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Biomedical subjects

Monica C Panelli

Publications and source records attributed to Monica C Panelli.

At least 19 recordsLinked to original sources
PubMed2007

Sequential gene profiling of basal cell carcinomas treated with imiquimod in a placebo-controlled study defines the requirements for tissue rejection.

BACKGROUND: Imiquimod is a Toll-like receptor-7 agonist capable of inducing complete clearance of basal cell carcinoma (BCC) and other cutaneous malignancies. We hypothesized that the characterization of the early transcriptional events induced by imiquimod may provide insights about immunological events preceding acute tissue and/or tumor rejection. RESULTS: We report a paired analysis of adjacent punch biopsies obtained pre- and post-treatment from 36 patients with BCC subjected to local application of imiquimod (n = 22) or vehicle cream (n = 14) in a blinded, randomized protocol. Four treatments were assessed (q12 applications for 2 or 4 days, or q24 hours for 4 or 8 days). RNA was amplified and hybridized to 17.5 K cDNA arrays. All treatment schedules similarly affected the transcriptional profile of BCC; however, the q12 x 4 days regimen, associated with highest effectiveness, induced the most changes, with 637 genes unequivocally stimulated by imiquimod. A minority of transcripts (98 genes) confirmed previous reports of interferon-alpha involvement. The remaining 539 genes portrayed additional immunological functions predominantly involving the activation of cellular innate and adaptive immune-effector mechanisms. Importantly, these effector signatures recapitulate previous observations of tissue rejection in the context of cancer immunotherapy, acute allograft rejection and autoimmunity. CONCLUSION: This study, based on a powerful and reproducible model of cancer eradication by innate immune mechanisms, provides the first insights in humans into the early transcriptional events associated with immune rejection. This model is likely representative of constant immunological pathways through which innate and adaptive immune responses combine to induce tissue destruction.

Aminoquinolines↗
PubMed2006

Comprehensive epitope mapping of the Epstein-Barr virus latent membrane protein-2 in normal, non tumor-bearing individuals.

Latent membrane protein (LMP)-2 is one of the Epstein-Barr virus (EBV)-encoded proteins consistently expressed by nasopharyngeal carcinoma (NPC). EBV-transformed lymphoblastoid cell lines (LCL) have been used in patients with NPC to induce LMP-2-recognizing T cell lines which have been in turn utilized for protein-wide mapping of T cell epitopes. However, comprehensive mapping of naturally recognized LMP-2 epitopes in non tumor-bearing individuals has not been reported. Here, we applied a low sensitivity epitope-defining technique for the identification of LMP-2 CTL responses detectable ex vivo in EBV-experienced individuals. This screening tool has been previously validated by analyzing memory CTL responses to Flu, cytomegalovirus (CMV), and the melanoma associated antigen gp100/Mel17. Peripheral blood monocytes (PBMC) from ten Caucasian and ten Chinese individuals were stimulated ex vivo with pools of nonamer (9-mer) peptides overlapping in a stepwise fashion each single amino acid of the LMP-2 sequence. No obvious differences were observed between the immune response of the two ethnic groups save for those related to the divergence in the ethnic prevalence of HLA haplotypes. Several novel and known LMP-2 epitopes were identified. Reactivity toward at least one LMP-2 epitope was detected in 18 of the 20 donors but no prevalent human leukocyte antigen (HLA)/epitope combination was observed confirming that LMP-2 reactivity in the context of common HLA alleles is more pleiotropic than that of FLU and CMV. We believe that the usefulness of these epitopes occurring naturally in non-cancer bearing patients as reagents for the immunization of patients with early or advanced stage NPC deserves further evaluation.

Amino Acid Sequence↗
PubMed2006

Comparison of proteomic profiles of serum, plasma, and modified media supplements used for cell culture and expansion.

BACKGROUND: The culture and expansion of human cells for clinical use requires the presence of human serum or plasma in culture media. Although these supplements have been extensively characterized in their chemical composition, only recently it has been possible to provide by high throughput protein analysis, a comprehensive profile of the soluble factors contributing to cell survival. This study analyzed and compared the presence of 100 proteins including chemokines, cytokines and soluble factors in six different types of media supplements: serum, plasma, recalcified plasma, heat inactivated serum, heat inactivated plasma and heat inactivated recalcified plasma. METHODS: Serum, plasma, recalcified plasma, and heat inactivated supplements were prepared from ten healthy subjects. The levels of 100 soluble factors were measured in each sample using a multiplexed ELISA assay and compared by Eisen hierarchical clustering analysis. RESULTS: A comparison of serum and plasma levels of soluble factors found that 2 were greater in plasma but 18 factors were greater in serum including 11 chemokines. The levels of only four factors differed between recalcified plasma and plasma. Heat inactivation had the greatest effect on soluble factors. Supervised Eisen hierarchical clustering indicated that the differences between heat inactivated supplements and those that were not were greater than the differences within these two groups. The levels of 36 factors differed between heat inactivated plasma and plasma. Thirty one of these factors had a lower concentration in heat inactivated plasma including 12 chemokines, 4 growth factors, 4 matrix metalloproteases, and 3 adhesion molecules. Heat inactivated decalcified plasma is often used in place of heat inactivated serum and the levels of 19 soluble factors differed between these two supplements. CONCLUSION: Our report provides a comprehensive protein profile of serum, plasma recalcified plasma, and heat inactivated supplements. This profile represents a qualitative and quantitative database that can aid in the selection of the appropriate blood derived supplement for human cell cultures with special requirements.

Journal Article↗
PubMed2006

MHC-peptide specificity and T-cell epitope mapping: where immunotherapy starts.

The evaluation and characterization of epitope-specific human leukocyte antigen (HLA)-restricted memory T-cell reactivity is an important step for the development of preventive vaccines and peptide-based immunotherapies for viral and tumor diseases. The past decade has witnessed the use of HLA-restricted peptides as tools to activate strong immune responses of naïve or memory T cells specifically. This has fuelled an active search for methodological approaches focusing on HLA and peptide associations. Here, we outline new perspective on the emerging opportunity of evaluating HLA and peptide restriction by using novel approaches, such as quantitative real-time PCR, that can identify epitope specificities that are potentially useful in clinical settings.

Animals↗
PubMed2006

Molecular signatures induced by interleukin-2 on peripheral blood mononuclear cells and T cell subsets.

Experimentally, interleukin-2 (IL-2) exerts complex immunological functions promoting the proliferation, survival and activation of T cells on one hand and inducing immune regulatory mechanisms on the other. This complexity results from a cross talk among immune cells which sways the effects of IL-2 according to the experimental or clinical condition tested. Recombinant IL-2 (rIL-2) stimulation of peripheral blood mononuclear cells (PBMC) from 47 donors of different genetic background induced generalized T cell activation and anti-apoptotic effects. Most effects were dependent upon interactions among immune cells. Specialized functions of CD4 and CD8 T cells were less dependent upon and often dampened by the presence of other PBMC populations. In particular, cytotoxic T cell effector function was variably affected with a component strictly dependent upon the direct stimulation of CD8 T cells in the absence of other PBMC. This observation may provide a roadmap for the interpretation of the discrepant biological activities of rIL-2 observed in distinct pathological conditions or treatment modalities.

Journal Article↗
PubMed2006

Common cancer biomarkers.

There is an increasing interest in complementing conventional histopathologic evaluation with molecular tools that could increase the sensitivity and specificity of cancer staging for diagnostic and prognostic purposes. This study strove to identify cancer-specific markers for the molecular detection of a broad range of cancer types. We used 373 archival samples inclusive of normal tissues of various lineages and benign or malignant tumors (predominantly colon, melanoma, ovarian, and esophageal cancers). All samples were processed identically and cohybridized with an identical reference RNA source to a custom-made cDNA array platform. The database was split into training (n = 201) and comparable prediction (n = 172) sets. Leave-one-out cross-validation and gene pairing analysis identified putative cancer biomarkers overexpressed by malignant lesions independent of tissue of derivation. In particular, seven gene pairs were identified with high predictive power (87%) in segregating malignant from benign lesions. Receiver operator characteristic curves based on the same genes could segregate malignant from benign tissues with 94% accuracy. The relevance of this study rests on the identification of a restricted number of biomarkers ubiquitously expressed by cancers of distinct histology. This has not been done before. These biomarkers could be used broadly to increase the sensitivity and accuracy of cancer staging and early detection of locoregional or systemic recurrence. Their selective expression by cancerous compared with paired normal tissues suggests an association with the oncogenic process resulting in stable expression during disease progression when the presently used differentiation markers are unreliable.

Biomarkers, Tumor↗
PubMed2006

Detection of human MCP-4/CCL13 isoforms by SELDI immunoaffinity capture.

Monocyte Chemoattractant Proteins 4 (MCP-4/CCL13) is a member of a distinct, structurally-related subclass of CC chemokines mainly involved in recruitment of eosinphils to inflammatory sites. Recent evidence demonstrates that serum level of this protein strongly increases following high dose IL-2 immunotherapy. The physiological form of human MCP-4/CCL13 has yet to be purified. Therefore, the primary structure of the biologically relevant (mature) form has not been established. By using SELDI immunoaffinity capture technology we describe two mature isoforms both present in serum before and after high-dose IL-2 immunotherapy.

Journal Article↗
PubMed2006

Inflammatory protein profile during systemic high dose interleukin-2 administration.

Systemic interleukin-2 (IL-2) administration induces an assortment of downstream effects whose biological and therapeutic significance remains unexplored mostly because of the methodological inability to globally address their complexity. Protein array analysis of sera from patients with renal cell carcinoma obtained prior and during high-dose IL-2 therapy had previously revealed extensive alterations in expression of the soluble factors analyzed, whose discovery was limited by the number of capture antibodies selected for protein detection. Here, we expanded the analysis to SELDI-TOF-MS and quantitative protein analysis (nephelometry). All cytokines/chemokines detected by protein arrays were below the SELDI detection limit, while novel IL-2-specific changes in expression of acute-phase reactants and high-density lipoprotein metabolites could be identified. Serum amyloid protein A (SAA) and C-reactive protein expression were consistently up-regulated after four doses of IL-2, while other proteins were down-regulated. These findings were confirmed by SELDI immunoaffinity capture and nephelometry. Immunoaffinity capture revealed different, otherwise undetectable, isoforms of SAA. A linear correlation between peak area by SELDI and protein concentration by nephelometry was observed. Overall distinct yet complementary information was obtained using different platforms, which may better illustrate complex phenomena such as the systemic response to biological response modifiers.

Biomarkers↗
PubMed2005

Delayed polarization of mononuclear phagocyte transcriptional program by type I interferon isoforms.

BACKGROUND: Interferon (IFN)-alpha is considered a key modulator of immunopathological processes through a signature-specific activation of mononuclear phagocytes (MPs). This study utilized global transcript analysis to characterize the effects of the entire type I IFN family in comparison to a broad panel of other cytokines on MP previously exposed to Lipopolysaccharide (LPS) stimulation in vitro. RESULTS: Immature peripheral blood CD14+ MPs were stimulated with LPS and 1 hour later with 42 separate soluble factors including cytokines, chemokines, interleukins, growth factors and IFNs. Gene expression profiling of MPs was analyzed 4 and 9 hours after cytokine stimulation. Four hours after stimulation, the transcriptional analysis of MPs revealed two main classes of cytokines: one associated with the alternative and the other with the classical pathway of MP activation without a clear polarization of type I IFNs effects. In contrast, after 9 hours of stimulation most type I IFN isoforms induced a characteristic and unique transcriptional pattern separate from other cytokines. These "signature" IFNs included; IFN-beta, IFN-alpha2b/alpha2, IFN-alphaI, IFN-alpha2, IFN-alphaC, IFN-alphaJ1, IFN-alphaH2, and INF-alpha4B and induced the over-expression of 44 genes, all of which had known functional relationships with IFN such as myxovirus resistance (Mx)-1, Mx-2, and interferon-induced hepatitis C-associated microtubular aggregation protein. A second group of type I IFNs segregated separately and in closer association with the type II IFN-gamma. The phylogenetic relationship of amino acid sequences among type I IFNs did not explain their sub-classification, although differences at positions 94 through 109 and 175 through 189 were present between the signature and other IFNs. CONCLUSION: Seven IFN-alpha isoforms and IFN-beta participate in the late phase polarization of MPs conditioned by LPS. This information broadens the previous view of the central role played by IFN-alpha in autoimmunity and tumor rejection by including and/or excluding an array of related factors likely to be heterogeneously expressed by distinct sub-populations of individuals in sickness or in response to biological therapy.

Journal Article↗
PubMed2005

Understanding the response to immunotherapy in humans.

Whether the efforts of the last decade aimed at the development of vaccines against tumor-specific antigens encountered success or failure is a matter of expectations. On the bright side, we could optimistically observe that anti-cancer-vaccines stand as an outstanding example of the successful implementation of modern biotechnology tools for the development of biologically sound therapeutics. In particular, vaccines against melanoma (the prototype model of tumor immunology in humans) can reproducibly induce cytotoxic T cell (CTL) responses exquisitely specific for cancer cells. This achievement trespasses the specificity of any other anti-cancer therapy. The skeptics, on the other end, might point out that immunization only rarely leads to cancer regression, labeling, therefore, this approach is ineffective. In our opinion this judgment stems from the naïve expectation that CTL induction is sufficient for an effective immune response. Here we propose that more needs to be understood about the mechanisms required for the induction of a therapeutically relevant immune response in humans. In particular, we will discuss the variables related to cancer heterogeneity, the weight of individual patients' polymorphism(s), the role of the T cell activation and differentiation and, finally, the complex relationship between immune and cancer cells within the tumor microenvironment.

Antigens, Neoplasm↗
PubMed2005

Polarized monocyte response to cytokine stimulation.

BACKGROUND: Mononuclear phagocytes (MPs) stand at the crossroads between the induction of acute inflammation to recruit and activate immune effector cells and the downmodulation of the inflammatory process to contain collateral damage. This decision is extensively modulated by the cytokine microenvironment, which includes a broad array of cytokines whose direct effect on MPs remains largely unexplored. Therefore, we tested whether polarized responses of MPs to pathogens are related to the influence of selected cytokines or represent a mandatory molecular switch through which most cytokines operate. RESULTS: Circulating CD14+ MPs were exposed to bacterial lipopolysaccharide (LPS) followed by exposure to an array of cytokines, chemokines and soluble factors involved in the immune response. Gene expression was studied by global transcript analysis. Two main classes of cytokines were identified that induced a classical or an alternative pathway of MP activation. Expression of genes affected by NFkappaB activation was most predictive of the two main classes, suggesting that this pathway is a fundamental target of cytokine regulation. As LPS itself induces a classical type of activation, the most dramatic modulation was observed toward the alternative pathway, suggesting that a broad array of cytokines may counteract the pro-inflammatory effects of bacterial components. CONCLUSIONS: This analysis is directly informative of the primary effect of individual cytokines on the early stages of LPS stimulation and, therefore, may be most informative of the way MP maturation may be polarized at the early stages of the immune response.

Adult↗
PubMed2005

Peritoneal and subperitoneal stroma may facilitate regional spread of ovarian cancer.

PURPOSE: Epithelial ovarian cancer (EOC) is characterized by early peritoneal involvement ultimately contributing to morbidity and mortality. To study the role of the peritoneum in fostering tumor invasion, we analyzed differences between the transcriptional repertoires of peritoneal tissue lacking detectable cancer in patients with EOC versus benign gynecologic disease. EXPERIMENTAL DESIGN: Specimens were collected at laparotomy from patients with benign disease (b) or malignant (m) ovarian pathology and comprised primary ovarian tumors, paired bilateral specimens from adjacent peritoneum and attached stroma (PE), subjacent stroma (ST), peritoneal washes, ascites, and peripheral blood mononuclear cells. Specimens were immediately frozen. RNA was amplified by in vitro transcription and cohybridized with reference RNA to a custom-made 17.5k cDNA microarray. RESULTS: Principal component analysis and unsupervised clustering did not segregate specimens from patients with benign or malignant pathology. Class comparison identified differences between benign and malignant PE and ST specimens deemed significant by permutation test (P = 0.027 and 0.012, respectively). A two-tailed Student's t test identified 402 (bPE versus mPE) and 663 (mST versus bST) genes differentially expressed at a significance level of P2 < or = 0.005 when all available paired samples from each patient were analyzed. The same comparison using one sample per patient reduced the pool of differentially expressed genes but retained permutation test significance for bST versus mST (P = 0.031) and borderline significance for bPE versus mPE (P = 0.056) differences. CONCLUSIONS: The presence of EOC may foster peritoneal implantation and growth of cancer cells by inducing factors that may represent molecular targets for disease control.

Computational Biology↗
PubMed2005

Gene profiling of immune responses against tumors.

Clinical trials of tumor-antigen-specific immunization have clearly shown that immune-mediated tumor rejection requires more than simple T cell-target cell interactions. In vivo generation of tumor-specific T cells is one of a series of steps necessary for the induction of clinically relevant immune responses. In recent years, high-throughput functional genomics exposed the complexity of tumor immune biology, which underlies the kaleidoscopic array of variables associated with cancer instability and immunogenetic variability in humans. In the quest to understand immune rejection, hypothesis-driven approaches have failed to take into account the intricacy of human pathology by relying mostly on hypotheses derived from experimental models rather than direct clinical observation. Future investigations should reframe scientific thinking when applied to humans, utilizing descriptive tools to generate novel hypotheses relevant to human disease.

Gene Expression Profiling↗
PubMed2005

Proteomic signature of myeloproliferation and neutrophilia: analysis of serum and plasma from healthy subjects given granulocyte colony-stimulating factor.

OBJECTIVE: Proteomic analysis could improve our understanding of the mechanisms and consequences of myeloproliferation. Healthy subjects treated with granulocyte colony-stimulating factor (G-CSF) were used as a model of myeloproliferation. METHODS: Levels of 80 soluble factors were measured by enzyme-linked immunosorbent assay before and after 5 days of G-CSF. Both serum and plasma levels were measured to generate a comprehensive profile and determine whether serum or plasma best portrays biological and physiological changes. RESULTS: Comparison of samples collected prior to G-CSF demonstrated that 44 factors differed between serum and plasma. Concentrations of several growth factors and chemokines were greater in serum than in plasma, while the opposite was true for several interleukins. Following G-CSF serum levels of 14 factors and plasma levels of 15 factors changed. Eleven increased in both serum and plasma, including cell adhesion molecules (vascular cell adhesion molecule-1, E-selectin, and L-selectin), matrix metalloproteases (MMP-1, -8, and -13), cytokine receptors (tumor necrosis factor receptor 1 and 2, and interleukin-2 receptor), the acute phase reactant, serum amyloid A, and a growth factor, hepatocyte growth factor. CONCLUSION: Some protein levels differ markedly in serum and plasma. Myeloproliferation is associated with changes in the levels of several proteases, adhesion molecules, and cytokines.

Blood Proteins↗
PubMed2004

Ontogeny and oncogenesis balance the transcriptional profile of renal cell cancer.

Global transcript analysis is increasingly used to describe cancer taxonomies beyond the microscopic reach of the eye. Diagnostic and prognostic portraits are formulated by ranking cancers according to transcriptional proximity. However, the role that distinct biological factors play in defining these portraits remains undefined. It is likely that the transcriptional repertoire of cancers depends, on one hand, on the anamnestic retention of their ontogenesis and, on the other, on the emergence of novel expression patterns related to oncogenesis. We compared the transcriptional profile of primary renal cell cancers (RCCs) with that of normal kidney tissue and several epithelial cancers of nonrenal origin to weigh the contribution that ontogeny and oncogenesis make in molding their genetic profile. Unsupervised global transcript analysis demonstrated that RCCs retain transcriptional signatures related to their ontogeny and cluster close to normal renal epithelium. When renal lineage-associated genes are removed from the analysis and cancer-specific genes are analyzed, RCCs segregate with other cancers with limited lineage specificity underlying a predominance of the oncogenic process over lineage specificity. However, a RCC-specific set of oncogenesis-related genes was identified and surprisingly shared by sarcomas. In summary, the transcriptional portrait of primary RCCs is largely dominated by ontogeny. Genes responsible for lineage specificity may represent poor molecular targets for immune or drug therapy. Most genes associated with oncogenesis are shared with other cancers and may represent better therapeutic targets. Finally, a small subset of genes is associated with lineage-specific oncogenesis, and these may provide information regarding the biological behavior of RCCs and facilitate diagnostic classification of RCCs.

Adenocarcinoma, Clear Cell↗
PubMed2004

Melanoma-restricted genes.

Human metastatic cutaneous melanoma has gained a well deserved reputation for its immune responsiveness. The reason(s) remain(s) unknown. We attempted previously to characterize several variables that may affect the relationship between tumor and host immune cells but, taken one at the time, none yielded a convincing explanation. With explorative purposes, high-throughput technology was applied here to portray transcriptional characteristics unique to metastatic cutaneous melanoma that may or may not be relevant to its immunogenic potential. Several functional signatures could be identified descriptive of immune or other biological functions. In addition, the transcriptional profile of metastatic melanoma was compared with that of primary renal cell cancers (RCC) identifying several genes co-coordinately expressed by the two tumor types. Since RCC is another immune responsive tumor, commonalities between RCC and melanoma may help untangle the enigma of their potential immune responsiveness. This purely descriptive study provides, therefore, a map for the investigation of metastatic melanoma in future clinical trials and at the same time may invite consideration of novel therapeutic targets.

Journal Article↗
PubMed2004

Selection and validation of endogenous reference genes using a high throughput approach.

BACKGROUND: Endogenous reference genes are commonly used to normalize expression levels of other genes with the assumption that the expression of the former is constant in different tissues and in different physiopathological conditions. Whether this assumption is correct it is, however, still matter of debate. In this study, we searched for stably expressed genes in 384 cDNA array hybridization experiments encompassing different tissues and cell lines. RESULTS: Several genes were identified whose expression was highly stable across all samples studied. The usefulness of 8 genes among them was tested by normalizing the relative gene expression against test genes whose expression pattern was known. The range of accuracy of individual endogenous reference genes was wide whereas consistent information could be obtained when information pooled from different endogenous reference genes was used. CONCLUSIONS: This study suggests that even when the most stably expressed genes in array experiments are used as endogenous reference, significant variation in test gene expression estimates may occur and the best normalization is achieved when data from several endogenous reference genes are pooled together to minimize minimal but significant variation among samples. We are presently optimizing strategies for the preparation of endogenous reference gene mixtures that could yield information comparable to that of data pooled from individual endogenous reference gene normalizations.

Carcinoma↗