND: Advances in the diagnosis and treatment of breast carcinoma have led to a multidisciplinary approach to management for patients with breast carcinoma. To assess the effect of this approach, the authors performed an evaluation for a cohort of patients examined in a multidisciplinary breast cancer center. METHODS: An analysis was performed for the records of 75 consecutive women with 77 breast lesions examined in consultation in a multidisciplinary breast cancer center between January and June 1998. Each patient's case was evaluated by a panel consisting of a medical oncologist, surgical oncologist, radiation oncologist, pathologist, diagnostic radiologist, and, when indicated, plastic surgeon. A comprehensive history and physical examination was performed, and the relevant mammograms, pathology slides, and medical records were reviewed. Treatment recommendations made before this evaluation were compared with the consensus recommendations made by the panel. RESULTS: For the 75 patients, the multidisciplinary panel disagreed with the treatment recommendations from the outside physicians in 32 cases (43%), and agreed in 41 cases (55%). Two patients (3%) had no treatment recommendation before consultation. For the 32 patients with a disagreement, the treatment recommendations were breast-conservation treatment instead of mastectomy (n = 13; 41%) or reexcision (n = 2; 6%); further workup instead of immediate definitive treatment (n = 10; 31%); treatment based on major change in diagnosis on pathology review (n = 3; 9%); addition of postmastectomy radiation treatment (n = 3; 9%); or addition of hormonal therapy (n = 1; 3%). CONCLUSIONS: The multidisciplinary breast cancer evaluation program provided an integrated program in which individual patients were evaluated by a team of physicians and led to a change in treatment recommendation for 43% (32 of 75) of the patients examined. This multidisciplinary program provided important second opinions for many patients with breast carcinoma.

    Title Calcium Ionophore Activation of Chronic Myelogenous Leukemia Progenitor Cells into Dendritic Cells is Mediated by Calcineurin Phosphatase.
    Date October 2000
    Journal Leukemia Research
    Excerpt

    We have previously demonstrated that Ph+ myeloid progenitor cells of patients with chronic myeloid leukemia (CML) can acquire characteristics of mature dendritic cells (DC) following calcium mobilization with calcium ionophore (A23187, CI). In this study we characterize the intracellular signaling pathway by which CI induces the acquisition of DC features in these leukemic cells. CI-induced activation of CML cells is attenuated by the calcineurin phosphatase inhibitor cyclosporin A (CsA) as well as the calmodulin (CaM) antagonist W-7. These cause ablation of both the CI-induced immunophenotypic expression of DC markers and immunostimulatory properties in mixed leukocyte responses (MLR). Minimal blocking effect was observed when Ca(2+)/CaM kinase II (281-301) inhibitor was added to the cultures. These findings suggest a Ca(2+)-dependent mechanism for the CI-induced activation of CML cells into antigen-presenting cells (APC), which is primarily mediated through the CaM/calcineurin pathway.

    Title Accuracy of Sentinel Lymph Node Biopsy in Patients with Large Primary Breast Tumors.
    Date September 2000
    Journal Cancer
    Excerpt

    BACKGROUND: Patients with large breast tumors are increasingly undergoing neoadjuvant treatment to downstage local disease; however, accurate staging of the axilla before the initiation of chemotherapy remains problematic. In the current study, the authors report on the accuracy of sentinel lymph node (SLN) biopsy in such patients to determine the feasibility of applying this technique before induction chemotherapy. METHODS: One hundred three patients with 104 tumors classified as American Joint Committee on Cancer (AJCC) T2 (tumor >/= 2 cm but </= 5 cm) or larger were recruited at the University of Pennsylvania and the Mayo Clinic. In the majority of cases, combined blue dye and radiotracer was used for SLN identification. After SLN identification, a completion axillary lymph node dissection was performed in 87 cases. The SLN was evaluated with hematoxylin and eosin and immunohistochemistry. RESULTS: The SLN was identified in 99% of cases. The overall rate of lymph node metastasis was 59% (95% exact confidence interval [95% CI], 49-68%) (61 of 104 cases). The SLN false-negative rate was 2% (95% exact CI, < 1-11.5%) (2 patients). In 56 tumors >/= 3 cm, 1 false-negative result (2% [95% exact CI, < 1-15%]) was identified, and the rate of lymph node metastasis was 62.5% (95% exact CI, 48. 5-75%) (35 of 56 tumors). Within 30 SLN positive patients with tumors >/= 3 cm and complete axillary lymph node dissection, 3 of 8 patients (37.5% [95% exact CI, 8.5-75.5%]) with micrometastasis (</= 2 mm) to the SLN had positive non-SLN compared with 21 of 22 patients (95.5% [95% exact CI, 77-100%]) with macrometastasis (> 2 mm) to the SLN (P = 0.002). CONCLUSIONS: SLN biopsy for patients with large breast tumors is technically feasible and highly accurate. SLN biopsy should be considered for the staging of clinically negative axilla in patients scheduled to receive neoadjuvant chemotherapy.

    Title Granulocyte-macrophage Colony-stimulating Factor, Interleukin-2, and Interleukin-12 Synergize with Calcium Ionophore to Enhance Dendritic Cell Function.
    Date August 2000
    Journal Journal of Immunotherapy (hagerstown, Md. : 1997)
    Excerpt

    The authors previously showed that monocytes treated with calcium ionophore (CI) acquire characteristics of mature dendritic cells (DC) in part through a calcineurin-dependent pathway. In this study, the authors evaluated the ability of granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin-2 (IL-2), and interleukin-12 (IL-12) alone or in combination with CI to induce DC characteristics in peripheral blood monocytes. Monocytes obtained by leukapheresis and countercurrent centrifugal elutriation were cultured with calcium, cytokines, or both, profiled by flow cytometry, and assessed for antigen uptake and sensitization of autologous CD8+ T cells to antigen. Monocytes treated with the combination of GM-CSF, IL-2, and IL-12 resulted in immunophenotypic and antigen uptake profiles typical of immature DC, including loss of surface CD14 expression, de novo low-level expression of B7.1, negligible CD83 expression, marked enhancement of CD40 and ICAM-1, and high major histocompatibility complex class I and II levels. A high level of antigen uptake by macro-pinocytosis was observed. In contrast, CI treatment significantly up-regulates B7.1, B7.2, CD40, CD54, and CD83 and substantially down-regulates CD14 and macro-pinocytosis, a profile consistent with mature DC. Many CI-induced modulations, but none resulting from cytokine treatment alone, were inhibited by the calcineurin phosphatase inhibitor cyclosporin A. Compared with monocytes treated with CI alone, combined treatment of monocytes with GM-CSF, IL-2, IL-12, and CI augmented B7.1 and CD83 expression and enhanced sensitization of autologous CD8+ T cells to melanoma-antigen-derived peptides. These results suggest that several independent pathways of DC activation can cooperatively enhance the function of monocyte-derived DC.

    Title Incidence of Sentinel Node Metastasis in Patients with Thin Primary Melanoma (< or = 1 Mm) with Vertical Growth Phase.
    Date July 2000
    Journal Annals of Surgical Oncology
    Excerpt

    BACKGROUND: Patients with thin primary melanomas (< or = 1 mm) generally have an excellent prognosis. However, the presence of a vertical growth phase (VGP) adversely impacts the survival rate. We report on the rate of occurrence of nodal metastasis in patients with thin primary melanomas with a VGP who are offered sentinel lymph node (SLN) biopsy. METHODS: Among 235 patients with clinically localized cutaneous melanomas who underwent successful SLN biopsy, 71 had lesions 1 mm or smaller with a VGP. The SLN was localized by using blue dye and a radiotracer. If negative for tumor by using hematoxylin and eosin staining, the SLN was further examined by immunohistochemistry. RESULTS: The rate of occurrence of SLN metastasis was 15.2% in patients with melanomas deeper than 1 mm and 5.6% in patients with thin melanomas. Three patients with thin melanomas and a positive SLN had low-risk lesions, based on a highly accurate six-variable multivariate logistic regression model for predicting 8-year survival in stage I/II melanomas. The fourth patient had a low- to intermediate-risk lesion based on this model. At the time of the lymphadenectomy, one patient had two additional nodes with metastasis. CONCLUSIONS: VGP in a melanoma 1 mm or smaller seems to be a risk factor for nodal metastasis. The risk of nodal disease may not be accurately predicted by the use of a multivariate logistic regression model that incorporates thickness, mitotic rate, regression, tumor-infiltrating lymphocytes, sex, and anatomical site. Patients with thin lesions having VGP should be evaluated for SLN biopsy and trials of adjuvant therapy when stage III disease is found.

    Title Mr Imaging in Patients with Nipple Discharge: Initial Experience.
    Date July 2000
    Journal Radiology
    Excerpt

    PURPOSE: To investigate the potential of magnetic resonance (MR) imaging in patients with nipple discharge. MATERIALS AND METHODS: Between February 1992 and December 1998, 23 patients with nipple discharge underwent contrast material-enhanced MR imaging at 1.5 T. Mammographic findings were negative in 22 of 23 patients and revealed asymmetry in one patient. Galactography was attempted in two patients, with negative findings in one patient and no success in the other. Fifteen of 23 patients underwent excisional biopsy-seven of 15 with MR imaging-guided localization, and one of 15 with mammographic localization. Eight of 23 patients were followed up clinically (range, 7-24 months; mean, 20 months). RESULTS: In 11 of the 15 (73%) patients who underwent excisional biopsy, MR imaging findings correlated with histopathologic findings. MR imaging demonstrated four of six benign papillomas and one of two fibroadenomas as circumscribed, enhancing subareolar masses. Findings of one MR imaging examination were negative, and benign tissue was found at excisional biopsy. MR imaging findings were suspicious in six of the seven patients with excisional biopsy findings of malignancy (regional enhancement [n = 2], ductal enhancement [n = 2], peripherally enhancing mass [n = 1], and spiculated mass [n = 1]). In one of the seven patients, a benign-appearing intraductal mass was identified at MR imaging; excisional biopsy revealed a benign papilloma with an adjacent focus of DCIS. CONCLUSION: MR imaging can help identify both benign and malignant causes of nipple discharge. It potentially offers a noninvasive alternative to galactography.

    Title Active Macromolecule Uptake by Lymph Node Antigen-presenting Cells: a Novel Mechanism in Determining Sentinel Lymph Node Status.
    Date May 2000
    Journal Annals of Surgical Oncology
    Excerpt

    BACKGROUND: Although sentinel lymph node (SLN) biopsy is a powerful staging tool for patients with melanoma and breast cancer, controversy remains regarding specific aspects of technique. We examined particle uptake by antigen-presenting cells (APCs) to determine if this mechanism is responsible for the differential retention of radioactivity in SLNs relative to nonsentinel lymph nodes (NSLNs). METHODS: Mapping was conducted in pigs injected with vital blue dye, fluoroscein isothiocyanate-labeled human serum albumin (FITC-HSA), and one of two 99mtechnetium-labeled tracers, i.e., human serum albumin, a small macromolecule, or unfiltered sulfur colloid, a mixture of small and large particles. Macromolecule uptake by APCs was studied in vitro by using FITC-HSA and measured by fluorescence-activated cell sorting (FACS). SLNs and NSLNs were analyzed by fluorescence microscopy or FACS, with counterstaining for leukocyte cell surface markers. RESULTS: Both radiotracers were effective. Cultured APCs rapidly took up FITC-HSA. Microscopy showed FITC-HSA in the subcapsular sinus of SLNs shortly after injection and subsequent distribution to interfollicular areas. FACS revealed increasing amounts of FITC-HSA in SLNs over time. Cells responsible for uptake were APCs, expressing major histocompatibility (locus) class II. CONCLUSIONS: This report establishes active macromolecule uptake as a mechanism that determines SLN status. This mechanism has important implications for performing SLN biopsy.

    Title Sentinel Lymph Node Biopsy with Metastasis: Can Axillary Dissection Be Avoided in Some Patients with Breast Cancer?
    Date January 2000
    Journal Journal of Clinical Oncology : Official Journal of the American Society of Clinical Oncology
    Excerpt

    PURPOSE: Recent studies have suggested that the sentinel lymph node (SLN) biopsy is an accurate alternative staging procedure for women with breast cancer. The goal of this study was to identify a subset of breast cancer patients in whom metastatic disease was confined only to the SLN. MATERIALS AND METHODS: From two institutions, we recruited 222 women with breast cancer for SLN biopsy. A SLN biopsy was performed in each patient, followed by an axillary dissection in 182 patients. Histologic and immunohistochemical cytokeratin stains were used on all SLNs. RESULTS: The SLN was identified in 220 (97. 8%) of the 225 biopsies. Evidence of metastatic breast cancer in the SLN was found in 60 (27.0%) of the 222 patients. Of these patients, 32 (53.3%) had evidence of tumor in the SLN only. By multivariate analysis, two factors were found to be significantly associated with a higher likelihood of tumor involvement in the non-SLNs: primary tumor size larger than 2.0 cm (P =.0004) and macrometastasis (> 2.0 mm) in the SLN (P =.002). Additional analysis revealed that none (0%; 95% confidence interval, 0% to 18.5%) of the 18 patients with primary tumors < or = 2.0 cm and micrometastasis to the SLN had remaining axillary lymph node involvement. CONCLUSION: The primary tumor size and metastasis size in the SLN are independent factors in predicting the incidence of tumor in the non-SLNs. Therefore, the SLN biopsy alone may be adequate for staging and/or therapy decision making in patients with primary breast tumors < or = 2.0 cm and micrometastasis in the SLN.

    Title Calcium Signaling Induces Acquisition of Dendritic Cell Characteristics in Chronic Myelogenous Leukemia Myeloid Progenitor Cells.
    Date October 1999
    Journal Proceedings of the National Academy of Sciences of the United States of America
    Excerpt

    Effective host T lymphocyte sensitization to malignant cells depends on successful antigen presentation. In this study, we examined the capacity of malignant myeloid progenitor cells of patients in the chronic phase of chronic myelogenous leukemia (CML) to acquire characteristics of activated dendritic cells (DCs) after intracellular calcium mobilization, thereby bypassing a need for third-party antigen-presenting cells. Treatment of purified CD33(+) CML cells from 15 patients with calcium ionophore (CI) consistently resulted in de novo expression of the costimulatory molecules CD80 (B7.1) and CD86 (B7.2), CD40 and the DC-specific activation marker CD83, as well as marked up-regulation of MHC class I and II molecules and the adhesion molecule CD54. Most of these changes occurred within 24 hr of treatment. Morphologically, CI-treated CML cells developed long dendritic projections similar to those seen in mature DCs. Functionally, CI-treated CML cells provided stimulation of allogeneic T lymphocytes 10- to 20-fold that of untreated CML cells or untreated monocytes. Fluorescent in situ hybridization of CI-activated CML cells confirmed their leukemic origin by displaying the typical bcr/abl fusion signal. No difference in bcr/abl translocation percentages between untreated and CI-treated CML nuclei was observed. These observations indicate that calcium mobilization may constitute a valuable approach for rapidly and reliably generating CML-derived DCs for immunotherapy of CML.

    Title Calcium Ionophore-treated Myeloid Cells Acquire Many Dendritic Cell Characteristics Independent of Prior Differentiation State, Transformation Status, or Sensitivity to Biologic Agents.
    Date September 1999
    Journal Blood
    Excerpt

    We previously reported that treatment of human peripheral blood monocytes or dendritic cells (DC) with calcium ionophore (CI) led to the rapid (18 hour) acquisition of many characteristics of mature DC, including CD83 expression. We therefore investigated whether less-mature myeloid cells were similarly susceptible to rapid CI activation. Although the promyelocytic leukemia line HL-60 was refractory to cytokine differentiation, CI treatment induced near-uniform overnight expression of CD83, CD80 (B7.1), and CD86 (B7. 2), as well as additional characteristics of mature DC. Several cytokines that alone had restricted impact on HL-60 could enhance CI-induced differentiation and resultant T-cell sensitizing capacity. In parallel studies, CD34(pos) cells cultured from normal donor bone marrow developed marked DC-like morphology after overnight treatment with either rhCD40L or CI, but only CI simultaneously induced upregulation of CD83, CD80, and CD86. This contrasted to peripheral blood monocytes, in which such upregulation could be induced with either CI or rhCD40L treatment. We conclude that normal and transformed myeloid cells at many stages of ontogeny possess the capacity to rapidly acquire many properties of mature DC in response to CI treatment. This apparent ability to respond to calcium mobilization, even when putative signal-transducing agents are inoperative, suggests strategies for implementing host antileukemic immune responses.

    Title 99mtc-human Serum Albumin: an Effective Radiotracer for Identifying Sentinel Lymph Nodes in Melanoma.
    Date July 1999
    Journal Journal of Nuclear Medicine : Official Publication, Society of Nuclear Medicine
    Excerpt

    Sentinel lymph node (SLN) biopsy has emerged as a novel approach for identifying patients with melanoma and regional nodal micrometastasis who may benefit from full nodal basin resection. To identify the pattern of tumor lymphatic drainage and the SLN, lymphoscintigraphy has been performed using primarily 99mTc-sulfur colloid (SC). In this study, we compare the efficacy of SLN biopsy using 99mTc-human serum albumin (HSA) with SLN biopsy after SC-based lymphoscintigraphy. METHODS: One hundred and six patients with localized cutaneous melanoma were studied. Lymphoscintigraphy was performed after intradermal injection of HSA in 85 patients and SC in 21 patients. Four patients underwent lymphoscintigraphy twice, once with SC and once with HSA. Dynamic images were acquired for up to 1 h, followed by high-count images of the SLN in various projections so that the most likely site was marked on the skin for biopsy. Intraoperatively, blue dye was injected around the primary site. Twenty-four patients underwent SLN dissection directed by preoperative lymphoscintigraphy and vital blue dye mapping; in the remaining 80 patients, a gamma probe was added intraoperatively to the localization procedure. Two patients underwent mapping with gamma probe alone. RESULTS: Draining lymphatic basins and nodes were identified by lymphoscintigraphy in all patients. The SLN was identified in 95% of patients when both blue dye and intraoperative gamma probe were used. When 99mTc-HSA was used for imaging, 98% of the SLNs ultimately identified were radiolabeled, and 82% were both hot and blue. Of the SLN recovered with SC, all the nodes were radiolabeled; however, there was only 58% hot and blue concordance. Greater numbers of SLNs were removed in the SC group (median 2.0 versus 1.0, P = 0.02); however, the incidence of micrometastasis was statistically similar in both HSA and SC cohorts. In the 4 patients examined with both tracers, SLN mapping was similar. CONCLUSION: Although SC has been the radiotracer of choice for SLN mapping in melanoma, HSA appears to be a suitable alternative, with identical success rates. In fact, the higher concordance between hot and blue nodes using HSA suggests superiority of this tracer for this purpose.

    Title Calcium Mobilization in Human Myeloid Cells Results in Acquisition of Individual Dendritic Cell-like Characteristics Through Discrete Signaling Pathways.
    Date July 1999
    Journal Journal of Immunology (baltimore, Md. : 1950)
    Excerpt

    We have shown previously that calcium ionophore (CI) treatment of various myeloid origin cells results in rapid acquisition of properties associated with mature, activated dendritic cells. These properties include increased CD83 and costimulatory molecule expression, tendencies to form dendritic processes, loss of CD14 expression by monocytes, and typically an enhanced capacity to sensitize T lymphocytes to Ag. We here analyze the intracellular signaling pathways by which CI induces acquisition of such properties. Thapsigargin, which raises intracellular Ca2+ levels by antagonizing its sequestration, induced immunophenotypic and morphologic changes that paralleled CI treatment. CI-induced activation was broadly attenuated by the Ca2+ chelating compound EGTA and by calmodulin antagonists trifluoperazine dimaleate and W-7. However, antagonists of signaling pathways downstream to calmodulin displayed more selective inhibitory effects. Calcineurin antagonists cyclosporin A and the FK-506 analogue, ascomycin, diminished costimulatory molecule and CD83 expression, as well as formation of dendritic processes in CI-treated myeloid cells, and strongly attenuated the T cell allosensitizing capacity of CI-treated HL-60 cells. These calcineurin antagonists displayed minimal effect on CI-induced CD14 down-regulation in monocytes. In contrast, the calmodulin-dependent protein kinase antagonists, K252a and KT5926, while displaying only modest effects on CI-induced costimulatory molecule and CD83 expression, strongly blocked CD14 down-regulation. These results are consistent with a Ca2+-dependent mechanism for CI-induced differentiation of myeloid cells, and indicate that multiple discrete signaling pathways downstream to calcium mobilization and calmodulin activation may be essential in regulating this process.

    Title Immunohistochemistry with Pancytokeratins Improves the Sensitivity of Sentinel Lymph Node Biopsy in Patients with Breast Carcinoma.
    Date April 1999
    Journal Cancer
    Excerpt

    BACKGROUND: Sentinel lymph node (SLN) biopsy is being investigated as a staging procedure for breast carcinoma. The authors evaluated whether immunohistochemical (IHC) analysis improves the sensitivity of this procedure. METHODS: Forty-four women with breast carcinoma were recruited for SLN biopsy. Preoperative lymphoscintigraphy was followed by intraoperative localization using a handheld gamma probe and blue dye. After SLN identification, an immediate complete axillary lymph node dissection was performed in all patients. All lymph nodes were subjected to routine histology (hematoxylin and eosin [H&E]) and IHC using antibody to cytokeratins. RESULTS: The SLN was identified in 41 of 43 patients (95%). Successful SLN identification was independent of biopsy technique (open surgical [95%] vs. fine-needle aspiration/core needle biopsy [96%]). Twelve of 41 patients (29%) had evidence of lymph node metastasis in the SLN by routine histology. Of the twenty-nine patients with H&E negative SLN, 3 were found to have metastasis by IHC for a conversion rate of 10%. Fifteen of 41 patients (37%) had evidence of metastasis in SLN. All 26 patients with H&E and IHC negative SLN had negative nonsentinel lymph nodes by routine histology and IHC (100% negative predictive value). All patients with tumors < 2 cm and micrometastasis to the SLN had no additional lymph node disease, in contrast to patients with lesions > 2 cm or patients with macrometastasis to the SLN (P = 0.007). CONCLUSIONS: These results confirm that SLN biopsy is extremely accurate for patients with breast carcinoma, even after open surgical biopsy. IHC analysis or serial sectioning of SLN improves the sensitivity of this staging technique.

    Title Calcium Ionophore-treated Peripheral Blood Monocytes and Dendritic Cells Rapidly Display Characteristics of Activated Dendritic Cells.
    Date November 1997
    Journal Journal of Immunology (baltimore, Md. : 1950)
    Excerpt

    Human peripheral blood contains a small subpopulation of immature dendritic cells (iDC) distinguished from circulating monocytes by their low expression of CD14. We utilized leukapheresis and countercurrent centrifugal elutriation to obtain myeloid origin mononuclear cell (MOMC) fractions of monocytes and iDC for study. These subpopulations were ultrastructurally and immunophenotypically similar before culture. After a 20- to 96-h culture either alone, with recombinant human granulocyte-monocyte CSF, or with endotoxin, greater up-regulation of costimulatory molecule expression was observed among iDC than among monocytes, and only iDC expressed the activation molecule CD83. Treatment with rhIL-4 caused many MOMC to develop morphologic properties of dendritic cells within 96 h, but costimulatory molecule up-regulation and CD14 down-regulation were heterogeneous, and CD83 expression was infrequent. In contrast, calcium ionophore (CI) treatment induced rapid and consistent effects in MOMC from both healthy volunteers and cancer patients, including down-regulated CD14 expression, acquisition of dendritic cell morphologic properties, up-regulated MHC and costimulatory molecule expression, and de novo CD83 expression. Many such effects occurred within 20 h of treatment. CI treatment activated purified CD14+ monocytes and also enhanced the spontaneous activation of purified CD14-/dim iDC in culture. Unfractionated MOMC, purified monocytes, and purified iDC displayed equivalently enhanced T cell-sensitizing efficiency following CI treatment. CD4+ T cell sensitization to keyhole limpet hemocyanin and CD8+ T cell sensitization to MART-1 melanoma-associated peptide were achieved in a single culture stimulation. Therefore, circulating monocytes and iDC can be induced by CI to manifest properties of activated DC, providing large numbers of efficient, nontransformed autologous APC for T cell sensitization strategies.

    Title Propagation of Mouse and Human T Cells with Defined Antigen Specificity and Function.
    Date August 1995
    Journal Ciba Foundation Symposium
    Excerpt

    Difficulties maintaining fully functional CD4+ T cells in culture have historically limited the study of their role in tumour rejection as well as other clinical applications. As the therapeutic value of current antitumour CD8+ T cell adoptive therapy becomes better defined, a strong impetus exists to determine optimal conditions for culturing antitumour CD4+ T cells. Our goal is to promote broadly polyclonal, antigen-specific CD4+ T cell responses of either Th1 or Th2 character for use in antitumour therapy or allograft facilitation, respectively. Similar obstacles exist in murine and human cultures: (1) during even brief periods of culture CD4+ T cells develop high 'background' reactivity to class II-positive antigen-presenting cells; (2) maintenance of antigen specificity as evidenced by cytokine secretion and short-term proliferation assays is insufficient to ensure bulk numerical expansion; (3) Th1-type CD4+ T cells often lose their potential for antigen-specific secretion of interleukin 2 on re-stimulation (though remain inducible by 12-O-tetradecanoylphorbol 13-acetate/ionomycin); (4) during prolonged culture selection pressure favours CD4+ subpopulations that recognize artifactual antigens such as culture medium proteins; (5) even with optimal culture conditions, cultured CD4+ T cells may function differently in vivo to uncultured CD4+ T cells. We have devised various strategies to surmount these obstacles by use of selected cytokines, antigen-presenting cells and timely culture manoeuvres.

    Title Tumor Promoters Stimulate the Formation of 1,2-diacylglycerol in Murine Peritoneal Macrophages: a Possible Mechanism for Stimulating Superoxide Anion Radical Production.
    Date January 1990
    Journal Cancer Letters
    Excerpt

    The formation of 1,2-diacylglycerol (DAG), a known stimulator of superoxide anion radical (O2-.) production in inflammatory cells, was assessed in murine peritoneal macrophages following treatment in vitro with tumor promoters. Addition of phorbol-12-myristate-13-acetate (PMA, 1-100 ng/ml) to resident peritoneal macrophage cultures from CD-1 female mice resulted in a 3- to 7-fold increase in [3H]DAG formation. The response was observed from 15 min to 2 h following the addition of PMA. At concentrations at which they stimulate O2-. production, PMA and other tumor promoters such as mezerein, phorbol-12,13-dibutyrate and 4-O-methyl-PMA stimulated the formation of [3H]DAG. Similar results were obtained when thioglycollate-elicited macrophages were used. Concurrent with the formation of [3H]DAG was a release of [3H]choline equivalents from the resident peritoneal macrophages treated with tumor promoters. The calcium ionophore A23187 did not stimulate O2-. production of [3H]DAG formation in resident peritoneal macrophages. These results demonstrate that tumor promoters stimulate the accumulation of DAGs in murine peritoneal macrophages at concentrations at which they stimulate O2-. production and suggest a mechanism by which tumor promoters such as mezerein, which are weak activators of protein kinase C, may indirectly stimulate O2-. production.

    Title Arachidonic Acid Potentiates Superoxide Anion Radical Production by Murine Peritoneal Macrophages Stimulated with Tumor Promoters.
    Date November 1989
    Journal Carcinogenesis
    Excerpt

    The role of arachidonic acid (AA) metabolism in the stimulation of oxygen radical production by murine peritoneal macrophages treated with tumor promoters was assessed. In vivo administration of the phospholipase A2 inhibitor dibromacetophenone, the anti-inflammatory steroid fluocinolone acetonide or the lipoxygenase inhibitor nordihydroguiaretic acid just prior to i.p. injection of phorbol-12-myristate-13-acetate (PMA, 100 ng) into unmanipulated CD-1 female mice resulted in a dose-dependent decrease in the number of peritoneal exudate cells (PEC) producing superoxide anion radical (O2) as assessed by the reduction of nitroblue tetrazolium, i.e. the formation of formazan-positive PEC. The cycloxygenase inhibitor indomethacin had no effect on the number of formazan-positive PEC caused by PMA treatment. The ability of PMA, phorbol-12,13-dibutyrate mezerein, phorbol-12,13-diacetate and 4-O-Me-PMA to stimulate the production of oxygen radicals by murine peritoneal macrophages correlated with their ability to stimulate the release of [3H]AA equivalents from the macrophages. The calcium ionophore A23187 which stimulated significant [3H]AA equivalent release did not stimulate superoxide anion radical production by the macrophages. PMA administered i.p. to SENCAR mice increased the number of formazan-positive PEC 4-to 5-fold compared with similarly treated C57BL/6 mice. PMA also stimulated the release of twice the amount of [3H]AA equivalents from peritoneal macrophages from SENCAR mice compared with that released by macrophages from C57BL/6 mice. The addition of low concentrations of AA (1-10 microM) in vitro to casein-elicited murine peritoneal macrophages treated with low concentrations of PMA (1 ng/ml) resulted in a 2-fold potentiation of the amount of superoxide anion radical produced compared with PMA treatment alone as assessed by the reduction of cytochrome c. These results demonstrate that AA and/or a lipoxygenase product can potentiate the production of oxygen radicals by murine peritoneal macrophages treated with tumor promoters.

    Title Tumor Promoters Differ in Their Ability to Stimulate Superoxide Anion Radical Production by Murine Peritoneal Exudate Cells Following in Vivo Administration.
    Date June 1989
    Journal Carcinogenesis
    Excerpt

    The production of superoxide anion radicals by murine peritoneal exudate cells (PECs) following intraperitoneal (i.p.) injection of mouse skin tumor promoters was assessed in vitro by the reduction of nitroblue tetrazolium (NBT). Phorbol-12-myristate-13-acetate (PMA) or mezerein when administered i.p. to CD-1 female mice at a dose of 0.1 microgram caused an inflammatory response in the peritoneal cavity. PMA (0.1 microgram) also caused an increase in the number of PEC reducing NBT, i.e. formazan-positive PECs (20% positive cells) as compared to vehicle control from 15 min through 2 h following i.p. administration to unmanipulated mice. In the same system, dose-response studies demonstrated that 4-O-methyl-phorbol myristate acetate (4-O-Me-PMA) and mezerein were approximately 50 times less active than PMA, while phorbol dibutyrate (PDB), phorbol diacetate (PDA), phorbol and the calcium ionophore A23187 did not cause an increase in the number of formazan-positive PECs at the doses used. When the same compounds (0.1 microgram) were administered to mice which had been pretreated i.p. with thioglycollate 6 days prior to the experiment, PMA, PDB, 4-O-Me-PMA, mezerein and A23187 all caused an increase in the number of formazan-positive PECs 2 h after treatment. The PEC which reduced NBT in response to tumor promoters were predominantly adherent and esterase positive cells, suggesting they were macrophages. These results indicate that in vivo administration of tumor promoters results in the production of superoxide anion radicals by murine peritoneal macrophages and that the physiological state of the macrophages is important in the response to tumor promoters.(ABSTRACT TRUNCATED AT 250 WORDS)

    Title Reengineering Dendritic Cell-based Anti-cancer Vaccines.
    Date
    Journal Immunological Reviews
    Excerpt

    Despite initial enthusiasm, dendritic cell (DC)-based anti-cancer vaccines have yet to live up to their promise as one of the best hopes for generating effective anti-tumor immunity. One of the principal reasons for the generally disappointing results achieved thus far could be that the full potential of DCs has not been effectively exploited. Here, we argue that dramatic improvements in vaccine efficacy will probably require a careful re-evaluation of current vaccine design. The formulation of new strategies must take into account the natural history of DCs, particularly their role in helping the immune system deal with infection. Equally critical is the emerging importance of soluble factors, notably interleukin-12, in modulating the quality of immune responses. Vaccines should also be designed to recruit helper T cells and antibody-producing B cells rather than simply cytotoxic T lymphocytes. Finally, the judicious selection of tumor, target antigen, and disease stage best suited for treatment should serve as the foundation of trial designs. Our discussion addresses a recent clinical vaccine trial to treat early breast cancer, where many elements of this new strategy were put into practice.

    Title Differential Production of Il-23 and Il-12 by Myeloid-derived Dendritic Cells in Response to Tlr Agonists.
    Date
    Journal Journal of Immunology (baltimore, Md. : 1950)
    Excerpt

    The recently delineated role for IL-23 in enhancing Th-17 activity suggests that regulation of its expression is distinct from that of IL-12. We hypothesized that independent TLR-mediated pathways are involved in the regulation of IL-12 and IL-23 production by myeloid-derived dendritic cells (DCs). The TLR 2 ligand, lipoteichoic acid (LTA), the TLR 4 ligand, LPS, and the TLR 7/8 ligand, resimiquod (R848), induced production of IL-23 by DCs. None of these TLR ligands alone induced significant IL-12 production, except when combined with IFN-gamma or other TLR ligands. Notably, IL-23 production in response to single TLR ligands was inhibited by IL-4. DCs treated with single TLR agonists induced IL-17A production by allogeneic and Ag-specific memory CD4(+) T cells, an effect that was abrogated by IL-23 neutralization. Moreover, these DCs stimulated IL-17A production by tumor peptide-specific CD8(+) T cells. In contrast, DCs treated with dual signals induced naive and memory Th1 responses and enhanced the functional avidity of tumor-specific CD8(+) T cells. These results indicate that distinct microbial-derived stimuli are required to drive myeloid DC commitment to IL-12 or IL-23 production, thereby differentially polarizing T cell responses.

    Title Effect of Multiple Activation Stimuli on the Generation of Th1-polarizing Dendritic Cells.
    Date
    Journal Human Immunology
    Excerpt

    It was originally reported that only a small fraction of total matured dendritic cells (DCs) produced interleukin (IL)-12, but it has never been determined whether different combinations of activating signals now shown to maximize secreted IL-12 do so through increasing output by the same IL-12 producers, or by recruiting additional cytokine-secreting cells. We therefore tested all combinations of bacterial lipopolysaccharide (LPS) (TLR4 ligand), R848 (TLR8 ligand), interferon (IFN)-γ, and CD40L for activating human monocyte-derived dendritic cells (DC), and determined by intracellular flow cytometry that enhanced IL-12 secretion was accomplished in large part by markedly increasing the proportion of cells producing IL-12, with the triple and quadruple combinations recruiting the most DC. This optimization requirement for multiple signals was not reflected in differential Toll-like receptor (TLR) expression by the cells. Interestingly, DCs activated with single TLR ligands plus IFN-γ were capable of responding with a second burst of IL-12 upon later CD40L stimulation, whereas DCs activated with R848 plus LPS were not, despite the trend of the latter for superior polarization of naive T cells toward IFN-γ-secreting Th1. These results have implications for the biology of IL-12-secreting DCs and choice of activation regimen for prospective use in DC-based immunotherapy.


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