Francesco Marincola - Academia.edu (original) (raw)

Papers by Francesco Marincola

Research paper thumbnail of Workshop on cancer biometrics: identifying biomarkers and surrogates of cancer in patients: a meeting held at the Masur Auditorium, National Institutes of Health

Journla of Immunotherapy

The current excitement about molecular targeted therapies has driven much of the recent dialog in... more The current excitement about molecular targeted therapies has driven much of the recent dialog in cancer diagnosis and treatment. Particularly in the biologic therapy of cancer, identifiable antigenic T-cell targets restricted by MHC molecules and the related novel stress molecules such as MICA/B and Letal allow a degree of precision previously unknown in cancer therapy. We have previously held workshops on immunologic monitoring and angiogenesis monitoring. This workshop was designed to discuss the state of the art in identification of biomarkers and surrogates of tumor in patients with cancer, with particular emphasis on assays within the blood and tumor. We distinguish this from immunologic monitoring in the sense that it is primarily a measure of the tumor burden as opposed to the immune response to it. Recommendations for intensive investigation and targeted funding to enable such strategies were developed in seven areas: genomic analysis; detection of molecular markers in peri...

Research paper thumbnail of Cancer vaccines: from research to clinical practice

Research paper thumbnail of Identification of immune dominant cytomegalovirus epitopes using quantitative real-time polymerase chain reactions to measure interferon-gamma production by peptide-stimulated peripheral blood mononuclear cells

Journla of Immunotherapy

The identification of HLA restricted immune dominant cytotoxic T cell (CTL) epitopes limits immun... more The identification of HLA restricted immune dominant cytotoxic T cell (CTL) epitopes limits immune therapy. Cytomegalovirus (CMV) disease remains a significant cause of morbidity after allogeneic stem cell transplantation. Adoptive immune therapy using CTLs stimulated with immune dominant CMV pp65 peptides may be effective in preventing CMV disease, but immune dominant CMV peptides have been identified for only a few HLA class I molecules. The purpose of this study was to use a novel molecular system to establish a rapid and precise method to identify new HLA-restricted CMV epitopes. Cytomegalovirus pp65 peptides expected to bind to the HLA-24 molecule were identified with a computer algorithm. Five candidate peptides were screened by direct ex vivo stimulation of peripheral blood mononuclear cells (PBMCs) from CMV-seropositive HLA-A*2402 individuals, and quantitative real time PCR (qRT-PCR) was used to evaluate CTL responses by measuring interferon-gamma (IFN-gamma) transcripts. On...

Research paper thumbnail of New HLA-A, -B, and -C locus-specific primers for PCR amplification from cDNA: application in clinical immunology

Journal of Immunological Methods

The individual cellular immune response to intracellular antigens is modeled by the highly polymo... more The individual cellular immune response to intracellular antigens is modeled by the highly polymorphic major histocompatibility complex (HLA) class I molecules. The epitopes presented and the T cell repertoire that recognizes them depend on the HLA constitution of the individual. Therefore, to monitor and to modify an individual's HLA class I-driven cellular immune response, it is necessary to know the HLA class I alleles of the person and the possible epitopes of the target antigen presented by those alleles. In particular, this is necessary in order to design peptide-based vaccines and immune therapies for the treatment of diseases caused by viruses, intracellular parasites or cancer, and to monitor the immune response during those treatments. We describe a new set of HLA-A, -B, and -C locus-specific primers for the polymerase chain reaction (PCR) amplification of the whole coding sequence of these genes from complementary DNA (cDNA). We describe their use for typing and for t...

Research paper thumbnail of The matrix protein pp65(341-350): a peptide that induces ex vivo stimulation and in vitro expansion of CMV-specific CD8+ T cells in subjects bearing either HLA-A*2402 or A*0101 allele

Transfusion

The stimulation of PBMNCs with HLA Class I restricted synthetic peptides derived from CMV phospho... more The stimulation of PBMNCs with HLA Class I restricted synthetic peptides derived from CMV phosphorylated matrix protein 65 (pp65) evokes CMV-specific cytotoxic T lymphocyte (CTL) activity, a necessary condition for initiating adoptive immunotherapy against CMV-related diseases in immune-compromised patients. It was previously demonstrated that the CMV decamer (10-mer) peptide pp65(341-350), QYDPVAALFF, was able to induce CMV-specific CTLs in HLA-A*2402 CMV-seropositive individuals. We investigated the ability of the peptide pp65(341-350) to reactivate memory CD8+ T cells in CMV-seropositive subjects bearing either the HLA-A24 or A1 allele. CTL responses were measured by IFN-gamma mRNA expression and IFN-gamma protein production as well as cytotoxic activity. In this study it was found that peptide pp65(341-350) induced a specific reactivation of memory CD8+ T cells from CMV-seropositive donors expressing either HLA-A*2402 and/or HLA-A*0101. Moreover, a pp65(341-350)-specific selecti...

Research paper thumbnail of CMV pp65 and IE-1 T cell epitopes recognized by healthy subjects

Journal of Translational Medicine

Adoptive immune and vaccine therapies have been used to prevent cytomegalovirus (CMV) disease in ... more Adoptive immune and vaccine therapies have been used to prevent cytomegalovirus (CMV) disease in recipients of hematopoietic progenitor cell transplants, but the nature of T cell responses to CMV have not been completely characterized. Peptide pools and individual peptides derived from the immune-dominant CMV proteins pp65 and IE-1 and antigen-specific, cytokine flow cytometry were used to characterize the prevalence and frequency of CD4+ and CD8+ memory T cells in 20 healthy CMV-seropositive subjects. CD8+ T cell responses to pp65 were detected in 35% of subjects and to IE-1 in 40% of subjects. CD4+ T cell responses to pp65 were detected in 50% of subjects, but none were detected to IE-1. Several new IE-1 HLA class I epitopes were identified, including 4 restricted to HLA-C antigens. One region of IE-1 spanning amino acids 300 to 327 was rich in class I epitopes. The HLA class I restrictions of IE-1 peptides were more promiscuous than those of pp65 peptides. Since naturally occurri...

Research paper thumbnail of Consensus nomenclature for CD8 + T cell phenotypes in cancer

OncoImmunology, 2015

Whereas preclinical investigations and clinical studies have established that CD8(+) T cells can ... more Whereas preclinical investigations and clinical studies have established that CD8(+) T cells can profoundly affect cancer progression, the underlying mechanisms are still elusive. Challenging the prevalent view that the beneficial effect of CD8(+) T cells in cancer is solely attributable to their cytotoxic activity, several reports have indicated that the ability of CD8(+) T cells to promote tumor regression is dependent on their cytokine secretion profile and their ability to self-renew. Evidence has also shown that the tumor microenvironment can disarm CD8(+) T cell immunity, leading to the emergence of dysfunctional CD8(+) T cells. The existence of different types of CD8(+) T cells in cancer calls for a more precise definition of the CD8(+) T cell immune phenotypes in cancer and the abandonment of the generic terms "pro-tumor" and…

Research paper thumbnail of Functional Heterogeneity of

Research paper thumbnail of cDNA arrays and the enigma of melanoma immune responsiveness

The Cancer Journal

Melanoma is a cancer characterized by predisposition to natural and induced immune rejection. The... more Melanoma is a cancer characterized by predisposition to natural and induced immune rejection. The occurrence of this phenomenon, however, remains sporadic and capricious while there is no clear understanding of the chain of events responsible. With the progression of the understanding of the molecular immunology of melanoma during the last decade, several theories suggesting immune tolerance (inadequate immune response) and/or immune escape (adaptation of cancer cells to a vigorous immune pressure) as possible modulators of melanoma growth have emerged. As the number of these hypotheses, mostly based on molecular or pre-clinical models, has increased exponentially, the relevance of each individual one has been conversely decreasing leaving the observer without confidence that a theory likely to explain this phenomenon can be soon formulated. In this review, we will discuss a direct approach whereby new theories might be identified by direct observation of human phenomena as they occ...

Research paper thumbnail of Use of laser scanning cytometry to study tumor microenvironment

Histology and histopathology

The study of phenomena occurring in the tumor microenvironment is a challenging task because of t... more The study of phenomena occurring in the tumor microenvironment is a challenging task because of technical difficulties, particularly when dealing with hypocellular specimens. Laser scanning cytometry (LSC) is a new laboratory technology that has been recently introduced to overcome the limitations of other traditional technologies. By combining the properties and the advantages of flow cytometry (FC) and immunohistochemistry (IHC), LSC allows the investigator to obtain objective information on DNA content, protein expression and cellular localization is combination with morphological features. It has been already shown that LSC results are reliable compared to more traditional technologies, and its implementation in the clinical routine is under way. Its use in oncology, which is rapidly expanding, spans from apoptosis analysis to DNA content quantitation and tumor cell phenotyping. Here we describe the technology underlying this novel fluorescence-based device, review its use in on...

Research paper thumbnail of The dual role of IL10

Classification of cytokines as pro- versus anti-inflammatory might not apply to the pleiotropic e... more Classification of cytokines as pro- versus anti-inflammatory might not apply to the pleiotropic effects of interleukin-10 (IL-10). Several reports suggest that IL-10 enhances the function of natural killer cells, which leads, through pathogen destruction, to increased antigen availability. In addition, by inhibiting the maturation of antigen-presenting cells (APCs), IL-10 preserves their ability for antigen uptake while simultaneously hampering their migration to draining lymph nodes. This review suggests that this ‘antigen-loading’ phase might constitute an important component of the innate immune reaction to a pathogen. Additional proinflammatory stimuli might subsequently lead to maturation of ‘loaded’ APCs that could migrate to draining lymph nodes or recruit and activate adaptive immune effectors locally.

Research paper thumbnail of A global approach to tumor immunology

Cellular & molecular immunology

Biological and clinical advances in the understanding of tumor immunology suggest that immune res... more Biological and clinical advances in the understanding of tumor immunology suggest that immune responsiveness of human tumors is a complex biological phenomenon that could be best studied by a real-time comparison of tumor/host interactions in the tumor microenvironment through a high-throughput discovery-driven approach. This conclusion is derived from our recognition that too many hypotheses or, in other words, no solid single hypothesis exist, based on experimental results, to further drive experimentation in human subjects. Functional genomic studies entertained during the last few years consolidated the belief that in humans the interactions between tumor and immune cells are too complex to be approached exclusively with a hypothesis driven method. We believe that immune cells suit cancer cells in a Yin and Yang balance by opposing and yet mutually depending on each other. Indeed, immune infiltration in tumors may play a dual role modulating in different circumstances cancer cel...

Research paper thumbnail of The enigma of tumor immune responsiveness: A high-throughput genomics question?

Tissue Antigens

Much interest is currently being directed towards serotonin (5-HT) receptors of type 2C (5-HT ) b... more Much interest is currently being directed towards serotonin (5-HT) receptors of type 2C (5-HT ) because of their possible 2C involvement in the control of different activities, such as the composition of the cerebrospinal fluid, locomotion, feeding, neuronal excitability and anxiety. The limited information regarding their distribution in the human brain prompted us to investigate, and to 3 characterize the binding of [ H]mesulergine, a HT antagonist, in autopsy samples from 24 subjects. 2C 3 The results showed that the [ H]mesulergine binding represented 95% of the total binding and equilibrium saturation binding experiments resulted in a single straight line, consistent with the presence of one site only. The area with the highest density of 3 [ H]mesulergine binding was the choroid plexus, followed at a significantly lower level by the hippocampus, substantia nigra, basal 3 ganglia, amygdala, hypothalamus and prefrontal cortex. The pharmacological profile of the [ H]mesulergine binding was consistent with that of 5-HT receptors, since the most effective displacers were ritanserin, mesulergine and mianserine, followed by clozapine, 2C ketanserine and m-CPP, while other compounds had a negligible or no effect. 3 These findings, showing a wide distribution of [ H]mesulergine binding sites in the human brain, could provide anatomical bases for the different functions attributable to 5-HT receptors in humans.

Research paper thumbnail of Tumor microenvironment: what have we learned studying the immune response in this puzzling battlefield?

Tumori

Recent developments hallmark the progress in the understanding of tumor immunology and related th... more Recent developments hallmark the progress in the understanding of tumor immunology and related therapeutic strategies. The administration of interleukin-2 (IL-2) to patients with cancer has shown that immune manipulation can mediate the regression of established cancers. The identification of the genes encoding cancer antigens and the development of means for effectively immunizing against these antigens has opened new avenues for the development of active immunization of patients with cancer. However, an efficient immune response against tumor comprises an intricate molecular network still poorly understood. Only when the code governing immune responsiveness of cancer will be deciphered, new therapeutic strategies could be designed to fit biologically defined mechanisms of immune rejection of cancer. In this review, we propose that the mechanisms regulating tumor rejection in response to vaccination will be more efficiently identified by following the evolution of treatment induced...

Research paper thumbnail of Gene expression profiling of anticancer immune responses

Current opinion in molecular therapeutics

Anticancer immune responses can be enhanced by immune manipulation, however, the biological mecha... more Anticancer immune responses can be enhanced by immune manipulation, however, the biological mechanism responsible for these immune responses remains largely unexplained. Conventional immunology researchers have extensively studied specific interactions between immune and cancer cells, and additional investigations have identified co-factors that may enhance the effectiveness of such interactions. As the molecular understanding of individual interactions increases, it is becoming apparent that no single mechanism can explain the phenomenon of tumor rejection. The contribution of several components of the innate and adaptive immune response is likely to be required for successful tumor rejection. These components may be variably recruited and activated by molecules with immune modulatory properties being produced by tumor and bystander cells within the tumor micro-environment. Such complexity can only be appreciated and solved by high-throughput tools capable of providing a global vie...

Research paper thumbnail of Integromics approaches to dissect genetic and functional heterogeneities between Caucasian and Asian immune responses

Research paper thumbnail of From the “Delayed Allergy Reaction” to the “Immunologic Constant of Rejection”

Immunologic Signatures of Rejection, 2010

This book collects salient observational data, derived predominantly from human studies, regardin... more This book collects salient observational data, derived predominantly from human studies, regarding the mechanism(s) of rejection in various pathologic conditions; the premise is that immune rejection, better defined as “immune-mediated, tissue-specific destruction” (TSD) (Wang et al. 2008), comprises a broad range of ­phenomena ranging from tumor regression, to clearance of pathogen through destruction of infected cells, autoimmunity, allograft rejection by the host and host vs. graft reactions. Like different hands can turn on or off a switch, distinct ­mechanisms can trigger TSD, however, a convergent pathway is ultimately observed when TSD occurs consisting of a dramatic switch from chronic to acute inflammation (Mantovani et al. 2008).

Research paper thumbnail of Different maturation cocktails provide dendritic cells with different chemoattractive properties

Journal of Translational Medicine, 2015

ABSTRACT Dendritic cells (DC) are currently implemented as immunotherapeutic strategy for the tre... more ABSTRACT Dendritic cells (DC) are currently implemented as immunotherapeutic strategy for the treatment of tumor patients based on their central role in the immune system. Despite good results were obtained in vitro and in animal models, their clinical use has provided limited success suggesting the requirement to optimise the protocol for their production. A cDNA array was performed on FastDC obtained from the differentiation of human peripheral blood monocytes stimulated with the clinical gold standard or with two alternative maturation cocktails combining interferon (IFN)γ and ligands for different toll like receptors (TLR). A stronger modulation of the DC transcriptome with respect to immature DC was found in alternatively stimulated DC when compared to DC stimulated with the clinical gold standard. A major class of molecules differentially expressed using distinct DC stimulation protocols were chemokines. Validation of their differential expression pattern at the mRNA and protein level confirmed the secretion of inflammatory chemokines by the alternative DC. Functional analyses of the chemotactic properties of DC "wash out" supernatants highlighted the ability of alternative, but not of gold standard DC to efficiently recruit immune cells with a prevalence of monocytes. Effector cells belonging to the innate as well as adaptive immunity were also attracted and the interaction with alternative DC resulted in enhanced secretion of IFNγ and induction of cytotoxic activity. Using leukocytes from cancer patients, it was demonstrated that the monocyte-attracting activity targeted cells with an inflammatory phenotype characterised by high levels of HLA-DR expression. Despite other classes of immune modulatory genes differently expressed in the alternative DC require to be investigated and characterised regarding their functional consequences, the reduced maturation state and chemoattractive properties of the gold standard versus alternative DC clearly promote the necessity to change the clinically used maturation cocktail of DC in order to improve the outcome of patients treated with DC-based vaccines.

Research paper thumbnail of 2015: The Year of Anti-PD-1/PD-L1s Against Melanoma and Beyond

Research paper thumbnail of Lineage relationship of CD8+ T cell subsets is revealed by progressive changes in the epigenetic landscape

Cellular and Molecular Immunology, 2015

To better elucidate epigenetic mechanisms that correlate with the dynamic gene expression program... more To better elucidate epigenetic mechanisms that correlate with the dynamic gene expression program observed upon T-cell differentiation, we investigated the genomic landscape of histone modifications in naive and memory CD8 1 T cells. Using a ChIP-Seq approach coupled with global gene expression profiling, we generated genome-wide histone H3 lysine 4 (H3K4me3) and H3 lysine 27 (H3K27me3) trimethylation maps in naive, T memory stem cells, central memory cells, and effector memory cells in order to gain insight into how histone architecture is remodeled during T cell differentiation. We show that H3K4me3 histone modifications are associated with activation of genes, while H3K27me3 is negatively correlated with gene expression at canonical loci and enhancers associated with T-cell metabolism, effector function, and memory. Our results also reveal histone modifications and gene expression signatures that distinguish the recently identified T memory stem cells from other CD8 1 T-cell subsets. Taken together, our results suggest that CD8 1 lymphocytes undergo chromatin remodeling in a progressive fashion. These findings have major implications for our understanding of peripheral T-cell ontogeny and the formation of immunological memory.

Research paper thumbnail of Workshop on cancer biometrics: identifying biomarkers and surrogates of cancer in patients: a meeting held at the Masur Auditorium, National Institutes of Health

Journla of Immunotherapy

The current excitement about molecular targeted therapies has driven much of the recent dialog in... more The current excitement about molecular targeted therapies has driven much of the recent dialog in cancer diagnosis and treatment. Particularly in the biologic therapy of cancer, identifiable antigenic T-cell targets restricted by MHC molecules and the related novel stress molecules such as MICA/B and Letal allow a degree of precision previously unknown in cancer therapy. We have previously held workshops on immunologic monitoring and angiogenesis monitoring. This workshop was designed to discuss the state of the art in identification of biomarkers and surrogates of tumor in patients with cancer, with particular emphasis on assays within the blood and tumor. We distinguish this from immunologic monitoring in the sense that it is primarily a measure of the tumor burden as opposed to the immune response to it. Recommendations for intensive investigation and targeted funding to enable such strategies were developed in seven areas: genomic analysis; detection of molecular markers in peri...

Research paper thumbnail of Cancer vaccines: from research to clinical practice

Research paper thumbnail of Identification of immune dominant cytomegalovirus epitopes using quantitative real-time polymerase chain reactions to measure interferon-gamma production by peptide-stimulated peripheral blood mononuclear cells

Journla of Immunotherapy

The identification of HLA restricted immune dominant cytotoxic T cell (CTL) epitopes limits immun... more The identification of HLA restricted immune dominant cytotoxic T cell (CTL) epitopes limits immune therapy. Cytomegalovirus (CMV) disease remains a significant cause of morbidity after allogeneic stem cell transplantation. Adoptive immune therapy using CTLs stimulated with immune dominant CMV pp65 peptides may be effective in preventing CMV disease, but immune dominant CMV peptides have been identified for only a few HLA class I molecules. The purpose of this study was to use a novel molecular system to establish a rapid and precise method to identify new HLA-restricted CMV epitopes. Cytomegalovirus pp65 peptides expected to bind to the HLA-24 molecule were identified with a computer algorithm. Five candidate peptides were screened by direct ex vivo stimulation of peripheral blood mononuclear cells (PBMCs) from CMV-seropositive HLA-A*2402 individuals, and quantitative real time PCR (qRT-PCR) was used to evaluate CTL responses by measuring interferon-gamma (IFN-gamma) transcripts. On...

Research paper thumbnail of New HLA-A, -B, and -C locus-specific primers for PCR amplification from cDNA: application in clinical immunology

Journal of Immunological Methods

The individual cellular immune response to intracellular antigens is modeled by the highly polymo... more The individual cellular immune response to intracellular antigens is modeled by the highly polymorphic major histocompatibility complex (HLA) class I molecules. The epitopes presented and the T cell repertoire that recognizes them depend on the HLA constitution of the individual. Therefore, to monitor and to modify an individual's HLA class I-driven cellular immune response, it is necessary to know the HLA class I alleles of the person and the possible epitopes of the target antigen presented by those alleles. In particular, this is necessary in order to design peptide-based vaccines and immune therapies for the treatment of diseases caused by viruses, intracellular parasites or cancer, and to monitor the immune response during those treatments. We describe a new set of HLA-A, -B, and -C locus-specific primers for the polymerase chain reaction (PCR) amplification of the whole coding sequence of these genes from complementary DNA (cDNA). We describe their use for typing and for t...

Research paper thumbnail of The matrix protein pp65(341-350): a peptide that induces ex vivo stimulation and in vitro expansion of CMV-specific CD8+ T cells in subjects bearing either HLA-A*2402 or A*0101 allele

Transfusion

The stimulation of PBMNCs with HLA Class I restricted synthetic peptides derived from CMV phospho... more The stimulation of PBMNCs with HLA Class I restricted synthetic peptides derived from CMV phosphorylated matrix protein 65 (pp65) evokes CMV-specific cytotoxic T lymphocyte (CTL) activity, a necessary condition for initiating adoptive immunotherapy against CMV-related diseases in immune-compromised patients. It was previously demonstrated that the CMV decamer (10-mer) peptide pp65(341-350), QYDPVAALFF, was able to induce CMV-specific CTLs in HLA-A*2402 CMV-seropositive individuals. We investigated the ability of the peptide pp65(341-350) to reactivate memory CD8+ T cells in CMV-seropositive subjects bearing either the HLA-A24 or A1 allele. CTL responses were measured by IFN-gamma mRNA expression and IFN-gamma protein production as well as cytotoxic activity. In this study it was found that peptide pp65(341-350) induced a specific reactivation of memory CD8+ T cells from CMV-seropositive donors expressing either HLA-A*2402 and/or HLA-A*0101. Moreover, a pp65(341-350)-specific selecti...

Research paper thumbnail of CMV pp65 and IE-1 T cell epitopes recognized by healthy subjects

Journal of Translational Medicine

Adoptive immune and vaccine therapies have been used to prevent cytomegalovirus (CMV) disease in ... more Adoptive immune and vaccine therapies have been used to prevent cytomegalovirus (CMV) disease in recipients of hematopoietic progenitor cell transplants, but the nature of T cell responses to CMV have not been completely characterized. Peptide pools and individual peptides derived from the immune-dominant CMV proteins pp65 and IE-1 and antigen-specific, cytokine flow cytometry were used to characterize the prevalence and frequency of CD4+ and CD8+ memory T cells in 20 healthy CMV-seropositive subjects. CD8+ T cell responses to pp65 were detected in 35% of subjects and to IE-1 in 40% of subjects. CD4+ T cell responses to pp65 were detected in 50% of subjects, but none were detected to IE-1. Several new IE-1 HLA class I epitopes were identified, including 4 restricted to HLA-C antigens. One region of IE-1 spanning amino acids 300 to 327 was rich in class I epitopes. The HLA class I restrictions of IE-1 peptides were more promiscuous than those of pp65 peptides. Since naturally occurri...

Research paper thumbnail of Consensus nomenclature for CD8 + T cell phenotypes in cancer

OncoImmunology, 2015

Whereas preclinical investigations and clinical studies have established that CD8(+) T cells can ... more Whereas preclinical investigations and clinical studies have established that CD8(+) T cells can profoundly affect cancer progression, the underlying mechanisms are still elusive. Challenging the prevalent view that the beneficial effect of CD8(+) T cells in cancer is solely attributable to their cytotoxic activity, several reports have indicated that the ability of CD8(+) T cells to promote tumor regression is dependent on their cytokine secretion profile and their ability to self-renew. Evidence has also shown that the tumor microenvironment can disarm CD8(+) T cell immunity, leading to the emergence of dysfunctional CD8(+) T cells. The existence of different types of CD8(+) T cells in cancer calls for a more precise definition of the CD8(+) T cell immune phenotypes in cancer and the abandonment of the generic terms "pro-tumor" and…

Research paper thumbnail of Functional Heterogeneity of

Research paper thumbnail of cDNA arrays and the enigma of melanoma immune responsiveness

The Cancer Journal

Melanoma is a cancer characterized by predisposition to natural and induced immune rejection. The... more Melanoma is a cancer characterized by predisposition to natural and induced immune rejection. The occurrence of this phenomenon, however, remains sporadic and capricious while there is no clear understanding of the chain of events responsible. With the progression of the understanding of the molecular immunology of melanoma during the last decade, several theories suggesting immune tolerance (inadequate immune response) and/or immune escape (adaptation of cancer cells to a vigorous immune pressure) as possible modulators of melanoma growth have emerged. As the number of these hypotheses, mostly based on molecular or pre-clinical models, has increased exponentially, the relevance of each individual one has been conversely decreasing leaving the observer without confidence that a theory likely to explain this phenomenon can be soon formulated. In this review, we will discuss a direct approach whereby new theories might be identified by direct observation of human phenomena as they occ...

Research paper thumbnail of Use of laser scanning cytometry to study tumor microenvironment

Histology and histopathology

The study of phenomena occurring in the tumor microenvironment is a challenging task because of t... more The study of phenomena occurring in the tumor microenvironment is a challenging task because of technical difficulties, particularly when dealing with hypocellular specimens. Laser scanning cytometry (LSC) is a new laboratory technology that has been recently introduced to overcome the limitations of other traditional technologies. By combining the properties and the advantages of flow cytometry (FC) and immunohistochemistry (IHC), LSC allows the investigator to obtain objective information on DNA content, protein expression and cellular localization is combination with morphological features. It has been already shown that LSC results are reliable compared to more traditional technologies, and its implementation in the clinical routine is under way. Its use in oncology, which is rapidly expanding, spans from apoptosis analysis to DNA content quantitation and tumor cell phenotyping. Here we describe the technology underlying this novel fluorescence-based device, review its use in on...

Research paper thumbnail of The dual role of IL10

Classification of cytokines as pro- versus anti-inflammatory might not apply to the pleiotropic e... more Classification of cytokines as pro- versus anti-inflammatory might not apply to the pleiotropic effects of interleukin-10 (IL-10). Several reports suggest that IL-10 enhances the function of natural killer cells, which leads, through pathogen destruction, to increased antigen availability. In addition, by inhibiting the maturation of antigen-presenting cells (APCs), IL-10 preserves their ability for antigen uptake while simultaneously hampering their migration to draining lymph nodes. This review suggests that this ‘antigen-loading’ phase might constitute an important component of the innate immune reaction to a pathogen. Additional proinflammatory stimuli might subsequently lead to maturation of ‘loaded’ APCs that could migrate to draining lymph nodes or recruit and activate adaptive immune effectors locally.

Research paper thumbnail of A global approach to tumor immunology

Cellular & molecular immunology

Biological and clinical advances in the understanding of tumor immunology suggest that immune res... more Biological and clinical advances in the understanding of tumor immunology suggest that immune responsiveness of human tumors is a complex biological phenomenon that could be best studied by a real-time comparison of tumor/host interactions in the tumor microenvironment through a high-throughput discovery-driven approach. This conclusion is derived from our recognition that too many hypotheses or, in other words, no solid single hypothesis exist, based on experimental results, to further drive experimentation in human subjects. Functional genomic studies entertained during the last few years consolidated the belief that in humans the interactions between tumor and immune cells are too complex to be approached exclusively with a hypothesis driven method. We believe that immune cells suit cancer cells in a Yin and Yang balance by opposing and yet mutually depending on each other. Indeed, immune infiltration in tumors may play a dual role modulating in different circumstances cancer cel...

Research paper thumbnail of The enigma of tumor immune responsiveness: A high-throughput genomics question?

Tissue Antigens

Much interest is currently being directed towards serotonin (5-HT) receptors of type 2C (5-HT ) b... more Much interest is currently being directed towards serotonin (5-HT) receptors of type 2C (5-HT ) because of their possible 2C involvement in the control of different activities, such as the composition of the cerebrospinal fluid, locomotion, feeding, neuronal excitability and anxiety. The limited information regarding their distribution in the human brain prompted us to investigate, and to 3 characterize the binding of [ H]mesulergine, a HT antagonist, in autopsy samples from 24 subjects. 2C 3 The results showed that the [ H]mesulergine binding represented 95% of the total binding and equilibrium saturation binding experiments resulted in a single straight line, consistent with the presence of one site only. The area with the highest density of 3 [ H]mesulergine binding was the choroid plexus, followed at a significantly lower level by the hippocampus, substantia nigra, basal 3 ganglia, amygdala, hypothalamus and prefrontal cortex. The pharmacological profile of the [ H]mesulergine binding was consistent with that of 5-HT receptors, since the most effective displacers were ritanserin, mesulergine and mianserine, followed by clozapine, 2C ketanserine and m-CPP, while other compounds had a negligible or no effect. 3 These findings, showing a wide distribution of [ H]mesulergine binding sites in the human brain, could provide anatomical bases for the different functions attributable to 5-HT receptors in humans.

Research paper thumbnail of Tumor microenvironment: what have we learned studying the immune response in this puzzling battlefield?

Tumori

Recent developments hallmark the progress in the understanding of tumor immunology and related th... more Recent developments hallmark the progress in the understanding of tumor immunology and related therapeutic strategies. The administration of interleukin-2 (IL-2) to patients with cancer has shown that immune manipulation can mediate the regression of established cancers. The identification of the genes encoding cancer antigens and the development of means for effectively immunizing against these antigens has opened new avenues for the development of active immunization of patients with cancer. However, an efficient immune response against tumor comprises an intricate molecular network still poorly understood. Only when the code governing immune responsiveness of cancer will be deciphered, new therapeutic strategies could be designed to fit biologically defined mechanisms of immune rejection of cancer. In this review, we propose that the mechanisms regulating tumor rejection in response to vaccination will be more efficiently identified by following the evolution of treatment induced...

Research paper thumbnail of Gene expression profiling of anticancer immune responses

Current opinion in molecular therapeutics

Anticancer immune responses can be enhanced by immune manipulation, however, the biological mecha... more Anticancer immune responses can be enhanced by immune manipulation, however, the biological mechanism responsible for these immune responses remains largely unexplained. Conventional immunology researchers have extensively studied specific interactions between immune and cancer cells, and additional investigations have identified co-factors that may enhance the effectiveness of such interactions. As the molecular understanding of individual interactions increases, it is becoming apparent that no single mechanism can explain the phenomenon of tumor rejection. The contribution of several components of the innate and adaptive immune response is likely to be required for successful tumor rejection. These components may be variably recruited and activated by molecules with immune modulatory properties being produced by tumor and bystander cells within the tumor micro-environment. Such complexity can only be appreciated and solved by high-throughput tools capable of providing a global vie...

Research paper thumbnail of Integromics approaches to dissect genetic and functional heterogeneities between Caucasian and Asian immune responses

Research paper thumbnail of From the “Delayed Allergy Reaction” to the “Immunologic Constant of Rejection”

Immunologic Signatures of Rejection, 2010

This book collects salient observational data, derived predominantly from human studies, regardin... more This book collects salient observational data, derived predominantly from human studies, regarding the mechanism(s) of rejection in various pathologic conditions; the premise is that immune rejection, better defined as “immune-mediated, tissue-specific destruction” (TSD) (Wang et al. 2008), comprises a broad range of ­phenomena ranging from tumor regression, to clearance of pathogen through destruction of infected cells, autoimmunity, allograft rejection by the host and host vs. graft reactions. Like different hands can turn on or off a switch, distinct ­mechanisms can trigger TSD, however, a convergent pathway is ultimately observed when TSD occurs consisting of a dramatic switch from chronic to acute inflammation (Mantovani et al. 2008).

Research paper thumbnail of Different maturation cocktails provide dendritic cells with different chemoattractive properties

Journal of Translational Medicine, 2015

ABSTRACT Dendritic cells (DC) are currently implemented as immunotherapeutic strategy for the tre... more ABSTRACT Dendritic cells (DC) are currently implemented as immunotherapeutic strategy for the treatment of tumor patients based on their central role in the immune system. Despite good results were obtained in vitro and in animal models, their clinical use has provided limited success suggesting the requirement to optimise the protocol for their production. A cDNA array was performed on FastDC obtained from the differentiation of human peripheral blood monocytes stimulated with the clinical gold standard or with two alternative maturation cocktails combining interferon (IFN)γ and ligands for different toll like receptors (TLR). A stronger modulation of the DC transcriptome with respect to immature DC was found in alternatively stimulated DC when compared to DC stimulated with the clinical gold standard. A major class of molecules differentially expressed using distinct DC stimulation protocols were chemokines. Validation of their differential expression pattern at the mRNA and protein level confirmed the secretion of inflammatory chemokines by the alternative DC. Functional analyses of the chemotactic properties of DC "wash out" supernatants highlighted the ability of alternative, but not of gold standard DC to efficiently recruit immune cells with a prevalence of monocytes. Effector cells belonging to the innate as well as adaptive immunity were also attracted and the interaction with alternative DC resulted in enhanced secretion of IFNγ and induction of cytotoxic activity. Using leukocytes from cancer patients, it was demonstrated that the monocyte-attracting activity targeted cells with an inflammatory phenotype characterised by high levels of HLA-DR expression. Despite other classes of immune modulatory genes differently expressed in the alternative DC require to be investigated and characterised regarding their functional consequences, the reduced maturation state and chemoattractive properties of the gold standard versus alternative DC clearly promote the necessity to change the clinically used maturation cocktail of DC in order to improve the outcome of patients treated with DC-based vaccines.

Research paper thumbnail of 2015: The Year of Anti-PD-1/PD-L1s Against Melanoma and Beyond

Research paper thumbnail of Lineage relationship of CD8+ T cell subsets is revealed by progressive changes in the epigenetic landscape

Cellular and Molecular Immunology, 2015

To better elucidate epigenetic mechanisms that correlate with the dynamic gene expression program... more To better elucidate epigenetic mechanisms that correlate with the dynamic gene expression program observed upon T-cell differentiation, we investigated the genomic landscape of histone modifications in naive and memory CD8 1 T cells. Using a ChIP-Seq approach coupled with global gene expression profiling, we generated genome-wide histone H3 lysine 4 (H3K4me3) and H3 lysine 27 (H3K27me3) trimethylation maps in naive, T memory stem cells, central memory cells, and effector memory cells in order to gain insight into how histone architecture is remodeled during T cell differentiation. We show that H3K4me3 histone modifications are associated with activation of genes, while H3K27me3 is negatively correlated with gene expression at canonical loci and enhancers associated with T-cell metabolism, effector function, and memory. Our results also reveal histone modifications and gene expression signatures that distinguish the recently identified T memory stem cells from other CD8 1 T-cell subsets. Taken together, our results suggest that CD8 1 lymphocytes undergo chromatin remodeling in a progressive fashion. These findings have major implications for our understanding of peripheral T-cell ontogeny and the formation of immunological memory.