Induction of robust type-I CD8+ T-cell responses in WHO grade 2 low-grade glioma patients receiving peptide-based vaccines in combination with poly-ICLC.

PURPOSE: WHO grade 2 low-grade gliomas (LGG) with high risk factors for recurrence are mostly lethal despite current treatments. We conducted a phase I study to evaluate the safety and immunogenicity of subcutaneous vaccinations with synthetic peptides for glioma-associated antigen (GAA) epitopes in HLA-A2(+) adults with high-risk LGGs in the following three cohorts: (i) patients without prior progression, chemotherapy, or radiotherapy (RT); (ii) patients without prior progression or chemotherapy but with prior RT; and (iii) recurrent patients. EXPERIMENTAL DESIGN: GAAs were IL13Rα2, EphA2, WT1, and Survivin. Synthetic peptides were emulsified in Montanide-ISA-51 and given every 3 weeks for eight courses with intramuscular injections of poly-ICLC, followed by q12 week booster vaccines. RESULTS: Cohorts 1, 2, and 3 enrolled 12, 1, and 10 patients, respectively. No regimen-limiting toxicity was encountered except for one case with grade 3 fever, fatigue, and mood disturbance (cohort 1). ELISPOT assays demonstrated robust IFNγ responses against at least three of the four GAA epitopes in 10 and 4 cases of cohorts 1 and 3, respectively. Cohort 1 patients demonstrated significantly higher IFNγ responses than cohort 3 patients. Median progression-free survival (PFS) periods since the first vaccine are 17 months in cohort 1 (range, 10-47+) and 12 months in cohort 3 (range, 3-41+). The only patient with large astrocytoma in cohort 2 has been progression-free for more than 67 months since diagnosis. CONCLUSION: The current regimen is well tolerated and induces robust GAA-specific responses in WHO grade 2 glioma patients. These results warrant further evaluations of this approach. Clin Cancer Res; 21(2); 286-94. ©2014 AACR.

Antigen-specific immune responses and clinical outcome after vaccination with glioma-associated antigen peptides and polyinosinic-polycytidylic acid stabilized by lysine and carboxymethylcellulose in children with newly diagnosed malignant brainstem and nonbrainstem gliomas.

PURPOSE: Diffuse brainstem gliomas (BSGs) and other high-grade gliomas (HGGs) of childhood carry a dismal prognosis despite current treatments, and new therapies are needed. Having identified a series of glioma-associated antigens (GAAs) commonly overexpressed in pediatric gliomas, we initiated a pilot study of subcutaneous vaccinations with GAA epitope peptides in HLA-A2-positive children with newly diagnosed BSG and HGG. PATIENTS AND METHODS: GAAs were EphA2, interleukin-13 receptor alpha 2 (IL-13Rα2), and survivin, and their peptide epitopes were emulsified in Montanide-ISA-51 and given every 3 weeks with intramuscular polyinosinic-polycytidylic acid stabilized by lysine and carboxymethylcellulose for eight courses, followed by booster vaccinations every 6 weeks. Primary end points were safety and T-cell responses against vaccine-targeted GAA epitopes. Treatment response was evaluated clinically and by magnetic resonance imaging. RESULTS: Twenty-six children were enrolled, 14 with newly diagnosed BSG treated with irradiation and 12 with newly diagnosed BSG or HGG treated with irradiation and concurrent chemotherapy. No dose-limiting non-CNS toxicity was encountered. Five children had symptomatic pseudoprogression, which responded to dexamethasone and was associated with prolonged survival. Only two patients had progressive disease during the first two vaccine courses; 19 had stable disease, two had partial responses, one had a minor response, and two had prolonged disease-free status after surgery. Enzyme-linked immunosorbent spot analysis in 21 children showed positive anti-GAA immune responses in 13: to IL-13Rα2 in 10, EphA2 in 11, and survivin in three. CONCLUSION: GAA peptide vaccination in children with gliomas is generally well tolerated and has preliminary evidence of immunologic and clinical responses. Careful monitoring and management of pseudoprogression is essential.

Phase II study of induction fixed-dose rate gemcitabine and bevacizumab followed by 30 Gy radiotherapy as preoperative treatment for potentially resectable pancreatic adenocarcinoma.

BACKGROUND: Eighty percent of patients with resected pancreatic ductal carcinoma (PDC) experience treatment failure within 2 years. We hypothesized that preoperative fixed-dose rate (FDR) gemcitabine (GEM) combined with the angiogenesis inhibitor bevacizumab (BEV) and accelerated 30 Gy radiotherapy (RT) would improve outcomes among patients with potentially resectable PDC. METHODS: This phase II trial tested induction FDR GEM (1,500 mg/m(2)) plus BEV (10 mg/kg IV) every 2 weeks for three cycles followed by accelerated RT (30 Gy in 10 fractions) plus BEV directed at gross tumor volume plus a 1-2 cm vascular margin. Subjects underwent laparoscopy and resection after day 85. Therapy was considered effective if the complete pathologic response rate exceeded 10 % and the margin-negative resection rate exceeded 80%. RESULTS: Fifty-nine subjects were enrolled; 29 had potential portal vein involvement. Two grade 4 (3.4%) and 19 grade 3 toxicities (32.8%) occurred. Four subjects manifested radiographic progression, and 10 had undetected carcinomatosis. Forty-three pancreatic resections (73%) were performed, including 19 portal vein resections (44%). Margin-negative outcomes were observed in 38 (88%, 95% confidence interval [CI] 75-96), with one complete pathologic response (2.3%; 95% CI 0.1-12). There were seven (6 grade 3; 1 grade 4) wound complications (13%). Median overall survival for the entire cohort was 16.8 months (95% CI 14.9-21.3) and 19.7 months (95% CI 16.5-28.2) after resection. CONCLUSIONS: Induction therapy with FDR GEM and BEV, followed by accelerated BEV/RT to 30 Gy, was well tolerated. Although both effectiveness criteria were achieved, survival outcomes were equivalent to published regimens.

LOH in the HLA class I region at 6p21 is associated with shorter survival in newly diagnosed adult glioblastoma.

PURPOSE: Glioblastoma (GBM) shows downregulated expression of human leukocyte antigen (HLA) class I, thereby escaping from cytotoxic T cells and limiting the efficacy of immunotherapy. Loss of heterozygosity (LOH) of HLA class I (6p21) and/or ß-2 microglobulin (B2m) (15q21) regions represents irreversible downregulation. In this study, we examined the prevalence of these LOH events and their relations with overall survival in GBM. EXPERIMENTAL DESIGN: In a cross-sectional analysis on 60 adult patients with GBM, DNA from formalin-fixed, paraffin-embedded specimens were evaluated for 10 microsatellite regions of HLA class I, B2m, HLA class II, HLA class III, and 6q by PCR as well as immunohistochemical evaluation of HLA class I expression and CD8(+) T-cell infiltration. RESULTS: LOH in HLA class I, B2m, HLA class II, HLA class III, and 6q regions was present in 41.4%, 18.2%, 9.4%, 77.8%, and 36.0% of informative cases, respectively. LOH of HLA class I was associated with shorter overall survival (HR = 4.89, P = 0.0078). HLA class I was downregulated in 22% to 43% of cases based on immunohistochemistry. Cases that displayed negative staining were significantly younger. HLA class I expression correlated with intratumoral CD8(+) T-cell infiltration. CONCLUSION: LOH in the HLA class I region is frequent in adult GBMs. The association of shorter survival with LOH in this region suggests a crucial role for these genes in immunosurveillance.

MUC1 vaccine for individuals with advanced adenoma of the colon: a cancer immunoprevention feasibility study.

Cancer vaccines based on human tumor-associated antigens (TAA) have been tested in patients with advanced or recurrent cancer, in combination with or following standard therapy. Their immunogenicity and therapeutic efficacy has been difficult to properly evaluate in that setting characterized by multiple highly suppressive effects of the tumor and the standard therapy on the patient's immune system. In animal models of human cancer, vaccines administered in the prophylactic setting are most immunogenic and effectively prevent cancer development and progression. We report results of a clinical study that show that in patients without cancer but with a history of premalignant lesions (advanced colonic adenomas, precursors to colon cancer), a vaccine based on the TAA MUC1 was highly immunogenic in 17 of 39 (43.6%) of vaccinated individuals, eliciting high levels of anti-MUC1 immunoglobulin G (IgG) and long-lasting immune memory. Lack of response in 22 of 39 individuals was correlated with high levels of circulating myeloid-derived suppressor cells (MDSC) prevaccination. Vaccine-elicited MUC1-specific immune response and immune memory were not associated with significant toxicity. Our study shows that vaccines based on human TAAs are immunogenic and safe and capable of eliciting long-term memory that is important for cancer prevention. We also show that in the premalignant setting, immunosuppressive environment (e.g., high levels of MDSC) might already exist in some individuals, suggesting an even earlier premalignant stage or preselection of nonimmunosuppressed patients for prophylactic vaccination.

Pitfalls in retrospective analyses of biomarkers: a case study with metastatic melanoma patients.

BACKGROUND: Reliable prognostic biomarkers of survival and response to treatment are clearly important in oncology, and many studies have been carried out with the objective of identifying new prognostic biomarkers. Retrospective analysis of blood banked from patients is a frequently used paradigm for these studies. We describe a new study of the association of serum biomarker level with overall survival in melanoma patients, and the problems encountered in carrying it out. METHODS: Blood samples from 56 patients with stage IV metastatic melanoma were drawn prior to initiation of any treatment for their disease. Sera from the samples were stored for up to 94 months at -80°C, and were subsequently thawed at the same time and tested by multiplex Luminex assay for 30 analytes (cytokines, chemokines and growth factors). Cox regression analysis was used to assess the association between these analytes and time-to-death. RESULTS: Of the 30 analytes, 17 were associated with survival, most strongly so, and in all cases, a higher analyte level was associated with increased survival. In addition, the correlations of the levels of all possible pairs of analytes were all positive and in almost all cases highly significant. However, these results are artifacts that arise from the combination of two peculiarities of the data: the apparent decrease in analyte level with storage time, and the uniformly shorter storage times of the samples from censored patients than the storage times of the samples from patients who died. CONCLUSIONS: All retrospective studies can have hidden biases, and thus investigators should not claim new findings before examining the data in detail with the goal of determining whether the findings could be spurious. There were several suspicious findings in our initial analyses: too many analytes found significant, too many very small p-values, a uniformly positive association of analyte level with survival, and a uniformly positive correlation between analyte levels. We were convinced that these findings must be artifacts, and further analyses showed that the findings could be explained by an apparent decrease of analyte level storage time.

Phase II study of irinotecan and cetuximab given every 2 weeks as second-line therapy for advanced colorectal cancer.

BACKGROUND: Irinotecan and weekly cetuximab (I+C) is a standard second-line regimen for metastatic colorectal cancer (mCRC). This study investigated the safety and efficacy of every 2 weeks I+C in patients with mCRC. PATIENTS AND METHODS: Patients with mCRC refractory to first-line fluoropyrimidine/oxaliplatin regimens and not previously treated with I+C were eligible. Response rate (RR) was the primary endpoint. Cetuximab 500 mg/m(2) and irinotecan 180 mg/m(2) were administered intravenously (I.V.) on day 1 every 2 weeks. RESULTS: Patient characteristics (n = 31): male (n = 17), median age 62; Eastern Cooperative Oncology Group (ECOG) performance status (PS) ≤1 (n = 30), and PS = 2 (n = 1). Median number of cycles = 3 (range, 1-22). I+C doses were modified in 18 and 12 patients, respectively. Grade 3/4 adverse events: acneiform rash (n = 6); neutropenia (n = 6); and diarrhea (n = 5); there was one grade 5 respiratory failure, possibly related to therapy. Two patients had a partial response, 11 had stable disease, and 18 had progressive disease resulting in an overall RR of 6% and disease control rate of 41.9%. Median overall survival (OS) was 9.3 months (95% CI, 5.1-15), and time to progression (TTP) was 2.4 months (95% CI, 1.3-4.6). K-ras and BRAF mutations were detected in 39% and 9%, respectively, of the patients tested. There was a trend toward longer TTP among patients with wild-type K-ras and BRAF (2.6 vs. 1.7 months; P = 0.16), and OS was significantly longer in those patients (14.1 vs. 5.5 months; P = 0.04). CONCLUSIONS: The RR and TTP were lower than expected and may reflect the reduced dose intensity due to toxicities. While the OS was consistent with previous publications, the efficacy of this combination was not demonstrated.

Multiplex serum biomarker assessments: technical and biostatistical issues.

BACKGROUND: Identification of predictive and prognostic biomarkers for patients with disease and undergoing different therapeutic options is a very active area of investigation. Many of these studies seek biomarkers among circulating proteins accessed in blood. Many levels of standardization in materials and procedures have been identified which can impact the resulting data. METHODS: Here, we have observed unexpected variability in levels of commonly tested analytes in serum which were processed and stored under standardized conditions. We have identified apparent changes in cytokine, chemokine and growth factor levels detected by multiplex Luminex assay in melanoma patient and healthy donor serum samples, over storage time at -80°C. Controls included Luminex kit standards, multiplexed cytokine standards and WHO cytokine controls. Data were analyzed by Wilcoxon rank-sum testing and Spearman's test for correlations. RESULTS: The interpretation of these changes is confounded by lot-to-lot kit standard curve reagent changes made by a single manufacturer of Luminex kits. CONCLUSIONS: This study identifies previously unknown sources of variation in a commonly used biomarker assay, and suggests additional levels of controls needed for identification of true changes in circulating protein levels.

A phase I/II study of capecitabine given on a week on/week off schedule combined with bevacizumab and oxaliplatin for patients with untreated advanced colorectal cancer.

BACKGROUND: A week on/week off capecitabine schedule with oxaliplatin/bevacizumab was evaluated in patients with metastatic colorectal cancer (mCRC). PATIENTS AND METHODS: Forty patients were required. The projected median progression-free survival (PFS) was 12 months (81% power, 1-sided level 0.1 log-rank test). Capecitabine dose was 2500 mg/m(2)/day on days 1-7 (n = 11) and was increased to 3000 mg/m(2)/day (n = 29) in combination with oxaliplatin (85 mg/m(2)) and bevacizumab (5 mg/kg). Cycles were repeated every 2 weeks. RESULTS: Patient characteristics included Eastern Cooperative Oncology Group (ECOG) performance status 0 (n = 24) or 1 (n = 15); median age of 62 years (range, 38-81 years). Median cycles administered were 7 (range, 125), corresponding to 3.5 months' treatment duration. Pertinent grade 3/4 toxicities seen were diarrhea (18%), hand-foot syndrome (10%), and peripheral neuropathy (10%). Bowel perforation in 1 patient (3%) and 1 death due to a cerebral hemorrhage (3%) were noted. Response rate (RR) was 38% (1 complete and 14 partial responses). Median PFS was 8.6 months (95% confidence interval [CI], 4.7-10.2 months). Median overall survival was 17.2 months (95% CI, 10.4-24.2 months). CONCLUSION: The first US experience of capecitabine to our knowledge (3000 mg/m(2) on days 1-7) in combination with oxaliplatin/bevacizumab in mCRC does not appear to have advantages compared with current standard first-line mCRC treatment regimens.

Recommendations from the iSBTc-SITC/FDA/NCI Workshop on Immunotherapy Biomarkers.

PURPOSE: To facilitate development of innovative immunotherapy approaches, especially for treatment concepts exploiting the potential benefits of personalized therapy, there is a need to develop and validate tools to identify patients who can benefit from immunotherapy. Despite substantial effort, we do not yet know which parameters of antitumor immunity to measure and which assays are optimal for those measurements. EXPERIMENTAL DESIGN: The iSBTc-SITC (International Society for Biological Therapy of Cancer-Society for Immunotherapy of Cancer), FDA (Food and Drug Administration), and NCI (National Cancer Institute) partnered to address these issues for immunotherapy of cancer. Here, we review the major challenges, give examples of approaches and solutions, and present our recommendations. RESULTS AND CONCLUSIONS: Although specific immune parameters and assays are not yet validated, we recommend following standardized (accurate, precise, and reproducible) protocols and use of functional assays for the primary immunologic readouts of a trial; consideration of central laboratories for immune monitoring of large, multi-institutional trials; and standardized testing of several phenotypic and functional potential potency assays specific to any cellular product. When reporting results, the full QA (quality assessment)/QC (quality control) should be conducted and selected examples of truly representative raw data and assay performance characteristics should be included. Finally, to promote broader analysis of multiple aspects of immunity, and gather data on variability, we recommend that in addition to cells and serum, RNA and DNA samples be banked (under standardized conditions) for later testing. We also recommend that sufficient blood be drawn to allow for planned testing of the primary hypothesis being addressed in the trial, and that additional baseline and posttreatment blood is banked for testing novel hypotheses (or generating new hypotheses) that arise in the field.

Induction of CD8+ T-cell responses against novel glioma-associated antigen peptides and clinical activity by vaccinations with {alpha}-type 1 polarized dendritic cells and polyinosinic-polycytidylic acid stabilized by lysine and carboxymethylcellulose in patients with recurrent malignant glioma.

PURPOSE: A phase I/II trial was performed to evaluate the safety and immunogenicity of a novel vaccination with α-type 1 polarized dendritic cells (αDC1) loaded with synthetic peptides for glioma-associated antigen (GAA) epitopes and administration of polyinosinic-polycytidylic acid [poly(I:C)] stabilized by lysine and carboxymethylcellulose (poly-ICLC) in HLA-A2(+) patients with recurrent malignant gliomas. GAAs for these peptides are EphA2, interleukin (IL)-13 receptor-α2, YKL-40, and gp100. PATIENTS AND METHODS: Twenty-two patients (13 with glioblastoma multiforme [GBM], five with anaplastic astrocytoma [AA], three with anaplastic oligodendroglioma [AO], and one with anaplastic oligoastrocytoma [AOA]) received at least one vaccination, and 19 patients received at least four vaccinations at two αDC1 dose levels (1 × or 3 × 10(7)/dose) at 2-week intervals intranodally. Patients also received twice weekly intramuscular injections of 20 µg/kg poly-ICLC. Patients who demonstrated positive radiologic response or stable disease without major adverse events were allowed to receive booster vaccines. T-lymphocyte responses against GAA epitopes were assessed by enzyme-linked immunosorbent spot and HLA-tetramer assays. RESULTS: The regimen was well-tolerated. The first four vaccines induced positive immune responses against at least one of the vaccination-targeted GAAs in peripheral blood mononuclear cells in 58% of patients. Peripheral blood samples demonstrated significant upregulation of type 1 cytokines and chemokines, including interferon-α and CXCL10. Nine (four GBM, two AA, two AO, and one AOA) achieved progression-free status lasting at least 12 months. One patient with recurrent GBM demonstrated sustained complete response. IL-12 production levels by αDC1 positively correlated with time to progression. CONCLUSION: These data support safety, immunogenicity, and preliminary clinical activity of poly-ICLC-boosted αDC1-based vaccines.

A fluorinated dendrimer-based nanotechnology platform: new contrast agents for high field imaging.

OBJECTIVES: We aimed to develop a directly detected magnetic resonance imaging (MRI) contrast agent for use with high fields based on a nanoscale fluorinated dendrimer-based platform for F MRI and overcome some of the problems with F MRI. MATERIALS AND METHODS: The dendrimers were prepared in a convergent manner by making the appropriate dendron, followed by coupling to a central core. The dendrons were prepared by attaching 3 equivalents of the fluorinated amino acid to the 3 carboxylic acids of the repeat branch unit followed by deprotection of the amine branch point, and either coupling to another repeat branch unit (increasing the generation G) or used directly allowing the precise growth of the dendrimer. The size of the dendrimers was determined by diffusion nuclear magnetic resonance (NMR) spectroscopy. The toxicity of the dendrimers was measured using the MTT assay. Fluorine longitudinal relaxation time measurements were performed on a Bruker ACP-500 NMR using a saturation recovery experiment at 470.59 MHz frequency. Healthy 150 g Sprague-Dawley female rats were imaged using a dendrimer solution. RESULTS: The size of the dendrimers is generally less than 3 nm, 2 orders of magnitude smaller than the size of the perfluorocarbon nanoparticles (about 200 nm). The longitudinal relaxation time, T1, decreases with increasing dendrimer generation. A significant improvement in relaxation rate and signal-to-noise ratio can be achieved by either the chemical modification of the dendrimer with a gadolinium-chelate or by the physical addition of exogenous contrast agent. Although the dendrimers with fluorine in the surface layer are toxic, this toxicity is easily reduced by burying the fluorine further into the dendrimer interior. (19)F MR images of the rat using the dendrimer solution were rapidly obtained at 7 Tesla, the strong contrast in the heart generated by the dendrimer can be seen. CONCLUSIONS: A novel fluorinated dendrimer-based nanotechnology platform in (19)F MRI and a new bifunctional DOTA chelate were prepared and characterized. We introduce 2 methods for reducing the (19)F longitudinal relaxation time: (a) Increasing the generation; (b) covalent and noncovalent introduction of Gd(III)-chelates. A new bifunctional Gd(III)-chelate is presented. The investigations of imaging on rats suggest potential importance of the dendrimers in (19)F MRI application.

Prevention of mammary carcinogenesis in MMTV-neu mice by cruciferous vegetable constituent benzyl isothiocyanate.

Benzyl isothiocyanate (BITC), a constituent of edible cruciferous vegetables, inhibits growth of human breast cancer cells in culture. The present study provides in vivo evidence for efficacy of BITC for prevention of mammary cancer in MMTV-neu mice. Administration of BITC at 1 and 3 mmol/kg diet for 25 weeks markedly suppressed the incidence and/or burden of mammary hyperplasia and carcinoma in female MMTV-neu mice without causing weight loss or affecting neu protein level. For example, cumulative incidence of hyperplasia/carcinoma was significantly lower in mice fed BITC-supplemented diets compared with control mice (P = 0.01 by Fisher's test). The BITC-mediated prevention of mammary carcinogenesis correlated with suppression of cell proliferation and increased apoptosis. The average number of Ki-67-positive cells in the carcinoma lesions of 3 mmol BITC group was lower by approximately 21% (P < 0.05) compared with tumors from control mice. Apoptotic bodies in the mammary tumor were higher by about 2- to 2.5-fold in the 1 and 3 mmol BITC treatment groups (P < 0.05) compared with control group. The BITC administration also resulted in overexpression of E-cadherin and infiltration of CD3(+) T-cells in the tumor. Although BITC treatment increased cytotoxicity of natural killer (NK) cells in vitro, dietary feeding of BITC failed to augment NK cell lytic activity in an ex vivo assay. The present study demonstrating efficacy of BITC against mammary cancer in an animal model provides impetus to determine its activity in a clinical setting.

Dicer-regulated microRNAs 222 and 339 promote resistance of cancer cells to cytotoxic T-lymphocytes by down-regulation of ICAM-1.

The RNase III endonuclease Dicer plays a key role in generation of microRNAs (miRs). We hypothesized that Dicer regulates cancer cell susceptibility to immune surveillance through miR processing. Indeed, Dicer disruption up-regulated intercellular cell adhesion molecule (ICAM)-1 and enhanced the susceptibility of tumor cells to antigen-specific lysis by cytotoxic T-lymphocytes (CTLs), while expression of other immunoregulatory proteins examined was not affected. Blockade of ICAM-1 inhibited the specific lysis of CTLs against Dicer-disrupted cells, indicating a pivotal role of ICAM-1 in the interaction between tumor cells and CTL. Both miR-222 and -339 are down-regulated in Dicer-disrupted cells and directly interacted with the 3' untranslated region (UTR) of ICAM-1 mRNA. Modulation of Dicer or these miRs inversely correlated with ICAM-1 protein expression and susceptibility of U87 glioma cells to CTL-mediated cytolysis while ICAM-1 mRNA levels remained stable. Immunohistochemical and in situ hybridization analyses of 30 primary glioblastoma tissues demonstrated that expression of Dicer, miR-222, or miR-339 was inversely associated with ICAM-1 expression. Taken together, Dicer is responsible for the generation of the mature miR-222 and -339, which suppress ICAM-1 expression on tumor cells, thereby down-regulating the susceptibility of tumor cells to CTL-mediated cytolysis. This study suggests development of novel miR-targeted therapy to promote cytolysis of tumor cells.

Humoral immune response to abnormal MUC1 in subjects with colorectal adenoma and cancer.

Colorectal cancer (CRC) expresses a hypoglycosylated (abnormal) form of MUC1 different than MUC1 expressed in normal colon, which elicits antibodies in patients with CRC. This form of MUC1 is expressed in other abnormal but non-malignant lesions in the colon, such as adenomatous polyps, precursors to CRC. Estimates of the prevalence of anti-MUC1 antibodies in subjects with these lesions are lacking. We evaluated IgM and IgG anti-MUC1 antibodies in 148 subjects with non-advanced adenomas (NAA), advanced adenomas (AA), colorectal cancer, hyperplastic polyps (HPP), and normal controls. We hypothesized that the prevalence of anti-MUC1 antibodies would increase along the adenoma-carcinoma sequence as more dysplastic tissues express more abnormal MUC1. Anti-MUC1 IgM was found in 5/47 (10.6%) of normals, 5/45 (11.1%) of NAA, 7/47 (14.9%) of AA, and 4/20 (20.0%) of CRC (p=0.70). The prevalence of anti-MUC1 IgG was 8/47 (17.0%) of normals, 14/45 (31.1%) of NAA, 14/47 (29.8%) of AA, and 6/20 (30.0%) of CRC (p=0.36). We found no significant differences in the prevalence of anti-MUC1 antibodies between subjects along the adenoma-carcinoma sequence. However, in an exploratory analysis, the median normalized anti-MUC1 IgG OD level of the combined abnormal groups (NAA, AA, CRC) was significantly higher than the normals (0.045 OD vs. 0.030 OD; p=0.017). Our data support further studies into the potential role of anti-MUC1 immunity in preventing progression of premalignant colon lesions to colon cancer.

A phase I study of hyperthermic isolated hepatic perfusion with oxaliplatin in the treatment of unresectable liver metastases from colorectal cancer.

Isolated hepatic perfusion (IHP) is a proven approach for regional delivery of chemotherapy in patients with unresectable primary and metastatic tumors of the liver. Most trials of IHP have utilized melphalan as the active drug in the perfusate. We performed a phase I trial to evaluate the efficacy, safety, and maximum tolerated dose (MTD) of oxaliplatin delivered by hyperthermic isolated hepatic perfusion. A phase I dose-escalation trial of hyperthermic IHP with oxaliplatin in patients with unresectable metastatic colorectal cancer scheduled to undergo placement of a hepatic arterial infusion (HAI) pump was carried out. Thirteen patients were enrolled between November 2003 and September 2006. Dose-limiting veno-occlusive disease was observed at 60 mg/m(2). At the MTD of 40 mg/m(2) only minor transient liver dysfunction was observed. Ultrafilterable platinum area under the curve and maximum concentration delivered by IHP increased nonlinearly with dose as did platinum concentrations in liver biopsies obtained at the end of the 60 min IHP. Seventy-seven percent of patients had a >50% decrease in serum carcinoembryonic antigen (CEA) after IHP. The overall response rate to the combined IHP and HAI therapy was 66%. One patient had a durable complete response (>4 years). We conclude that hyperthermic IHP with oxaliplatin was safe and feasible at a dose of 40 mg/m(2). The ability to obtain complete vascular isolation with open IHP was confirmed. The response rate in this small phase I study was encouraging.

Regulation of antigen presentation machinery in human dendritic cells by recombinant adenovirus.

Recombinant adenoviral vectors (AdV) are potent vehicles for antigen engineering of dendritic cells (DC). DC engineered with AdV to express full length tumor antigens are capable stimulators of antigen-specific polyclonal CD8+ and CD4+ T cells. To determine the impact of AdV on the HLA class I antigen presentation pathway, we investigated the effects of AdV transduction on antigen processing machinery (APM) components in human DC. Interactions among AdV transduction, maturation, APM regulation and T cell activation were investigated. The phenotype and cytokine profile of DC transduced with AdV was intermediate, between immature (iDC) and matured DC (mDC). Statistically significant increases in expression were observed for peptide transporters TAP-1 and TAP-2, and HLA class I peptide-loading chaperone ERp57, as well as co-stimulatory surface molecule CD86 due to AdV transduction. AdV transduction enhanced the expression of APM components and surface markers on mDC, and these changes were further modulated by the timing of DC maturation. Engineering of matured DC to express a tumor-associated antigen stimulated a broader repertoire of CD8+ T cells, capable of recognizing immunodominant and subdominant epitopes. These data identify molecular changes in AdV-transduced DC (AdV/DC) that could influence T cell priming and should be considered in design of cancer vaccines.

A systematic approach to biomarker discovery; preamble to "the iSBTc-FDA taskforce on immunotherapy biomarkers".

The International Society for the Biological Therapy of Cancer (iSBTc) has initiated in collaboration with the United States Food and Drug Administration (FDA) a programmatic look at innovative avenues for the identification of relevant parameters to assist clinical and basic scientists who study the natural course of host/tumor interactions or their response to immune manipulation. The task force has two primary goals: 1) identify best practices of standardized and validated immune monitoring procedures and assays to promote inter-trial comparisons and 2) develop strategies for the identification of novel biomarkers that may enhance our understating of principles governing human cancer immune biology and, consequently, implement their clinical application. Two working groups were created that will report the developed best practices at an NCI/FDA/iSBTc sponsored workshop tied to the annual meeting of the iSBTc to be held in Washington DC in the Fall of 2009. This foreword provides an overview of the task force and invites feedback from readers that might be incorporated in the discussions and in the final document.

Garlic constituent diallyl trisulfide prevents development of poorly differentiated prostate cancer and pulmonary metastasis multiplicity in TRAMP mice.

Identification of agents that are nontoxic but can delay onset and/or progression of prostate cancer, which is the second leading cause of cancer-related deaths among men in the United States, is highly desirable. We now show that p.o. gavage of garlic constituent diallyl trisulfide (DATS; 1 and 2 mg/day, thrice/week for 13 weeks beginning at age 8 weeks) significantly inhibits progression to poorly differentiated prostate carcinoma and pulmonary metastasis multiplicity in transgenic adenocarcinoma of mouse prostate (TRAMP) mice without any side effects. There was a trend of a decrease in average wet weights of the urogenital tract and prostate gland in 1 and 2 mg DATS-treated mice compared with controls ( approximately 25-46% decrease in DATS-treated mice compared with controls). The incidence and the area of the dorsolateral prostate occupied by the poorly differentiated carcinoma were significantly lower in both 1 and 2 mg DATS-treated mice compared with control mice. In addition, DATS administration resulted in a statistically significant decrease in pulmonary metastasis multiplicity compared with controls (P = 0.002). The dorsolateral prostate from DATS-treated TRAMP mice exhibited decreased cellular proliferation in association with induction of cyclinB1 and securin protein levels, and suppression of the expression of neuroendocrine marker synaptophysin. However, DATS administration did not have any appreciable effect on apoptosis induction, angiogenesis, or natural killer and dendritic cell function. In conclusion, the results of the present study show, for the first time, that DATS administration prevents progression to invasive carcinoma and lung metastasis in TRAMP mice.

A phase I study of 17-allylamino-17-demethoxygeldanamycin combined with paclitaxel in patients with advanced solid malignancies.

BACKGROUND: 17-Allylamino-17-demethoxygeldanamycin (17-AAG) inhibits heat shock protein 90, promotes degradation of oncoproteins, and exhibits synergy with paclitaxel in vitro. We conducted a phase I study in patients with advanced malignancies to determine the recommended phase II dose of the combination of 17-AAG and paclitaxel. METHODS: Patients with advanced solid malignancies that were refractory to proven therapy or without any standard treatment were included. 17-AAG (80-225 mg/m2) was given on days 1, 4, 8, 11, 15, and 18 of each 4-week cycle to sequential cohorts of patients. Paclitaxel (80-100 mg/m2) was administered on days 1, 8, and 15. Pharmacokinetic studies were conducted during cycle 1. RESULTS: Twenty-five patients were accrued to five dose levels. The median number of cycles was 2. Chest pain (grade 3), myalgia (grade 3), and fatigue (grade 3) were dose-limiting toxicities at dose level 4 (225 mg/m2 17-AAG and 80 mg/m2 paclitaxel). None of the six patients treated at dose level 3 with 17-AAG (175 mg/m2) and paclitaxel (80 mg/m2) experienced dose-limiting toxicity. Disease stabilization was noted in six patients, but there were no partial or complete responses. The ratio of paclitaxel area under the concentration to time curve when given alone versus in combination with 17-AAG was 0.97 +/- 0.20. The ratio of end-of-infusion concentration of 17-AAG (alone versus in combination with paclitaxel) was 1.14 +/- 0.51. CONCLUSIONS: The recommended phase II dose of twice-weekly 17-AAG (175 mg/m2) and weekly paclitaxel (80 mg/m2/wk) was tolerated well. There was no evidence of drug-drug pharmacokinetic interactions.