Identification of Near-Pan-neutralizing Antibodies against HIV-1 by Deconvolution of Plasma Humoral Responses.

Anti-HIV-1 envelope broadly neutralizing monoclonal antibodies (bNAbs) isolated from memory B cells may not fully represent HIV-1-neutralizing profiles measured in plasma. Accordingly, we characterized near-pan-neutralizing antibodies extracted directly from the plasma of two "elite neutralizers." Circulating anti-gp120 polyclonal antibodies were deconvoluted using proteomics to guide lineage analysis of bone marrow plasma cells. In both subjects, a single lineage of anti-CD4-binding site (CD4bs) antibodies explained the plasma-neutralizing activity. Importantly, members of these lineages potently neutralized 89%-100% of a multi-tier 117 pseudovirus panel, closely matching the specificity and breadth of the circulating antibodies. X-ray crystallographic analysis of one monoclonal, N49P7, suggested a unique ability to bypass the CD4bs Phe43 cavity, while reaching deep into highly conserved residues of Layer 3 of the gp120 inner domain, likely explaining its extreme potency and breadth. Further direct analyses of plasma anti-HIV-1 bNAbs should provide new insights for developing antibody-based antiviral agents and vaccines.

Altered Response Pattern following AZD5582 Treatment of SIV-Infected, ART-Suppressed Rhesus Macaque Infants.

The "shock and kill" strategy for HIV-1 cure incorporates latency-reversing agents (LRA) in combination with interventions that aid the host immune system in clearing virally reactivated cells. LRAs have not yet been investigated in pediatric clinical or preclinical studies. Here, we evaluated an inhibitor of apoptosis protein (IAP) inhibitor (IAPi), AZD5582, that activates the noncanonical NF-κB (ncNF-κB) signaling pathway to reverse latency. Ten weekly doses of AZD5582 were intravenously administered at 0.1 mg/kg to rhesus macaque (RM) infants orally infected with SIVmac251 at 4 weeks of age and treated with a triple ART regimen for over 1 year. During AZD5582 treatment, on-ART viremia above the limit of detection (LOD, 60 copies/mL) was observed in 5/8 infant RMs starting at 3 days post-dose 4 and peaking at 771 copies/mL. Of the 135 measurements during AZD5582 treatment in these 5 RM infants, only 8 were above the LOD (6%), lower than the 46% we have previously reported in adult RMs. Pharmacokinetic analysis of plasma AZD5582 levels revealed a lower Cmax in treated infants compared to adults (294 ng/mL versus 802 ng/mL). RNA-Sequencing of CD4+ T cells comparing pre- and post-AZD5582 dosing showed many genes that were similarly upregulated in infants and adults, but the expression of key ncNF-κB genes, including NFKB2 and RELB, was significantly higher in adult RMs. Our results suggest that dosing modifications for this latency reversal approach may be necessary to maximize virus reactivation in the pediatric setting for successful "shock and kill" strategies. IMPORTANCE While antiretroviral therapy (ART) has improved HIV-1 disease outcome and reduced transmission, interruption of ART results in rapid viral rebound due to the persistent latent reservoir. Interventions to reduce the viral reservoir are of critical importance, especially for children who must adhere to lifelong ART to prevent disease progression. Here, we used our previously established pediatric nonhuman primate model of oral SIV infection to evaluate AZD5582, identified as a potent latency-reversing agent in adult macaques, in the controlled setting of daily ART. We demonstrated the safety of the IAPi AZD5582 and evaluate the pharmacokinetics and pharmacodynamics of repeated dosing. The response to AZD5582 in macaque infants differed from what we previously showed in adult macaques with weaker latency reversal in infants, likely due to altered pharmacokinetics and less inducibility of infant CD4+ T cells. These data supported the contention that HIV-1 cure strategies for children are best evaluated using pediatric model systems.

Novel prosperous computational estimations for greenhouse gas adsorptive control by zeolites using machine learning methods.

To predict CO2 adsorptive capture, as a vital environmental issue, using different zeolites including 5A, 13X, T-Type, SSZ-13, and SAPO-34, different models have been developed by implementing artificial intelligence algorithms. Hybrid adaptive neuro-fuzzy inference system (Hybrid-ANFIS), particle swarm optimization-adaptive neuro-fuzzy inference system (PSO-ANFIS) and the least-squares support vector machine (LSSVM) modeling optimized with the coupled simulated annealing (CSA) optimization have been employed for the models. The developed models, validated by utilizing various graphical and statistical methods exhibited that the Hybrid-ANFIS model estimations for the gas adsorption on 5A, T-Type, SSZ-13, and SAPO-34 zeolites with average absolute relative deviation (AARD) % of 8.21, 1.92, 4.99 and 2.26, and PSO ANFIS model estimations for the gas adsorption on zeolite 13X with an AARD of 4.85% were in good agreement with corresponding experimental data. It could be deduced that the proposed models were more prosperous and efficient in favor of the design and analysis of adsorption processes than previous ones.

SMAC Mimetic Plus Triple-Combination Bispecific HIVxCD3 Retargeting Molecules in SHIV.C.CH505-Infected, Antiretroviral Therapy-Suppressed Rhesus Macaques.

The "shock-and-kill" human immunodeficiency virus type 1 (HIV-1) cure strategy involves latency reversal followed by immune-mediated clearance of infected cells. We have previously shown that activation of the noncanonical NF-κB pathway using an inhibitor of apoptosis (IAP), AZD5582, reverses HIV/simian immunodeficiency virus (SIV) latency. Here, we combined AZD5582 with bispecific HIVxCD3 DART molecules to determine the impact of this approach on persistence. Rhesus macaques (RMs) (n = 13) were infected with simian/human immunodeficiency virus SHIV.C.CH505.375H.dCT, and triple antiretroviral therapy (ART) was initiated after 16 weeks. After 42 weeks of ART, 8 RMs received a cocktail of 3 HIVxCD3 DART molecules having human A32, 7B2, or PGT145 anti-HIV-1 envelope (Env) specificities paired with a human anti-CD3 specificity that is rhesus cross-reactive. The remaining 5 ART-suppressed RMs served as controls. For 10 weeks, a DART molecule cocktail was administered weekly (each molecule at 1 mg/kg of body weight), followed 2 days later by AZD5582 (0.1 mg/kg). DART molecule serum concentrations were well above those considered adequate for redirected killing activity against Env-expressing target cells but began to decline after 3 to 6 weekly doses, coincident with the development of antidrug antibodies (ADAs) against each of the DART molecules. The combination of AZD5582 and the DART molecule cocktail did not increase on-ART viremia or cell-associated SHIV RNA in CD4+ T cells and did not reduce the viral reservoir size in animals on ART. The lack of latency reversal in the model used in this study may be related to low pre-ART viral loads (median, <105 copies/ml) and low preintervention reservoir sizes (median, <102 SHIV DNA copies/million blood CD4+ T cells). Future studies to assess the efficacy of Env-targeting DART molecules or other clearance agents to reduce viral reservoirs after latency reversal may be more suited to models that better minimize immunogenicity and have a greater viral burden.IMPORTANCE The most significant barrier to an HIV-1 cure is the existence of the latently infected viral reservoir that gives rise to rebound viremia upon cessation of ART. Here, we tested a novel combination approach of latency reversal with AZD5582 and clearance with bispecific HIVxCD3 DART molecules in SHIV.C.CH505-infected, ART-suppressed rhesus macaques. We demonstrate that the DART molecules were not capable of clearing infected cells in vivo, attributed to the lack of quantifiable latency reversal in this model with low levels of persistent SHIV DNA prior to intervention as well as DART molecule immunogenicity.

Latency Reversal 2.0: Giving the Immune System a Seat at the Table.

PURPOSE OF REVIEW: For most people living with HIV (PLWH), treatment with effective antiretroviral therapy (ART) results in suppression of viremia below the limit of detection of clinical assays, immune reconstitution, reduced immune activation, avoidance of opportunistic infections, and progression to AIDS. However, ART alone is not curative, and HIV persists in a non-replicating, latent form. In this review, we provide a historical perspective on non-specific latency reversal approaches (LRA 1.0) and summarize recent advances in latency reversal strategies that target specific signaling pathways within CD4+ T cells or other immune cells to induce expression of latent HIV (immune-based latency reversal, or LRA 2.0). RECENT FINDINGS: The HIV reservoir is primarily composed of latently infected CD4+ T cells carrying integrated, replication-competent provirus that can give rise to rebound viremia if ART is stopped. Myeloid lineage cells also contribute to HIV latency in certain tissues; we focus here on CD4+ T cells as a sufficient body of evidence regarding latency reversal in myeloid cells is lacking. The immunomodulatory LRA 2.0 approaches we describe include pattern recognition receptor agonists, immune checkpoint inhibitors, non-canonical NF-kB stimulation, and transient CD8+ lymphocyte depletion, along with promising combination strategies. We highlight recent studies demonstrating robust latency reversal in nonhuman primate models. While significant strides have been made in terms of virus reactivation from latency, initial hopes for latency reversal alone to result in a reduction of infected cells, through viral cytopathic effect or an unboosted immune system, have not been realized and it seems clear that even effective latency reversal strategies will need to be paired with an approach that facilitates immune recognition and clearance of cells containing reactivated virus.

Rigorous prognostication and modeling of gas adsorption on activated carbon and Zeolite-5A.

Gas adsorption on various adsorbents is of highly important issue for the separation of gas mixtures in many industrial processes. In this work, estimation of pure gases (CH4, N2, CO2, H2, C2H4) adsorption on activated carbon (AC) and CO2, CH4, N2 on Zeolite-5A adsorbent were studied by developing four different computing techniques, namely MLP-ANN, ANFIS, LSSVM, and PSO-ANFIS for a broad range of experimental data found in the literature. Temperature, pressure, pore size (only for AC) and kinetic diameter of adsorbed gases are considered as the inputs and the gas adsorption as the output parameters of the developed models. We also used several statistical and graphical tools to assess the accuracy and applicability of the proposed models. The results of the study suggest the reliability and validity of all the models developed for estimating the equilibrium adsorption of gases on the adsorbents. Also, it is found that of all the models developed, the ANN model estimates experimental data of the gas adsorption on AC more accurately due to its values of R2 and AARD%, 0.9865 and 0.8948, respectively. Besides, PSO-ANFIS is the best model to prognosticate gas adsorption on zeolite 5A with R2 and AARD%, 0.9897 and 0.9551, respectively.

Application of machine learning models to improve the prediction of pesticide photodegradation in water by ZnO-based photocatalysts.

Pesticide pollution has been posing a significant risk to human and ecosystems, and photocatalysis is widely applied for the degradation of pesticides. Machine learning (ML) emerges as a powerful method for modeling complex water treatment processes. For the first time, this study developed novel ML models that improved the estimation of the photocatalytic degradation of various pesticides using ZnO-based photocatalysts. The input parameters encompassed the source of light, mass proportion of dopants to Zn, initial pesticide concentration (C0), pH of the solution, catalyst dosage and irradiation time. Additionally, physicochemical properties such as the molecular weight of the dopants and pesticides, as well as the water solubility of both dopants and pesticides, were considered. Notably, the numerical data were extracted from the literature via relevant tables (directly) or graphs (indirectly) using the web-based tool WebPlotDigitizer. Four ML models including multi-layer perceptron artificial neural network (MLP-ANN), particle swarm optimization-adaptive neuro fuzzy inference system (PSO-ANFIS), radial basis function (RBF), and coupled simulated annealing-least squares support vector machine (CSA-LSSVM) were developed. In comparison, RBF showed the best accuracy of modeling among all models, with the highest determination coefficient (R2) of 0.978 and average absolute relative deviation (AARD) of 4.80%. RBF model was effective in estimating the photocatalytic degradation of pesticides except for 2-chlorophenol, triclopyr and lambda-cyhalothrin, where CSA-LSSVM model demonstrated superior performance. Dichlorvos was completely degraded by ZnO photocatalyst under visible light. The sensitivity analysis by relevancy factor exhibited that light irradiation time and initial pesticide concentration were the most important parameters influencing photocatalytic degradation of pesticides positively and negatively, respectively. The new ML models provide a powerful tool for predicting pesticide degradation in wastewater treatment, which will reduce photochemical experiments and promote sustainable development.

AZD5582 plus SIV-specific antibodies reduce lymph node viral reservoirs in antiretroviral therapy-suppressed macaques.

The main barrier to HIV cure is a persistent reservoir of latently infected CD4+ T cells harboring replication-competent provirus that fuels rebound viremia upon antiretroviral therapy (ART) interruption. A leading approach to target this reservoir involves agents that reactivate latent HIV proviruses followed by direct clearance of cells expressing induced viral antigens by immune effector cells and immunotherapeutics. We previously showed that AZD5582, an antagonist of inhibitor of apoptosis proteins and mimetic of the second mitochondrial-derived activator of caspases (IAPi/SMACm), induces systemic reversal of HIV/SIV latency but with no reduction in size of the viral reservoir. In this study, we investigated the effects of AZD5582 in combination with four SIV Env-specific Rhesus monoclonal antibodies (RhmAbs) ± N-803 (an IL-15 superagonist) in SIV-infected, ART-suppressed rhesus macaques. Here we confirm the efficacy of AZD5582 in inducing SIV reactivation, demonstrate enhancement of latency reversal when AZD5582 is used in combination with N-803 and show a reduction in total and replication-competent SIV-DNA in lymph-node-derived CD4+ T cells in macaques treated with AZD5582 + RhmAbs. Further exploration of this therapeutic approach may contribute to the goal of achieving an HIV cure.

Brain microglia serve as a persistent HIV reservoir despite durable antiretroviral therapy.

Brain microglia (MG) may serve as a human immunodeficiency virus 1 (HIV) reservoir and ignite rebound viremia following cessation of antiretroviral therapy (ART), but they have yet to be proven to harbor replication-competent HIV. Here, we isolated brain myeloid cells (BrMCs) from nonhuman primates and rapid autopsy of people with HIV (PWH) on ART and sought evidence of persistent viral infection. BrMCs predominantly displayed microglial markers, in which up to 99.9% of the BrMCs were TMEM119+ MG. Total and integrated SIV or HIV DNA was detectable in the MG, with low levels of cell-associated viral RNA. Provirus in MG was highly sensitive to epigenetic inhibition. Outgrowth virus from parietal cortex MG in an individual with HIV productively infected both MG and PBMCs. This inducible, replication-competent virus and virus from basal ganglia proviral DNA were closely related but highly divergent from variants in peripheral compartments. Phenotyping studies characterized brain-derived virus as macrophage tropic based on the ability of the virus to infect cells expressing low levels of CD4. The lack of genetic diversity in virus from the brain suggests that this macrophage-tropic lineage quickly colonized brain regions. These data demonstrate that MG harbor replication-competent HIV and serve as a persistent reservoir in the brain.

HIV Reservoirs: Modeling, Quantification, and Approaches to a Cure.

Biomedical research in animal models depends heavily on nonhuman primates (NHP) (Phillips et al., Am J Primatol 76(9):801-827, 2014). In their physiology, neurobiology, and, most importantly, their susceptibility to infectious diseases and subsequent immune responses, NHPs have many parallels with humans (Rhesus Macaque Genome Sequencing and Analysis Consortium et al., Science 316(5822):222-234, 2007). Different species of NHPs have served as important animal models for numerous infectious diseases spanning a wide range of pathogens (Gardner and Luciw, ILAR J 49(2):220-255, 2008). As a result of recognizing their utility in HIV research, NHPs have contributed to groundbreaking studies of disease pathogenesis, vaccination, and curative research (London et al., Lancet 2(8355):869-873, 1983; Henrickson et al., Lancet 1 (8321):388-390, 1983). Many African NHPs are considered natural hosts for SIV in which SIV infection is usually nonprogressive and does not cause acquired immunodeficiency syndrome (AIDS) (Chahroudi et al., Science 335(6073):1188-1193, 2012; Taaffe et al., J Virol 84(11):5476-5484, 2010). However, cross-species transmission of SIV strains to other NHPs or to humans (nonnatural hosts) leads to progressive disease and AIDS (Paiardini et al., Annu Rev Med 60:485-495, 2009). In particular, SIV infection of Asian rhesus macaques recapitulates many features of HIV infection in humans and therefore has become a widely used approach for contemporary HIV research into virus persistence and cure strategies (Gardner and Luciw, FASEB J 3(14):2593-2606, 1989). There are multiple factors that should be considered in HIV/SIV studies using NHPs including the particular monkey species and geographic background, age and sex, certain genetic properties, virus strain, route and dose of infection, interventional treatments, and prespecified study outcomes. Here, we discuss consideration of these factors to address specific questions in HIV cure research.

Evaluation of CO2 Absorption by Amino Acid Salt Aqueous Solution Using Hybrid Soft Computing Methods.

Amino acid salt (AAs) aqueous solutions have recently exhibited a great potential in CO2 absorption from various gas mixtures. In this work, four hybrid machine learning methods were developed to evaluate 626 CO2 and AAs equilibrium data for different aqueous solutions of AAs (potassium sarcosinate, potassium l-asparaginate, potassium l-glutaminate, sodium l-phenylalanine, sodium glycinate, and potassium lysinate) gathered from reliable references. The models are the hybrids of the least squares support vector machine and coupled simulated annealing optimization algorithm, radial basis function neural network (RBF-NN), particle swarm optimization-adaptive neuro-fuzzy inference system, and hybrid adaptive neuro-fuzzy inference system. The inputs of the models are the CO2 partial pressure, temperature, mass concentration in the aqueous solution, molecular weight of AAs, hydrogen bond donor count, hydrogen bond acceptor count, rotatable bond count, heavy atom count, and complexity, and the CO2 loading capacity of AAs aqueous solution is considered as the output of the models. The accuracies of the models' results were verified through graphical and statistical analyses. RBF-NN performance is promising and surpassed that of other models in estimating the CO2 loading capacities of AAs aqueous solutions.

Relationship between Young's Modulus and Planar Density of Unit Cell, Super Cells (2 × 2 × 2), Symmetry Cells of Perovskite (CaTiO3) Lattice.

Calcium titanate-CaTiO3 (perovskite) has been used in various industrial applications due to its dopant/doping mechanisms. Manipulation of defective grain boundaries in the structure of perovskite is essential to maximize mechanical properties and stability; therefore, the structure of perovskite has attracted attention, because without fully understanding the perovskite structure and diffracted planes, dopant/doping mechanisms cannot be understood. In this study, the areas and locations of atoms and diffracted planes were designed and investigated. In this research, the relationship between Young's modulus and planar density of unit cell, super cells (2 × 2 × 2) and symmetry cells of nano CaTiO3 is investigated. Elastic constant, elastic compliance and Young's modulus value were recorded with the ultrasonic pulse-echo technique. The results were C11 = 330.89 GPa, C12 = 93.03 GPa, C44 = 94.91 GPa and E = 153.87 GPa respectively. Young's modulus values of CaTiO3 extracted by planar density were calculated 162.62 GPa, 151.71 GPa and 152.21 GPa for unit cell, super cells (2 × 2 × 2) and symmetry cells, respectively. Young's modulus value extracted by planar density of symmetry cells was in good agreement with Young's modulus value measured via ultrasonic pulse-echo.

X-ray Diffraction Analysis and Williamson-Hall Method in USDM Model for Estimating More Accurate Values of Stress-Strain of Unit Cell and Super Cells (2 × 2 × 2) of Hydroxyapatite, Confirmed by Ultrasonic Pulse-Echo Test.

Taking into account X-ray diffraction, one of the well-known methods for calculating the stress-strain of crystals is Williamson-Hall (W-H). The W-H method has three models, namely (1) Uniform deformation model (UDM); (2) Uniform stress deformation model (USDM); and (3) Uniform deformation energy density model (UDEDM). The USDM and UDEDM models are directly related to the modulus of elasticity (E). Young's modulus is a key parameter in engineering design and materials development. Young's modulus is considered in USDM and UDEDM models, but in all previous studies, researchers used the average values of Young's modulus or they calculated Young's modulus only for a sharp peak of an XRD pattern or they extracted Young's modulus from the literature. Therefore, these values are not representative of all peaks derived from X-ray diffraction; as a result, these values are not estimated with high accuracy. Nevertheless, in the current study, the W-H method is used considering the all diffracted planes of the unit cell and super cells (2 × 2 × 2) of Hydroxyapatite (HA), and a new method with the high accuracy of the W-H method in the USDM model is presented to calculate stress (σ) and strain (ε). The accounting for the planar density of atoms is the novelty of this work. Furthermore, the ultrasonic pulse-echo test is performed for the validation of the novelty assumptions.

Elimination of SHIV Infected Cells by Combinations of Bispecific HIVxCD3 DART® Molecules.

Bispecific HIVxCD3 DART molecules that co-engage the viral envelope glycoprotein (Env) on HIV-1-infected cells and the CD3 receptor on CD3+ T cells are designed to mediate the cytolysis of HIV-1-infected, Env-expressing cells. Using a novel ex vivo system with cells from rhesus macaques (RMs) infected with a chimeric Simian-Human Immunodeficiency Virus (SHIV) CH505 and maintained on ART, we tested the ability of HIVxCD3 DART molecules to mediate elimination of in vitro-reactivated CD4+ T cells in the absence or presence of autologous CD8+ T cells. HIVxCD3 DART molecules with the anti-HIV-1 Env specificities of A32 or 7B2 (non-neutralizing antibodies) or PGT145 (broadly neutralizing antibody) were evaluated individually or combined. DART molecule-mediated antiviral activity increased significantly in the presence of autologous CD8+ T cells. In this ex vivo system, the PGT145 DART molecule was more active than the 7B2 DART molecule, which was more active than the A32 DART molecule. A triple combination of the DART molecules exceeded the activity of the individual PGT145 DART molecule. Modified quantitative virus outgrowth assays confirmed the ability of the DART molecules to redirect RM CD3+ T cells to eliminate SHIV-infected RM CD4+ T cells as demonstrated by the decreased propagation of in vitro infection by the infected cells pre-incubated with DART molecules in presence of effector CD8+ T cells. While mediating cytotoxic activity, DART molecules did not increase proinflammatory cytokine production. In summary, combination of HIVxCD3 DART molecules that have broadly-neutralizing and non-neutralizing anti-HIV-1 Env specificities can leverage the host immune system for treatment of HIV-1 infection but will require appropriate reactivation of the latent reservoir.

Measurement Modulus of Elasticity Related to the Atomic Density of Planes in Unit Cell of Crystal Lattices.

Young's modulus (E) is one of the most important parameters in the mechanical properties of solid materials. Young's modulus is proportional to the stress and strain values. There are several experimental and theoretical methods for gaining Young's modulus values, such as stress-strain curves in compression and tensile tests, electromagnetic-acoustic resonance, ultrasonic pulse echo and density functional theory (DFT) in different basis sets. Apparently, preparing specimens for measuring Young's modulus through the experimental methods is not convenient and it is time-consuming. In addition, for calculating Young's modulus values by software, presumptions of data and structures are needed. Therefore, this new method for gaining the Young's modulus values of crystalline materials is presented. Herein, the new method for calculating Young's modulus of crystalline materials is extracted by X-ray diffraction. In this study, Young's modulus values were gained through the arbitrary planes such as random (hkl) in the research. In this study, calculation of Young's modulus through the relationship between elastic compliances, geometry of the crystal lattice and the planar density of each plane is obtained by X-ray diffraction. Sodium chloride (NaCl) with crystal lattices of FCC was selected as the example. The X-ray diffraction, elastic stiffness constant and elastic compliances values have been chosen by the X'Pert software, literature and experimental measurements, respectively. The elastic stiffness constant and Young's modulus of NaCl were measured by the ultrasonic technique and, finally, the results were in good agreement with the new method of this study. The aim of the modified Williamson-Hall (W-H) method in the uniform stress deformation model (USDM) utilized in this paper is to provide a new approach of using the W-H equation, so that a least squares technique can be applied to minimize the sources of errors.

Broadly Neutralizing Antibodies against HIV: Back to Blood.

After years of continuous exposure to HIV envelope antigens, a minority of HIV-infected individuals develop a cognate polyclonal humoral response comprising very potent and extremely cross-reactive neutralizing antibodies [broadly neutralizing antibodies (bNAbs)]. Isolated bNAbs derived from memory B cell pools have been the focus of intense studies over the past decade. However, it is not yet known how to translate the features of bNAbs into practical HIV prevention methods. In this review, we attempt to seek insights from emerging information about the human broadly neutralizing plasma response as well as its frequency, clonal composition, specificity, potency, and commonality among infected subjects. We also consider how this information points to selecting and prioritizing certain epitope targets and strategies for HIV vaccine design.

Induction of Fc-Mediated Effector Functions Against a Stabilized Inner Domain of HIV-1 gp120 Designed to Selectively Harbor the A32 Epitope Region.

Recent clinical trials and studies using nonhuman primates (NHPs) suggest that antibody-mediated protection against HIV-1 will require α-HIV envelope humoral immunity beyond direct neutralization to include Fc-receptor (FcR) mediated effector functions such as antibody-dependent cellular cytotoxicity (ADCC). There is also strong evidence indicating that the most potent ADCC response in humans is directed toward transitional non-neutralizing epitopes associated with the gp41-interactive face of gp120, particularly those within the first and second constant (C1-C2) region (A32-like epitopes). These epitopes were shown to be major targets of ADCC responses during natural infection and have been implicated in vaccine-induced protective immunity. Here we describe the immunogenicity of ID2, an immunogen consisting of the inner domain of the clade A/E 93TH057 HIV-1 gp120 expressed independently of the outer domain (OD) and stabilized in the CD4-bound conformation to harbor conformational A32 region epitopes within a minimal structural unit of HIV-1 Env. ID2 induced A32-specific antibody responses in BALB/c mice when injected alone or in the presence of the adjuvants Alum or GLA-SE. Low α-ID2 titers were detected in mice immunized with ID2 alone whereas robust responses were observed with ID2 plus adjuvant, with the greatest ID2 and A32-specific titers observed in the GLA-SE group. Only sera from groups immunized in the presence of GLA-SE were capable of mediating significant ADCC using NKr cells sensitized with recombinant BaL gp120 as targets and human PBMCs as effectors. A neutralization response to a tier 2 virus was not observed. Altogether, our studies demonstrate that ID2 is highly immunogenic and elicits A32-specific ADCC responses in an animal host. The ID2 immunogen has significant translational value as it can be used in challenge studies to evaluate the role of non-neutralizing antibodies directed at the A32 subregion in HIV-1 protection.

Identification of different Escherichia coli pathotypes in north and north-west provinces of Iran.

BACKGROUND AND OBJECTIVES: Diarrhea is one of the most prevalent diseases in the world, specially in developing countries. One of the most important causative agents of bacterial diarrhea is diarrheagenic Escherichia coli (DEC) which causes gastroenteritis and this group involving enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC), enteroaggregative E. coli (EAEC), enterohemoragic E. coli (EHEC), enteroinvasive E. coli (EIEC), diffusely adherence E. coli (DAEC). The aim of this study was to identify different E. coli pathotypes in north and north-west of Iran, among the clinical isolates. MATERIALS AND METHODS: In this study for identification of E. coli, 170 fecal samples were cultured on MacConkey agar and identified by biochemical tests. Samples with E. coli characteristics were selected (145 samples) and their genomes were purified by phenol-chloroform method. After extraction of genomes, lt and sta genes identified by PCR for ETEC, eae gene for atypical and eae and bfp for typical EPEC, AA region for EAEC, stx1 and stx2 and eae genes for EHEC (stx1 or stx2 or both for STEC) and invE for EIEC. RESULTS: Finally 10 samples identified as ETEC (%5.88), 18 (%10.58) EPEC, 6 (%3.52) EHEC and 12 (7.05%) samples were STEC. None of the samples were positive for EAEC and EIEC. CONCLUSION: The results obtained in this study showed that ETEC, EPEC, EHEC and STEC are prevalent bacterial agents in north and north-west of Iran. Complementary studies to identify these pathotypes in other seasons can help to adopt necessary policies against outbreaks in Iran.

Dendritic cell based immunotherapy using tumor stem cells mediates potent antitumor immune responses.

Cancer stem cells (CSCs) are demonstrated to be usually less sensitive to conventional methods of cancer therapies, resulting in tumor relapse. It is well-known that an ideal treatment would be able to selectively target and kill CSCs, so as to avoid the tumor reversion. The aim of our present study was to evaluate the effectiveness of a dendritic cell (DC) based vaccine against CSCs in a mouse model of malignant melanoma. C57BL/6 mouse bone marrow derived DCs pulsed with a murine melanoma cell line (B16F10) or CSC lysates were used as a vaccine. Immunization of mice with CSC lysate-pulsed DCs was able to induce a significant prophylactic effect by a higher increase in lifespan and obvious depression of tumor growth in tumor bearing mice. The mice vaccinated with DCs loaded with CSC-lysate were revealed to produce specific cytotoxic responses to CSCs. The proliferation assay and cytokine (IFN-γ and IL-4) secretion of mice vaccinated with CSC lysate-pulsed DCs also showed more favorable results, when compared to those receiving B16F10 lysate-pulsed DCs. These findings suggest a potential strategy to improve the efficacy of DC-based immunotherapy of cancers.