Phagocytosis by an HIV antibody is associated with reduced viremia irrespective of enhanced complement lysis.

Increasingly, antibodies are being used to treat and prevent viral infections. In the context of HIV, efficacy is primarily attributed to dose-dependent neutralization potency and to a lesser extent Fc-mediated effector functions. It remains unclear whether augmenting effector functions of broadly neutralizing antibodies (bNAbs) may improve their clinical potential. Here, we use bNAb 10E8v4 targeting the membrane external proximal region (MPER) to examine the role of antibody-mediated effector and complement (C') activity when administered prophylactically against SHIV challenge in rhesus macaques. With sub-protective dosing, we find a 78-88% reduction in post-acute viremia that is associated with 10E8v4-mediated phagocytosis acting at the time of challenge. Neither plasma nor tissue viremic outcomes in vivo is improved with an Fc-modified variant of 10E8v4 enhanced for C' functions as determined in vitro. These results suggest that effector functions inherent to unmodified 10E8v4 contribute to efficacy against SHIVSF162P3 in the absence of plasma neutralizing titers, while C' functions are dispensable in this setting, informing design of bNAb modifications for improving protective efficacy.

Non-neutralizing antibodies targeting the immunogenic regions of HIV-1 envelope reduce mucosal infection and virus burden in humanized mice.

Antibodies are principal immune components elicited by vaccines to induce protection from microbial pathogens. In the Thai RV144 HIV-1 vaccine trial, vaccine efficacy was 31% and the sole primary correlate of reduced risk was shown to be vigorous antibody response targeting the V1V2 region of HIV-1 envelope. Antibodies against V3 also were inversely correlated with infection risk in subsets of vaccinees. Antibodies recognizing these regions, however, do not exhibit potent neutralizing activity. Therefore, we examined the antiviral potential of poorly neutralizing monoclonal antibodies (mAbs) against immunodominant V1V2 and V3 sites by passive administration of human mAbs to humanized mice engrafted with CD34+ hematopoietic stem cells, followed by mucosal challenge with an HIV-1 infectious molecular clone expressing the envelope of a tier 2 resistant HIV-1 strain. Treatment with anti-V1V2 mAb 2158 or anti-V3 mAb 2219 did not prevent infection, but V3 mAb 2219 displayed a superior potency compared to V1V2 mAb 2158 in reducing virus burden. While these mAbs had no or weak neutralizing activity and elicited undetectable levels of antibody-dependent cellular cytotoxicity (ADCC), V3 mAb 2219 displayed a greater capacity to bind virus- and cell-associated HIV-1 envelope and to mediate antibody-dependent cellular phagocytosis (ADCP) and C1q complement binding as compared to V1V2 mAb 2158. Mutations in the Fc region of 2219 diminished these effector activities in vitro and lessened virus control in humanized mice. These results demonstrate the importance of Fc functions other than ADCC for antibodies without potent neutralizing activity.

Polyfunctional Tier 2-Neutralizing Antibodies Cloned following HIV-1 Env Macaque Immunization Mirror Native Antibodies in a Human Donor.

Vaccine efforts to combat HIV are challenged by the global diversity of viral strains and shielding of neutralization epitopes on the viral envelope glycoprotein trimer. Even so, the isolation of broadly neutralizing Abs from infected individuals suggests the potential for eliciting protective Abs through vaccination. This study reports a panel of 58 mAbs cloned from a rhesus macaque (Macaca mulatta) immunized with envelope glycoprotein immunogens curated from an HIV-1 clade C-infected volunteer. Twenty mAbs showed neutralizing activity, and the strongest neutralizer displayed 92% breadth with a median IC50 of 1.35 µg/ml against a 13-virus panel. Neutralizing mAbs predominantly targeted linear epitopes in the V3 region in the cradle orientation (V3C) with others targeting the V3 ladle orientation (V3L), the CD4 binding site (CD4bs), C1, C4, or gp41. Nonneutralizing mAbs bound C1, C5, or undetermined conformational epitopes. Neutralization potency strongly correlated with the magnitude of binding to infected primary macaque splenocytes and to the level of Ab-dependent cellular cytotoxicity, but did not predict the degree of Ab-dependent cellular phagocytosis. Using an individualized germline gene database, mAbs were traced to 23 of 72 functional IgHV alleles. Neutralizing V3C Abs displayed minimal nucleotide somatic hypermutation in the H chain V region (3.77%), indicating that relatively little affinity maturation was needed to achieve in-clade neutralization breadth. Overall, this study underscores the polyfunctional nature of vaccine-elicited tier 2-neutralizing V3 Abs and demonstrates partial reproduction of the human donor's humoral immune response through nonhuman primate vaccination.

An HIV Vaccine Targeting the V2 Region of the HIV Envelope Induces a Highly Durable Polyfunctional Fc-Mediated Antibody Response in Rhesus Macaques.

The HIV vaccine field now recognizes the potential importance of generating polyfunctional antibodies (Abs). The only clinical HIV vaccine trial to date to show significant efficacy (RV144) found that reduced infection rates correlated with the level of nonneutralizing Abs specific for the V2 region of the envelope glycoprotein. We have conducted a comprehensive preclinical reverse vaccinology-based vaccine program that has included the design and production and testing of numerous scaffolded V2 region immunogens. The most immunogenic vaccine regimen in nonhuman primates among those studied as part of this program consisted of a cocktail of three immunogens presenting V2 from different viruses and clades in the context of different scaffolds. Presently we demonstrate that the V2-specific Ab response from this regimen was highly durable and functionally diverse for the duration of the study (25 weeks after the final immunization). The total IgG binding response at this late time point exhibited only an ∼5× reduction in potency. Three immunizations appeared essential for the elicitation of a strong Ab-dependent cellular cytotoxicity (ADCC) response for all animals, as opposed to the Ab-dependent cellular phagocytosis (ADCP) and virus capture responses, which were comparably potent after only 2 immunizations. All functionalities measured were highly durable through the study period. Therefore, testing this vaccine candidate for its protective capacity is warranted.IMPORTANCE The only HIV vaccine trial for which protective efficacy was detected correlated this efficacy with V2-specific Abs that were effectively nonneutralizing. This result has fueled a decade of HIV vaccine research focused on designing an HIV vaccine capable of eliciting V2-focused, polyfunctional Abs that effectively bind HIV and trigger various leukocytes to kill the virus and restrict viral spread. From the numerous vaccine candidates designed and tested as part of our V2-focused preclinical vaccine program, we have identified immunogens and a vaccine regimen that induces a highly durable and polyfunctional V2-focused Ab response in rhesus macaques, described herein.

Modified Adenovirus Prime-Protein Boost Clade C HIV Vaccine Strategy Results in Reduced Viral DNA in Blood and Tissues Following Tier 2 SHIV Challenge.

Designing immunogens and improving delivery methods eliciting protective immunity is a paramount goal of HIV vaccine development. A comparative vaccine challenge study was performed in rhesus macaques using clade C HIV Envelope (Env) and SIV Gag antigens. One group was vaccinated using co-immunization with DNA Gag and Env expression plasmids cloned from a single timepoint and trimeric Env gp140 glycoprotein from one of these clones (DNA+Protein). The other group was a prime-boost regimen composed of two replicating simian (SAd7) adenovirus-vectored vaccines expressing Gag and one Env clone from the same timepoint as the DNA+Protein group paired with the same Env gp140 trimer (SAd7+Protein). The env genes were isolated from a single pre-peak neutralization timepoint approximately 1 year post infection in CAP257, an individual with a high degree of neutralization breadth. Both DNA+Protein and SAd7+Protein vaccine strategies elicited significant Env-specific T cell responses, lesser Gag-specific responses, and moderate frequencies of Env-specific TFH cells. Both vaccine modalities readily elicited systemic and mucosal Env-specific IgG but not IgA. There was a higher frequency and magnitude of ADCC activity in the SAd7+Protein than the DNA+Protein arm. All macaques developed moderate Tier 1 heterologous neutralizing antibodies, while neutralization of Tier 1B or Tier 2 viruses was sporadic and found primarily in macaques in the SAd7+Protein group. Neither vaccine approach provided significant protection from viral acquisition against repeated titered mucosal challenges with a heterologous Tier 2 clade C SHIV. However, lymphoid and gut tissues collected at necropsy showed that animals in both vaccine groups each had significantly lower copies of viral DNA in individual tissues compared to levels in controls. In the SAd7+Protein-vaccinated macaques, total and peak PBMC viral DNA were significantly lower compared with controls. Taken together, this heterologous Tier 2 SHIV challenge study shows that combination vaccination with SAd7+Protein was superior to combination DNA+Protein in reducing viral seeding in tissues in the absence of protection from infection, thus emphasizing the priming role of replication-competent SAd7 vector. Despite the absence of correlates of protection, because antibody responses were significantly higher in this vaccine group, we hypothesize that vaccine-elicited antibodies contribute to limiting tissue viral seeding.

Insights into the dynamics of memory, effector and apoptotic cytotoxic T lymphocytes in channel catfish, Ictalurus punctatus.

In this study, we used the channel catfish model clonal TS32.15 alloantigen-specific cytotoxic T cell (CTL) line to examine the dynamics of memory CTL expansion and senescence in teleosts. Although TS32.15 has been routinely cultured to study catfish CTL responses and killing mechanisms, little is known about the dynamics of the CTLs in these cultures. Here we show that this cell line consists of small non-cytotoxic T cells and larger granular effector T cells and that their ratios vary with time after stimulation. Small CTLs, when exposed to their irradiated targets, replicate and differentiate to morphologically distinct cytotoxic effectors, which do not replicate. After lysing target cells, or with prolonged absence of stimulation, the effector cells transition to a non-cytolytic senescent stage or become apoptotic. In addition, we demonstrate that natural IgM in catfish serum binds lipids, including PIP2, on early apoptotic CTLs, and that these IgM+ CTL can be cleared by catfish head kidney-derived macrophages.

Risk factors for the development of pleural empyema in children.

Pediatric pleural empyema has increased substantially over the past 20 years and reasons for this rise remain not fully explained. We investigated potential risk factors for the development of empyema in children by examining a cohort of patients with community-acquired pneumonia. Demographic, clinical, and socioeconomic characteristics, use of Ibuprofen prior to presentation and selected potential epidemiological risk factors were analyzed. Data were collected from a prospective etiological study of radiologically confirmed pneumonia in hospitalized children aged ≤16 years. One hundred sixty children were enrolled; 56% were male and 69% aged <5 years. Empyema complication developed in 40 (25%) children. Children with empyema were more frequently prescribed Ibuprofen prior to admission to hospital than those without (82% vs. 46.2%; OR 1.94, 97.5% credible interval 0.80-3.18). Bacterial infection was strongly associated with the development of empyema (OR 3.34, 97.5% credible interval 1.70-5.14). In contrast age, sex, maternal age, parental smoking, level of socioeconomic status, nursery attendance, asthma, household characteristics (bedrooms and number of occupants) were not significantly different between groups. In conclusion, children with pneumonia who developed empyema had more often received Ibuprofen prior to hospitalization and confirmed bacterial infection. We suggest a population-based study involving both primary and secondary care settings would help to investigate the role of Ibuprofen use in modulating the course of disease in children with pneumonia.

Use of somatostatin analogues to treat chylothorax in a child with Generalised Lymphatic Dysplasia.

Generalised Lymphatic Dysplasia is a rare condition that may be associated with significant chylothoraces. The management of such effusions is often challenging. We present the case of a 15-year-old girl with bilateral chylothoraces and lymphoedema of her limbs. A clinical diagnosis of Generalised Lymphatic Dysplasia was made and long-term treatment with somatostatin analogues (somatostatin initially followed by monthly octreotide) was initiated. Over 12 months there was symptomatic benefit with some objective improvement in lung function and no adverse effects. After a year of treatment there was some reaccumulation of fluid, however this did not require any intervention. This is the first paediatric report of the use of somatostatin analogues to manage chylothorax in Generalised Lymphatic Dysplasia and we conclude that they represent a potentially useful treatment modality. Experience is only anecdotal however and further studies are required to establish an evidence base with regard to efficacy and safety.

Bronchomalacia occurring in monozygotic twins--further information about its inheritance.

UNLABELLED: We report left main-stem bronchomalacia occurring in monozygotic twins that became symptomatic during infancy. This phenomenon has not previously been reported to occur in twins. Little is known about the aetiology of primary bronchomalacia, but this occurrence suggests that there may be a significant genetic component in some cases. CONCLUSION: Left main-stem bronchomalacia occurring in monozygotic twins suggests that there may be a significant genetic component in some cases and provides further information about the inheritance of this condition.

Chronic respiratory complications in pediatric heart transplant recipients.

BACKGROUND: The frequency and spectrum of chronic respiratory complications in pediatric heart transplant recipients have not been extensively studied. The aim of this study was to describe the chronic respiratory complications in 126 consecutive pediatric heart transplant recipients. METHODS: Retrospective review of medical records. RESULTS: Between 1987 and 2005, 126 (64 males and 62 females) heart transplantations were performed at Freeman Hospital, Newcastle upon Tyne, United Kingdom. The median age at transplantation was 7.4 years (range, 0.1-17) and the median length of follow-up was 6.8 years (range, 0-18.2). Twenty-four patients have died, and 36 have been transferred to adult follow-up, leaving 66 under pediatric follow-up. Chronic respiratory complications have been documented in 33 children (50%). Bronchiectasis has been identified in 10 children, and 12 further children have had recurrent lower respiratory tract infections (without bronchiectasis) requiring long-term antibiotic prophylaxis. Of those with infectious complications, 81% underwent transplantation before 4 years and had deficiency of pneumococcal-specific antibody response. Obstructive sleep apnea has occurred in 5 children, sub-glottic stenosis has occurred in 3, and significant compression of the left main stem bronchus related to a disproportionately large donor heart has occurred in 2. One child had marked mosaic attenuation on chest computed tomography scan indicative of small airways disease. CONCLUSION: Chronic respiratory complications are common in pediatric heart transplant recipients. The respiratory prognosis for this complex group of patients is usually good, but long-term follow-up by both a respiratory pediatrician and an immunologist is frequently required.