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1.
Ann Oncol ; 30(1): 44-56, 2019 01 01.
Article in English | MEDLINE | ID: mdl-30395155

ABSTRACT

Background: Treatment with immune checkpoint blockade (ICB) with agents such as anti-programmed cell death protein 1 (PD-1), anti-programmed death-ligand 1 (PD-L1), and/or anti-cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) can result in impressive response rates and durable disease remission but only in a subset of patients with cancer. Expression of PD-L1 has demonstrated utility in selecting patients for response to ICB and has proven to be an important biomarker for patient selection. Tumor mutation burden (TMB) is emerging as a potential biomarker. However, refinement of interpretation and contextualization is required. Materials and methods: In this review, we outline the evolution of TMB as a biomarker in oncology, delineate how TMB can be applied in the clinic, discuss current limitations as a diagnostic test, and highlight mechanistic insights unveiled by the study of TMB. We review available data to date studying TMB as a biomarker for response to ICB by tumor type, focusing on studies proposing a threshold for TMB as a predictive biomarker for ICB activity. Results: High TMB consistently selects for benefit with ICB therapy. In lung, bladder and head and neck cancers, the current predictive TMB thresholds proposed approximate 200 non-synonymous somatic mutations by whole exome sequencing (WES). PD-L1 expression influences response to ICB in high TMB tumors with single agent PD-(L)1 antibodies; however, response may not be dependent on PD-L1 expression in the setting of anti-CTLA4 or anti-PD-1/CTLA-4 combination therapy. Disease-specific TMB thresholds for effective prediction of response in various other malignancies are not well established. Conclusions: TMB, in concert with PD-L1 expression, has been demonstrated to be a useful biomarker for ICB selection across some cancer types; however, further prospective validation studies are required. TMB determination by selected targeted panels has been correlated with WES. Calibration and harmonization will be required for optimal utility and alignment across all platforms currently used internationally. Key challenges will need to be addressed before broader use in different tumor types.


Subject(s)
Antineoplastic Agents, Immunological/therapeutic use , Biomarkers, Tumor/genetics , Immunotherapy/methods , Mutation , Neoplasms/drug therapy , Neoplasms/genetics , Humans , Neoplasms/immunology , Neoplasms/pathology , Prognosis
2.
J Intern Med ; 283(6): 568-577, 2018 06.
Article in English | MEDLINE | ID: mdl-29464806

ABSTRACT

BACKGROUND: Use of the immune checkpoint inhibitor ipilimumab is sometimes complicated by ipilimumab-associated colitis (Ipi-AC), an immune-mediated colitis that mimics inflammatory bowel disease. OBJECTIVE: We sought to characterize the histopathologic and immunophenotypic features of Ipi-AC and to directly compare these features to ulcerative colitis (UC). METHODS: This is a retrospective cohort study of 22 patients with Ipi-AC, 12 patients with treatment-naïve UC and five controls with diarrhoea but normal endoscopic findings. Immunohistopathologic features were described, and quantitative immunohistochemistry (IHC) was performed for CD4, CD8, CD20, CD138 and FOXP3. RESULTS: Endoscopic findings in both the Ipi-AC and UC groups included ulcerated, oedematous and erythematous mucosa. Involvement of the GI tract was more diffuse in Ipi-AC. As compared to UC, a smaller proportion of Ipi-AC biopsies had basal plasmacytosis (14% for Ipi-AC vs. 92% for UC, P < 0.0001) and crypt distortion (23% for Ipi-AC vs. 75% for UC, P = 0.003), whereas Ipi-AC biopsies had more apoptotic bodies in the left colon (17.6 ± 15.3 for Ipi-AC vs. 8.2 ± 4.2 for UC, P = 0.011). Cryptitis, ulcerations and crypt abscesses were common in both groups. Biopsy specimens from Ipi-AC had a lower density of CD20-positive lymphocytes than UC (275.8 ± 253.3 cells mm-2 for Ipi-AC vs. 1173.3 ± 1158.2 cells mm-2 for UC, P = 0.022) but had a similar density of CD4, CD8, CD138 and FOXP3-positive cells. CONCLUSIONS: Ipi-AC is a distinct pathologic entity with notable clinical and histopathological differences compared to UC. These findings provide insights into the pathophysiology of immune-related adverse events (iAEs) from ipilimumab therapy.


Subject(s)
Antineoplastic Agents, Immunological/adverse effects , Colitis/chemically induced , Ipilimumab/adverse effects , Adult , Colitis/immunology , Colitis/pathology , Colitis, Ulcerative/immunology , Colitis, Ulcerative/pathology , Diarrhea/etiology , Female , Humans , Immunohistochemistry , Intestinal Mucosa/pathology , Male , Middle Aged , Neoplasms/drug therapy , Retrospective Studies
3.
J Viral Hepat ; 20(3): 219-24, 2013 Mar.
Article in English | MEDLINE | ID: mdl-23383661

ABSTRACT

The effects of genomic changes in hepatitis B virus (HBV) on the occurrence of hepatocellular carcinoma (HCC) are still unclear, especially in relation to the genotype of HBV. In this study, we examined the effects of genomic changes in HBV of genotype C2 on the development of HCC. A total of 318 patients with HBV-associated HCC and 234 patients with chronic hepatitis B (CHB) were studied. All of HCC cases were diagnosed histologically and treated with surgical resection. The whole of the X, S, basal core promoter (BCP) and precore regions of the viral genome from sera or liver tissues were sequenced. All subjects had HBV of genotype C2. The prevalence of the T1653 mutation in the X region and the A1896 mutation in the precore region of HBV was significantly higher in the HCC group than in the control CHB group (22% vs 11%, P = 0.003; 50% vs 23%, P < 0.001, respectively). Moreover, the T1762/A1764 mutations in the BCP region in combination with either T1653 or A1896 were more common in the HCC compared with the CHB group (BCP+X1653: 18% vs 11%, P = 0.05; BCP+PC, 40% vs 15%, P < 0.001, respectively). In multivariate analysis, T1653 and A1896 were revealed to be independent risk factors for HCC development. G1896A in the precore region and C1653T mutation in the X region of genotype C2 HBV are important risk factors for HCC development. Also, the A1762T/G1764A double mutation may act in synergy with C1653T to increase the risk of HCC in patients chronically infected with HBV genotype C2.


Subject(s)
Carcinoma, Hepatocellular/virology , DNA, Viral/genetics , Hepatitis B virus/genetics , Hepatitis B, Chronic/complications , Hepatitis B, Chronic/virology , Point Mutation , Adult , Aged , Aged, 80 and over , DNA, Viral/chemistry , Female , Genotype , Hepatitis B virus/physiology , Humans , Male , Middle Aged , Sequence Analysis, DNA
4.
Cytogenet Genome Res ; 118(2-4): 148-56, 2007.
Article in English | MEDLINE | ID: mdl-18000365

ABSTRACT

Karyotype analysis can provide clues to significant genes involved in the genesis and growth of pancreas cancer. The genome of pancreas cancer is complex, and G-band analysis cannot resolve many of the karyotypic abnormalities seen. We studied the karyotypes of 15 recently established cell lines using molecular cytogenetic tools. Comparative genomic hybridization (CGH) analysis of all 15 lines identified genomic gains of 3q, 8q, 11q, 17q, and chromosome 20 in nine or more cell lines. CGH confirmed frequent loss of chromosome 18, 17p, 6q, and 8p. 14/15 cell lines demonstrated loss of chromosome 18q, either by loss of a copy of chromosome 18 (n = 5), all of 18q (n = 7) or portions of 18q (n = 2). Multicolor FISH (Spectral Karyotyping, or SKY) of 11 lines identified many complex structural chromosomal aberrations. 93 structurally abnormal chromosomes were evaluated, for which SKY added new information to 67. Several potentially site-specific recurrent rearrangements were observed. Chromosome region 18q11.2 was recurrently involved in nine cell lines, including formation of derivative chromosomes 18 from a t(18;22) (three cell lines), t(17;18) (two cell lines), and t(12;18), t(15;18), t(18;20), and ins(6;18) (one cell line each). To further define the breakpoints involved on chromosome 18, YACs from the 18q11.2 region, spanning approximately 8 Mb, were used to perform targeted FISH analyses of these lines. We found significant heterogeneity in the breakpoints despite their G-band similarity, including multiple independent regions of loss proximal to the already identified loss of DPC4 at 18q21.


Subject(s)
Adenocarcinoma/genetics , Chromosome Aberrations , Pancreatic Neoplasms/genetics , Adenocarcinoma/pathology , Cell Line, Tumor , Chromosome Banding , Chromosomes, Human, Pair 18 , Chromosomes, Human, Pair 8 , Humans , In Situ Hybridization, Fluorescence , Karyotyping , Metaphase , Nucleic Acid Hybridization , Pancreatic Neoplasms/pathology
5.
Cytotherapy ; 7(1): 46-56, 2005.
Article in English | MEDLINE | ID: mdl-16040383

ABSTRACT

BACKGROUND: GM-CSF-secreting, allogeneic cell-based cancer vaccines have shown promise for the treatment of a variety of solid tumors. We have now applied this approach to breast cancer. The aim of these studies was to optimize expansion parameters, qualify the manufacturing process, and establish expected outcomes for cGMP-compliant manufacturing of two GM-CSF-secreting breast tumor cell lines. METHODS: The variables affecting the efficiency of expanding and formulating two allogeneic GM-CSF-secreting cell lines, 2T47D-V and 3SKBR3-7, were systematically evaluated. Production criteria investigated included alternative cell culture vessels (flasks vs. cell factories), centrifugation time and speed variables for large volume cell concentration, cell seeding density, the minimal concentration of FBS required for maximal cell expansion, and the dose and timing of irradiation in relation to cryopreservation. RESULTS: These studies demonstrate that, in comparison with standard 150-cm2 tissue culture flasks, Nunc 10-Stack Cell Factories are a more efficient and practical cell culture vessel for vaccine cell line manufacture. Centrifugation optimization studies using the COBE 2991 Cell Processor established that a speed of 2000 r.p.m. (450 g) for 2 min reliably concentrated the cells while maintaining acceptable viability and bioactivity. Radiation studies established that lethal irradiation prior to cryopreservation does not compromise the quality of the product, as measured by post-thaw cell viability and GM-CSF cell line-specific secretion levels. Finally, studies aimed at optimizing the production of one vaccine cell line, 3SKBR3-7, demonstrated that seeding the cells at a higher density and maintaining them in half the initial concentration of FBS maximized the yield of bioactive cells, resulting in significant cost savings. DISCUSSION: A manufacturing process that simultaneously maximizes cell yield, minimizes cell manipulation and maintains vaccine cell potency is critical for producing cell-based cancer vaccines in an academic setting. These studies define a feasible, reproducible and cost-effective methodology for production of a GM-CSF-secreting breast cancer vaccine that is cGMP compliant.


Subject(s)
Breast Neoplasms/therapy , Cancer Vaccines/chemical synthesis , Carcinoma/therapy , Granulocyte-Macrophage Colony-Stimulating Factor/chemical synthesis , Academic Medical Centers/economics , Academic Medical Centers/methods , Academic Medical Centers/standards , Breast Neoplasms/immunology , Cancer Vaccines/economics , Cancer Vaccines/radiation effects , Carcinoma/immunology , Cell Culture Techniques/economics , Cell Culture Techniques/methods , Cell Line, Tumor , Cost-Benefit Analysis , Cryopreservation/methods , Female , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Granulocyte-Macrophage Colony-Stimulating Factor/therapeutic use , Guideline Adherence , Humans , Laboratories/economics , Laboratories/standards , Radiation Dosage , Transplantation, Homologous/economics , Transplantation, Homologous/immunology , Transplantation, Homologous/methods
6.
Endocr Relat Cancer ; 12(1): 1-17, 2005 Mar.
Article in English | MEDLINE | ID: mdl-15788636

ABSTRACT

Optimizing standard treatment modalities for breast cancer has improved the outlook for women afflicted with it, but the fact that 40% still ultimately die from the disease highlights the need for new therapies. Remarkable advances in molecular immunology and biotechnology have created a unique opportunity for developing active vaccination strategies that engage the patient's own immune system in the fight against breast cancer. Early clinical trials have established the safety and bioactivity of some breast cancer vaccine approaches, with a hint of clinical response. They have also highlighted the importance of elucidating the pharmacodynamic interactions between established therapies for breast cancer, such as tamoxifen, aromatase inhibitors, chemotherapy, the HER-2/neu-specific monoclonal antibody trastuzumab (Herceptin), and breast cancer vaccines. Preclinical studies have simultaneously defined the importance of developing targeted approaches for circumventing established immune tolerance to breast cancer during the vaccination process. The first strategies targeting the negative influence of CD4(+)CD25(+)T regulatory cells and the CTLA-4 signaling pathway are just entering clinical testing in combination with tumor vaccines. Developing the most potent approach for activating antitumor immunity while maintaining the efficacy of standard approaches to breast cancer management will ensure that active immunotherapy is successfully integrated into the standard of care.


Subject(s)
Breast Neoplasms/immunology , Breast Neoplasms/therapy , Cancer Vaccines , Immunity/physiology , Antineoplastic Agents/therapeutic use , Female , Humans , Immunotherapy
8.
Clin Cancer Res ; 7(12): 3862-8, 2001 Dec.
Article in English | MEDLINE | ID: mdl-11751476

ABSTRACT

PURPOSE: Effective new markers of pancreatic carcinoma are urgently needed. In a previous analysis of gene expression in pancreatic adenocarcinoma using serial analysis of gene expression (SAGE), we found that the tag for the mesothelin mRNA transcript was present in seven of eight SAGE libraries derived from pancreatic carcinomas but not in the two SAGE libraries derived from normal pancreatic duct epithelial cells. In this study, we evaluate the potential utility of mesothelin as a tumor marker for pancreatic adenocarcinoma. EXPERIMENTAL DESIGN: Mesothelin mRNA expression was evaluated in pancreatic adenocarcinomas using reverse-transcription PCR (RT-PCR) and in situ hybridization, whereas mesothelin protein expression was evaluated by immunohistochemistry. RESULTS: Using an online SAGE database (http://www.ncbi.nlm.gov/SAGE), we found the tag for mesothelin to be consistently present in the mesothelioma, ovarian cancer, and pancreatic cancer libraries but not in normal pancreas libraries. Mesothelin mRNA expression was confirmed by in situ hybridization in 4 of 4 resected primary pancreatic adenocarcinomas and by RT-PCR in 18 of 20 pancreatic cancer cell lines, whereas mesothelin protein expression was confirmed by immunohistochemistry in all 60 resected primary pancreatic adenocarcinomas studied. The adjacent normal pancreas in these 60 cases did not label, or at most only rare benign pancreatic ducts showed weak labeling for mesothelin. CONCLUSIONS: Mesothelin is a new marker for pancreatic adenocarcinoma identified by gene expression analysis. Mesothelin overexpression in pancreatic adenocarcinoma has potential diagnostic, imaging, and therapeutic implications.


Subject(s)
Adenocarcinoma/genetics , Antigens, Neoplasm/genetics , Biomarkers, Tumor/analysis , Gene Expression Regulation, Neoplastic , Membrane Glycoproteins/genetics , Pancreatic Neoplasms/genetics , Adenocarcinoma/chemistry , Adenocarcinoma/pathology , Antigens, Neoplasm/analysis , GPI-Linked Proteins , Humans , Immunohistochemistry , In Situ Hybridization , Membrane Glycoproteins/analysis , Mesothelin , Online Systems , Pancreatic Neoplasms/chemistry , Pancreatic Neoplasms/pathology , Reverse Transcriptase Polymerase Chain Reaction , Transcription, Genetic , Tumor Cells, Cultured
9.
Cancer J ; 7(4): 324-37, 2001.
Article in English | MEDLINE | ID: mdl-11561608

ABSTRACT

Pancreatic cancer remains one of the most difficult cancers to treat; very few effective therapies are available, with surgery being the sole chance for cure-yet surgery is not a viable option for most pancreatic cancer patients. Immunotherapy has the potential to provide a non-cross-resistant mechanism of antitumor activity that can be integrated with surgery, radiation therapy, and chemotherapy. However, the inherent instability of the tumorgenome as well as tumor tolerance mechanisms are significant practical obstacles that must be overcome if immune-based approaches for pancreatic cancer can achieve its promise. Recent advances in both tumor immunology and vaccine design have already resulted in promising preliminary data from phase I studies, and additional trials are already in progress. This article summarizes some of the progress and challenges in immunotherapy research.


Subject(s)
Immunotherapy/methods , Pancreatic Neoplasms/therapy , B7-1 Antigen , Cancer Vaccines/immunology , Cancer Vaccines/therapeutic use , Forecasting , Humans , Immune Tolerance , Immunity, Cellular , Immunotherapy/classification , Pancreas/immunology , Pancreatic Neoplasms/immunology , Tumor Necrosis Factor Receptor Superfamily, Member 7
10.
Curr Opin Investig Drugs ; 2(1): 133-5, 2001 Jan.
Article in English | MEDLINE | ID: mdl-11527005

ABSTRACT

Recent advances in our understanding of immune function with regard to the generation of a potent antitumor response have resulted in a renewed interest in cancer vaccines and point to a role for immunotherapy in the treatment of cancer. Currently, the majority of vaccine strategies for the treatment of solid malignancies focus on the generation of cytotoxic T lymphocytes (CTL) that destroy tumor cells that express a given target protein, or antigen. However, antibody therapies have already been successful against some cancers. Current humoral immunotherapy typically involves the passive infusion of monoclonal antibodies, which usually target a specific tumor-encoded antigen. However, vaccines can be engineered to induce humoral immunity. By focusing on the cellular arm of the immune response at the expense of humoral immunity (or the converse), we may have inadvertently limited the potential efficacy of our anticancer vaccines. This article seeks to explore the notion that a vaccine designed to optimally activate both arms of the immune system may well generate an antitumor immune response greater than the sum of the two individual effector mechanisms alone.


Subject(s)
Neoplasms/immunology , Antibodies, Monoclonal/therapeutic use , Antibody Formation , Antigen Presentation , B-Lymphocytes/immunology , Cancer Vaccines/immunology , Humans , Immunity, Cellular , Neoplasms/therapy , T-Lymphocytes/immunology
11.
Gene Ther ; 8(13): 969-76, 2001 Jul.
Article in English | MEDLINE | ID: mdl-11438831

ABSTRACT

In this study we analyzed two ways of retargeting of Ad-vectors to human pancreatic carcinoma with the aim of enhancing the gene transfer efficiency. First, we analyzed the expression of the epidermal growth factor receptor (EGFR) on primary, as well as established pancreatic carcinoma cells by flow cytometry which revealed high expression levels of EGFR on the surface of these cells. We showed that EGFR-retargeted entry pathway using a bispecific fusion protein formed by a recombinant soluble form of truncated Coxsackie and Adenovirus Receptor (sCAR) genetically fused with human EGF (sCAR-EGF) redirects them to the EGFR leading to an enhanced gene transfer efficiency to pancreatic carcinoma cells. Since flow cytometry revealed absence of CAR expression, but the presence of at least one of both alphav integrins on the pancreatic carcinoma cells, a second way of targeting was investigated using a genetically modified Ad vector which has an RGD (Arg-Gly-Asp)-containing peptide inserted into the HI-loop of the fiber knob. This RGD targeted Ad (AdlucRGD) revealed efficient CAR-independent infection by allowing binding to cellular integrins resulting in a dramatic enhancement of gene transfer. These findings have direct relevance for Ad-vector based gene therapy strategies for pancreatic carcinoma.


Subject(s)
ErbB Receptors/metabolism , Gene Targeting/methods , Gene Transfer Techniques , Genetic Therapy/methods , Pancreatic Neoplasms/metabolism , Adenoviridae/genetics , Genetic Vectors , Humans , Integrins/metabolism , Neoplasm Proteins/metabolism , Pancreatic Neoplasms/pathology , Pancreatic Neoplasms/virology , Receptors, Vitronectin/metabolism , Tumor Cells, Cultured
12.
Cancer Res ; 61(11): 4320-4, 2001 Jun 01.
Article in English | MEDLINE | ID: mdl-11389052

ABSTRACT

Serial analysis of gene expression (SAGE) can be used to quantify gene expression in human tissues. Comparison of gene expression levels in neoplastic tissues with those seen in nonneoplastic tissues can, in turn, identify novel tumor markers. Such markers are urgently needed for highly lethal cancers like pancreatic adenocarcinoma, which typically presents at an incurable, advanced stage. The results of SAGE analyses of a large number of neoplastic and nonneoplastic tissues are now available online, facilitating the rapid identification of novel tumor markers. We searched an online SAGE database to identify genes preferentially expressed in pancreatic cancers as compared with normal tissues. SAGE libraries derived from pancreatic adenocarcinomas were compared with SAGE libraries derived from nonneoplastic tissues. Three promising tags were identified. Two of these tags corresponded to genes (lipocalin and trefoil factor 2) previously shown to be overexpressed in pancreatic carcinoma, whereas the third tag corresponded to prostate stem cell antigen (PSCA), a recently discovered gene thought to be largely restricted to prostatic basal cells and prostatic adenocarcinomas. PSCA was expressed in four of the six pancreatic cancer SAGE libraries, but not in the libraries derived from normal pancreatic ductal cells. We confirmed the overexpression of the PSCA mRNA transcript in 14 of 19 pancreatic cancer cell lines by reverse transcription-PCR, and using immunohistochemistry, we demonstrated PSCA protein overexpression in 36 of 60 (60%) primary pancreatic adenocarcinomas. In 59 of 60 cases, the adjacent nonneoplastic pancreas did not label for PSCA. PSCA is a novel tumor marker for pancreatic carcinoma that has potential diagnostic and therapeutic implications. These results establish the validity of analyses of SAGE databases to identify novel tumor markers.


Subject(s)
Biomarkers, Tumor/biosynthesis , Membrane Glycoproteins/biosynthesis , Neoplasm Proteins/biosynthesis , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/immunology , Adenocarcinoma/genetics , Adenocarcinoma/immunology , Antigens, Neoplasm , Biomarkers, Tumor/genetics , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/immunology , GPI-Linked Proteins , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Humans , Immunohistochemistry , Membrane Glycoproteins/genetics , Neoplasm Proteins/genetics , Reverse Transcriptase Polymerase Chain Reaction , Trefoil Factor-2 , Tumor Cells, Cultured
13.
J Immunol ; 167(1): 147-55, 2001 Jul 01.
Article in English | MEDLINE | ID: mdl-11418643

ABSTRACT

There is accumulating evidence that CD4(+) T cell responses are important in antitumor immunity. Accordingly, we generated CD4(+) T cells against the murine CT26 colon cancer. Three of three independent CT26-specific CD4(+) hybridomas were found to recognize the high m.w. precursor of the env gene product gp90. The CD4(+) response was completely tumor specific in that the same glycoprotein expressed by other tumors was not recognized by the CT26-specific hybridomas. The recognition of gp90 by the hybridomas was strictly dependent on the conformation of gp90. Different procedures that disrupted the conformation of the glycoprotein, such as disulfide bond reduction and thermal denaturation, completely abrogated recognition of gp90 by all three hybridomas. In CT26 cells, but not in other tumor cells tested, a large proportion of gp90 was retained in the endoplasmic reticulum, mostly bound to the endoplasmic reticulum chaperone, calreticulin. Although calreticulin was not essential for the stimulation of the gp90-specific hybridomas, most of the antigenic form of gp90 was bound to it. The antigenicity of gp90 correlated well with calreticulin binding, reflecting the fact that specificity of binding of calreticulin to its substrate required posttranslational modifications that were also necessary for the generation of this tumor-specific CD4(+) epitope.


Subject(s)
Antigens, Neoplasm/metabolism , Calcium-Binding Proteins/metabolism , Endoplasmic Reticulum/metabolism , Histocompatibility Antigens Class II/metabolism , Immunodominant Epitopes/metabolism , Ribonucleoproteins/metabolism , Animals , Antigens, Neoplasm/chemistry , CD4-Positive T-Lymphocytes/immunology , Calreticulin , Endoplasmic Reticulum/immunology , Epitopes, T-Lymphocyte/immunology , Female , Gene Products, env/metabolism , Hot Temperature , Hybridomas/metabolism , Leukemia Virus, Murine/immunology , Lymphocyte Activation , Mice , Mice, Inbred BALB C , Mice, SCID , Mice, Transgenic , Protein Binding/immunology , Protein Conformation , Protein Denaturation , Tumor Cells, Cultured
15.
Cancer Res ; 61(9): 3689-97, 2001 May 01.
Article in English | MEDLINE | ID: mdl-11325840

ABSTRACT

Tumor-specific immune tolerance limits the effectiveness of cancer vaccines. In addition, tumor vaccines alone have a limited potential for the treatment of measurable tumor burdens. This highlights the importance of identifying more potent cancer vaccine strategies for clinical testing. We tested immune-modulating doses of chemotherapy in combination with a granulocyte/macrophage-colony stimulating factor (GM-CSF)-secreting, HER-2/neu (neu)-expressing whole-cell vaccine as a means to treat existing mammary tumors in antigen-specific tolerized neu transgenic mice. Earlier studies have shown that neu transgenic mice exhibit immune tolerance to the neu-expressing tumors similar to what is observed in patients with cancer. We found that cyclophosphamide, paclitaxel, and doxorubicin, when given in a defined sequence with a GM-CSF-secreting, neu-expressing whole-cell vaccine, enhanced the vaccine's potential to delay tumor growth in neu transgenic mice. In addition, we showed that these drugs mediate their effects by enhancing the efficacy of the vaccine rather than via a direct cytolytic effect on cancer cells. Furthermore, paclitaxel and cyclophosphamide appear to amplify the T helper 1 neu-specific T-cell response. These findings suggest that the combined treatment with immune-modulating doses of chemotherapy and the GM-CSF-secreting neu vaccine can overcome immune tolerance and induce an antigen-specific antitumor immune response. These data provide the immunological rationale for testing immune-modulating doses of chemotherapy in combination with tumor vaccines in patients with cancer.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/pharmacology , Cancer Vaccines/immunology , Genes, erbB-2/immunology , Granulocyte-Macrophage Colony-Stimulating Factor/immunology , Immune Tolerance/immunology , 3T3 Cells , Animals , Cancer Vaccines/administration & dosage , Cancer Vaccines/genetics , Combined Modality Therapy , Cyclophosphamide/administration & dosage , Doxorubicin/administration & dosage , Drug Administration Schedule , Drug Synergism , Epitopes, T-Lymphocyte/immunology , Female , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Humans , Immune Tolerance/genetics , Lymphocyte Activation/immunology , Mammary Neoplasms, Experimental/genetics , Mammary Neoplasms, Experimental/immunology , Mammary Neoplasms, Experimental/therapy , Mice , Mice, Transgenic , Paclitaxel/administration & dosage , Rats , T-Lymphocytes/immunology , Th1 Cells/immunology
16.
Curr Opin Mol Ther ; 3(1): 77-84, 2001 Feb.
Article in English | MEDLINE | ID: mdl-11249735

ABSTRACT

Cancer vaccines are on the threshold of taking their place alongside the more traditional cancer treatment modalities of surgery, radiation therapy and chemotherapy. The toxicology and immunopharmacology of therapeutic cancer vaccines, particularly those that secrete granulocyte macrophage colony stimulating factor (GM-CSF), are currently under active clinical investigation. Interestingly, drugs traditionally used for tumor cytoreduction can have both positive and negative effects on host immunity. Exploration of the potential pharmacodynamic interactions of antineoplastic drugs with GM-CSF-secreting vaccines has revealed that low doses of some chemotherapeutics can augment the antitumor immunity induced by GM-CSF-secreting vaccines. These interactions will require thorough preclinical evaluation to maximize the clinical impact of this type of therapeutic cancer vaccine.


Subject(s)
Antineoplastic Agents/administration & dosage , Cancer Vaccines/administration & dosage , Neoplasms/drug therapy , Neoplasms/therapy , Adjuvants, Immunologic/administration & dosage , Adjuvants, Immunologic/adverse effects , Animals , Antineoplastic Agents/adverse effects , Cancer Vaccines/adverse effects , Combined Modality Therapy , Cyclophosphamide/administration & dosage , Cyclophosphamide/adverse effects , Doxorubicin/administration & dosage , Doxorubicin/adverse effects , Granulocyte-Macrophage Colony-Stimulating Factor/adverse effects , Granulocyte-Macrophage Colony-Stimulating Factor/therapeutic use , Humans , Immune Tolerance , Models, Biological , Paclitaxel/administration & dosage , Paclitaxel/adverse effects
17.
Cancer Res ; 61(3): 880-3, 2001 Feb 01.
Article in English | MEDLINE | ID: mdl-11221874

ABSTRACT

HER-2/neu (neu) transgenic mice (neu-N mice), which express the nontransforming rat proto-oncogene, demonstrate immunological tolerance to neu that is similar to what is encountered in patients with neu-expressing breast cancer. We have shown previously that a significant increase in neu-specific T cells, but no induction of neu-specific antibody, is seen after neu-specific vaccination in neu-N mice. In contrast, a significant induction of both neu-specific T-cell and antibody responses is found in nontoleragenic FVB/N mice after vaccination. These mice are fully protected from a s.c. challenge with NT cells, a mammary tumor cell line derived from a spontaneous tumor that arose in a neu-N mouse, whereas neu-N mice are not. In this study, we demonstrate that CD4+ T cell-depleted FVB/N mice show no induction of neu-specific IgG after vaccination and are unable to reject an NT challenge (0 of 10 mice were tumor free). Conversely, the depletion of natural killer cells has no effect on vaccine-mediated tumor rejection (100% of mice were tumor free). In CD8+ T cell-depleted animals, where vaccine-induced neu-specific IgG titers were normal, NT growth was delayed, but only 10% of mice remained tumor free, demonstrating that neu-specific IgG alone is insufficient for protection from NT challenge. To directly assess the necessity for the combination of neu-specific cellular and humoral immune responses, severe combined immunodeficient mice were given an adoptive transfer of CTLs plus IgG derived from FVB/N mice. Animals that were given CTLs that recognized an irrelevant antigen plus neu-specific IgG developed tumors at a rate similar to CD8+ T cell-depleted FVB/N mice. Animals receiving an adoptive transfer of neu-specific CTLs plus control IgG derived from naive FVB/N mice were only partially protected from NT challenge (50% of animals were tumor free). However, only animals receiving the combination of neu-specific CTLs and neu-specific IgG were fully protected from NT challenge (100% of animals were tumor free). These studies specifically define the immunological requirements for the eradication of neu-expressing tumors in this model system, demonstrating that both cellular and humoral neu-specific responses are necessary for protection from an NT challenge. These data suggest that vaccines optimized to induce maximal T- and B-cell immunity to neu, and possibly to similar putative tumor-rejection antigens, may lead to more potent in vivo antitumor immunity.


Subject(s)
B-Lymphocytes/immunology , Mammary Neoplasms, Experimental/immunology , Receptor, ErbB-2/immunology , T-Lymphocytes/immunology , Vaccination , 3T3 Cells/immunology , 3T3 Cells/metabolism , Animals , Female , Immune Tolerance , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunotherapy, Adoptive , Lymphocyte Activation/immunology , Mammary Neoplasms, Experimental/metabolism , Mammary Neoplasms, Experimental/prevention & control , Mice , Mice, Inbred BALB C , Mice, SCID , Neoplasm Transplantation , Receptor, ErbB-2/biosynthesis , T-Lymphocyte Subsets/immunology
18.
J Clin Oncol ; 19(1): 145-56, 2001 Jan 01.
Article in English | MEDLINE | ID: mdl-11134207

ABSTRACT

PURPOSE: Allogeneic granulocyte-macrophage colony-stimulating factor (GM-CSF)-secreting tumor vaccines can cure established tumors in the mouse, but their efficacy against human tumors is uncertain. We have developed a novel GM-CSF-secreting pancreatic tumor vaccine. To determine its safety and ability to induce antitumor immune responses, we conducted a phase I trial in patients with surgically resected adenocarcinoma of the pancreas. PATIENTS AND METHODS: Fourteen patients with stage 1, 2, or 3 pancreatic adenocarcinoma were enrolled. Eight weeks after pancreaticoduodenectomy, three patients received 1 x 10(7) vaccine cells, three patients received 5 x 10(7) vaccine cells, three patients received 10 x 10(7) vaccine cells, and five patients received 50 x 10(7) vaccine cells. Twelve of 14 patients then went on to receive a 6-month course of adjuvant radiation and chemotherapy. One month after completing adjuvant treatment, six patients still in remission received up to three additional monthly vaccinations with the same vaccine dose that they had received originally. RESULTS: No dose-limiting toxicities were encountered. Vaccination induced increased delayed-type hypersensitivity (DTH) responses to autologous tumor cells in three patients who had received >or= 10 x 10(7) vaccine cells. These three patients also seemed to have had an increased disease-free survival time, remaining disease-free at least 25 months after diagnosis. CONCLUSION: Allogeneic GM-CSF-secreting tumor vaccines are safe in patients with pancreatic adenocarcinoma. This vaccine approach seems to induce dose-dependent systemic antitumor immunity as measured by increased postvaccination DTH responses against autologous tumors. Further clinical evaluation of this approach in patients with pancreatic cancer is warranted.


Subject(s)
Adenocarcinoma/therapy , Cancer Vaccines/therapeutic use , Granulocyte-Macrophage Colony-Stimulating Factor/therapeutic use , Pancreatic Neoplasms/therapy , Adenocarcinoma/immunology , Adenocarcinoma/pathology , Aged , Cancer Vaccines/adverse effects , Cancer Vaccines/pharmacokinetics , Combined Modality Therapy , Consumer Product Safety , Disease-Free Survival , Dose-Response Relationship, Immunologic , Female , Granulocyte-Macrophage Colony-Stimulating Factor/adverse effects , Granulocyte-Macrophage Colony-Stimulating Factor/pharmacokinetics , Humans , Hypersensitivity, Delayed/pathology , Male , Middle Aged , Pancreatic Neoplasms/immunology , Pancreatic Neoplasms/pathology
19.
Curr Protoc Hum Genet ; Chapter 13: Unit 13.8, 2001 May.
Article in English | MEDLINE | ID: mdl-18428256

ABSTRACT

This unit describes the use of retroviral vectors that can be successfully employed for gene transfer into both primary tumor cultures and established cell lines. The unit includes procedures for assaying the stability of the vaccine following gene transfer. Techniques for maintaining the retroviral producer lines and titering the retroviral vectors are also described. A protocol for frozen storage of the vaccine (transduced tumor cells) is provided. In addition, methods are described for characterizing the vaccine cells following gene transfer. Directions for testing the expression of the transferred gene in the transfected tumor line are also given. The final protocol provides suggestions for designing in vivo animal experiments and discusses what has to be observed in a clinical setting.


Subject(s)
Cancer Vaccines/genetics , Neoplasms/therapy , Animals , Cytokines/biosynthesis , Cytokines/genetics , Gene Transfer Techniques , Genetic Vectors , Genetics, Medical , Humans , Mice , Neoplasms/genetics , Neoplasms/immunology , Retroviridae/genetics , T-Lymphocytes/immunology , Tumor Cells, Cultured
20.
Int J Radiat Oncol Biol Phys ; 48(4): 1089-96, 2000 Nov 01.
Article in English | MEDLINE | ID: mdl-11072167

ABSTRACT

PURPOSE: (1) To determine the toxicity of an intensified postoperative adjuvant regimen for periampullary adenocarcinoma (pancreatic and nonpancreatic) utilizing concurrent 5-fluorouracil (5-FU), leucovorin (LV), dipyridamole (DPM), and mitomycin-C (MMC) combined with split-course locoregional external beam radiotherapy (EBRT) to 50 Gy. This was followed by 4 cycles of the same chemotherapy as adjuvant therapy. (2) To determine preliminary estimates of the overall and disease-free survival associated with the use of this regimen. (3) To compare the toxicities and early survival results of patients treated with the current regimen to those of patients who completed our prior trial of concurrent chemoradiation infusion with 5-FU/LV chemotherapy and regional nodal and prophylactic hepatic irradiation. METHODS: Postpancreaticoduodenectomy, patients received every 4 weeks bolus administration of 5-FU, (400 mg/m(2)), and LV, (20 mg/m(2), Days l-3), DPM (75 mg p.o., 4 times per day, Days 0-3, and every 8 weeks), MMC, (10 mg/m(2); maximum of 20 mg, Day l during EBRT). This was followed by 4 months of the same chemotherapy, beginning 1 month following the completion of EBRT. EBRT consisted of split-course 5000 cGy/20 fractions with a 2-week planned rest after the first 10 fractions (2500 cGy). RESULTS: From 4/96 to 6/99, 45 patients were enrolled and treated. Their experience constitutes the basis of this analysis. There were 29 patients with pancreatic cancer and 16 with nonpancreatic periampullary cancer. Seventeen patients had tumors of 3 cm or more, and 39 patients had at least 1 histologically involved lymph node. Thirteen patients had a histologically positive margin of resection. The mean time to start of treatment was 63 days following surgery. During chemoradiation therapy there were no Grade 3 or worse nonhematologic toxicities and 47% Grade 3 or Grade 4 hematologic toxicities of short duration. Following chemoradiation, during chemotherapy treatment only, there was one Grade 3 hepatic and one Grade 3 pulmonary toxicity which was nondebilitating (2% each case) and 42% Grade 3 or 4 hematologic toxicity. There were 2 episodes of neutropenic fever requiring admission and no treatment-related mortalities. One patient developed a mild case of HUS, which responded to standard management. One patient developed persistent shortness of breath (nondebilitating), and another patient had occasional dyspnea on exertion, both occurring after all therapy. The majority of patients complained of increased fatigue (Grade 1-2), greatest during the combined therapy and improving post all treatment. As of 6/23/99, 20 of 45 patients have relapsed, 13 in the liver. Twelve patients have died. Median follow-up for surviving patients is 14.3 months. Disease-free survival at 12 months following surgery is 66% (as compared to 25% in our prior study), and the median disease-free survival is 17 months (as compared to 8. 3 months in our prior study). Median survival has not yet been reached, but will be greater than 17 months. CONCLUSION: With a 14.3-month median follow-up, acute toxicity has been acceptable and manageable. Observed relapses were seen 9-13 months following surgical resection. Early survival analysis suggests a trend toward increased median disease-free survival (8.3 vs. 17 months), especially for patients with nonpancreatic periampullary adenocarcinoma.


Subject(s)
Adenocarcinoma/therapy , Pancreatic Neoplasms/therapy , Adenocarcinoma/mortality , Adenocarcinoma/pathology , Adult , Aged , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Chemotherapy, Adjuvant , Dipyridamole/administration & dosage , Dipyridamole/adverse effects , Disease-Free Survival , Duodenal Neoplasms/mortality , Duodenal Neoplasms/pathology , Duodenal Neoplasms/therapy , Female , Fluorouracil/administration & dosage , Fluorouracil/adverse effects , Follow-Up Studies , Humans , Leucovorin/administration & dosage , Leucovorin/adverse effects , Male , Middle Aged , Mitomycin/administration & dosage , Mitomycin/adverse effects , Neoplasm Recurrence, Local , Pancreatic Neoplasms/mortality , Pancreatic Neoplasms/pathology , Pancreaticoduodenectomy , Radiotherapy Dosage , Radiotherapy, Adjuvant , Time Factors
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