ABSTRACT
Background: The prevalence and mortality of sepsis in Internal Medicine Units (IMUs) is poorly understood as most of the data derive from studies conducted in Intensive Care Units. Aim of SEpsis Management in INternal medicine Apulia (SEMINA) study was to determine the prevalence of sepsis and the characteristics and outcomes of patients with Sepsis-3 criteria admitted in Apulia's Internal Medicine Units for over six months. Methods: The SEpsis Management in INternal medicine of Apulia study was a prospective, multicentre, observational study. Adult admissions to the 13 Apulia Region's Internal Medicine Units between November 15, 2018 and May 15, 2019 were screened for sepsis according to the Sepsis-3 criteria. Medical data were collected in electronic case report form. Results: Out of 7,885 adult patients of the Internal Medicine Units, 359 (4.55%) fulfilled the inclusion criteria, and 65 of them (18.1%) met the septic shock criteria. The patients enrolled were elderly, suffering from chronic poly-pathologies and from cognitive and functional impairment. The respiratory system was the most common site of infection and the most common pathogens isolated from blood cultures were Staphylococcus spp., E. coli, Klebsiella spp., Enterococcus spp. and Acinetobacter spp. The in-hospital fatality rate was 31.2% and was significantly higher for septic shock. Sequential Organ Failure Assessment score, dementia and infections from Acinetobacter spp. were independent risk factors for mortality. Conclusions: A high prevalence of sepsis and a high fatality rate were detected in Apulia Region's Internal Medicine Units. The high fatality rate observed in our study could be related to the underlying diseases and to the vulnerability of elderly patients admitted to our Internal Medicine Units.
Subject(s)
Sepsis , Shock, Septic , Adult , Aged , Humans , Hospital Mortality , Intensive Care Units/statistics & numerical data , Prospective Studies , Sepsis/epidemiology , Sepsis/microbiology , Sepsis/therapy , Shock, Septic/epidemiology , Shock, Septic/microbiology , Shock, Septic/therapy , PrevalenceABSTRACT
OBJECTIVE: The aim of the present study was to assess the association among anxiety, depression, stress, social support and emotional abilities with adherence and healthcare spending in type 2 diabetic patients. PATIENTS AND METHODS: Sixty-four patients were enrolled and completed: Interpersonal Processes of Care (IPC), 20-item Toronto Alexithymia Scale (TAS-20), Rapid Stress Assessment Scale (RSAS), Morisky Medication Adherence Scale (MMAS-4), International Physical Activity Questionnaire (IPAQ)-Short Form and a socio-anamnestic questionnaire regarding also the healthcare spending. RESULTS: Mathematical linear regressions models were performed showing the predictive effects of: anxiety and social support scores (RSAS) on adherence levels (respectively p =. 019; p =. 016); adherence levels on anxiolytic use (p =.04); aggressiveness scores (RSAS) on the number of general check-ups (p =.031); TAS-20 and physician-patient communication (IPC) on the number of hospitalization days (respectively p=.001; p=.008); physician patient decision making (IPC) scores on physical activity (IPAQ) levels (p=.025); physical activity (IPAQ) on the number of medical examinations (p=.039). CONCLUSIONS: An association among psychosocial impairment, adherence and healthcare spending was found. Future studies should investigate the effect of a brief psychological intervention in increasing adherence levels and reducing the healthcare spending in this clinical population.
Subject(s)
Diabetes Mellitus, Type 2/psychology , Health Expenditures/statistics & numerical data , Medication Adherence/statistics & numerical data , Social Support , Aged , Aged, 80 and over , Anxiety/diagnosis , Anxiety/economics , Anxiety/epidemiology , Anxiety/psychology , Depression/diagnosis , Depression/economics , Depression/epidemiology , Depression/psychology , Diabetes Mellitus, Type 2/drug therapy , Diabetes Mellitus, Type 2/economics , Female , Humans , Male , Middle Aged , Psychometrics/statistics & numerical data , Self Report/statistics & numerical data , Stress, Psychological/diagnosis , Stress, Psychological/economics , Stress, Psychological/epidemiology , Stress, Psychological/psychologyABSTRACT
The current study examined whether alexithymia is involved in colon cancer as has been found in breast and uterine cancer research. The TAS-20 was administered before examination to patients who underwent colon cancer screening. Histological data were obtained from the biopsies. Gender, age, education, smoking and drinking habits, and personal and family histories of neoplastic colonic disease were controlled for in the analysis. Both adenoma and adenocarcinoma patients had higher TAS-20 scores than negative cases, and both high and intermediate levels of alexithymia were implicated. Difficulty identifying feelings and externally oriented thinking were the most prominent facets related to the disease process.
Subject(s)
Affective Symptoms/epidemiology , Affective Symptoms/psychology , Colonic Neoplasms/epidemiology , Colonic Neoplasms/psychology , Colonoscopy , Mass Screening/methods , Comorbidity , Female , Humans , Male , Middle Aged , Psychiatric Status Rating Scales/statistics & numerical data , Risk Factors , Rome/epidemiology , Surveys and QuestionnairesABSTRACT
Weak immunogenicity of chronic lymphocytic leukemia (CLL) cells may contribute to disease progression and inhibit effective immunotherapy. Accordingly, agents that enhance the immunogenicity of CLL cells may be useful in immunotherapeutic approaches to this disease. Since Toll-like receptors (TLRs) are major regulators of innate immunity and initiation of adaptive immunity, we studied the effects of viral pathogen associated molecular pattern agonists (that are recognized by TLRs) on the costimulatory phenotype and function of CLL cells. CLL cells (especially those with high endogenous expression of CD38) responded to TLR7-activating imidazoquinolines and guanosine analogs by increasing costimulatory molecule expression, producing inflammatory cytokines, and becoming more sensitive to killing by cytotoxic effectors. Additional activation of protein kinase C pathways increased the ability to stimulate T-cell proliferation, blocked phosphorylation of the transcription factor, signal transducer and activator of transcription (STAT)3, and resulted in the acquisition of a dendritic cell surface phenotype by TLR7-activated CLL cells. Normal B cells also responded to TLR7 activation by increasing costimulatory molecule expression and cytokine production. These findings suggest a potential role for TLR7 agonists in CLL immunotherapy.
Subject(s)
Imidazoles/pharmacology , Immunologic Factors/pharmacology , Leukemia, Lymphocytic, Chronic, B-Cell/drug therapy , Leukemia, Lymphocytic, Chronic, B-Cell/metabolism , Phorbol Esters/pharmacology , Quinolines/pharmacology , Toll-Like Receptor 7/metabolism , Adult , Aged , Aged, 80 and over , B-Lymphocytes/drug effects , B-Lymphocytes/metabolism , Chemokines/biosynthesis , Cytokines/biosynthesis , Female , Humans , Leukemia, Lymphocytic, Chronic, B-Cell/immunology , Male , Middle Aged , STAT3 Transcription Factor/drug effects , STAT3 Transcription Factor/metabolism , Sensitivity and Specificity , Toll-Like Receptor 7/drug effects , Tumor Cells, CulturedABSTRACT
In this review, we discuss the ongoing development of a new treatment option for genital herpes (GH), the disease caused by herpes simplex virus (HSV) types I and II. Following infection, the virus establishes a latent infection in peripheral neurons, which periodically activates to cause recurrent skin lesions or asymptomatic shedding in the anogenital area. A new class of drugs, the immune response modifiers (IRMs), modulates the immune system against viral infection. This approach is currently being tested as a treatment for GH. We first review the effectiveness of treatment of other viral diseases with imiquimod, the first IRM to be licensed (Aldara, imiquimod 5% cream), and one used for the treatment of external anogenital warts. We then focus on resiquimod, an analog of imiquimod, which shows early promise as a new treatment option for GH. The evidence from in vitro and in vivo studies, in particular the guinea pig model of GH, describing the effectiveness and mode of action of this novel immunopharmacological agent is presented. Resiquimod stimulates specific cells of the innate immune system (including monocytes/macrophages, dendritic cells (DC) and B lymphocytes) to produce cytokines (in particular IFN-alpha, IL-12, TNF-alpha and IFN-gamma) that initiate and drive the development of the Th1 acquired immune response against HSV-infected cells. Recent results from clinical trials and in vivo studies in animal models are consistent with the hypothesis that the development of HSV-specific cell-mediated immunity may prove to be the key in providing a long-lasting protection against GH recurrences.
Subject(s)
Adjuvants, Immunologic/therapeutic use , Herpes Genitalis/drug therapy , Herpes Genitalis/immunology , Adjuvants, Immunologic/chemistry , Adjuvants, Immunologic/pharmacology , Aminoquinolines/chemistry , Aminoquinolines/pharmacology , Aminoquinolines/therapeutic use , Animals , Herpes Genitalis/pathology , Herpes Genitalis/virology , Herpesvirus 1, Human/drug effects , Herpesvirus 2, Human/drug effects , Humans , Imidazoles/chemistry , Imidazoles/pharmacology , Imidazoles/therapeutic use , Imiquimod , RecurrenceABSTRACT
The effect of resiquimod (R-848), an immune-response modifier that is similar to imiquimod, on recurrent herpes simplex virus (HSV) was evaluated using the guinea pig model of genital herpes. Guinea pigs were intravaginally infected with HSV-2 and then were randomized on day 14 to receive nothing or 0.1 mL/kg per dose of subcutaneous resiquimod, given either daily, every other day, or weekly from days 15-35. During a 3-week course of therapy, recurrences in all 3 treated groups were reduced by >80%, compared with the control group. After therapy, recurrences remained significantly (P<.05) decreased in all 3 groups for the next 3 weeks. The group treated weekly developed the fewest recurrences. Significant increases in interleukin-2 levels, produced by incubation of mononuclear cells with HSV-2 antigens, but not in circulating antibody also were detected in the treated groups. Resiquimod treatment may offer significant advantages to present antiviral therapies for the control of recurrent genital herpes.
Subject(s)
Herpes Genitalis/prevention & control , Herpesvirus 2, Human/immunology , Imidazoles/therapeutic use , Immunologic Factors/therapeutic use , Animals , Antibodies, Viral/biosynthesis , Antigens, Viral/immunology , Cells, Cultured , Drug Administration Schedule , Drug Evaluation, Preclinical , Female , Guinea Pigs , Herpes Genitalis/immunology , Herpes Genitalis/virology , Imidazoles/administration & dosage , Immunologic Factors/administration & dosage , Interferons/biosynthesis , Interleukin-2/biosynthesis , Leukocytes, Mononuclear/immunology , Random Allocation , Secondary Prevention , Time FactorsABSTRACT
Members of the imidazoquinoline molecule family, including imiquimod and resiquimod (R-848), have potent antiviral and antitumor activities. Imiquimod cream (5%) (Aldara) is currently indicated for treatment of external genital and perianal warts. Previous characterization of these compounds has focused upon their ability to activate monocytes and dendritic cells, but recent studies have shown that resiquimod also stimulates B lymphocytes to proliferate and express an activated phenotype. This suggests that resiquimod could potentially serve as an effective vaccine adjuvant in stimulating a humoral immune response. This study shows that resiquimod mimics effects of the T-dependent CD40 signal in both mouse and human B cell lines. Resiquimod, like CD40, stimulates antibody secretion, cytokine production, protection from apoptosis, and CD80 upregulation. In addition, it shows synergy with signals delivered by the B cell antigen receptor and heightens CD40-mediated B cell activation, demonstrating that resiquimod can enhance antigen-specific responses in B lymphocytes.
Subject(s)
B-Lymphocytes/drug effects , B-Lymphocytes/immunology , CD40 Antigens/immunology , Imidazoles/pharmacology , Immunologic Factors/pharmacology , Lymphocyte Activation/drug effects , Adjuvants, Immunologic/pharmacology , Animals , Antibodies, Monoclonal , Apoptosis/drug effects , B-Lymphocytes/cytology , B-Lymphocytes/metabolism , B7-1 Antigen/metabolism , Cell Division/drug effects , Cell Line , CpG Islands/immunology , Drug Synergism , Humans , Immunoglobulin M/immunology , Immunoglobulin M/metabolism , Interleukin-6/metabolism , Lipopolysaccharides/immunology , Mice , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Antigen, B-Cell/immunology , Tumor Necrosis Factor-alpha/metabolism , Up-Regulation/drug effectsABSTRACT
The imidazoquinoline R-848, originally identified as a highly effective antiviral agent, has recently been shown to be capable of potent B lymphocyte activation. The B cell-activating properties of R-848 are strikingly similar to the effects of the CD40 ligand CD154. The present study demonstrates that this similarity extends to the intracellular signaling pathways triggered by the compound, although both overlapping and distinct mechanisms of signaling were seen. Like CD40 ligation, R-848 stimulated activation of the stress-activated protein kinases c-Jun kinase and p38 and activated the NF-kappaB family of transcription factors. Both R-848- and CD40-mediated B cell differentiation were dependent upon NF-kappaB activation, although the relative importance of individual NF-kappaB family members appeared to differ between R-848- and CD40-mediated signals. Both signals were partially dependent upon induction of TNF-alpha and IL-6, and the cytoplasmic adaptor molecule TNF receptor-associated factor 2 is involved in both R-848- and CD40-mediated differentiation.
Subject(s)
Adjuvants, Immunologic/pharmacology , B-Lymphocytes/drug effects , B-Lymphocytes/immunology , Imidazoles/pharmacology , Lymphocyte Activation/drug effects , Animals , Antibody-Producing Cells/drug effects , Antibody-Producing Cells/immunology , Antibody-Producing Cells/metabolism , B-Lymphocytes/enzymology , B-Lymphocytes/metabolism , Cell Differentiation/drug effects , Cell Differentiation/immunology , Cell Line , Enzyme Activation/drug effects , Female , Humans , Interleukin-6/physiology , JNK Mitogen-Activated Protein Kinases , Mice , Mice, Inbred C57BL , Mice, Knockout , Mitogen-Activated Protein Kinases/metabolism , NF-kappa B/metabolism , Proteins/physiology , Receptors, Tumor Necrosis Factor/physiology , Signal Transduction/immunology , TNF Receptor-Associated Factor 2 , Transcriptional Activation/drug effects , Transcriptional Activation/immunology , Tumor Necrosis Factor-alpha/physiology , p38 Mitogen-Activated Protein KinasesABSTRACT
R-848 and imiquimod belong to a class of immune response modifiers that are potent inducers of cytokines, including IFN-alpha, TNF-alpha, IL-12, and IFN-gamma. Many of these cytokines can affect the acquired immune response. This study examines the effects of R-848 on aspects of acquired immunity, including immunoglobulin secretion, in vivo cytokine production, and Ag-specific T cell cytokine production. Results are compared with those of Th1 CpG ODN. R-848 and CpG ODN are effective at skewing immunity in the presence of Alum toward a Th1 Ab response (IgG2a) and away from a Th2 Ab response (IgE). R-848 and CpG ODN are also capable of initiating an immune response in the absence of additional adjuvant by specifically enhancing IgG2a levels. Both R-848 and imiquimod showed activity when given subcutaneously or orally, indicating that the compound mechanism was not through generation of a depot effect. Although CpG ODN behaves similarly to R-848, CpG ODN has a distinct cytokine profile, is more effective than R-848 when given with Alum in the priming dose, and is active only when given by the same route as the Ag. The mechanism of R-848's adjuvant activity is linked to cytokine production, where increases in IgG2a levels are associated with IFN-alpha, TNF-alpha, IL-12, and IFN-gamma induction, and decreases in IgE levels are associated with IFN-alpha and TNF-alpha. Imiquimod also enhances IgG2a production when given with Ag. The above results suggest that the imidazoquinolines R-848 and imiquimod may be attractive compounds for use as vaccine adjuvants and in inhibiting pathological responses mediated by Th2 cytokines.
Subject(s)
Adjuvants, Immunologic , Imidazoles/immunology , Oligodeoxyribonucleotides/immunology , Administration, Oral , Animals , Cell Separation , Cytokines/analysis , Female , Immunization, Secondary , Immunoglobulin E/immunology , Immunoglobulin G/immunology , Interferon-gamma/immunology , Mice , Mice, Inbred BALB C , Ovalbumin/immunology , Spleen/cytology , Spleen/immunology , Th1 Cells/immunology , Th2 Cells/immunology , VaccinationABSTRACT
Imiquimod and R-848 are members of a family of immune response modifiers that stimulate cytokine production in monocyte/macrophages and dendritic cell cultures. This study evaluated the effects of the imidazoquinolines, imiquimod and R-848, on B lymphocyte activation. Both agents induced proliferation of murine T-cell-depleted and highly purified splenic B cell preparations as well as purified human B cells. Resting and activated B cells responded to these agents, with activated cells responding more efficiently. B cells from the LPS-hyporesponsive C3H/HeJ mice and guanosine-hyporesponsive SJL mice proliferated in response to imiquimod and R-848, indicating a different mechanism of action than lipopolysaccharide and guanine nucleosides. B cells were also stimulated by imiquimod and R-848 to produce increased immunoglobulin levels. Increased expression of a number of B cell activation markers were seen following imiquimod or R-848 stimulation. Finally, R-848 was shown to act as a vaccine adjuvant enhancing OVA-specific IgG2a levels while suppressing total IgE. These results indicate that R-848 and imiquimod are potent activators of B lymphocytes and are capable of augmenting antigen-specific immunoglobulin production.
Subject(s)
Adjuvants, Immunologic/pharmacology , Aminoquinolines/pharmacology , B-Lymphocytes/immunology , Imidazoles/pharmacology , Lymphocyte Activation/drug effects , Animals , Antibody Specificity , Antigens, CD/immunology , B-Lymphocytes/drug effects , B7-2 Antigen , Cells, Cultured , Guanosine/immunology , Humans , Imiquimod , Immunoglobulin Class Switching , Immunoglobulin E/biosynthesis , Immunoglobulin E/immunology , Immunoglobulin G/biosynthesis , Immunoglobulin G/immunology , Lipopolysaccharides/immunology , Membrane Glycoproteins/immunology , Mice , Mice, Inbred Strains , Ovalbumin/immunology , Spleen/cytology , Spleen/immunologyABSTRACT
Imiquimod (IQ) has been successfully used in treatment of genital warts. In clinical settings, patients responded well but wart reduction rates varied. Our aim was to find a correlation between clinical responses and pretreatment (constitutive) levels of genes that might be involved in the molecular action of IQ. Since IQ is a cytokine inducer, we analyzed levels of expression of genes of the JAK/STAT signaling pathway and their inhibitors as well as interferon response factors (IRFs) in pretreatment biopsy specimens from complete responders (99 to 100% wart reduction rate) versus incomplete responders (75 to 92% wart reduction rate) by reverse transcription-PCR. We found that mRNA levels of signal transducer and activator of transcription 1 (STAT1) and IRF1 were higher in complete responders than in incomplete responders. Incomplete responders expressed larger amounts of STAT3, IRF2, and protein inhibitor of activated STAT1 (PIAS1) mRNAs compared to complete responders before IQ treatment. We hypothesize that high-level expression of STAT1 and IRF1 is advantageous for a better IQ response. The observed differences in constitutive mRNA levels of these genes may be the consequence of alterations in cellular differentiation and/or variable expression of endogenous interferons. Previous in vitro studies showed that keratinocyte differentiation coordinates the balance between positive and negative signals along the JAK/STAT pathway by regulating the IRF1:IRF2 and STAT1:PIAS1 ratios and thus affecting induction of IQ-inducible genes. Specifically, differentiation supports constitutive expression of STAT1 and IRF1 mRNAs but not expression of IRF2 and PIAS1. Our data are in good agreement with studies that showed the importance of STAT1 in cytokine induction and activation of interferon-responsive genes by IQ.
Subject(s)
Aminoquinolines/pharmacology , Condylomata Acuminata/genetics , DNA-Binding Proteins/genetics , Gene Expression Regulation/drug effects , Interferon Inducers/pharmacology , Trans-Activators/genetics , Aminoquinolines/therapeutic use , Biopsy , Cell Differentiation/drug effects , Condylomata Acuminata/drug therapy , Condylomata Acuminata/immunology , Condylomata Acuminata/metabolism , DNA-Binding Proteins/metabolism , Double-Blind Method , Humans , Imiquimod , Interferon Inducers/therapeutic use , Interferons/genetics , Interferons/metabolism , Keratinocytes/drug effects , Keratinocytes/physiology , Protein Inhibitors of Activated STAT , Proteins/genetics , Proteins/metabolism , RNA, Messenger/metabolism , STAT1 Transcription Factor , STAT3 Transcription Factor , Trans-Activators/metabolismABSTRACT
OBJECTIVES: Imiquimod (IQ) is used clinically for the topical treatment of external genital warts. IQ is an immune response modifier and induces the expression of interferon-alpha and other cytokines in human Peripheral Blood Monocytes (PBMC). Trophoblasts have been previously shown to express inflammatory cytokines upon lipopolysaccharide (LPS) stimulation. The objective of this study was to evaluate the ability of IQ to induce transcription of cytokines in trophoblasts. METHODS: A transformed human first trimester trophoblast cell line, HTR-8/SVneo, was cultured in DMEM containing IQ at concentrations of 0 to 5.0 microg/ml. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) viability assays were conducted to control for any drug-induced cell death. Total RNA was isolated from trophoblasts at 0, 8 and 24 hours of culture and Reverse Transcription-Polymerase Chain Reaction (RT-PCR) was conducted using specific amplimers for the inflammatory cytokines interleukin (IL)-1alpha, IL-1beta, IL-6 and IL-8. RT-PCR of beta-actin was performed to control for equal RNA loading. RESULTS: RT-PCR was unable detect an increase in either IL-1alpha, IL-1beta, IL-6 or IL-8 mRNA in first trimester trophoblasts cultured in the presence of 0 to 5.0 microg/mL of IQ for up to 24 hours. RT-PCR confirmed equal RNA loading and MTT viability assays did not show loss of cell viability at concentrations of IQ up to 5.0 microg/ml. CONCLUSIONS: IQ, at the concentrations tested, did not induce the transcriptional expression of inflammatory cytokines in human first trimester trophoblasts. These data suggest that IQ would not induce the expression of inflammatory cytokines in placental trophoblasts.
Subject(s)
Aminoquinolines/pharmacology , Cytokines/biosynthesis , Cytokines/drug effects , Interferon Inducers/pharmacology , RNA/analysis , Trophoblasts/metabolism , Cell Line , Cell Survival/drug effects , Cell Survival/physiology , Dose-Response Relationship, Drug , Female , Humans , Imiquimod , Pregnancy , Pregnancy Trimester, First , Reverse Transcriptase Polymerase Chain Reaction , Sensitivity and Specificity , Trophoblasts/cytologyABSTRACT
Langerhans cells are bone marrow derived dendritic cells that represent the major antigen-presenting cells in the skin. Langerhans cells take up and process antigen within the epidermis and present processed antigen to T lymphocyte in the regional lymph nodes and thus form an integral part of the cutaneous immune response. The cutaneous immune response can be modified by a number of pharmacologic agents, including corticosteroids, cyclosporine, and retinoids as well as physical agents, such as ultraviolet light. For the most part these agents act by suppressing immune function. A topical immune response modifier, imiquimod has been shown to enhance the cutaneous immune response. Imiquimod has anti-viral and anti-tumor effects in animal models and has been approved for the topical treatment of external genital and perianal warts in humans. The biologic activity of imiquimod in part is due to its effect as a cytokine inducer. Preliminary data suggested that imiquimod could have an effect on Langerhans cells. In order to clarify this effect on Langerhans cells, we examined Langerhans cell morphology and migration in imiquimod-treated skin. The density of Ia + cells decreased 2 d after treatment, falling to approximately 43% by day 10. The Ia positive in cells remaining in the skin appeared larger and more dendritic suggesting an activated state. ATPase staining of epidermal sheet confirmed the decreased number of Langerhans cells. To clarify status of Langerhans cells, the activation of B7 was examined. Activation of B7-1 or B7-2 was not detected. Imiquimod, however, did enhance Langerhans cell migration from skin to draining lymph nodes. This enhanced Langerhans cell migration was also associated with an enhanced allergic contact hypersensitivity. These results suggest that the mechanism of modulation of immune response by imiquimod is in part due to effects on Langerhans cells.
Subject(s)
Adjuvants, Immunologic/pharmacology , Aminoquinolines/pharmacology , Langerhans Cells/drug effects , Adenosine Triphosphatases/metabolism , Administration, Topical , Animals , Antibody Formation/drug effects , Cell Count/drug effects , Cell Movement/drug effects , Cytokines/genetics , Dermatitis, Contact/physiopathology , Epidermal Cells , Histocompatibility Antigens Class II/analysis , Imiquimod , Langerhans Cells/cytology , Langerhans Cells/enzymology , Langerhans Cells/immunology , Langerhans Cells/physiology , Mice , Mice, Inbred C57BL , RNA, Messenger/metabolism , Up-RegulationABSTRACT
Imiquimod (R-837) and its more potent derivative (R-848) are imidazoquinolines that have adjuvant activity in cultured human mononuclear cells. Its mechanism of action on epidermal antigen-presenting cells is not known. The purpose of the present investigation was to determine whether imiquimod and R-848 affect human epidermal Langerhans' cells' (LC) in vitro maturation. Pulse incubations (6-16 h) of cultured unfractionated epidermal cells or highly enriched LC suspensions with either imiquimod or R-848 (0. 05-5.0 microg/ml of culture medium) reproducibly enhanced their ability to induce T-cell proliferation in a primary mixed lymphocyte reaction. There was a 30 to 300% increase in T-lymphocyte proliferation induced by either imiquimod- or R-848-treated LC when compared to control, untreated LC. IFN-gamma secretion by T-lymphocytes stimulated by imiquimod- or R-848-treated LC was increased compared to control, untreated LC. After a 6-h incubation, phenotypic analysis of control-, imiquimod-, or R-848-treated LC indicated that such antigen-presenting cells were in an "intermediate" state of maturation (CD1a(+), HLA-DR, DP, DQ(bright+), CD40(low+), CD86(high+), and CD80(low+)). RNase protection assays demonstrated that either imiquimod or R-848 treatments increased steady-state transcripts encoding for IL-12 p40, IL-1beta, TNF-alpha, and IL-1 receptor antagonist by LC. These data indicate that imiquimod and R-848 dissociate the functional maturation (cytokine-mediated) and phenotypic maturation of epidermal LC. These data warrant further exploration for the use of imidazoquinoline-treated LC or other DC subsets for processing and presentation of viral peptides to Th-lymphocytes as a novel vaccine strategy to induce protective antiviral responses.
Subject(s)
Adjuvants, Immunologic/pharmacology , Aminoquinolines/pharmacology , Imidazoles/pharmacology , Immunologic Factors/pharmacology , Langerhans Cells/cytology , Antigen Presentation/drug effects , Cell Division/drug effects , Cell Division/genetics , Cells, Cultured , Cytokines/genetics , Gene Expression/drug effects , Humans , Imiquimod , Lymphocyte Culture Test, Mixed , Phenotype , Th1 Cells/cytologyABSTRACT
Agents that enhance dendritic cell maturation can enhance T-cell activation and therefore may improve the efficiency of vaccines or improve cellular immunotherapy. Previously, we demonstrated that a novel low-molecular-weight synthetic immune response modifier, R-848, induces IL-12 and IFN-alpha secretion from monocytes and macrophages. Here we report that R-848 induces the maturation of human monocyte-derived dendritic cells. Characteristic of dendritic cell maturation, R-848 treatment induces cell surface expression of CD83 and increases cell surface expression of CD80, CD86, CD40, and HLA-DR. Additionally, R-848 induces cytokine (IL-6, IL-12, TNF-alpha, IFN-alpha) and chemokine (IL-8, MIP-1alpha, MCP-1) secretion from dendritic cells. Most significantly, R-848 enhances dendritic cell antigen presenting function, as measured by increased T-cell proliferation and T-cell cytokine secretion in both allogeneic and autologous T-cell systems. Consequently, low-molecular-weight synthetic molecules such as R-848 and its derivatives may be useful as vaccine adjuvants or as ex vivo stimulators of dendritic cells for cellular immunotherapy.
Subject(s)
Dendritic Cells/drug effects , Dendritic Cells/immunology , Imidazoles/pharmacology , T-Lymphocytes/immunology , Antigens, CD/biosynthesis , Antigens, Surface/biosynthesis , Cell Differentiation/drug effects , Cell Differentiation/immunology , Cell Division/drug effects , Cells, Cultured , Chemokines/biosynthesis , Chemokines/metabolism , Cytokines/biosynthesis , Cytokines/metabolism , Dendritic Cells/metabolism , Dose-Response Relationship, Drug , Flow Cytometry , Humans , Lipopolysaccharides/pharmacology , Lymphocyte Activation/drug effects , Lymphocyte Activation/immunology , Monocytes/cytology , T-Lymphocytes/cytology , T-Lymphocytes/metabolism , Tumor Necrosis Factor-alpha/biosynthesis , Tumor Necrosis Factor-alpha/metabolismABSTRACT
The mechanism of action of imiquimod 5% cream applied topically to patients with genital warts was evaluated in a double-blind, placebo-controlled study. Imiquimod (16 patients) or placebo (three patients) was applied three times per week for up to 16 weeks. All imiquimod-treated patients had a > or =75% reduction in total wart area while only one of three placebo-treated patients had a similar reduction. Wart biopsies were taken at prestudy, week 6, and end of treatment. Polymerase chain reaction (PCR) for human papillomavirus (HPV) DNA and reverse transcriptase (RT)-PCR for messenger (m)RNAs were used to identify cytokines, cellular markers, viral gene products, and cell cycle markers in these biopsies. Treatment with imiquimod, an immune response modifier, stimulated significant increases in mRNA for interferon (IFN)-alpha, IFN-gamma and 2',5' oligoadenylate synthetase (2',5'-AS) as well as a tendency towards increases in tumor necrosis factor (TNF)-alpha and interleukin-12 p40. Significant increases in mRNA for CD4 and a trend toward increases in CD8 were also observed in imiquimod-treated patients, suggesting activation of a cell mediated immune response. Imiquimod administration was also associated with a significant decrease in viral load as measured by HPV DNA and L1 mRNA. The effects on HPV markers were accompanied by an apparent decrease in mRNA expression for markers of cell proliferation and an increase in mRNA for markers of keratinocyte differentiation and tumor suppressors.
Subject(s)
Aminoquinolines/therapeutic use , Condylomata Acuminata/drug therapy , Genital Diseases, Female/drug therapy , Genital Diseases, Male/drug therapy , Interferon Inducers/therapeutic use , Adolescent , Adult , Antigens, CD/genetics , Antigens, CD/metabolism , Cell Differentiation , Cell Division , Condylomata Acuminata/immunology , Condylomata Acuminata/virology , Cytokines/genetics , Cytokines/metabolism , Double-Blind Method , Female , Genital Diseases, Female/immunology , Genital Diseases, Female/virology , Genital Diseases, Male/immunology , Genital Diseases, Male/virology , Humans , Imiquimod , Keratinocytes/pathology , Male , Middle Aged , Papillomaviridae/genetics , Papillomaviridae/isolation & purification , Papillomaviridae/physiology , Polymerase Chain Reaction , RNA, Messenger/metabolism , Viral LoadABSTRACT
Imiquimod (S-26308, R-837) (1-(2-methylpropyl)-1H-imidazo[4,5-c]quinolin-4 amine), an immune response modifier, demonstrates potent antiviral and antitumor activity in animal models (see structure in Fig. 1). The drug exhibits no direct antiviral or antiproliferative activity when tested in a number of cell culture systems. Imiquimod's activity was discovered while screening for anti-herpes virus activity. One of the first analogs in the series, S-25059 was tested in the early 1980's and due to slight toxicity, caused slightly reduced herpes cytopathology in Vero cell cultures. Follow-up testing in herpes infected guinea pigs showed complete protection toward lesion development. Activity of these drugs results primarily from interferon alpha (IFN-alpha) induction and other cytokine induction. At least part of the cytokine induction is mediated through NF-kappaB activation. These cytokines stimulate several other aspects of the innate immune response. In addition, imiquimod stimulates acquired immunity, in particular the cellular arm which is important for control of viral infections and various tumors. This effect is mediated by drug induced IFN-alpha and Interleukin-12 (IL-12) and IFN-gamma induced by these cytokines. Imiquimod is expected to be effective where exogenous IFN-alpha has shown utility and where enhancement of cell-mediated immunity is needed. The following is a brief review of the preclinical pharmacology of imiquimod and the clinical results of genital wart trials. The mechanism of action of topically applied imiquimod will likely lead to benefits in several other chronic virus infections and tumors of the skin. Two other reviews on imiquimod that focus mainly on the clinical results have been published (Beutner & Geisse, 1997; Slade, Owens, Tomai & Miller, 1998).
Subject(s)
Adjuvants, Immunologic/pharmacology , Aminoquinolines/pharmacology , Antiviral Agents/pharmacology , Adjuvants, Immunologic/administration & dosage , Administration, Topical , Aminoquinolines/administration & dosage , Animals , Antiviral Agents/administration & dosage , Humans , ImiquimodABSTRACT
Imiquimod is an oral inducer of interferon (IFN) and several other proinflammatory cytokines and has been successfully used topically as an antiviral agent for the treatment of genital warts. We have investigated the molecular mechanisms by which imiquimod induces the expression of IFNs, IFN-stimulated genes (ISGs), and proinflammatory cytokines in vivo, using mice deficient in various components of the IFN signaling system. Mice deficient in the transcription factor interferon regulatory factor 1 (IRF-1) or in the serine/threonine protein kinase PKR responded normally to imiquimod, producing high levels of circulating IFN and induction of several ISGs. On the other hand, when mice deficient in STAT-1 were treated, a 32-fold reduction in the level of circulating IFN was observed, together with a lack of induction of 2-5 oligo adenylate synthetase (2-5 OAS) and IRF-1 genes. Interestingly, there was also a lack of induction of interleukin-6 (IL-6) gene expression, although tumor necrosis factor was induced and readily detected in serum. In mice deficient in the type I IFN receptor, imiquimod induced levels of IFN similar to those in control mice, but again, neither 2-5 OAS, IRF-1, nor IL-6 genes were induced in mutant mice. Our results suggest that STAT-1 plays a critical role in the mechanism of gene activation by imiquimod. Moreover, induction of IL-6 gene expression appears to be dependent on components of the IFN signaling cascade.
Subject(s)
Aminoquinolines/pharmacology , DNA-Binding Proteins/immunology , Interferon Inducers/pharmacology , Interferons/biosynthesis , Interleukin-6/biosynthesis , Trans-Activators/immunology , Administration, Oral , Animals , DNA-Binding Proteins/biosynthesis , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Imiquimod , Interferon Regulatory Factor-1 , Interferons/immunology , Interleukin-6/genetics , Interleukin-6/immunology , Mice , Phosphoproteins/biosynthesis , Phosphoproteins/genetics , Phosphoproteins/immunology , RNA, Messenger/biosynthesis , STAT1 Transcription Factor , Trans-Activators/metabolismABSTRACT
The purpose of this work was to develop a topical formulation of imiquimod, a novel immune response modifier, to induce local cytokine production for the treatment of external genital and perianal warts. A pH-solubility profile and titration data were used to calculate a pKa of 7.3, indicative of a weak base. Solubility experiments were conducted to identify a solvent that dissolves imiquimod to achieve a 5% formulation concentration. Studies to select surfactants, preservatives, and viscosity-enhancing excipients to formulate an oil-in-water cream indicated that fatty acids were the preferred solvent for topical imiquimod formulations, and isostearic acid (ISA) was selected. A relationship existed between the fatty acid composition of four commercially available ISA sources and the solubility of imiquimod. A combination of polysorbate 60, sorbitan monostearate, and xanthan gum was used to produce a physically stable cream. The preservative system included parabens and benzyl alcohol to meet the USP criteria for preservative activity. An in vitro method was developed to demonstrate that imiquimod was released from the formulation. Topical application of the formulation induced local cytokine activity in mice.
Subject(s)
Adjuvants, Immunologic/pharmacology , Aminoquinolines/pharmacology , Adjuvants, Immunologic/administration & dosage , Adjuvants, Immunologic/chemistry , Administration, Topical , Aminoquinolines/administration & dosage , Aminoquinolines/chemistry , Animals , Chemistry, Pharmaceutical , Cytokines/biosynthesis , Emulsions , Excipients , Gels , Hydrogen-Ion Concentration , Imiquimod , Male , Mice , Mice, Hairless , Molecular Structure , Ointments , Skin/metabolism , Solubility , Stearic Acids , Surface-Active Agents , Suspensions , TemperatureABSTRACT
Cytokines produced by antigen-presenting cells are known to affect the development and cytokine profile of T cells. The immune response modifiers imiquimod and R-848 were previously shown to stimulate human and mouse cultures to secrete interferon-alpha. Results from the present study demonstrate that R-848 and imiquimod are capable of inducing interleukin-12 and interferon-gamma in mouse and human cell cultures. Both CD4(+) and CD8(+) T lymphocytes were responsible for producing IFN-gamma following stimulation with R-848. Macrophages were required for induction of interferon-gamma by R-848 and the cytokines IFN-alpha and IL-12 mediated this response. R-848 and imiquimod were also found to inhibit IL-4 and IL-5 production in mouse and human culture systems. The inhibition of IL-5 in response to R-848 is seen in cultures containing CD4(+) lymphocytes and macrophages and is mediated in part by IFN-alpha. These data suggest that imiquimod and R-848 may have clinical utility in diseases where cell-mediated immune responses are important and in diseases associated with overexpression of IL-4 or IL-5 such as atopic disease.