Interplay between gain and loss in arrays of nonlinear plasmonic nanoparticles: toward parametric downconversion and amplification.

With the help of a theoretical model and finite-difference time-domain (FDTD) simulations based on the hydrodynamic-Maxwell model, we examine the effect of difference-frequency generation (DFG) in an array of L-shaped metal nanoparticles (MNPs) characterized by intrinsic plasmonic nonlinearity. The outcomes of the calculations reveal the spectral interplay between gain and loss in the vicinity of the fundamental frequency of the localized surface plasmon resonances. Subsequently, we identify different array thicknesses and pumping regimes facilitating parametric amplification and spontaneous parametric downconversion. Our results suggest that the parametric amplification regime becomes feasible on a scale of hundreds of nanometers and spontaneous parametric downconversion on the scale of tens of nanometers, opening up new exciting opportunities for developing building blocks of photonic metasurfaces.

Human IgG lacking effector functions demonstrate lower FcRn-binding and reduced transplacental transport.

We have previously generated human IgG1 antibodies that were engineered for reduced binding to the classical Fcγ receptors (FcγRI-III) and C1q, thereby eliminating their destructive effector functions (constant region G1Δnab). In their potential use as blocking agents, favorable binding to the neonatal Fc receptor (FcRn) is important to preserve the long half-life typical of IgG. An ability to cross the placenta, which is also mediated, at least in part, by FcRn is desirable in some indications, such as feto-maternal alloimmune disorders. Here, we show that G1Δnab mutants retain pH-dependent binding to human FcRn but that the amino acid alterations reduce the affinity of the IgG1:FcRn interaction by 2.0-fold and 1.6-fold for the two antibodies investigated. The transport of the modified G1Δnab mutants across monolayers of human cell lines expressing FcRn was approximately 75% of the wild-type, except that no difference was observed with human umbilical vein endothelial cells. G1Δnab mutation also reduced transport in an ex vivo placenta model. In conclusion, we demonstrate that, although the G1Δnab mutations are away from the FcRn-binding site, they have long-distance effects, modulating FcRn binding and transcellular transport. Our findings have implications for the design of therapeutic human IgG with tailored effector functions.

Igg Subclasses Targeting the Flagella of Salmonella enterica Serovar Typhimurium Can Mediate Phagocytosis and Bacterial Killing.

Invasive non-typhoidal Salmonella are a common cause of invasive disease in immuno-compromised individuals and in children. Multi-drug resistance poses challenges to disease control, with a critical need for effective vaccines. Flagellin is an attractive vaccine candidate due to surface exposure and high epitope copy number, but its potential as a target for opsonophacytic antibodies is unclear. We examined the effect of targeting flagella with different classes of IgG on the interaction between Salmonella Typhimurium and a human phagocyte-like cell line, THP-1. We tagged the FliC flagellar protein with a foreign CD52 mimotope (TSSPSAD) and bacteria were opsonized with a panel of humanised CD52 antibodies with the same antigen-binding V-region, but different constant regions. We found that IgG binding to flagella increases bacterial phagocytosis and reduces viable intracellular bacterial numbers. Opsonisation with IgG3, followed by IgG1, IgG4, and IgG2, resulted in the highest level of bacterial uptake and in the highest reduction in the intracellular load of viable bacteria. Taken together, our data provide proof-of-principle evidence that targeting flagella with antibodies can increase the antibacterial function of host cells, with IgG3 being the most potent subclass. These data will assist the rational design of urgently needed, optimised vaccines against iNTS disease.

Human IgG1 antibodies suppress angiogenesis in a target-independent manner.

Aberrant angiogenesis is implicated in diseases affecting nearly 10% of the world's population. The most widely used anti-angiogenic drug is bevacizumab, a humanized IgG1 monoclonal antibody that targets human VEGFA. Although bevacizumab does not recognize mouse Vegfa, it inhibits angiogenesis in mice. Here we show bevacizumab suppressed angiogenesis in three mouse models not via Vegfa blockade but rather Fc-mediated signaling through FcγRI (CD64) and c-Cbl, impairing macrophage migration. Other approved humanized or human IgG1 antibodies without mouse targets (adalimumab, alemtuzumab, ofatumumab, omalizumab, palivizumab and tocilizumab), mouse IgG2a, and overexpression of human IgG1-Fc or mouse IgG2a-Fc, also inhibited angiogenesis in wild-type and FcγR humanized mice. This anti-angiogenic effect was abolished by Fcgr1 ablation or knockdown, Fc cleavage, IgG-Fc inhibition, disruption of Fc-FcγR interaction, or elimination of FcRγ-initated signaling. Furthermore, bevacizumab's Fc region potentiated its anti-angiogenic activity in humanized VEGFA mice. Finally, mice deficient in FcγRI exhibited increased developmental and pathological angiogenesis. These findings reveal an unexpected anti-angiogenic function for FcγRI and a potentially concerning off-target effect of hIgG1 therapies.

The INNs and outs of antibody nonproprietary names.

An important step in drug development is the assignment of an International Nonproprietary Name (INN) by the World Health Organization (WHO) that provides healthcare professionals with a unique and universally available designated name to identify each pharmaceutical substance. Monoclonal antibody INNs comprise a -mab suffix preceded by a substem indicating the antibody type, e.g., chimeric (-xi-), humanized (-zu-), or human (-u-). The WHO publishes INN definitions that specify how new monoclonal antibody therapeutics are categorized and adapts the definitions to new technologies. However, rapid progress in antibody technologies has blurred the boundaries between existing antibody categories and created a burgeoning array of new antibody formats. Thus, revising the INN system for antibodies is akin to aiming for a rapidly moving target. The WHO recently revised INN definitions for antibodies now to be based on amino acid sequence identity. These new definitions, however, are critically flawed as they are ambiguous and go against decades of scientific literature. A key concern is the imposition of an arbitrary threshold for identity against human germline antibody variable region sequences. This leads to inconsistent classification of somatically mutated human antibodies, humanized antibodies as well as antibodies derived from semi-synthetic/synthetic libraries and transgenic animals. Such sequence-based classification implies clear functional distinction between categories (e.g., immunogenicity). However, there is no scientific evidence to support this. Dialog between the WHO INN Expert Group and key stakeholders is needed to develop a new INN system for antibodies and to avoid confusion and miscommunication between researchers and clinicians prescribing antibodies.

Clearance of human IgG1-sensitised red blood cells in vivo in humans relates to the in vitro properties of antibodies from alternative cell lines.

We previously produced a recombinant version of the human anti-RhD antibody Fog-1 in the rat myeloma cell line, YB2/0. When human, autologous RhD-positive red blood cells (RBC) were sensitised with this IgG1 antibody and re-injected, they were cleared much more rapidly from the circulation than had been seen earlier with the original human-mouse heterohybridoma-produced Fog-1. Since the IgG have the same amino acid sequence, this disparity is likely to be due to alternative glycosylation that results from the rat and mouse cell lines. By comparing the in vitro properties of YB2/0-produced Fog-1 IgG1 and the same antibody produced in the mouse myeloma cell line NS0, we now have a unique opportunity to pinpoint the cause of the difference in ability to clear RBC in vivo. Using transfected cell lines that express single human FcγR, we showed that IgG1 made in YB2/0 and NS0 cell lines bound equally well to receptors of the FcγRI and FcγRII classes but that the YB2/0 antibody was superior in FcγRIII binding. When measuring complexed IgG binding, the difference was 45-fold for FcγRIIIa 158F, 20-fold for FcγRIIIa 158V and approximately 40-fold for FcγRIIIb. The dissimilarity was greater at 100-fold in monomeric IgG binding assays with FcγRIIIa. When used to sensitise RBC, the YB2/0 IgG1 generated 100-fold greater human NK cell antibody-dependent cell-mediated cytotoxicity and had a 103-fold advantage over the NS0 antibody in activating NK cells, as detected by CD54 levels. In assays of monocyte activation and macrophage adherence/phagocytosis, where FcγRI plays major roles, RBC sensitised with the two antibodies produced much more similar results. Thus, the alternative glycosylation profiles of the Fog-1 antibodies affect only FcγRIII binding and FcγRIII-mediated functions. Relating this to the in vivo studies confirms the importance of FcγRIII in RBC clearance.

Severity and interference of chronic pain in methadone-maintained outpatients.

OBJECTIVES: Patients with opioid use disorder maintained on methadone report more chronic pain than the general population. The current study characterized chronic pain in patients with opioid use disorder. DESIGN: A one-time self-report survey. SETTING: The Addiction Treatment Services methadone-maintenance clinic, located on the campus of Johns Hopkins Bayview Medical Center in Baltimore MD. SUBJECTS: A convenience sample of 227 methadone-maintained patients. METHODS: Participants completed a one-time self-report administration of the brief pain inventory (BPI) and a demographic survey; additional treatment variables were obtained directly from clinic records. RESULTS: Sixty percent of the sample endorsed chronic pain. Patients with pain were significantly older, had a higher mean methadone dose, and provided more benzodiazepine-positive urine samples. Pain was primarily located in the lower extremities (59%) and back (51%), and mean BPI severity and interference subscale scores were 5.7 and 5.4 out of 10, respectively. Logistic regressions indicated that age (P < 0.001) and methadone dose (P < 0.001) were significantly associated with having pain and that pain was a significant predictor of benzodiazepine use (P = 0.01). Only 13% (N = 18) of patients with pain were receiving pain management, and few were being treated with any nonopioid adjuvant analgesics. Yet patients who did receive treatment reported a mean 51% improvement in their pain, indicating they are not treatment refractory. CONCLUSIONS: Results suggest there is a large discrepancy in the percent of patients who may need treatment for pain and those receiving treatment for pain and that more efforts should be made to provide standard pain management techniques to patients with opioid use disorder to reduce their overall level of pain and potentially improve their overall treatment outcomes.

Low-affinity FcγR interactions can decide the fate of novel human IgG-sensitised red blood cells and platelets.

G1Δnab is a mutant human IgG1 constant region with a lower ability to interact with FcγR than the natural IgG constant regions. Radiolabelled RBCs and platelets sensitised with specific G1Δnab Abs were cleared more slowly from human circulation than IgG1-sensitised counterparts. However, non-destructive splenic retention of G1Δnab-coated RBCs required investigation and plasma radioactivities now suggest this also occurred for platelets sensitised with an IgG1/G1Δnab mixture. In vitro assays with human cells showed that G1Δnab-sensitised RBCs did not cause FcγRI-mediated monocyte activation, FcγRIIIa-mediated antibody-dependent cell-mediated cytotoxicity (ADCC) or macrophage phagocytosis although they did adhere to macrophages. Thus, FcγRII was implicated in the adhesion despite the Δnab mutation reducing the already low-affinity binding to this receptor class. Additional contacts via P-selectin enhance the interaction of sensitised platelets with monocytes and this system provided evidence of FcγRII-dependent activation by G1Δnab. These results emphasise the physiological relevance of low-affinity interactions: It appears that FcγRII interactions of G1Δnab allowed splenic retention of G1Δnab-coated RBCs with inhibitory FcγRIIb binding preventing RBC destruction and that FcγRIIb engagement by G1Δnab on IgG1/G1Δnab-sensitised platelets overcame activation by IgG1. Considering therapeutic blocking Abs, G1Δnab offers lower FcγR binding and a greater bias towards inhibition than IgG2 and IgG4 constant regions.

Targeting systemic inflammation in patients with obesity-related pain: How best to prevent acute pain from becoming chronic?

The best approach is to find the individual risk factors and known predictors and manage them early on.

Borderline personality disorder and chronic pain: a practical approach to evaluation and treatment.

Patients with chronic pain present a spectrum of complexity that can be overwhelming for the individual practitioner. These patients require thoughtful care and a comprehensive treatment plan. This complexity should be acknowledged, not avoided, and the patient should be engaged, not shunned. A practical approach will assist in developing expertise and proceeding empathically. The presence of a superimposed personality disorder significantly increases the difficulty of caring for these patients. Studies investigating the prevalence of borderline personality disorder in patients with chronic pain averaged 30 %, highlighting the importance of being able to effectively treat this patient population. Appropriate management of these patients should focus on a collaboration to practice productive behaviors despite intense emotional distress. Longitudinal research provides a foundation for an optimistic prognosis that can be enhanced with this rehabilitative approach.

Quantification of the effects of antibodies on the extra- and intracellular dynamics of Salmonella enterica.

Antibodies are known to be essential in controlling Salmonella infection, but their exact role remains elusive. We recently developed an in vitro model to investigate the relative efficiency of four different human immunoglobulin G (IgG) subclasses in modulating the interaction of the bacteria with human phagocytes. Our results indicated that different IgG subclasses affect the efficacy of Salmonella uptake by human phagocytes. In this study, we aim to quantify the effects of IgG on intracellular dynamics of infection by combining distributions of bacterial numbers per phagocyte observed by fluorescence microscopy with a mathematical model that simulates the in vitro dynamics. We then use maximum likelihood to estimate the model parameters and compare them across IgG subclasses. The analysis reveals heterogeneity in the division rates of the bacteria, strongly suggesting that a subpopulation of intracellular Salmonella, while visible under the microscope, is not dividing. Clear differences in the observed distributions among the four IgG subclasses are best explained by variations in phagocytosis and intracellular dynamics. We propose and compare potential factors affecting the replication and death of bacteria within phagocytes, and we discuss these results in the light of recent findings on dormancy of Salmonella.

From stigmatized neglect to active engagement.

Chronic pain and substance abuse are common problems. Each entity represents a significant and independent burden to the patients affected by them, the healthcare system caring for them, and society at large supporting them. If the two problems occur together, all of these burdens and their consequences are magnified. Traditional treatments fail a substantial percentage of even the most straightforward cases. Clearly, new approaches are required for the most complex of cases. Success is possible only if multiple disciplines provide integrated care that incorporates all of the principles of substance abuse and chronic pain rehabilitation treatment into one package. While experience provides the foundation for implementing these programs, research that documents the methods behind successful outcomes will be needed to sustain support for them.

A behaviorist perspective.

Chronic pain is a sensory experience that produces suffering and functional impairment and is the result of both sensory input as well as secondary adaptation of the nervous system. The sensitization of the nervous system to pain is influenced by physical activity (or inactivity) and medication exposure. Medication taking and physical activity are behaviors that are increased or decreased by positive and negative reinforcement. Patients often have comorbid psychiatric conditions at presentation, including addictions, mood disorders, personality vulnerabilities and life circumstances that amplify their disability and impede their recovery. Behavioral conditioning contributes to chronic pain disorders in the form of both classical (Pavlov) and operant (Skinner) conditioning that increases the experience of pain, the liability to ongoing injury, the central amplification of pain, the use of reinforcing medications such as opiates and benzodiazepines, and behaviors associated with disability. The term 'abnormal illness behavior' has been used to describe behaviors that are associated with illness but are not explained physiologically. Behavioral conditioning often amplifies these abnormal behaviors in patients with chronic pain. Addiction can also be seen as a behavior that is reinforced and conditioned. The same factors that amplify abnormal illness behaviors also increase the liability to addiction. Psychiatric comorbidities also complicate and amplify abnormal illness behaviors and addictive behaviors and further contribute to the disability of chronic pain patients. Model interventions that reinforce healthy behaviors and extinguish illness behaviors are effective in patients with addictions and chronic pain. Maladaptive behaviors including addictive behaviors can be used as targets for classical and operant conditioning techniques, and these techniques are demonstrably effective in patients with chronic pain and addictions.

Optimizing treatment with opioids and beyond.

Patients with both chronic pain and substance use disorders offer special challenges and opportunities. They represent a large number of patients with significant costs to themselves and society that translate into poor outcome. The challenges in defining addiction in patients with chronic pain, particularly in those treated with chronic opioid therapy, have distracted the healthcare community from designing effective treatment programs. Traditional treatment programs for chronic pain disorders or substance use disorders are incapable of addressing the issues of the patients' 'other' problem. Treatment devolves to prescribing opioid medications with the belief that both disorders will be treated at least in part, which is deemed better than receiving no treatment at all. Patients are actually concerned about the risks of this type of treatment, and even if it did offer significant benefits, physicians demonstrate a lack of knowledge and skill in administering opioids to these patients. The inadequate treatment of either chronic pain or addiction interferes with the treatment of the other condition and necessitates the design of new treatment paradigms. A new approach to patients with both chronic pain and addiction should start with an evaluation and formulation of these patients to determine the different domains that contribute to their disability (diseases, dimensions, behaviors, life stories). A comprehensive formulation provides the appropriate platform for the implementation of an integrated program of therapy for both conditions that can be intensified to provide more, rather than less, care for the patient that does not meet the goals of functional rehabilitation.

Human IgG isotypes and activating Fcγ receptors in the interaction of Salmonella enterica serovar Typhimurium with phagocytic cells.

Several classes and multiple subclasses of immunoglobulins are produced towards protein and polysaccharide antigens in response to Salmonella infection and play a key role in protection against systemic disease. The targeting of Salmonella to Fc receptors (FcR) on phagocytes is a key step in the antibody-mediated antibacterial functions of host cells. We wished to compare the relative efficiency of different human IgG subclasses, which targeted the Salmonella enterica OmpA surface protein in modulating the interaction of bacteria with human phagocytes. To this end, we developed a novel system by tagging OmpA with a foreign CD52 mimotope (TSSPSAD) and opsonizing the bacteria with a panel of humanized CD52 antibodies that share the same antigen-binding V-region, but have constant regions of different subclasses. Our data revealed that opsonization with all the IgG subclasses increases Salmonella uptake by human phagocytes. IgG3 resulted in the highest level of bacterial uptake and the highest average bacterial load per infected cell, which was closely followed by IgG1, then IgG4 and lastly IgG2. Phagocytosis mediated by IgG1, IgG3 and IgG4 had a higher dependency on FcγRI than FcγRIIA, whereas IgG2-mediated phagocytosis required FcγRIIA more than FcγRI. The results show that IgG binding to OmpA increases the uptake of Salmonella by human phagocytic cells and that the efficiency of this process depends both on the subclass of the IgG and the type of FcR that is available for antibody binding.

Expression of human FcgammaRIIIa as a GPI-linked molecule on CHO cells to enable measurement of human IgG binding.

The efficacy of a therapeutic IgG molecule may be as dependent on the optimisation of the constant region to suit its intended indication as on the selection of its variable regions. A crucial effector function to be maximised or minimised is antibody-dependent cell-mediated cytotoxicity by natural killer cells. Traditional assays of ADCC activity suffer from considerable inter-donor and intra-donor variability, which makes the measurement of antibody binding to human FcgammaRIIIa, the key receptor for ADCC, an attractive alternative method of assessment. Here, we describe the development of cell lines and assays for this purpose. The transmembrane receptor, FcgammaRIIIa, requires co-expression with signal transducing subunits to prevent its degradation, unlike the homologous receptor FcgammaRIIIb that is expressed as a GPI-anchored molecule. Therefore, to simplify the production of cell lines as reliable assay components, we expressed FcgammaRIIIa as a GPI-anchored molecule. Separate, stable CHO cell lines that express either the 158F or the higher-affinity 158V allotype of FcgammaRIIIa were isolated using fluorescence-activated cell sorting. The identities of the expressed receptors were confirmed using a panel of monoclonal antibodies that distinguish between subclasses and allotypes of FcgammaRIII and the cell lines were shown to have slightly higher levels of receptor than FcgammaRIII-positive peripheral blood mononuclear cells. Because the affinity of FcgammaRIIIa for IgG is intermediate amongst the receptors that bind IgG, we were able to use these cell lines to develop flow cytometric assays to measure the binding of both complexed and monomeric immunoglobulin. Thus, by choosing the appropriate method, weakly- or strongly-binding IgG can be efficiently compared. We have quantified the difference in the binding of wildtype IgG1 and IgG3 molecules to the two functional allotypes of the receptor and report that the FcgammaRIIIa-158V-antibody interaction is 3- to 4-fold stronger that the interaction with FcgammaRIIIa-158F. Overall, these robust assays should be valuable for batch-testing clinical material as well as providing tools for improving the design of therapeutic IgG.

Psychiatric issues in chronic pain.

Chronic pain requires comprehensive care. While interdisciplinary approaches are recommended, the role of psychiatrists is often misunderstood. Psychiatrists should be involved with the care of patients with chronic pain as early as possible to maximize outcome. Psychiatrists offer an expertise that specifically addresses important deficiencies in the care of patients with chronic pain: 1) the lack of a detailed formulation, 2) the lumping of all psychopathology, and 3) the failure to effectively use psychopharmacologic treatments. This review provides a framework for formulating the diagnoses and treatments of patients with chronic pain.

Assessment and management of chronic pain in individuals seeking treatment for opioid dependence disorder.

OBJECTIVE: To review the problem of chronic pain in patients with substance use disorders, focusing on the prevalence of chronic pain in patients with substance dependence disorders, especially prescription opioid dependence, associated comorbidities, and the impact on drug abuse treatment response. METHOD: We identified relevant articles using PubMed from 1987 to 2008. Additional articles were obtained from the reference lists of key reviews of relevant topics. Studies were included if they investigated the relation between chronic pain and substance use disorders. Of particular interest were articles that proposed integrated treatment for both problems. RESULTS: The high prevalence of chronic pain syndromes was only recently explored in patients seeking treatment for drug abuse. The presence of chronic pain increases the risk of poor response to substance abuse treatment and an increased likelihood of multiple comorbidities that further add to the negative impact experienced by patients with substance dependence disorders. Substance abuse treatment programs offering integrated medical and psychiatric care for these comorbidities improve outcomes, with stepped care approaches offering the best treatment by tailoring the level of care to the individual patient's needs. CONCLUSIONS: Substance abuse treatment programs should expand their services to address the comorbidities likely to pose barriers to successful drug rehabilitation. Given the high prevalence and negative impact of chronic pain, new pain management services should be integrated within the drug treatment program and adapted as patients demonstrate the need for more intensive treatment. If applied to the problem of chronic pain, a model substance abuse treatment program of integrated stepped care would improve outcomes for patients with both devastating disorders.

Morphine versus mexiletine for treatment of postamputation pain: a randomized, placebo-controlled, crossover trial.

BACKGROUND: Stump and phantom pains are debilitating sequelae of amputations that are often resistant to treatment. The efficacy of pharmacologic therapies, including opioids and sodium channel blockers, for postamputation pain is uncertain. METHODS: The authors conducted a double-blind, randomized, placebo-controlled, crossover study in adult patients with postamputation pain of 6 months or longer and greater than 3 on a 0-10 numeric pain rating scale. Each of the three treatment periods (morphine, mexiletine, or placebo) included a 1-week drug-free interval followed by 4-week titration, 2-week maintenance, and 2-week drug-taper phases. The primary outcome measure was change in average pain intensity from the drug-free baseline to the last week of maintenance. RESULTS: Sixty amputees were enrolled; data were analyzed from 56 subjects for one drug period, 45 subjects for two drug periods, and 35 subjects who completed all three drug periods. The mean morphine and mexiletine dosages were 112 and 933 mg, respectively. Morphine treatment provided lower pain scores compared with placebo and mexiletine (P = 0.0003). The mean percent pain relief during treatment with placebo, mexiletine, and morphine was 19, 30, and 53%, respectively (P < 0.0001, morphine vs. placebo and mexiletine). The numbers needed to treat to obtain 50% and 33% decreases in pain intensity with morphine were 5.6 and 4.5, respectively. Treatment with morphine was associated with a higher rate of side effects. CONCLUSIONS: Therapy with morphine, but not mexiletine, resulted in a decrease in intensity of postamputation pain but was associated with a higher rate of side effects and no improvement in self-reported levels of overall functional activity and pain-related interference in daily activities.