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Background: Male breast cancer accounts for < 1% of breast cancers. Sentinel lymph node (SLN) operation is commonly used in the evaluation of female breast cancer patients. The purpose of this study was to determine whether SLN operation is as feasible and accurate in male patients compared with female patients., Study Design: Between 1999 and 2005, 30 men and 2,784 women underwent SLN operation. Clinical and pathologic data were reviewed and statistical analysis performed., Results: Men presented at an older age (p = 0.005) and with larger tumors than women (p = 0.04). The SLN was identified in 100% of men and in 98.3% of women (p = NS). The mean number of SLNs harvested was 3.5 in men and 3.0 in women (p = NS). The incidence of positive SLNs was higher in men (37.0% versus 22.3%), although this did not reach statistical significance (p = 0.1). In patients with a positive SLN there were additional non-SLNs positive in 62.5% of men, compared with 20.7% in women (p = 0.01). The median size of the largest lymph node metastasis was 10 mm in men and 3 mm in women (p = 0.03)., Conclusions: SLN operation in clinically node-negative men is feasible and accurate. Male breast cancer patients present at an older age and with larger tumors than female breast cancer patients. Male patients have higher nodal tumor burden reflected in a larger size of nodal metastasis and increased risk of harboring additional disease in axillary lymph nodes when the SLN is positive. Intraoperative SLN evaluation should be considered in the surgical management of male breast cancer.

Background: The risk of developing breast cancer rises with increasing age. The very elderly population (80 years of age and greater) is often excluded from both clinical trials and retrospective analyses. We performed a retrospective review of the very elderly population treated at our institution in order to assess treatment patterns and the safety of therapy in an older population., Data Sources: In this institutional experience at a comprehensive cancer center, we retrospectively reviewed the charts of patients 80 years and older diagnosed with a new breast cancer between September 1, 1989, and September 1, 2004., Results: Two hundred thirteen patients were identified for this study. Median age was 83 (range 80-97). Median survival was 7.28 years, with a median follow up of 4 years for patients still alive at the end of the study period. Ninety-eight percent of patients (208/213) received 1 or more components of recommended multimodality treatment. Five patients refused all treatment. Overall, complications affected 12% of patients who received treatment (26/208). There were 2 deaths, 1 after surgery and 1 related to chemotherapy. The majority, 69% (18/26), of the documented complications were classified as minor. Surgery resulted in complications in 6% (11/188) of patients. Five percent (5/112) of patients who received radiation suffered adverse effects. Chemotherapy-related complications affected 30% (6/18) of treated patients. Hormonal agents resulted in complications in 3% (3/112) of patients. No correlation between the American Society of Anesthesiologists score and incidence of complication was observed (P = .58)., Conclusions: Very elderly patients can be safely treated with surgery and radiation in accordance with accepted recommendations for their stage of breast cancer. Treatment with surgery and/or radiation should be considered despite age and moderate comorbidity in order to affect locoregional control. Chemotherapy results in a significant incidence of complications and should be cautiously implemented in this age group. A prospective trial is necessary to assess the necessity of aggressive multimodality therapy in this very elderly population.

Background: Breast-conservation therapy (BCT), including wide local excision and postoperative irradiation, is considered standard treatment for early-stage invasive ductal carcinoma (IDC). The use of BCT in patients with invasive lobular carcinoma (ILC) has been questioned because of concerns regarding ipsilateral breast recurrence and risk of bilateral breast cancer. We evaluated our institutional experience with BCT and compared treatment outcomes for ILC with those for IDC., Methods: A review of our BCT database revealed 84 patients with ILC and 1,126 with IDC with stage I or II disease treated with BCT and radiation between 1976 and 1999. We evaluated local-regional recurrence, disease-specific survival, and contralateral breast cancer rates in both groups., Results: The 5- and 10-year local-regional recurrence rates for the ILC group were 1% and 7%, respectively, and 4% and 9%, respectively, for the IDC group (P = .70). There were no significant differences in the 5- and 10-year disease-specific survival rates between the groups. Contralateral breast cancer occurred in 11.3% of patients with IDC and 11.9% of patients with ILC., Conclusions: BCT achieves similar local-regional control and survival outcomes in selected patients with ILC or IDC. Breast-conservation therapy is an appropriate treatment strategy for patients with early-stage invasive lobular carcinoma.

Background: Patients with invasive cancer identified at the time of prophylactic mastectomy (PM) will require axillary lymph node dissection for staging; therefore, many surgeons advocate sentinel lymph node (SLN) surgery at the time of PM. The current study investigates the invasive cancer rate in PM and evaluates factors associated with invasive cancer to guide SLN surgery use., Methods: Patients undergoing PM at the M. D. Anderson Cancer Center between January 2000 and July 2005 were identified from a prospective database. Clinical, radiographic, and pathologic data were collected., Results: A total of 409 patients (436 PM cases) were identified; 382 underwent contralateral PM (CPM) and 27 underwent bilateral PM (BPM). Cancer was identified in 22 of 436 PM cases (5%). Of these, 14 patients (64%) had ductal carcinoma in situ (DCIS). Only 8 patients (1.8%) had invasive cancer, with a mean tumor size of 5 mm (range, 2-9 mm). There was no difference in the occult cancer rate between CPM and BPM. No cases of invasive cancer were identified in the 23 patients with BRCA mutations. Significantly increased risk of invasive cancer in the PM breast was seen in postmenopausal patients (3.7%; P = .007), patients age >60 years (7.5%; P = .008), and patients with history of invasive lobular carcinoma (9.7%; P = .0002) or lobular carcinoma in situ (LCIS) (7.7%; P = .008)., Conclusions: The frequency of cancer in PM is very low and the majority represents DCIS. Therefore, routine use of SLN surgery in all patients undergoing PM is not warranted. However, patients at higher risk for whom SLN surgery should be considered include older women and patients with a history of lobular cancer or LCIS., ((c) 2006 American Cancer Society.)

Objective: To determine the effect of preoperative chemotherapy on the volume of tissue excised and the number of breast operations in patients undergoing breast-conserving therapy (BCT)., Summary Background Data: Preoperative chemotherapy is increasingly being used for breast cancer and increases rates of BCT. Its impact on the extent of surgery and the number of surgical procedures in BCT has never been fully defined. The extent of surgery in BCT directly affects cosmesis., Methods: We reviewed the records of 509 consecutive patients with T1-T3, N0-N2 breast cancer who were treated in prospective randomized clinical trials of chemotherapy between 1998 and 2005. We analyzed the final surgical procedure (BCT or mastectomy), the number of operations, and, in patients who underwent BCT, re-excision rates, and the total volume of breast tissue excised [4Pi/3(width/2 x length/2 x height/2)]., Results: A total of 241 patients underwent BCT, and 268 patients underwent mastectomy. Among BCT patients who had initial tumor size >2.0 cm, patients who received preoperative chemotherapy had significantly smaller volumes of breast tissue excised compared with patients who received postoperative chemotherapy (113 cm vs. 213 cm, P = 0.004). The re-excision rate and total number of breast operations did not significantly differ between the groups. Among BCT patients who had initial tumor size < or = 2 cm, preoperative chemotherapy had no impact on volume of breast tissue excised, re-excision rate, or number of breast operations (P > 0.05)., Conclusions: Among patients treated with BCT for larger breast tumors, patients treated with preoperative chemotherapy have less extensive resection, with no change in rates of re-excision.

The anti-HER2 antibody trastuzumab (Herceptin) has been used to treat patients with breast cancers that overexpress HER2. We have demonstrated that p27(Kip1) upregulation is one of the key events that cause G(1) arrest upon trastuzumab treatment. Here, we have examined the effect of trastuzumab on expression of CDK2, Rb, E2F, NPAT and histone H4 in breast cancer cells that overexpress HER2. Trastuzumab treatment dramatically inhibited the kinase activity and expression of CDK2, whereas the kinase activity and expression of CDK4 were not affected. Unlike the p27(Kip1) upregulation that occurs primarily through post-translational mechanisms, CDK2 was downregulated primarily at a transcriptional level as shown by Northern blotting and real-time RT-PCR analyses. With a decrease in CDK2 activity, trastuzumab decreased the kinase activity of cyclin E but had little effect on cyclin E protein level. Overexpression of wild-type cyclin E or its lower molecular weight forms did not influence the response to trastuzumab. Levels and activities of CDK6, cyclin A, and cyclin D1 were all suppressed by trastuzumab. As a result, trastuzumab inhibited Rb phosphorylation that associates with CDK2, cyclin E, CDK6, cyclin A, or cyclin D1. As predicted from these changes, trastuzumab decreased the DNA-binding activity of E2F, decreased the level of NPAT protein, and decreased the level of histone H4 mRNA. Blockade of the PI3K pathway with LY294002 produced similar effects to trastuzumab treatment on expression of each of these genes. Taken together, treatment of breast cancer cells that overexpress HER2 with the anti-HER2 antibody trastuzumab inhibits CDK2, Rb phosphorylation, E2F activity, NPAT, and histone H4 via PI3K signaling that are needed for both DNA and histone synthesis during progression from G(1) phase to S phase of the cell cycle.

Background: Patients with locally recurrent breast cancer can also harbor regional nodal disease. Sentinel lymph node (LN; SLN) surgery is an accepted method for LN evaluation in early-stage breast cancer. We sought to evaluate the feasibility of SLN surgery in patients with locally recurrent breast cancer., Patients and Methods: Patients undergoing SLN surgery at the time of treatment for recurrent breast cancer were identified. Clinical, pathologic, and operative details of initial and recurrent breast cancer events were analyzed., Results: From 2001 to 2005, 21 patients underwent SLN surgery for locally recurrent breast cancer. Previous breast procedures included segmental mastectomy in 17 patients and mastectomy in 4 patients. Previous axillary procedures included axillary LN dissection in 12 patients, SLN surgery in 5 patients, and no axillary surgery in 4 patients. Twelve patients had received breast irradiation. The SLN was identified and excised in 13 patients (62%). Six patients had drainage to nodal basins outside of the axilla (internal mammary and/or contralateral axilla). As the number of axillary nodes removed at primary surgery increased, the incidence of alternative lymphatic drainage increased: no previous axillary surgery, 0; 1-10 LNs removed, 30%; > 10 LNs removed, 50%., Conclusion: Sentinel LN surgery is feasible in patients with locally recurrent breast cancer regardless of previous axillary node surgery or radiation. The incidence of alternative lymphatic drainage is increased in patients with > 10 axillary LNs removed at original operation or when radiation was part of the previous treatment. Sentinel LN surgery is a tool for guiding local-regional management of patients with locally recurrent breast cancer.

Purpose: Few biologic markers have been studied as prognostic factors in recurrent rectal carcinoma patients. We sought to determine the influence of clinical, pathologic, and biologic (p53, bcl-2, and ki-67) variables on survival after curative resection of locally recurrent rectal cancer., Methods: Retrospective review of patients with locally recurrent rectal cancer who received surgery with curative intent., Results: From 1988 to 1998, 134 patients with locally recurrent rectal cancer underwent operative exploration. Curative resection was performed in 85 patients. Median follow-up was 43 (range, 1.3-149) months. On multivariate analysis, negative predictors of overall survival included an elevated carcinoembryonic antigen level (P=0.02; hazard ratio 2.41; 95 percent confidence interval, 1.19-4.89) and an R1 resection margin (P = 0.01; hazard ratio, 2.81; 95 percent confidence interval, 1.27-6.21). In 26 patients for whom biologic variables were available, p53, bcl-2, and ki-67 did not significantly impact disease-specific survival or overall survival. Five-year disease-specific survival, overall survival, and pelvic control rates were 46, 36, and 51 percent respectively. Of the 50 patients who relapsed, time to second local recurrence was longer than time to development of metastasis (median, 16.5 vs. 9 months). Median survival for patients with metastatic recurrence was 26.l vs. 41.5 months for those with a subsequent local recurrence alone., Conclusions: Approximately two-thirds of patients with locally recurrent rectal cancer can be resected for cure. Preoperative carcinoembryonic antigen and an R0 resection margin were the only significant predictors of overall survival. p53, bcl-2, and ki-67 did not impact survival outcomes.

We show that atypical PKCiota, which plays a critical role in the establishment and maintenance of epithelial cell polarity, is genomically amplified and overexpressed in serous epithelial ovarian cancers. Furthermore, PKCiota protein is markedly increased or mislocalized in all serous ovarian cancers. An increased PKCiota DNA copy number is associated with decreased progression-free survival in serous epithelial ovarian cancers. In a Drosophila in vivo epithelial tissue model, overexpression of persistently active atypical PKC results in defects in apical-basal polarity, increased Cyclin E protein expression, and increased proliferation. Similar to the Drosophila model, increased PKCiota proteins levels are associated with increased Cyclin E protein expression and proliferation in ovarian cancers. In nonserous ovarian cancers, increased PKCiota protein levels, particularly in the presence of Cyclin E, are associated with markedly decreased overall survival. These results implicate PKCiota as a potential oncogene in ovarian cancer regulating epithelial cell polarity and proliferation and suggest that PKCiota is a novel target for therapy.

Background: In patients with breast carcinoma, ipsilateral breast tumor recurrence (IBTR) after breast-conserving therapy (BCT) is an independent predictor of systemic recurrence and disease-specific survival (DSS). However, only a subgroup of patients with IBTR develop systemic recurrences. Therefore, the management of isolated IBTR remains controversial. The objective of the current study was to identify determinants of systemic recurrence and DSS after IBTR., Methods: The medical records of 120 women who underwent BCT for Stage 0-III breast carcinoma between 1971 and 1996 and who subsequently developed isolated IBTR were reviewed. Clinicopathologic factors were studied using univariate and multivariate analyses for their association with DSS and the development of systemic recurrence after IBTR., Results: The median time to IBTR was 59 months. At a median follow-up of 80 months after IBTR, 45 patients (37.5%) had a systemic recurrence. Initial lymph node status was the strongest predictor of systemic recurrence according to the a univariate analysis (P = 0.001). Other significant factors included lymphovascular invasion (LVI) in the primary tumor, time to IBTR < or = 48 months, clinical and pathologic IBTR tumor size > 1 cm, LVI in the recurrent tumor, and skin involvement at IBTR. In a multivariate logistic regression analysis, initially positive lymph node status (relative risk [RR], 5.3; 95% confidence interval [95% CI], 1.4-20.1; P = 0.015) and skin involvement at IBTR (RR, 15.1; 95% CI, 1.5-153.8; P = 0.022) remained independent predictors of systemic recurrence. The 5-year and 10-year DSS rates after IBTR were 78% and 68%, respectively. In a multivariate Cox proportional hazards model analysis, only LVI in the recurrent tumor was found to be an independent predictor of DSS (RR, 4.6; 95% CI, 1.5-14.1; P = 0.008)., Conclusions: Patients who initially had lymph node-positive disease or skin involvement or LVI at IBTR represented especially high-risk groups that warranted consideration for aggressive, systemic treatment and novel, targeted therapies after IBTR. Determinants of prognosis after IBTR should be taken into account when evaluating the need for further systemic therapy and designing risk-stratified clinical trials., ((c) 2005 American Cancer Society.)

The low molecular weight (LMW) isoforms of cyclin E are unique to cancer cells. In breast cancer, such alteration of cyclin E is a very strong predictor of poor patient outcome. Here we show that alteration in binding properties of these LMW isoforms to CDK2 and the CDK inhibitors (CKIs), p21 and p27, results in their functional hyperactivity. The LMW forms of cyclin E are severalfold more effective at binding to CDK2. Additionally, compared with the full-length cyclin E-CDK2 complexes, the LMW cyclin E-CDK2 complexes are significantly more resistant to inhibition by p21 and p27, despite equal binding of the CKIs to the LMW complexes. When both the full-length and the LMW cyclin E are co-expressed, p27 preferentially binds to the LMW forms yet is unable to inhibit the CDK2 activity. Thus, the LMW forms of cyclin E may contribute to tumorigenesis through their resistance to the inhibitory activities of p21 and p27 while sequestering these CKIs from the full-length cyclin E.

We have previously shown that the low molecular weight (LMW) forms (trunk 1 and trunk 2) of cyclin E are biochemically hyperactive and induce G1/S progression in normal epithelial cells. Here we investigate the biologic consequences of LMW cyclin E expression in ovarian cancer cells. Using a panel of ovarian carcinoma tumors we find that cyclin E overexpression is invariably due to the presence of LMW forms and that expression of these forms appears to correlate with more advanced grade and stage of disease. Despite similar expression of p21 and p27, cyclin E overexpressing tumors have higher kinase function. Using an isogenic ovarian cancer model, we find that clones that overexpress the trunk 1 (T1) protein have a 10-fold increase in cyclin E kinase function, a 20% increase in S-phase fraction, a 10-15% decrease in doubling time and a 20% increase in colony formation compared to parental cells that express only the FL cyclin E protein. T1 clones were resistant to G1 arrest but more sensitive to cisplatin. Therefore, in ovarian tumors, the presence of LMW cyclin E forms confers altered biologic properties. Our data provides a potential mechanism for the poor prognosis of patients with LMW cyclin E expressing tumors.

Background: Ultrasonography and fine-needle aspiration (FNA) are used to evaluate the breast and regional nodes in breast cancer patients. We sought to identify factors influencing the sensitivity of ultrasonography for detection of nodal metastasis., Methods: Patients with a clinically negative axilla who underwent axillary ultrasonography and sentinel lymph node biopsy were included., Results: Of 208 patients, axillary ultrasonography was negative in 180 (86%) and suspicious or indeterminate in 28 (14%). FNA was performed in 22 patients whose findings were indeterminate or suspicious, and 3 were positive for malignancy. Final pathological examinations revealed positive nodes in 53 patients: 39 (22%) of 180 with negative ultrasonographic findings and 14 (50%) of 28 with indeterminate or suspicious ultrasonographic findings (P =.001). Excisional biopsy was more common for patients with indeterminate or suspicious findings on preoperative ultrasonography (P =.038). There were no significant differences in tumor size, histological features, size of nodal metastasis, or number of positive nodes between patients whose ultrasonography findings were negative and those whose findings were indeterminate or suspicious., Conclusions: Ultrasonographically suggested nodal metastasis is associated with the finding of nodal disease on final pathological examination. No significant clinicopathologic criteria were found to impact sensitivity of ultrasonography; however, excisional biopsy for diagnosis may be a confounding variable in subsequent axillary ultrasonography.

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.

Cyclin E in complex with CDK2 is a positive regulator of the G1 to S phase transition of the cell cycle and is responsible for cells passing the restriction point, committing the cell to another round of cell division. Cyclin E is overexpressed and proteolytically cleaved into low molecular weight (LMW) isoforms in breast cancer cell lines and tumor tissues compared to normal cells and tissues. These alterations in cyclin E are linked to poor prognosis in breast cancer patients. In order to evaluate the biological effects of the LMW cyclin E, immortalized mammary epithelial cells, 76NE6, were stably transfected with each of the three cyclin E constructs. Our results reveal that the LMW forms of cyclin E (T1 and T2) are biologically functional, as their overexpression in the immortalized cells increases the ability of these cells to enter S and G2/M phase by 2 fold over full length or vector-alone transfected cells, concomitant with an increased rate of cell proliferation. In addition, these LMW isoforms are biochemically hyperactive, shown by their ability to phosphorylate substrates such as histone H1 4 fold more in cells transfected with T1 or T2 versus cells transfected with the full length form. These results suggest that overexpression of the LMW forms of cyclin E is mitogenic, stimulating the cells to progress through the cell cycle much more efficiently than the full length cyclin E.

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.

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., (Copyright 2003 American Cancer Society.DOI 10.1002/cncr.11490)

Over the past 30 years, several important surgery clinical trials examining the issues of excision margins, ELND, and regional therapy have given rise to the current practice guidelines for the care of melanoma patients. The development of the SLN selective lymphadenectomy concept and the increasing use of molecular staging techniques have presented new challenges in this field. Clinical trials have been initiated to address these latest issues. In the next decade, these trials should help guide the integration of the new technologies into the surgical management of melanoma.

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.

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.

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.

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.

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.

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.

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.

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.

Unlabelled: 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.

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.

Interleukin 1 (IL-1) is a primary mediator of a wide variety of immunologic and inflammatory responses, including reactions to microbial infections. To study this cytokine in an animal model, we have developed specific and sensitive radioimmunoassays for the quantitation of rabbit IL-1 alpha and IL-1 beta. The sensitivity (limit of detection at 95% confidence level) of our assay for IL-1 alpha and 1 beta was 20-40 and 40-80 pg/ml, respectively. Recovery of IL-1 from tissues ranged from 75 to 107%, with a mean of 95% for IL-1 alpha and 89% (range 19-98) for IL-1 beta. We employed these assays in in vivo and in vitro studies. In an in vivo model, we measured the amount of rabbit IL-1 alpha and 1 beta protein present in brain, kidney, liver, lung, muscle, and spleen at various times after the injection of endotoxin. IL-1 was found in all tissues studied but largely in the spleen; IL-1 levels were transient, reaching peak levels by 4 h after injection of endotoxin and rapidly decreasing to low levels by 24 h. In similar in vitro studies, IL-1 alpha levels reached peak elevation 6 h after addition of endotoxin, whereas IL-1 beta was maximal at 24 h. IL-1 alpha was detected in all tissues; IL-1 beta was observed primarily in lung, kidney, and spleen. These studies establish the presence of IL-1 in various tissues during endotoxemia.

Taurolidine (Geistlich Pharm, AG, Wolhusen, Switzerland), a derivative of the amino acid taurine, is commonly used in some parts of the world as an adjunctive therapy for various infections. Its mechanism of action is thought to be related to its antimicrobial properties, including its ability to interfere with some of the biological activities of endotoxin (lipopolysaccharide, LPS). For example, taurolidine has been shown to protect animals against endotoxic shock and death. In this study we examined the ability of taurolidine to block LPS-induced tumor necrosis factor (TNF) and interleukin 1 (IL-1) synthesis in human peripheral blood mononuclear cells (PBMC) from 27 donors. We observed a dose-dependent reduction in the synthesis of these two cytokines when taurolidine was preincubated with LPS before being added to PBMC. This reduction was independent of the molar ratio of taurolidine to LPS but was related to the concentration of taurolidine present in the PBMC cultures. There was a 80 to 90% reduction in total IL-1 and TNF synthesis induced by LPS at concentrations of taurolidine of 40 to 100 micrograms/mL; the vehicle was without effect. Following a 30-min preincubation with PBMC, taurolidine could be washed from the cells and still suppress cytokine synthesis induced by LPS. Using release of lactic acid dehydrogenase, 100 micrograms/mL of taurolidine was not toxic for PBMC. Taurolidine also reduced IL-1 and TNF synthesis induced by the Staphylococcus aureus-derived toxic shock syndrome toxin-1 as well as that induced by nontoxic heat-killed Staphylococcus epidermidis organisms.(ABSTRACT TRUNCATED AT 250 WORDS)