Randomized controlled trial of the Valencia model of waking hypnosis plus CBT for pain, fatigue, and sleep management in patients with cancer and cancer survivors.

BACKGROUND: This study evaluated the efficacy of an intervention combining the Valencia model of waking hypnosis with cognitive-behavioral therapy (VMWH-CBT) in managing cancer-related pain, fatigue, and sleep problems in individuals with active cancer or who were post-treatment survivors. We hypothesized that four sessions of VMWH-CBT would result in greater improvement in participants' symptoms than four sessions of an education control intervention. Additionally, we examined the effects on several secondary outcome domains that are associated with increases in these symptoms (depression, pain interference, pain catastrophizing, and cancer treatment distress). METHODS: The study design was a randomized controlled crossover clinical trial comparing the VMWH-CBT intervention with education control. Participants (N = 44) received four sessions of both treatments, in a counterbalanced order (n = 22 per order condition). RESULTS: Participants were 89% female (N = 39) with mean age of 61 years (SD = 12.2). They reported significantly greater improvement after receiving the active treatment relative to the control condition in all the outcome measures. Treatment gains were maintained at 3-month follow-up. CONCLUSIONS: This study supports the beneficial effects of the VMWH-CBT intervention relative to a control condition and that treatment gains remain stable. VMWH-CBT-trained clinicians should be accessible for managing symptoms both during and after cancer treatment, though the findings need to be replicated in larger samples of cancer survivors.

SWOG S0800 (NCI CDR0000636131): addition of bevacizumab to neoadjuvant nab-paclitaxel with dose-dense doxorubicin and cyclophosphamide improves pathologic complete response (pCR) rates in inflammatory or locally advanced breast cancer.

SWOG S0800, a randomized open-label Phase II clinical trial, compared the combination of weekly nab-paclitaxel and bevacizumab followed by dose-dense doxorubicin and cyclophosphamide (AC) with nab-paclitaxel followed or preceded by AC as neoadjuvant treatment for HER2-negative locally advanced breast cancer (LABC) or inflammatory breast cancer (IBC). Patients were randomly allocated (2:1:1) to three neoadjuvant chemotherapy arms: (1) nab-paclitaxel with concurrent bevacizumab followed by AC; (2) nab-paclitaxel followed by AC; or (3) AC followed by nab-paclitaxel. The primary endpoint was pathologic complete response (pCR) with stratification by disease type (non-IBC LABC vs. IBC) and hormone receptor status (positive vs. negative). Overall survival (OS), event-free survival (EFS), and toxicity were secondary endpoints. Analyses were intent-to-treat comparing bevacizumab to the combined control arms. A total of 215 patients were accrued including 11 % with IBC and 32 % with triple-negative breast cancer (TNBC). The addition of bevacizumab significantly increased the pCR rate overall (36 vs. 21 %; p = 0.019) and in TNBC (59 vs. 29 %; p = 0.014), but not in hormone receptor-positive disease (24 vs. 18 %; p = 0.41). Sequence of administration of nab-paclitaxel and AC did not affect the pCR rate. While no significant differences in OS or EFS were seen, a trend favored the addition of bevacizumab for EFS (p = 0.06) in TNBC. Overall, Grade 3-4 adverse events did not differ substantially by treatment arm. The addition of bevacizumab to nab-paclitaxel prior to dose-dense AC neoadjuvant chemotherapy significantly improved the pCR rate compared to chemotherapy alone in patients with triple-negative LABC/IBC and was accompanied by a trend for improved EFS. This suggests reconsideration of the role of bevacizumab in high-risk triple-negative locally advanced breast cancer.

Germline genetic variants in ABCB1, ABCC1 and ALDH1A1, and risk of hematological and gastrointestinal toxicities in a SWOG Phase III trial S0221 for breast cancer.

Hematological and gastrointestinal toxicities are common among patients treated with cyclophosphamide and doxorubicin for breast cancer. To examine whether single-nucleotide polymorphisms (SNPs) in key pharmacokinetic genes were associated with risk of hematological or gastrointestinal toxicity, we analyzed 78 SNPs in ABCB1, ABCC1 and ALDH1A1 in 882 breast cancer patients enrolled in the SWOG trial S0221 and treated with cyclophosphamide and doxorubicin. A two-SNP haplotype in ALDH1A1 was associated with an increased risk of grade 3 and 4 hematological toxicity (odds ratio=1.44, 95% confidence interval=1.16-1.78), which remained significant after correction for multiple comparisons. In addition, four SNPs in ABCC1 were associated with gastrointestinal toxicity. Our findings provide evidence that SNPs in pharmacokinetic genes may have an impact on the development of chemotherapy-related toxicities. This is a necessary first step toward building a clinical tool that will help assess risk of adverse outcomes before undergoing chemotherapy.

The role of bisphosphonates as adjuvant therapy for breast cancer.

Bone is the most common site of distant recurrence in breast cancer. The development of skeletal metastases involves complex interactions between the cancer cells and the bone microenvironment. The presence of tumor in bone is associated with activation of osteoclasts, resulting in excessive bone resorption. Bisphosphonates are potent inhibitors of osteoclastic bone resorption with proven efficacy in reducing tumor-associated skeletal complications. Several studies have investigated the adjuvant, or preventive, use of these drugs in breast cancer. Laboratory experiments have shown that the development of bone metastases can be inhibited by bisphosphonates. Three randomized clinical trials of bisphosphonates in nonmetastatic breast cancer patients have yielded conflicting results with respect to development of osseous and visceral metastases and survival. Defining the potential role of these agents in adjuvant breast cancer treatment requires further investigation in randomized, large-scale, multicenter clinical trials. The data available to date provide a strong impetus for continued clinical and laboratory work with bisphosphonates in breast cancer.

Relation of body mass index to tumor markers and survival among young women with invasive ductal breast carcinoma.

BACKGROUND: Obesity has been shown to affect breast carcinoma prognosis, with the heaviest women having a higher mortality due to breast carcinoma. Few studies have focused on premenopausal women or the correlation of body mass index (BMI) to tumor characteristics related to prognosis. METHODS: The authors conducted a population-based follow-up study for mortality of 1177 women younger than 45 years of age who had invasive ductal breast carcinoma diagnosed from 1983 through 1992. Histologic slides and/or tumor tissue were collected for pathologic review, immunohistochemistry assays, and bivariate flow cytometric analysis. RESULTS: Women with breast carcinoma who were in the highest quartile of BMI were 2.5 times as likely (95% confidence interval [CI], 1.6-3.9) to die of their disease within 5 years of diagnosis compared with women in the lowest quartile of BMI. The tumors of the women in the highest quartile of BMI were more likely to be estrogen receptor negative (odds ratio [OR], 1.5; 95% CI, 1.0-2.2) and to have a high S-phase fraction (OR, 1.9; 95% CI, 1.2-3.1), high histologic grade (OR, 1.7; 95% CI, 1.0-2.9), high mitotic cell count (OR, 2.0; 95% CI, 1.2-3.1), and large tumor size (2 to < 5 cm: OR, 2.3; 95% CI, 1.5-3.1; or > or = 5 cm: OR, 2.7; 95% CI, 1.5-4.8) compared with the tumors of women whose BMI was in the first quartile. Relative to the large tumors (> or = 2 cm) in women in the lowest BMI quartile, the large tumors in women in the highest BMI quartile were more likely to express markers of high proliferation, indicating they may have grown faster than similar size tumors of the thinnest women. In a multivariate analysis including the tumor characteristics, obesity, as measured by being in the highest quartile of BMI, remained an independent prognostic factor for mortality (hazard ratio, 1.7; 95% CI, 1.0-2.9; P < 0.05. CONCLUSIONS: Our study results indicated that being in the highest quartile of BMI was a strong predictor of mortality in women with breast carcinoma diagnosed at a young age. The tumors of the heavy women were larger and more likely to have markers of high cellular proliferation than those of thinner women.

18fluorodeoxyglucose positron emission tomography to detect mediastinal or internal mammary metastases in breast cancer.

PURPOSE: To determine the prevalence of suspected disease in the mediastinum and internal mammary (IM) node chain by 18fluorodeoxyglucose (FDG) positron emission tomography (PET), compared with conventional staging by computed tomography (CT) in patients with recurrent or metastatic breast cancer. PATIENTS AND METHODS: We retrospectively evaluated intrathoracic lymph nodes using FDG PET and CT data in 73 consecutive patients with recurrent or metastatic breast cancer who had both CT and FDG PET within 30 days of each other. In reviews of CT scans, mediastinal nodes measuring 1 cm or greater in the short axis were considered positive. PET was considered positive when there were one or more mediastinal foci of FDG uptake greater than the mediastinal blood pool. RESULTS: Overall, 40% of patients had abnormal mediastinal or IM FDG uptake consistent with metastases, compared with 23% of patients who had suspiciously enlarged mediastinal or IM nodes by CT. Both FDG PET and CT were positive in 22%. In the subset of 33 patients with assessable follow-up by CT or biopsy, the sensitivity, specificity, and accuracy for nodal disease was 85%, 90%, and 88%, respectively, by FDG PET; 54%, 85%, and 73%, respectively, by prospective interpretation of CT; and 50%, 83%, and 70%, respectively, by blinded observer interpretation of CT. Among patients suspected of having only locoregional disease recurrence (n = 33), 10 had unsuspected mediastinal or IM disease by FDG PET. CONCLUSION: FDG PET may uncover disease in these nodal regions not recognized by conventional staging methods. Future prospective studies using histopathology for confirmation are needed to validate the preliminary findings of this retrospective study.

[18F]fluoroestradiol radiation dosimetry in human PET studies.

UNLABELLED: [18F]16alpha-fluoroestradiol (FES) is a PET imaging agent useful for the study of estrogen receptors in breast cancer. We estimated the radiation dosimetry for this tracer using data obtained in patient studies. METHODS: Time-dependent tissue concentrations of radioactivity were determined from blood samples and PET images in 49 patients (52 studies) after intravenous injection of FES. Radiation absorbed doses were calculated using the procedures of the MIRD committee, taking into account the variation in dose based on the distribution of activities observed in the individual patients. Effective dose equivalent was calculated using International Commission on Radiological Protection Publication 60 weights for the standard woman. RESULTS: The effective dose equivalent was 0.022 mSv/MBq (80 mrem/mCi). The organ that received the highest dose was the liver (0.13 mGy/MBq [470 mrad/mCi]), followed by the gallbladder (0.10 mGy/MBq [380 mrad/mCi]) and the urinary bladder (0.05 mGy/MBq [190 mrad/mCi]). CONCLUSION: The organ doses are comparable to those associated with other commonly performed nuclear medicine tests. FES is a useful estrogen receptor-imaging agent, and the potential radiation risks associated with this study are well within accepted limits.

Concurrent radiation therapy and paclitaxel or docetaxel chemotherapy in high-risk breast cancer.

PURPOSE: Evidence supports the inclusion of the taxanes in the treatment of breast cancer. A recent randomized trial has shown a survival advantage to the addition of paclitaxel in the adjuvant treatment of node-positive patients. Several studies have suggested diminished local control if adjuvant radiation is delayed, while in vitro and in vivo studies have demonstrated a benefit of concurrent administration of taxanes with radiation. For these reasons, we began in 1995 to administer radiation therapy concurrently with the taxanes in advanced breast cancer. This retrospective review examines the feasibility of such treatment. METHODS AND MATERIALS: Forty-four patients were treated with concurrent radiation and either paclitaxel (29 patients) or docetaxel (15 patients). One patient received both paclitaxel and docetaxel. Eighteen patients were treated for recurrent disease, 9 had received prior radiation. Toxicity was assessed by the RTOG scale for acute and late effects. RESULTS: Concurrent radiation and taxane chemotherapy was well tolerated. Nine patients (20%) experienced Grade 3 acute skin toxicity. This was more likely with docetaxel than paclitaxel (p = 0. 04). Among patients undergoing breast conservation, there were no Grade 3 toxicities. With a median follow-up of 11 months, 1 patient has developed breast fibrosis. CONCLUSION: Concurrent administration of both paclitaxel and docetaxel with radiation resulted in acceptable toxicity. Overall, the acute skin toxicity seen with docetaxel was more pronounced. However, among patients undergoing breast conservation the taxanes were both well tolerated. Further study is necessary to assess the impact of concurrent treatment on long-term outcome.

Dendritic cells lose ability to present protein antigen after stimulating antigen-specific T cell responses, despite upregulation of MHC class II expression.

Immature dendritic cells (DC) take up, process and present protein antigens; mature DC are specialized for stimulating primary T cell responses with increased expression of MHC class II and co-stimulatory molecules, but are incapable of processing and presenting soluble protein. The current study examined whether maturation of DC is triggered by T cell recognition of antigens presented by immature DC. Human DC derived from CD34+ progenitor cells by culture with granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-6 (IL-6) in serum-free medium could prime naive CD4+ T cells to keyhole limpet hemocyanin (KLH) and ovalbumin (OVA). The cultured DC retained the ability to prime T cells to native protein for at least 15 days. To test for changes in DC function after participation in an immune response, DC were co-cultured with either allogeneic or autologous CD4+ T cells. DC co-cultured with autologous T cells retained the ability to prime T cells to intact protein antigens. By contrast, DC which had previously stimulated an allogeneic T cell response lost ability to prime T cells to soluble proteins. However, such <> induced a MLR and stimulated peptide-specific primary CD4+ T cell responses. This indicated that <> did not die or lose the ability to prime, but lost the ability to process and present subsequent antigens. Following participation in T cell activation, DC increased surface expression of MHC class II, co-stimulatory molecules CD40 and B7.2, and the intercellular adhesion molecule-1 (ICAM-1). In addition, our data suggest that interferon gamma (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) are involved in this T cell-mediated DC maturation.

Monitoring the response of patients with locally advanced breast carcinoma to neoadjuvant chemotherapy using [technetium 99m]-sestamibi scintimammography.

BACKGROUND: Mammographic and physical examination assessments of the response of locally advanced breast carcinoma (LABC) to neoadjuvant therapy have been shown to be inaccurate. The authors studied the feasibility and accuracy of [technetium 99m]-sestamibi (MIBI) for monitoring the response of patients with LABC to neoadjuvant chemotherapy. METHODS: Patients receiving neoadjuvant chemotherapy for LABC underwent prone lateral scintimammography before therapy, after 2 months of therapy, and close to the completion of chemotherapy (presurgery) if chemotherapy continued for >3 months. Images were analyzed both qualitatively and quantitatively using the lesion-to-normal breast MIBI uptake ratio (L:N). Imaging results were compared with the clinical response and the pathologic response as determined from the posttherapy surgical specimen. RESULTS: A total of 32 patients (29 who were assessable for primary tumor response and 28 who were assessable for lymph node response) were included in the study. The mean change in the primary tumor L:N MIBI uptake ratio after 2 months of chemotherapy was -35% for clinical responders and +17% for nonresponders (P<0.001). Patients achieving a pathologic primary tumor macroscopic complete response (CR) had a mean change in uptake on the presurgical scan of -58% versus -18% for patients with a partial response (P<0.005). A decrease of > or =40% in the MIBI uptake ratio identified CRs with 100% sensitivity and 89% specificity. Pretherapy imaging predicted axillary lymph node metastases in 85% of patients ultimately found to have > or =1 positive lymph nodes at surgery, but was less accurate in identifying residual lymph node disease after therapy (55% sensitivity and 75% specificity). CONCLUSIONS: MIBI imaging accurately assessed the response to neoadjuvant chemotherapy in patients with LABC. Further studies are needed to determine the role of MIBI in this group of patients.

Infusional paclitaxel and weekly vinorelbine chemotherapy with concurrent filgrastim for metastatic breast cancer: high complete response rate in a phase I-II study of doxorubicin-treated patients.

PURPOSE: We investigated 96-hour paclitaxel infusion combined with weekly (days eight and 15) vinorelbine as salvage therapy for metastatic breast cancer in anthracycline-exposed patients. All patients received scheduled support with granulocyte colony-stimulating factor (G-CSF; filgrastim). Tumor response, toxicity, time to progression (TTP), and survival were assessed. PATIENTS AND METHODS: This single-center nonrandomized trial enrolled 32 patients. Anthracycline exposure and subsequent progression were common to all patients. Paclitaxel and vinorelbine were escalated over three dosing levels, stratified by liver function. RESULTS: Seven patients (22%) achieved a complete response and nine patients achieved a partial response for an overall response rate of 50%. The median TTP was 6.1 months, and median survival time was 14.1 months. Dose-limiting toxicity was neutropenia, with dose delay or reduction in seven of 32 patients. Febrile neutropenia requiring hospitalization was uncommon (three of 32 patients; 9%). There were no treatment-related deaths. Grade 3/4 thrombocytopenia occurred in two patients (6%), and 13 patients (41%) required RBC transfusions for anemia. Grade 3 nausea and vomiting was seen in one patient, who was found to be Addisonian. Despite potentially overlapping neurologic toxicities of the two agents, only two patients (6%) were removed from the study because of progressive peripheral neuropathy. CONCLUSION: Administration of 96-hour paclitaxel infusion and subsequent weekly vinorelbine with G-CSF support is well tolerated. The response rate, TTP, and survival data are encouraging for therapy given to anthracycline pretreated patients with metastatic breast cancer. If these results can be verified in multi-institution trials, this or a similar combination of drugs would merit investigation as first-line therapy in this patient population.

Value of combined FDG PET and MR imaging in the evaluation of suspected recurrent local-regional breast cancer: preliminary experience.

PURPOSE: To assess the performance and potential clinical effects of combined 2-[fluorine 18]fluoro-2-deoxy-D-glucose (FDG) positron emission tomography (PET) and magnetic resonance (MR) imaging of the axilla and brachial plexus in patients suspected of having local-regional breast cancer metastases. MATERIALS AND METHODS: Upper-body FDG PET and axillary and supraclavicular MR imaging were performed in 10 patients (age range, 45-71 years) with clinical findings suggestive of breast cancer metastases. Medical records were reviewed retrospectively. Imaging findings were correlated with clinical data and follow-up findings in all patients. Surgical findings were available in four patients. RESULTS: Nine patients had local-regional breast cancer metastases. MR imaging was diagnostic for tumor in five patients and was indeterminate in four patients with axillary or chest wall metastases. With FDG PET, metastatic tumor was positively identified in all nine patients. MR imaging was useful for determining the relationship of metastatic tumor to axillary and supraclavicular neurovascular structures. FDG PET helped confirm metastases in patients with indeterminate MR imaging findings and depicted unsuspected metastases outside the axilla. CONCLUSION: MR imaging and FDG PET are complementary in detecting and characterizing local-regional breast cancer metastases. Combined FDG PET and MR imaging provide useful treatment-planning data for patients clinically suspected of having recurrent axillary or supraclavicular breast cancer.

High-titer HER-2/neu protein-specific antibody can be detected in patients with early-stage breast cancer.

PURPOSE: To evaluate HER-2/neu-specific antibody immunity in patients with breast cancer, to determine the rate of occurrence of serum antibodies to HER-2/neu in patients with breast cancer, and to relate the presence of specific immunity to overexpression of HER-2/neu protein in primary tumor. METHODS: The antibody response to HER-2/neu protein was analyzed in 107 newly diagnosed breast cancer patients. Sera was analyzed for the presence of HER-2/neu-specific antibodies with a capture enzyme-linked immunosorbent assay (ELISA) and verified by Western blot. Sera from 200 volunteer blood donors was used as a control population. RESULTS: The presence of antibodies to HER-2/neu correlated with the presence of breast cancer. HER-2/neu antibodies at titers of > or = 1:100 were detected in 12 of 107 (11%) breast cancer patients versus none of 200 (0%) normal controls (P < .01). The presence of antibodies to HER-2/neu also correlated to overexpression of HER-2/neu protein in the patient's primary tumor. Nine of 44 (20%) patients with HER-2/neu-positive tumors had HER-2/neu-specific antibodies, whereas three of 63 (5%) patients with HER-2/neu-negative tumors had antibodies (P = .03). The antibody responses could be substantial. Titers of greater than 1:5,000 were detected in five of 107 (5%). CONCLUSION: The presence of HER-2/neu antibodies in breast cancer patients and the correlation with HER-2/neu-positive cancer implies that immunity to HER-2/neu develops as a result of exposure of patients to HER-2/neu protein expressed by their own cancer. These findings should stimulate further studies to develop the detection of immunity to oncogenic proteins as tumor markers, as well as the development and testing of vaccine strategies to induce and augment immunity to HER-2/neu for the treatment of breast cancer or prevention of recurrent disease.

Induction of multiple anti-c-erbB-2 specificities accompanies a classical idiotypic cascade following 2B1 bispecific monoclonal antibody treatment.

The bispecific monoclonal antibody (bsmAb) 2B1, targeting the extracellular domain of c-erbB-2, the protein product of the HER-2/neu proto-ocogene, and Fc gamma RIII (CD16), expressed by human natural killer cells, neutrophils and differentiated monocytes, mediates the specific cytotoxic activity of these effector cells to tumor cells. A group of 24 patients with c-erbB-2-overexpressing tumors were treated with intravenously administered 2B1 in a phase I clinical trial and followed after treatment to evaluate the diversity and extent of the 2B1-induced humoral immune responses. As expected, 17 of 24 patients developed human anti-(murine Ig) antibodies (HAMA) to whole 2B1 IgG in a range from 100 ng/ml to more than 50000 ng/ml; 10 of these patients (42%) had strong (at least 1000 ng/ml) HAMA responses, some of which were still detectable at day 191. These responses were usually associated with similar reactivity to the F(ab')2 fragments of the parental antibodies 520C9 (anti-c-erbB-2) and 3G8 (anti-CD16). We sought evidence of an idiotypic cascade induction, indicating a prolonged specific treatment-induced effect on at least one selected target of 2B1. Using competition-based enzyme-linked immunosorbent assays, specific anti-idiotypic antibodies (Ab2) were detectable against 520C9 in 11 patients and against 3G8 in 13 patients. Peak anti-idiotypic antibodies generally occurred 3-5 weeks from treatment initiation, with a downward trend thereafter. There was a statistically significant correlation among the induction of significant HAMA responses, anti-idiotypic antibody production and the development of antibodies to c-erbB-2. The anti-c-erbB-2 responses, which were distinct from anti-anti-idiotypic (Ab3) antibodies, were detected in the post-treatment sera of 6/16 patients examined. No obvious correlation could be made between the development of humoral immune responses, the dose received, and the clinical response. Future investigation involving 2B1 therapy will concentrate on investigating an association of these humoral responses to any c-erbB-2-specific cellular responses. Manipulations of 2B1 therapy effects that augment immunity to c-erbB-2 could provide additional avenues for immunotherapy with this and other bispecific antibodies.

Dose-intensive vinorelbine with concurrent granulocyte colony-stimulating factor support in paclitaxel-refractory metastatic breast cancer.

PURPOSE: We evaluated weekly single-agent intravenous (IV) vinorelbine as salvage therapy for metastatic breast cancer. After the first five patients, all received elective growth factor support with granulocyte colony-stimulating factor (G-CSF; filgrastim) in an attempt to maximize delivered dose-intensity (DDI). Objective tumor response, DDI, and toxicity were assessed, as well as time to progression (TTP) and survival. PATIENTS AND METHODS: This single-center nonrandomized trial enrolled 40 patients. Anthracycline exposure and subsequent progression were common to all patients, and 38 of 40 were paclitaxel-refractory. Vinorelbine was given initially at 30 mg/m2/wk, then at 35 mg/m2/wk in a phase I/II design, which involved first intermittent (6 days of 7) and then continuous (daily) administration of G-CSF at 5 micrograms/kg. RESULTS: The maximum-tolerated starting dose was 35 mg/m2/wk with continuous G-CSF support. The mean DDI was 27.7 mg/m2/wk for all patients. There were two complete responses (CRs) and eight partial responses (PRs) in 40 assessable patients for an overall response rate of 25% (95% confidence interval [CI], 13% to 41%). The median TTP was 13 weeks and median survival time 33 weeks. The dose-limiting toxicity was neutropenia, with dose delay or reduction required in 14 of 27 patients entered at 35 mg/m2. Febrile neutropenia that required hospitalization was unusual (three of 40 patients, 8%). There were no treatment-related deaths. Grade 3/4 thrombocytopenia occurred in nine patients (23%) and 26 patients (65%) required RBC transfusions for anemia. Seven patients (18%) had reversible grade 3/4 nonhematologic complications, primarily related to neurotoxicity. Grade > or = 3 mucositis was absent. CONCLUSION: Concurrent administration of weekly vinoralbine and daily G-CSF is feasible and permits an increase in DDI for vinorelbine of 43% to 76% over that reported in series without growth factor support. The response rate, TTP, and survival data are encouraging for therapy given to heavily pretreated patients with metastatic breast cancer. Vinorelbine is not cross-resistant with paclitaxel and should be considered for further trials in the dose-intensified mode made possible by G-CSF, alone or combined with other agents.

Granulocyte-macrophage colony-stimulating factor: an effective adjuvant for protein and peptide-based vaccines.

The current studies evaluate granulocyte-macrophage colony-stimulating factor (GM-CSF) as a vaccine adjuvant. An important issue for developing vaccine therapy for human malignancy is identifying adjuvants that can elicit T-cell responses to proteins and peptides derived from "self" tumor antigens. GM-CSF, in vitro, stimulates the growth of antigen-presenting cells such as dendritic cells and macrophages. Initial experiments examined whether GM-CSF injected into the skin of rats could affect the number or character of antigen presenting cells, measured as class II major histocompatability complex expressing cells, in lymph nodes draining the injection site. Intradermal (id) inoculation of GM-CSF every 24 hours for a total of five inoculations resulted in an increase of class II+ fluorescing cells that peaked at the fourth inoculation. Subcutaneous (sq) inoculation resulted in an increase of class II+ fluorescing cells that peaked following the second inoculation, then decreased over time. Using this schema for "conditioning" the inoculation site, GM-CSF was administered id or sq for five injections and a foreign antigen, tetanus toxoid (tt), was given at the beginning or the end of the immunization cycle. Id immunization was more effective than sq at eliciting tt specific immunity. In addition, GM-CSF id, administered as a single dose with antigen, compared favorably with complete Freund's adjuvant (CFA) and alum in eliciting tt specific antibody and cellular immunity. We have shown that immunity to rat neu (c-erbB-2) protein, an oncogenic self protein, can be generated in rats by immunization with peptides derived from the normal rat neu sequence plus CFA. The current study demonstrates that rat neu peptides inoculated with GM-CSF could elicit a strong delayed type hypersensitivity reaction (DTH) response, whereas peptides alone were non-immunogenic. GM-CSF was as effective as CFA in generating rat neu specific DTH responses after immunization with a neu peptide based vaccine. Soluble GM-CSF is a potent adjuvant for the generation of immune responses to foreign proteins as well as peptides derived from a self tumor antigen.

Peptide-based, but not whole protein, vaccines elicit immunity to HER-2/neu, oncogenic self-protein.

HER-2/neu, an overexpressed oncogenic protein, has been proposed as a human cancer vaccine target. HER-2/neu is a "self" protein, however, and methods of vaccine strategies that would be effective in immunizing patients to a "self" tumor Ag have not been established. Many of the tumor Ags defined in humans are nonmutated self proteins, e.g., MAGE, and overcoming tolerance may be key in the generation of effective anti-tumor immunity. One theory states that tolerance to self proteins is directed only to dominant epitopes of proteins and not to every portion of the protein. Accordingly, tolerance can be circumvented by immunization to peptide fragments, but not whole protein. The studies outlined here demonstrate rat neu-specific immunity could be elicited in rats by vaccination with immunogenic rat neu peptides, but not by immunization with the intact protein. A rat model was used since rat neu protein is 89% homologous to human HER-2/neu protein and has a similar tissue distribution and level of expression. Rats were immunized with groups of peptides derived from the amino acid sequence of the intracellular domain or extracellular domain of rat neu protein and both groups developed CD4+ T cell immunity and Ab immunity to rat neu peptides and protein. Animals immunized in a similar fashion with intact purified rat neu protein did not develop Ab or T cell immunity to rat neu. Furthermore, rats that developed neu-specific immunity showed no histopathologic evidence of autoimmunity directed against organs expressing basal levels of rat neu protein. These studies suggest an immunization strategy that might be effective in human cancer vaccines targeting self tumor Ag.

Generation of immunostimulatory dendritic cells from human CD34+ hematopoietic progenitor cells of the bone marrow and peripheral blood.

Dendritic antigen-presenting cells are considered to be the most effective stimulators of T cell immunity. The use of dendritic cells has been proposed to generate therapeutic T cell responses to tumor antigens in cancer patients. One limitation is that the number of dendritic cells in peripheral blood is exceedingly low. Dendritic cells originate from CD34+ hematopoietic progenitor cells (HPC) which are present in the bone marrow and in small numbers in peripheral blood. CD34+ HPC can be mobilized into the peripheral blood by in vivo administration of granulocyte-colony-stimulating factor. The aim of the current study was to determine whether functional dendritic cells could be elicited and grown in vitro from CD34+ HPC derived from bone marrow or granulocyte-colony-stimulating factor-mobilized peripheral blood. Culture of CD34+ HPC with granulocyte-macrophage-colony-stimulating factor and tumor necrosis factor alpha yielded a heterogeneous cell population containing cells with typical dendritic morphology. Phenotypic studies demonstrated a loss of the CD34 molecule over 1 week and an increase in cells expressing surface markers associated with dendritic cells, CD1a, CD80 (B7/BB1), CD4, CD14, HLA-DR, and CD64 (Fc gamma RI). Function was validated in experiments showing that cultured cells could stimulate proliferation of allogeneic CD4+ and CD8+ T lymphocytes. Antigen-presenting capacity was further confirmed in experiments showing that cultured cells could effectively stimulate tetanus toxoid-specific responses and HER-2/neu peptide-specific responses. The derivation and expansion of dendritic cells from cultured bone marrow or granulocyte-colony-stimulating factor-mobilized CD34+ HPC may provide adequate numbers for testing of dendritic cells in clinical studies, such as vaccine and T cell therapy trials.