text:
The 24th International AIDS Conference (AIDS 2022) officially opens on Friday in Montreal, but a Zoom press conference today shared news of results from several studies, including two case reports relevant to HIV cure research.
Jana Dickter from the City of Hope hospital in Los Angeles described another instance of "prolonged HIV-1 remission" after receipt of a stem cell transplant required to treat a life-threatening cancer, this time in a 66-year-old man. The stem cell donor was homozygous for the CCR5Δ32 mutation, which causes immune cells to be resistant to most HIV variants. Dickter emphasized the unique aspects compared to previously reported cases:
The person was older (63 at the time of the transplant procedure, 66 now)
Has been living with HIV longer (over 31 years)
Received a less immunosuppressive regimen during transplantation and a chemotherapy regimen that is better tolerated by older people
As explained in a press release from the City of Hope, the stem cell transplant was administered in early 2019 after a diagnosis of acute myelogenous leukemia (AML). Antiretroviral therapy (ART) was stopped in March 2021 and the man has now been followed for over 17 months with no HIV viral load rebound or HIV DNA detected. Antibody responses against HIV have waned and virus-specific T-cell responses are no longer detectable. The AML is in remission. Regrettably, the details of these results won't become available until Dickter presents at the conference on Monday August 1 (the abstract text is unavailable until tomorrow).
Dickter noted that the particular protocol employed at City of Hope is likely better suited to older people with HIV who require stem cell transplants to treat cancers, and may offer additional opportunities to attempt cures if appropriate CCR5Δ32 homozygote donors can be identified.
As with the New York City woman reported at CROI earlier this year, the researchers are being cautious and using the term "remission" to describe the case rather than cure, due to limited follow-up. It would be helpful for the HIV cure research field to come to consensus on how best to define "cure" in such cases, because news coverage tends not to adhere to these distinctions in terminology.
There's also an urgent need for consensus on how many cases of HIV cure/remission have been achieved by stem cell transplants from CCR5Δ32 homozygote donors: both the International AIDS Society and City of Hope press releases erroneously state that this represents the "fourth" such case but in fact, as reported for NBC News by Benjamin Ryan, there are five in total:
Timothy Ray Brown (aka The Berlin Patient)
Adam Castillejo (aka The London Patient)
The Düsseldorf Patient
The New York Patient
The City of Hope Patient
Details on each of the prior cases is included in TAG's latest Research Toward a Cure and Immune-Based Therapies Pipeline Report.
Dr. Núria Climent delivered the second report, about a case of "exceptional post-treatment control" in a woman with HIV who'd participated in a clinical trial in Barcelona. The Hospital Clínic-IDIBAPS have issued a press release about the findings. If I'm understanding correctly, it also involves an older individual: the woman was 59 at the time of study enrollment and so is in her 70s now.
The study was complicated, beginning by randomizing people with recent HIV infection to receive either ART or ART plus the immunosuppressant drug cyclosporine. There were a total of 20 participants, 19 cisgender men and the one cisgender woman (who was assigned to receive ART plus cyclosporine). Results from this initial part of the study were published in 2016.
The woman was then rolled over into another protocol that involved an analytical treatment interruption (ATI). A short course of GM-CSF and pegylated alpha interferon was given during the ATI, followed by another period of ART plus subcutaneous low-dose IL-2 before treatment was interrupted completely.
The crux of today's report is that the participant has now maintained an undetectable HIV viral load for more than 15 years off ART and has displayed a progressive decline in measures of the HIV reservoir during that time. HIV that's capable of replicating can still be detected at low levels, however, suggesting that control is being actively maintained by the immune system. In laboratory tests, the researchers identified "memory-like" natural killer cells and gamma-delta CD8 T cells as contributors to suppressing HIV replication (these represent relatively small subsets of immune system cells whose potential role in suppressing HIV hasn't received much attention historically).
The woman does not have any genetic factors that have been associated with immunological control of HIV infection, and Climent explained that she experienced quite severe symptoms during primary infection (which was diagnosed at Feibig stage 5).
In response to questions, Climent noted that the woman was the only study participant who experienced post-treatment control, so the role (if any) of the immune-based interventions that were given is unclear. The researchers have also not tested for the presence of antiretrovirals in blood samples, which is necessary to confirm that the viral load control has occurred post-treatment. The formal presentation at the conference will occur on Sunday July 31.
There's been at least one other report of extended post-treatment control in a person who received ART plus cyclosporine during early HIV infection; in that case it was a man described as "the Lausanne Patient" by researcher Guiseppe Pantaleo, who'd been off ART for over eight years at the time of the report back in 2009. HIV cure research might potentially benefit from compiling all the historical reports of post-treatment control and assessing their current status (if the information is available); so far the phenomenon appears rare but it has the potential to offer clues to help develop curative interventions.
AIDS 2022 will feature several sessions including HIV cure research in the coming days, listed below.
Pre-conference symposium: Pathways to an HIV cure: Research and advocacy priorities
Satellite: Africa HIV cure research: Strengthening industry-community engagement in clinical research
The view from the bench: Advances in HIV basic and translational research
Late Breaker Track A
Global Village: On the road to HIV cure gene therapy: Who can learn from whom?
Novel insights into the nature of the HIV reservoir and mechanisms of persistence
Sunday July 31
Approaches for HIV cure and vaccine research
Combining immunotherapeutic agents to achieve ARV-free remission of HIV
Responding to the virus: Advances in HIV immunology
Monday August 1
Late Breaker Track B
Finding the needle in the haystack: Progress in understanding the HIV reservoir
Tuesday August 2
Shake and bake: Promising strategies for HIV cure
July 27, 2022 in Analytical treatment interruptions, Conferences & meetings, Cure research, HIV remission, Natural killer cells, Post-treatment controllers, Stem cell transplantation | Permalink | Comments (2)
Significant Effects of a TLR7 Agonist Plus Broadly Neutralizing Antibody in the SHIV/Macaque Model
Among the most newsworthy presentations at CROI 2018 earlier this year was Dan Barouch's description of a study involving a toll-like receptor 7 (TLR7) agonist combined with a broadly neutralizing antibody (bNAb) in SHIV-infected macaques (see contemporary coverage by AIDSMap, i-Base and POZ Magazine). The results have now been published in the journal Nature.
The specific experimental candidates that were tested are vesatolimod (formerly known as GS-9620), a TLR7 agonist, and PGT121, a bNAb initially discovered by researchers supported by the International AIDS Vaccine Initiative but now licensed to Gilead Sciences for therapeutic development. Previous laboratory experiments using CD4 T cells isolated from people on ART found that the combination promoted the reversal of HIV latency and killing of virus-infected cells.
A key goal of Barouch's study was to establish whether bNAbs are capable of mediating depletion of the viral reservoir in vivo, in addition to displaying the direct antiviral activities that have been documented in other experiments (such as the trials described in the previous blog post).
A total of 44 macaques were infected with SHIV-SF162P3, a hybrid virus that combines elements of SIV and HIV. ART was initiated seven days later—extremely early after infection—leading to rapid suppression of SHIV replication that was maintained for 96 weeks without any viral load blips.
For the subsequent intervention phase, animals were divided into four groups of 11 and received, respectively: sham administrations (control group), vesatolimod alone, PGT121 alone, or the combination of vesatolimod and PGT121. Vesatolimod was given orally every other week between weeks 96 and 114 of the study, while PGT121 was administered five times via infusion from weeks 106-114. ART was then interrupted in all macaques at week 130.
The most dramatic finding was a significantly reduced incidence of SHIV viral load rebound in the group that received the dual vesatolimod/PGT121 combination. Out of the 11 macaques, six (55%) rebounded, compared to all 11 of the sham controls, 10 out of 11 (91%) assigned to vesatolimod alone and nine of 11 (82%) given PGT121.
Among the animals in the combination group that did rebound, viral loads were 2.6 logs lower at the peak and 1.5 logs lower at the set point compared to the controls. Additional analyses showed that levels of SHIV DNA in lymph nodes were significantly lower in the combination group prior to the ART interruption.
The interventions appeared to be well tolerated. Vesatolimod caused transient activation of CD4 T cells and natural killer cells, as well as transient increases in proinflammatory cytokines, which were evident the day after administration.
In an effort to assess whether a reservoir of replication-competent SHIV was still present in the animals that failed to rebound, experiments were conducted in which large numbers of cells were transferred into uninfected macaques. SHIV infection was not transmitted in any case. In contrast, cells from animals that experienced viral load rebounds uniformly transmitted SHIV infection.
CD8 T cells were also depleted from some macaques, as a test of whether SHIV was being actively contained by the immune system in the group that did not experience viral load rebounds. The depletion did not lead to the reappearance of SHIV viral load in these animals.
A computer modeling study indicated that the activation of natural killer (NK) cells by vesatolimod was the strongest correlate of delayed viral load rebound. The researchers suggest that this is consistent with a mechanism of action in which stimulation of innate immunity and CD4 T cells by vesatolimod causes virus-infected cells to express viral antigens, which are then recognized and bound to by PGT121, triggering killing of the infected cells by NK cells and/or other effector cells such as monocytes. This antibody-mediated recruitment of immune cells capable of delivering the coup de grâce to virus-infected cells is known as antibody-mediated cellular cytotoxicity (ADCC) or antibody-mediated cellular phagocytosis (ADCP).
In the discussion section of the paper, the researchers emphasize that the study represented an idealized circumstance due to the very early ART initiation, and that the effects of the interventions were most evident in animals with the lowest pre-ART viral loads. The implications for the more typical circumstance of chronic infection are therefore unclear.
They also note that the failure to transmit SHIV infection is not definitive evidence that all replication-competent virus has been cleared. This has recently been demonstrated by Ron Desrosiers in a study in which replication-competent SHIV was eventually detected in a macaque that had failed to transmit the virus in several cell transfer experiments.
Additionally, there is limited experience with the SHIV-SF162P3 challenge virus in the context of cure research and prior studies have not documented an ability of this virus to develop resistance to PGT121. More information on HIV resistance to PGT121 should emerge from ongoing clinical trials.
Despite these caveats, the evidence that the dual combination promoted clearance—or at least significant depletion—of the viral reservoir in some cases is encouraging.
Gilead Sciences ultimately aims to study vesatolimod combined with a derivative of PGT121 dubbed GS-9722 in people with HIV. GS-9722 is a version of PGT121 that has been modified in an effort to further enhance ADCC/ADCP activity, and it is currently being tested in a phase I trial (the trial is listed on the subscription-only website Adis Insight so details are scarce; there is no clinicaltrials.gov entry yet).
October 25, 2018 in Broadly neutralizing antibodies, Cure research, HIV remission, Immune-based therapies, Innate immunity, Natural killer cells | Permalink | Comments (0)
Post-AIDS 2018 Updates on HIV Cure Research
In the aftermath of the 22nd International AIDS Conference (AIDS 2018), which took place in Amsterdam in July, there has been some reflecting on the challenges facing the HIV cure research field. The presentations that garnered the most news coverage described disappointing study results, but there were also nuggets of novelty and encouragement to be found amidst the sea of data on offer (see previous post for links to relevant conference sessions, many of which now have video and/or slides available).
The RIVER Trial
The most widely reported findings came from a clinical trial in the UK known as RIVER (Research In Viral Eradication of HIV Reservoirs). Participants with primary HIV infection were randomized to receive a standard antiretroviral therapy (ART) combination plus the integrase inhibitor raltegravir or ART plus raltegravir along with a therapeutic HIV vaccine regimen and a short course of the HDAC inhibitor vorinostat (a candidate HIV latency-reversing agent). The primary purpose was to evaluate whether the vorinostat and vaccine combination—a version of the proposed "kick & kill" approach to depleting the HIV reservoir—had a significant effect on HIV reservoir measures compared to ART.
As described in detail in multiple online reports (e.g. see i-Base, AIDSMap, and the Imperial College website), the size of the HIV reservoir—as assessed by both total HIV DNA and the viral outgrowth assay—remained equivalent between the two arms. Presenter Sarah Fidler pointed out some possible caveats, such as the relatively short follow up time, but the data appear to rule out any significant effect by this particular kick & kill combination.
The researchers cited several potential explanations: the latency-reversing activity of vorinostat may be suboptimal, and there is uncertainty as to whether the HIV-specific T cell responses that were successfully induced by the vaccines were targeting parts of the virus most likely to be displayed by infected cells after latency reversal. As is emphasized in much of the coverage, the study itself should not be considered a failure because it provided a clear answer to the question it was designed to address. The interventions were also found to be safe and no participants withdrew.
The results echo a theme that has been sounding recently in HIV cure research: randomized controlled trials are required to rigorously evaluate the potential of candidate interventions, and positive results from single-arm exploratory trials (which tend to compare results to baseline values or historical controls) need to be interpreted with caution.
Studies published over the past year that have reached a similar conclusion include randomized controlled evaluations of a therapeutic vaccine combination and single doses of the candidate latency-reversing agent romidepsin.
Vedolizumab
The second piece of unwelcome news related to vedolizumab, an antibody that targets the α₄β₇ integrin, a protein involved in CD4 T cell trafficking to the gut that may also facilitate HIV entry into target cells. In 2016, an experiment in SIV-infected macaques generated excitement when administration of a version of vedolizumab adapted for macaques (see comment by Robert Reinhard below) was associated with control of viral load after ART interruption. Human trials were initiated relatively quickly because vedolizumab is already FDA approved as treatment for ulcerative colitis and Crohn's disease.
During a talk at the conference on HIV remission, Anthony Fauci from the National Institute of Allergy and Infectious Diseases (NIAID) provided a glimpse at the data from the first of these trials, and sadly the results did not mirror the published macaque study. While it appeared that one or two participants displayed some evidence of viral load containment after an analytical treatment interruption (ATI), the majority did not.
Fauci suggested the variations in viral load levels were similar to those his group has observed in placebo recipients in prior trials, and were not indicative of any effect from vedolizumab. At least two other clinical trials involving the antibody are ongoing, so additional results will be forthcoming.
In a separate presentation, Michele DiMascio put another dent in the optimism that had surrounded vedolizumab by reporting that an attempt to repeat the original results obtained in SIV-infected macaques had failed. This time, there was no evidence of antibody-induced SIV control. The reasons for the divergent outcomes are unclear, but may relate to the type of SIV used in the experiments, which has a mutation in the nef gene (update 3/21/19: the journal Science has now published an "editorial expression of concern" about the original study).
Reservoir Targeting
A number of presentations described novel approaches for targeting the HIV reservoir, highlighting the amount of work that is underway to try and improve upon the interventions tested to date.
Isa Munoz-Arias and colleagues from UCSF and Merck reported that a number of FDA-approved chemotherapeutic drugs have HIV latency-reversing activity in laboratory studies. In some cases the magnitude of the effect was demonstrated to be greater than the combination of bryostatin and romidepsin, which has previously been shown to be among the most potent latency-reversing strategies in vitro. The effects were not associated with significant T cell activation or CD4 T cell death (although it's important to note that this does not mean the drugs are without side effects - those are described on their labels).
A total of 12 FDA-approved chemotherapies were found to reverse HIV latency via a variety of novel pathways, suggesting new avenues for exploration beyond the current candidates, which primarily comprise HDAC inhibitors. The details of the presentation can be found on Jules Levin's NATAP website (which often does more to make information public than conferences themselves), and it was also covered by Simon Collins for i-Base.
The research group of Wen Kang debuted data from a small, uncontrolled pilot study of the HDAC inhibitor chidamide, which is approved in China as a cancer therapy. Kang described evidence that the drug had stimulated production of HIV RNA in seven individuals on ART, and may have slightly reduced HIV DNA levels. However in the Q&A after the talk Sharon Lewin noted that the various possible markers of activity that were analyzed did not appear to necessarily correlate with each other. A larger randomized controlled trial that should provide more definitive results is now ongoing.
Tim Henrich from UCSF followed up on work published earlier this year in PLoS Pathogens identifying CD30—a cell surface molecule known for its association with lymphoma—as preferentially expressed on HIV-infected CD4 T cells.
In the paper, the researchers describe an individual with HIV who exhibited undetectable viral RNA and DNA levels after receiving therapy for lymphoma including brentuximab vedotin, an anti-CD30 antibody-drug conjugate. Unfortunately the individual died after cancer recurrence so no further investigation was possible.
Henrich's conference abstract reports the identification of a second person with HIV who received brentuximab vedotin as part of treatment for lymphoma (now in remission). Three weeks after administration of the first dose, HIV RNA was reduced to undetectable from a previous level of 7,359 copies per million CD4 T cells (a greater than 3 log reduction). HIV DNA levels fell by 42%. The individual is now being followed longitudinally.
This research opens up the possibility of targeting CD30 as a means to deplete the HIV reservoir, and in addition to brentuximab vedotin there are also CD30-specific chimeric antigen receptor (CAR) T cells in development that are already being studied in clinical trials for cancer.
An interesting presentation by Sarah Joseph from the University of North Carolina at Chapel Hill outlined an effort to establish when viruses enter the latent reservoir. Using complex phylogenetic analyses (see the abstract for an example), Joseph uncovered evidence that in most individuals studied, the majority (~72%) of the replication-competent latent HIV reservoir was most closely related to viruses circulating in the year prior to ART initiation. In contrast, only 5% was derived from virus replicating during the first year of infection.
A paper published in the journal eLife in 2016, involving ten participants, reported similar results, although Joseph also pointed out that a much smaller study presented at the conference by Zabrina Brumme (subsequently published in PNAS) reached different conclusions, finding more diverse dates of establishment of the reservoir.
Joseph's data suggests that latently infected cells tend to be shorter-lived during untreated infection, with ART initiation triggering the formation of the bulk of the long-lived HIV reservoir. The implication for therapeutic strategies targeting the reservoir is that there might be a window of opportunity to intervene at the time ART is started (currently, almost all clinical trials involve testing candidate therapies in individuals after ART has suppressed viral load). This idea could potentially be explored in the SIV/macaque model.
HIV Control Off ART
Participants in the VISCONTI cohort represent the best known and most widely cited examples of post-treatment control of HIV replication. In 2013, when Asier Sáez-Cirión and colleagues published a detailed report in PLoS Pathogens, the cohort comprised 14 individuals. Updates have been fragmented since that time, occurring at various conferences, and as yet there have been no follow up publications as thorough as the original paper (at least that I'm aware of - for the sake of disclosure I should note that I chided the investigators about this situation in a public comment to the PLoS Pathogens article in 2017).
At AIDS 2018, Laurent Hocqueloux and colleagues presented a poster on factors associated with loss of post-treatment controller status that included an update on the VISCONTI cohort. The study also offered results from analyses of inflammatory biomarkers, which I've not seen described previously.
A total of 24 post-treatment controllers have now been added to the VISCONTI cohort; all started ART during primary infection and then interrupted after a median of 3.5 years of treatment. During subsequent follow up (current median of 12 years), five (21%) have restarted ART, four due to increasing viral load and one as a result of a head and neck cancer diagnosis.
These five participants were among seven who experienced one or more viral load measurements above 400 copies/ml during monitoring; none of the 17 who remained below this viral load level restarted ART (a highly statistically significant difference). The fact that the individual who developed cancer was in the former group, despite not reinitiating ART based on viral load criteria, may raise concern that prior exposure to detectable HIV viremia was a risk factor for cancer development. Whether remaining on ART would have been associated with lower risk is an unanswerable question. This conundrum illustrates that firm conclusions about the clinical benefits of post-treatment control compared to continuous ART cannot be drawn until post-treatment control can be induced in sufficient numbers of people to allow a randomized comparison.
Overall, CD4 counts and CD4:CD8 ratios have remained stable in the cohort with medians not significantly different between the time of ART interruption and last follow up. Individual plots are not shown in the poster, however, so it's unclear if declines occurred in participants who experienced viral load increases.
Encouragingly, the majority of VISCONTI cohort members have maintained viral loads below 50 copies/ml, and levels of three inflammatory biomarkers—IP-10, sCD163 and sCD14—were not significantly different between these post-treatment controllers and healthy HIV-negative individuals (see figure 6 in the poster).
In a separate oral presentation, Asier Sáez-Cirión showed evidence that certain immune response genes are associated with post-treatment control in the VISCONTI cohort, and may be mediating their effects—at least partly—through superior natural killer (NK) cell activity against HIV (unfortunately neither slides or video of this presentation are available on the conference website).
Lisa Chakrabarti described an investigation of potential mechanisms of HIV control in a different population: elite controllers participating in the ANRS CODEX cohort. The researchers focused on HIV Gag-specific CD4 T cell responses, and found that CCR5 expression was lower among elite controllers compared to a control group of people with HIV on ART.
The lower CCR5 expression by Gag-specific CD4 T cells was associated with reduced susceptibility to HIV infection, suggesting this may contribute to elite controller status. The results may offer support to efforts to protect virus-specific CD4 T cells from HIV infection using gene therapies that ablate CCR5 expression (or otherwise attempt to protect the cells from HIV entry), an approach being pursued by researchers at the defeatHIV collaboratory.
Analytical Treatment Interruptions (ATIs)
A controversial component of research working toward achieving post-treatment control of HIV is the use of ATIs. A systematic review of past studies involving ART interruptions could help shed light on the safest approaches to conducting ATIs in clinical trials, and Jillian Lau and colleagues from Alfred Hospital and Monash University in Melbourne, Australia delivered just such a review as a poster presentation at the conference. Their work is now in press at a journal and will hopefully be published soon.
Online Post-Conference Coverage
As is always the case, an array of excellent reporting has become available online since the conference. See below for a selection; please leave a comment if you know of coverage I've missed that should be included.
AIDS 2018 HIV Cure Research Highlights - Karine Dubé and Jeff Berry, Positively Aware
The Road Ahead for HIV Cure Research - Benjamin Ryan, POZ Magazine, September 6, 2018
Cure Updates from AIDS 2018 - hivcure.com.au/Doherty Institute
i-Base
NATAP
September 19, 2018 in Analytical treatment interruptions, Conferences & meetings, Cure research, Elite controllers, HIV remission, Inflammation, Natural killer cells, Post-treatment controllers | Permalink | Comments (1)
Elite Controllers: Sex Differences and Factors Associated with Loss of Immune Control
Over the past few months, several interesting papers addressing elite control of HIV infection have seen publication. The ability of elite controllers to maintain undetectable viral loads and relatively preserved CD4 T cell counts in the absence of ART has led them to be proposed as a model for a functional cure of HIV infection. But there is also evidence that many elite controllers exhibit elevated levels of inflammation compared to HIV-negative counterparts, and eventually experience disease progression, leading some researchers to call this proposition into question. The uncertain relevance of elite control to HIV cure research is prompting studies that attempt to parse the factors distinguishing individuals who preserve elite controller status from those who ultimately progress.
In the journal eBioMedcine, Wang Zhang and colleagues explore the expression of a variety of genes and proteins in a cohort of 19 elite controllers, compared to 32 individuals with progressive HIV infection and 23 healthy HIV-negative controls. Of particular note, nearly half the elite controllers (9) were women, allowing the researchers to compare results based on sex. The authors point out that several previous studies have reported that women tend to be overrepresented among elite controllers (e.g. see Crowell et al and de Azevedo et al).
A potentially important finding is that the gene expression profiles of female elite controllers were similar to HIV-negative females, but there were significant differences between male elite controllers and HIV-negative male controls. A potential implication is that women will be more likely to maintain elite controller status over time compared to men—this possibility will need to be investigated further.
Among the differences observed in elite controllers compared to individuals with progressive HIV infection were downregulation of the genes for CXCR6 and SIGLEC1. The authors explain that these differences could be associated with decreased susceptibility of CD4 T cells for HIV entry and reduced cell-to-cell virus transmission mediated by myeloid cells.
Levels of the chemokine CCL4 (MIP-1β) were found to be higher among elite controllers versus progressors, consistent with a previous study. Conversely, levels of the inhibitory immune cell receptor PD-1 (and its ligand PD1-L2) were significantly lower, echoing another recent paper reporting lower levels of multiple inhibitory receptors in elite controllers.
BMC Medicine has published an analysis by José Benito and colleagues that looks at possible contributors to loss of elite control. The researchers evaluated 36 elite controllers followed for approximately a decade on average, comparing multiple immunological parameters between those with stable CD4 T cell counts (n=22) and those exhibiting significant CD4 T cell decline during the follow up period (n=14). Interestingly, women made up 67% of the former group but only 31% of the latter, a statistically significant difference (p=0.04). The authors don't offer any comment on this apparent overrepresentation of women in the group of elite controllers with stable CD4 T counts.
Many immunological variables were found to differ between elite controllers and HIV-negative healthy controls, as well as between stable elite controllers and those experiencing CD4 T cell declines. Distinctions between elite controllers and HIV-negative healthy controls included lower levels of several T cell subsets: naïve, recent thymic emigrant, stem cell memory, and regulatory. Expression of the T cell costimulatory receptor CD28 was lower, whereas CD95—involved in apoptotic cell death—was increased.
Comparisons between the stable elite controllers and progressors revealed that the latter group had lower levels of naïve and recent thymic emigrant CD8 T cells, as well as higher levels of CD8 T cells with effector memory and senescent phenotypes. The finding suggests that CD8 T cells may have been differentiating at a higher rate in this group—in other words, naïve CD8 T cells were more frequently becoming activated and transitioning into memory cells (perhaps reflective of a more strenuous battle to keep HIV contained). The progressor group also showed increased expression levels of PD-1 in both total CD4 T cells and the central memory CD4 T cell subset.
In a separate study published in the Journal of Virology earlier this month, María Pernas and colleagues conducted a retrospective, longitudinal analysis of factors contributing to loss of viral load suppression in a cohort of elite controllers. A total of 31 elite controllers were included, defined based on having viral load below detectable levels (50 copies/ml) on three consecutive measures over a year of follow up.
Fourteen of these individuals subsequently experienced increases in viral load (two consecutive measures above the detection limit within a year), and were classified as transient controllers. The remaining 17 maintained undetectable viral loads and were classified as persistent controllers. Sex differences were not apparent in this study, with women making up 43% of the former group and 41% of the latter. The only significant differences were for time since diagnosis (averaging 8 years and 18 years, respectively) and sexual transmission as mode of HIV acquisition (71% vs. 35%).
The availability of samples prior to the viral load increases in the transient controller group allowed several factors associated with the loss of HIV suppression to be identified. CD8 T cells targeting the HIV Gag protein were found to be significantly less polyfunctional (assessed based on production of the cytokines IFN-γ, TNF-α and IL-2) and displayed a more activated phenotype a year prior to viral load becoming detectable. HIV genetic diversity within the env gene was also significantly higher, with a similar trend observed for the gag gene. In contrast, persistent controllers showed little or no evidence of ongoing viral evolution.
Inflammatory biomarkers were elevated in the transient controllers. and an analysis of 70 different cytokines and chemokines in plasma samples revealed that increased levels of RANTES and Platelet Derived Growth Factor (PDGF) AA were the best predictors of subsequent loss of elite controller status. RANTES in particular showed a strong association, being an average of four-fold higher in transient controllers.
In discussing their results, the authors emphasize the importance of focusing on examples of strict, persistent control of HIV if the goal is to identify "the right model of functional remission." They also suggest that factors strongly predictive of future viral load rebound—such as RANTES levels—might have the potential to help discriminate elite controllers likely to benefit from ART from those who may not require it.
Similar points are made in a commentary in EBioMedicine by Laura Tarancon-Diez and colleagues. The commentary accompanies a paper describing an elite controller who has maintained extremely low levels of HIV RNA and HIV DNA for a decade, without fully seroconverting on the Western Blot antibody assay. Evidence of polyfunctional HIV-specific CD8 T cell responses and strong antibody-mediated cellular cytotoxicity (ADCC) by natural killer cells is reported. The individual acquired HIV from a partner who was not able to control the same CRF02_AG virus variant. Neither partner possesses known favorable HLA alleles although both are heterozygous for the CCR5Δ32 mutation.
The commentary also cites work published last year from the French CODEX cohort, which described a subset of HIV controllers in whom viral load had never been detectable using routine assays during an average of 18 years of follow up. In a comparison with controllers who had experienced viral load blips, T cell counts were reported to be stable (as opposed to progressively declining), and this was accompanied by lower T cell activation and HIV DNA measurements. This study also showed a trend toward an overrepresentation of women in the persistently undetectable group (34 out of 52, 65%) versus the viral load blips group (90 out of 178, 51%, p=0.06).
Tarancon-Diez and colleagues advocate concentrating on "this very scarce proportion of individuals that are able to persistently control the virus" in order to inform the design of cure strategies aiming to induce long-term remission in the absence of ongoing ART.
Whether there might be particular features of the immune response (or other factors) in women that increase the likelihood of persistent elite control needs to be elucidated. Ongoing work is investigating possible differences related to HIV persistence in women receiving ART, with Eileen Scully presenting the latest findings from her research as a poster at the CROI 2018 conference. Scully has uncovered a number of significant sex-based associations, including higher expression of several antiviral genes.
As Zhang and colleagues argue in the closing section of their EBioMedicine paper: "Despite extensive data on the male and female difference on disease outcome, research does not sufficiently take gender into account. Altogether, our study showed that it would be important to carefully consider gender in cohort design for future transcriptomic and intervention studies on HIV-1 patients."
March 14, 2018 in Cure research, Elite controllers, HIV remission, HIV-specific T cell immunity, Natural killer cells | Permalink | Comments (0)
Recruiting Natural Killer Cells to Target HIV Persistence
An important goal in HIV cure research is the identification of immune responses that might be induced or enhanced to promote clearance of virus-infected cells. The main focus of this work has been on adaptive immunity—components of the immune system that can specifically recognize HIV, which include CD4 T cells, CD8 T cells, B cells and antibodies. But there is growing interest in cells considered part of the innate immune system, particularly natural killer (NK) cells. NK cells have the potential to destroy virus-infected cells by several mechanisms, including the identification of generic signs of cellular distress or infection, or via antibody-mediated recruitment to a target cell (known as antibody-dependent cellular cytotoxicity/ADCC).
Over the past few months a number of studies have been published that support the idea that NK cells can play an important role in controlling virus replication. In the journal Nature Medicine, Nicolas Huot and colleagues describe evidence that NK cells contribute to suppression of SIV replication in the lymph nodes of African Green Monkeys (AGMs), a host species that does not experience pathogenic consequences from the infection.
In experiments comparing nonpathogenic SIV infection of AGMs to pathogenic infection in macaques, NK cells were found to localize within and around lymph node B cell follicles—the major site of virus replication and persistence—in AGMs, but were scattered randomly in macaques, with no accumulation in follicles. NK cell numbers in lymph nodes also progressively declined in macaques, while being maintained at pre-infection levels in AGMs.
Additional analyses found that these differences were associated with an increased frequency of NK cells expressing CXCR5 (a receptor governing homing into follicles) and localized production of the cytokine IL-15 in the follicles of AGMs. The role of IL-15 was further confirmed by administration of an anti-IL-15 antibody, which depleted NK cells from the lymph nodes of the AGMs and led to a significant increase in SIV replication.
The authors write in their conclusion: "On the basis of our results, we anticipate that better comprehension of NK cell biology in lymphoid tissues, as provided here, will endorse the search for new NK cell–based immunotherapies for HIV infection."
A review published in the journal AIDS in November covers some of the potential NK cell–based immunotherapies that are under investigation. A panoply of clinical trials are testing broadly neutralizing antibodies (bNAbs), which may have the ability to promote NK cell-mediated ADCC. In some cases bNAbs are being evaluated in tandem with latency-reversing agents, with the aim of depleting the HIV reservoir.
Also cited is the toll-like receptor 9 (TLR9) agonist MGN1703, which researchers at the University of Aarhus have shown can promote NK cell activation in HIV-positive people on ART. The same research group has also reported that NK cell responses may have been linked to an HIV DNA decline in some participants in a trial of the latency-reversing agent panobinostat.
Other NK cell–based immunotherapies in clinical trials include ALT-803, a modified version of the cytokine IL-15. The research group of Timothy Schacker at the University of Minnesota is conducting a small study involving ALT-803 administration to HIV-positive individuals on ART. A paper published in the Journal of Virology late last year describes a transient anti-SIV effect of the compound in macaques that were not receiving ART. Schacker and colleagues have also recently launched a trial in which individuals will receive infusions of NK cells from matched donors in combination with the cytokine IL-2.
The effects of the cytokine alpha interferon on HIV persistence are a major topic of interest at the newly-funded BEAT-HIV Collaboratory led by Luis Montaner at the Wistar Institute in Philadelphia. Last month in Clinical Infectious Diseases, Stéphane Hua and colleagues presented evidence that NK cell activation associates with a decline in HIV DNA levels in HIV-positive individuals receiving alpha interferon for the treatment of hepatitis C.
In sum, after a long period in the shadow of the better-known components of the adaptive immune system, NK cells are now emerging as potentially important players in HIV cure research. Results from the ongoing trials should soon shed additional light on how they might be able to contribute.
January 18, 2018 in Broadly neutralizing antibodies, Cure research, Immune-based therapies, Innate immunity, Natural killer cells | Permalink | Comments (0)
HIV Remission News from IAS 2017
The 9th International AIDS Society Conference on HIV Science (IAS 2017) began on Sunday in Paris, and the topic of HIV remission has been the focus of several high profile presentations.
One of the first major news stories to emerge from the meeting involves a newly described example of prolonged HIV remission in a nine-year-old South African (see the conference abstract and poster discussion slides presented today by Avy Violari). The case was identified because the child was a participant in the Children with HIV Early Antiretroviral Therapy (CHER) trial, which took place from 2005-2011 in South Africa and evaluated immediate, time-limited (40- or 96-week) courses of antiretroviral therapy (ART) in perinatally infected infants compared to the strategy of deferring initiation until clinical or immunological signs of disease progression were evident (preliminary results from the study, demonstrating the benefits of immediate ART, were published in the New England Journal of Medicine in 2008).
The child was diagnosed by HIV DNA PCR 32 days after birth, and displayed pre-treatment viral load levels of over 750,000 copies/ml and 151,000 copies/ml on days 39 and 60, respectively. ART was initiated after 61 days, with viral load declining to below 50 copies/ml at week 24 of on-treatment follow up. ART was subsequently stopped at week 40. The CHER protocol mandated restarting ART if certain immunological and/or clinical criteria were met, but the child has remained healthy and maintained a high CD4 count throughout follow up.
When stored samples were later accessed in order to measure viral load, the levels were found to be persistently below 20 copies/ml. The first post-interruption sample was taken eight weeks after stopping ART, so it's not known if viral load failed to rebound or if there was a transient increase that was rapidly controlled. Viral load has now remained undetectable for 8.75 years and counting, making this one of the longest reported cases of HIV remission (there is a French teenager first reported in 2015 and described in a paper in The Lancet HIV last year who has contained viral load to undetectable levels for over 12 years, after receiving a more prolonged course of ART in early life).
The researchers have conducted multiple analyses to try and gain insight into the factors that may have contributed to the outcome. Measurement of the HIV reservoir using a sensitive test for total HIV DNA revealed similar levels after one year and at 9.5 years of age: around 5 copies per million peripheral blood mononuclear cells (PBMC). However, no replication-competent virus could be detected using two different culture methods. Virus sequencing showed that the child was infected with HIV-1 from clade C, the most prevalent strain in South Africa.
The ELISA HIV antibody test was negative but weak reactivity to Gag p40 and p24 was detectable by Western blot. One exception to the generally weak HIV-specific antibody responses was a high IgA2 (a type of mucosal antibody) response to the gp41 protein. In studies of cellular immunity, a low-level CD4 T cell response to the HIV Gag protein was discernible with a whole blood intracellular cytokine assay, but no HIV-specific CD8 T cells could be detected.
Additional parameters that were investigated included the density of CCR5 molecules on CD4 T cells, which was found to be lower than HIV-negative controls – it is perhaps possible that lower CCR5 density contributed to the maintenance of an HIV-specific CD4 T cell response by reducing the susceptibility of these cells to HIV infection. Encouragingly, levels of immune activation were similar to HIV-negative individuals and lower than those observed in elite controllers (in the latter group, elevated immune activation has been associated with disease progression despite persistently low viral loads).
PD-1 expression was found to be high on both CD4 and CD8 cells compared to HIV- individuals and elite controllers; the significance of this finding remains to be elucidated. None of the beneficial HLA alleles that have been associated with elite control were present, although the child was heterozygous at all HLA loci and this has been linked to a better prognosis compared to homozygosity.
The researchers note that other factors beyond the temporary course of ART likely contributed to the salutary outcome – the majority of participants in both the 40- and 96-week immediate ART arms of the CHER trial had met the criteria for restarting treatment by the time the study ended, and no other cases of HIV remission have yet been identified.
The hope is that additional analyses will uncover novel correlates of immune control and aid the design of interventions aiming to induce HIV remission in larger numbers of HIV-positive individuals.
The news coverage of the case—which has been extensive—appears to have generally been accurate, although the BBC erred by describing the child as "virtually cured" in their headline – it is not known if viral load might rebound at some point in the future, as has occurred in some other examples of HIV remission. There are articles describing the child as either a girl or a boy, but Amy Green from the South African Health News Service interviewed Avy Violari and reports that gender is not being disclosed in order to protect the individual's privacy.
The New York Times article, authored by Donald McNeil Jr., has some poor mistakes: the opening paragraph erroneously states that the ART was given at "high doses" when the CHER trial employed normal pediatric dosing. Later in the piece, individuals who are homozygous for the CCR5delta32 mutation, which confers a high degree of resistance to HIV infection, are mistakenly referred to as elite controllers (elite controllers are HIV-positive individuals who naturally contain HIV viral load to low levels in the absence of ART). Timothy Brown, the one individual considered cured of HIV infection, is said to have received a bone marrow transplant from an elite controller, which is not the case - his bone marrow donor was HIV-negative and homozygous for the CCR5delta32 mutation. One of these errors has since been corrected, but two remain at the time of writing.
The first mention of the South African HIV remission case at the IAS conference came yesterday in a presentation by Anthony Fauci, Director of the National Institute of Allergy and Infectious Diseases (NIAID) during the "Challenges and Opportunities in HIV Science" session – this appears to explain the timing of the NIAID news release about the study. Fauci's talk was titled: "Sustained ART-free HIV remission: opportunities and obstacles" and he offered some additional nuggets of news, as well as discussing issues to be considered when evaluating the remission cases that have been reported to date (slides are available for download, and video was posted to the IAS conference youtube channel on July 26).
In particular, he outlined two potential contributors to HIV remission: in some cases, the size of the HIV reservoir may be so small that after an ART interruption the few latently infected CD4 T cells that are present remain dormant for an extended period, before one or more cells is activated and starts producing HIV (triggering viral load rebound). Fauci referred to this phenomenon as "stochastic reactivation," and evidence suggests it may explain the period of remission documented in the Mississippi baby who was started on ART prior to developing detectable immune responses to HIV (it has also been posited to explain the adult examples of the Boston patients and an individual treated within days of infection at UCSF—see below).
In other instances—such as the South African child, the French teenager and the VISCONTI cohort participants—evidence suggests immunological control of HIV replication is contributing, although the exact mechanisms are unclear.
Fauci noted that both of these phenomena could also be operational simultaneously, but presently the short answer as to precisely what explains a given case of HIV remission is: "we don't know."
On the topic of interventions aiming to induce HIV remission, Fauci offered a glimpse of unpublished data from two studies. The first was a clinical trial conducted at the National Institutes of Health (NIH) Clinical Center that evaluated the effect of therapeutic vaccination in HIV-positive individuals started on ART within 12 weeks of a diagnosis of acute or early infection, who had maintained undetectable viral loads for over a year.
A total of 30 participants enrolled, with 15 randomized to receive a prime-boost HIV vaccine regimen developed by Profectus Biosciences (comprising DNA and vesicular stomatitis virus vectors) and 15 randomized to receive placebo immunizations. The final vaccination was administered at week 48 of the study and, eight weeks later, all participants underwent a 16-week analytical ART interruption.
The results demonstrated that there were no significant differences in time to viral load rebound to over 40 copies/ml (or to over 400 copies/ml) between the vaccine and placebo groups. Fauci emphasized the importance of the placebo group by showing that at the end of the ATI period, 20% of controls had viral loads less than 20 copies/ml, 27% were below 400 copies/ml, and 40% were below 2,000 copies/ml.
This finding may temper excitement about the single-arm study of therapeutic vaccination plus romidepsin that drew so much attention at CROI earlier this year – in that trial, the fact that 38% of participants maintained viral loads below 2,000 copies/ml at 4-6 months after an ATI was presented as evidence that the interventions had enhanced control of HIV replication, compared to historical studies of ART alone. Fauci suggested that preliminary clinical trials and animal studies using broadly neutralizing antibodies (bNAbs) offer evidence that they may have more potential for enhancing post-treatment control of viral load than therapeutic vaccines.
Fauci's own laboratory has reported that administering an antibody against the α₄β₇ integrin to SIV-infected macaques led to surprisingly robust control of SIV replication after an ART interruption, and in his presentation he showed the unpublished results of recent experiments aiming to shed light on the mechanisms involved. The approach that the researchers took was to deplete different types of immune cells in the animals controlling SIV viral load, then assess whether this led to an increase in viral replication. The experiments compared:
Antibodies to the CD8 receptor alpha chain, which deplete CD8 T cells, natural killer T cells (NKTs) and natural killer (NK) cells
Antibodies to the CD8 receptor beta chain, which deplete CD8 T cells
Antibodies to CD20, which deplete B cells
A transient rebound in viral load was only seen in recipients of antibodies to the CD8 receptor alpha chain, indicating that NKTs and NK cells are making an important contribution to the observed control of SIV replication. A clinical trial investigating the anti-HIV effects of the anti-α₄β₇ integrin antibody vedolizumab, which is FDA-approved for the treatment of ulcerative colitis and Crohn's disease, is ongoing at the NIH Clinical Center, and another is due to start soon at the Ottawa Hospital Research Institute in Canada.
Timothy Henrich from UCSF gave another presentation involving HIV remission today, in the same poster discussion session during which Avy Violari described the South African case (video of most of the session is posted on the IAS youtube channel, but Violari's talk is not included). Henrich reported details relating to an individual who was mentioned several times during CROI earlier this year (see the BSVC blog conference report). Both Henrich's abstract and slide presentation are available on the IAS conference website.
The individual in question was diagnosed within an estimated 10 days after acquiring HIV infection, because the exposure occurred in a short window between screening for a pre-exposure prophylaxis (PrEP) program and the baseline visit at which Truvada PrEP was first administered. When the baseline test results came back showing the presence of HIV, Truvada was quickly switched to a four-drug ART regimen including Truvada, darunavir and raltegravir. No HIV could be detected in blood or tissue samples during ART, although a low level of viremia was briefly detected in one out of 10 humanized mice administered a large volume of cells sampled from the individual (this novel test is known as the murine viral outgrowth assay).
After 34 months of continuous ART, an analytical treatment interruption was undertaken, and no viral load rebound occurred for 224 days, during which time no HIV DNA or RNA was detectable in plasma or sampled CD4 T cells. After this prolonged period of HIV remission, viral load rebounded and was initially detected at a level of 36 copies/ml. Six days later viral load had increased to 59,805 copies/ml and ART was immediately restarted. Sequencing of the HIV genome demonstrated that the rebounding virus was identical to that sampled at the time of acute infection.
In a collaboration with Alison Hill, mathematical modeling studies estimated the size of the HIV reservoir to have been approximately 200 latently infected cells prior to the ART interruption. Based on Hill's prior work, a latent HIV reservoir of this size confers only a small (~1%) chance of achieving a lifelong cure, due to the risk of stochastic reactivation of one or more of the latently infected cells.
Henrich and colleagues continue to evaluate enrollees in PrEP projects for evidence of recent HIV infection, and he showed a slide documenting that another individual has been started on ART very early as a result of the effort. Follow up of this second individual is ongoing.
In addition to several sessions and presentations related to HIV cure research at the main conference (see here for a roadmap), IAS hosted a pre-conference HIV Cure & Cancer Forum at the Institut Curie. The abstracts and many of the presentations have already been made available online, and a brief report from the meeting will follow in a separate blog post.
July 25, 2017 in Broadly neutralizing antibodies, Conferences & meetings, Cure research, Natural killer cells, Post-treatment controllers, Therapeutic vaccines | Permalink | Comments (0)
Targeting Latently Infected Cells via IFITM1
One of the holy grails in cure research is the discovery of markers that would allow the specific targeting of cells harboring latent HIV. The goal is to be able flag those cells for elimination while sparing their uninfected counterparts. So far, a number of cellular receptors have been identified as being more commonly—but not exclusively— expressed on latently infected cells, such as PD-1, LAG-3 and TIGIT. In the new issue of the open access journal JCI Insight, Rui André Saraiva Raposo and colleagues from the laboratory of Doug Nixon at The George Washington University add a new candidate to the mix: interferon-induced transmembrane protein 1 (IFITM1).
IFITM1 belongs to a family of proteins that respond to interferons and are involved in a variety of cellular processes, particularly immunological signaling. IFITM1 has also been implicated as an innate HIV restriction factor. Although not a household name, IFITM1 has attained sufficient celebrity to have its own Wikipedia entry.
Raposo and colleagues hypothesized that expression of IFITM1 might be altered by the presence of HIV in latently infected cells. Studying an in vitro model of HIV latency, they found that IFITM1 expression was increased fourfold in resting latent cells when compared with reactivated cells. Furthermore, significant killing of resting latently infected cells could be achieved with a combination of an anti-IFITM1 antibody and natural killer cells (via the mechanism of antibody dependent cellular cytotoxicity or ADCC).
A preliminary analysis of CD4 T cells sampled from HIV+ individuals on ART revealed a significant increase in IFITM1 expression compared to HIV- donors (2% vs. 0.9%), particularly among central memory CD4 T cells, the subset in which most latent HIV resides. This finding makes an inferential case that IFITM1 expression may be elevated in latently infected CD4 T cells from people on ART, but latently infected cells were not specifically identified and analyzed (their rarity makes such analyses challenging). As observed in the in vitro model, IFITM1-expressing CD4 T cells sampled from HIV+ individuals were sensitive to ADCC in the presence of an anti-IFITM1 antibody and natural killer cells.
The researchers note that the approach could be imperfect and potentially lead to the death of uninfected CD4 T cells, but argue it deserves to be explored:
"We acknowledge that residual IFITM1 expression in non-latently infected cells may lead to the death of bystander cells in the event this strategy translates into an eradication approach in humans. Nonetheless, we believe that an eradication strategy involving IFITM1 in conjugation with immune checkpoint molecules shown to contribute to HIV persistence during ART would be of significant value to effectively target and eliminate latently infected cells in vivo."
January 23, 2017 in Cure research, Immune-based therapies, Innate immunity, Natural killer cells | Permalink | Comments (0)
Innate Immunity and HIV DNA Declines in Panobinostat Recipients
Results from a phase I trial of the candidate latency reversing agent panobinostat were published last year in The Lancet HIV (see blog post from November 2014). The paper notes that in an exploratory analysis, a subset of four (out of a total of 15) participants experienced a significant decline in cell-associated HIV DNA levels of around 70-80%, although there was no detectable change in the trial cohort overall. Furthermore, the HIV DNA reduction in these four individuals was associated with a slightly longer time to viral load rebound during an analytical ART interruption. At the NIAID-sponsored Strategies for an HIV Cure meeting in October 2014, Matthias Lichterfeld reported that several measures of innate immunity correlated with the observed diminution of HIV DNA, and these analyses have now been published online in the Journal of Virology.
The most prominent finding is an association between natural killer (NK) cells expressing markers of enhanced functionality (including CD57, associated with cytotoxic activity) and reductions in HIV DNA during the trial. The researchers suggest the effect might be due to reversal of HIV latency causing up-regulatiion of receptors that activate NK cells on infected CD4 T cells and/or causing down-regulation of HLA class I receptors on these cells (both phenomena would promote NK cell-mediated killing of the CD4 T cells). Complementary correlations were seen with additional markers of innate immunity, including expression of interferon-stimulated genes and plasmacytoid dendritic cell frequencies. The IL28B "CC" genotype, which has been linked to superior innate immune function against hepatitis C, was also associated with a greater reduction of HIV DNA during panobinostat treatment.
HIV-specific CD8 T cell responses, which have been suggested to be important for mediating clearance of the HIV reservoir after latency reversal, did not correlate with HIV DNA levels. The researchers note that this could be due to the HIV-specific CD8 T cell dysfunction that is known to occur in chronic infection and the presence of escape mutations in the latent HIV reservoir that abrogate CD8 T cell recognition. No evidence of an inhibitory effect of panobinostat on HIV-specific CD8 T cells (a possibility raised by a prior laboratory study) was found. Interestingly, a larger HIV-specific CD4 T cell response at baseline (but not at any other timepoint) was associated with a greater HIV DNA decline during the trial; the explanation for this finding is not known but will be probed in future work.
The discussion section of the paper emphasizes that the research involves small numbers and exploratory subset analyses, so the results must be interpreted very cautiously. But the idea that promoting innate immunity may aid in the clearance of the HIV reservoir after latency reversal is encouraging, and will be explored further in ongoing and new clinical trials. For example, toll-like receptor (TLR) agonists may have the capacity to stimulate innate immunity, and several are now being tested in HIV-positive people on ART to assess any effects on the latent reservoir (see TAG's cure-related clinical trials page under latency-reversing agents).
J Virol. 2015 Jul 29. pii: JVI.01484-15. [Epub ahead of print]
Innate immune activity correlates with CD4 T cell-associated HIV-1 DNA decline during latency-reversing treatment with panobinostat.
Olesen R, Vigano S, Rasmussen T, Søgaard OS, Ouyang Z, Buzon M, Bashirova A, Carrington M, Palmer S, Brinkmann CR, Yu XG, Østergaard L, Tolstrup M, Lichterfeld M.
Pharmaceutical reactivation of dormant HIV-1 proviruses by histone deacetylase inhibitors (HDACi) represents a possible strategy to reduce the reservoir of HIV-1 infected cells in individuals treated with suppressive antiretroviral combination therapy (cART). However, effects of such latency-reversing agents on the viral reservoir size are likely to be influenced by host immune responses. Here, we analyzed immune factors associated with changes in proviral HIV-1 DNA levels during treatment with the potent HDACi panobinostat in a human clinical trial involving 15 cART-treated HIV-1 patients. We observed that the magnitude, breadth, and cytokine-secretion profile of HIV-1-specific CD8 T cell responses were unrelated to HIV-1 DNA changes in CD4 T cells during panobinostat treatment. In contrast, proportions of CD3- CD56+ total NK cells and CD16+ CD56dim NK cells were inversely correlated with HIV-1 DNA throughout the study, and changes of HIV-1 DNA during panobinostat treatment were negatively associated with corresponding changes in CD69+ NK cells. Decreasing levels of HIV-1 DNA during latency-reversing treatment were also related to proportions of plasmacytoid dendritic cells, to distinct gene expression patterns of interferon-stimulated genes and to expression of the IL28B "CC" genotype. Together, these data suggest that innate immune activity can critically modulate effects of latency-reversing agents on the viral reservoir and may represent a target for future immunotherapeutic interventions in HIV-1 eradication studies.
IMPORTANCE: Currently available antiretroviral drugs are highly effective in suppressing HIV-1 replication, but the virus persists despite treatment in a latent form that does not actively express HIV-1 gene products. One approach to eliminate these cells, colloquially termed as the "shock and kill" strategy, focuses on the use of latency-reversing agents that induce active viral gene expression in latently infected cells, followed by immune-mediated killing. Panobinostat, a histone deacetylase inhibitor, demonstrated potent activities in reversing HIV-1 latency in a recent pilot clinical trial, and reduce HIV-1 DNA in a subset of patients. Interestingly, we found that innate immune factors, such as natural killer cells, plasmacytoid dendritic cells and expression patterns of Interferon-stimulated genes, were most closely linked to a decline of HIV-1 DNA during treatment with panobinostat. These data suggest that innate immune activity may play an important role in reducing the residual reservoir of HIV-1 infected cells.
August 04, 2015 in Cure research, HDAC inhibitors, HIV-specific T cell immunity, Innate immunity, Latency reversing agents, Natural killer cells | Permalink | Comments (0)
Souped-Up IL-15 Sparks Natural Killer Cell Activity Against HIV
The cytokine interleukin-15 (IL-15) can exert a range of effects on the immune system, but is particularly important for promoting the proliferation and function of natural killer (NK) cells and CD8 T cells. Some observational studies in HIV have suggested salutary effects of higher IL-15 levels, in one case finding an association with delayed viral load rebound after ART interruption. But attempts at therapeutic administration of the cytokine to SIV-infected macaques have generally produced deleterious effects, including CD4 T cell depletion. More recently, IL-15 has been reexamined in the context of cure research, with laboratory studies demonstrating it can induce latently infected CD4 T cells to produce HIV. There is also increasing interest in therapeutic manipulation of NK cell responses as a means to eliminate the HIV reservoir, a task for which IL-15 may be well suited. A paper published last week in the Journal of Virology reports that an optimized form of IL-15 promoted potent inhibition of HIV by NK cells in humanized mice.
The version of IL-15 used in the study is described as a superagonist, comprising the cytokine bound to part of the receptor with which it interacts. This modified IL-15 is being developed as a cancer therapeutic under the name ALT-803 and, in mice, it has been shown to have superior biological activity (~25-fold greater) and a much longer half-life (~25 hours vs. ~40 minutes) compared to the unaltered cytokine. The new Journal of Virology paper reports that in a humanized mouse model, administration of ALT-803 either one or three days after HIV challenge reduced subsequent virus levels by over 90% (measured up to 21 days later). However, there was no significant effect if the delay in ALT-803 administration was extended to five days after challenge. The researchers used cell transfer experiments to confirm that NK cells, and not CD8 T cells, mediated the inhibition of acute HIV infection in this model. Discussing the results, they state: "to our knowledge, this study is the first reported demonstration that in vivo activation of NK cells can inhibit acute HIV-1 infection." They also note that the poor outcomes seen previously in the SIV model could be due to unmodified IL-15 producing less antiviral activity and more unwanted effects (e.g. CD4 T cell activation), or might reflect differential responsiveness in macaque compared to human cells.
A dose-escalation pilot trial of ALT-803 in HIV-positive individuals on ART is due to start soon at the University of Minnesota. The primary aim is to evaluate safety and tolerability, but the study will also measure HIV reservoir levels as a secondary endpoint. Due to evidence from animal studies that ALT-803 may be able to invoke NK cell activity against tumors, several phase I trials are already ongoing in people with cancers.
J Virol. 2015 Apr 1. pii: JVI.00563-15. [Epub ahead of print]
In Vivo Activation of Human NK cells by Treatment with an IL-15 Superagonist Potently Inhibits Acute In Vivo HIV-1 Infection in Humanized Mice.
Seay K, Church C, Zheng JH, Deneroff K, Ochsenbauer C, Kappes JC, Liu B, Jeng EK, Wong HC, Goldstein H.
Natural killer (NK) cells with anti-HIV-1 activity may inhibit HIV-1 replication and dissemination during acute HIV-1 infection. We hypothesized that the NK cell capacity to suppress acute in vivo HIV-1 infection would be augmented by activating them via treatment with an IL-15 superagonist, IL-15 bound to soluble IL-15Rα, an approach that potentiates human NK cell-mediated killing of tumor cells. In vitro stimulation of human NK cells with a recombinant IL-15 superagonist significantly induced their expression of cytotoxic effector molecules granzyme B and perforin, their degranulation upon exposure to K562 cells as indicated by cell-surface expression of CD107a and their capacity to lyse K562 cells and HIV-1-infected T cells. The impact of IL-15 superagonist-induced activation of human NK cells on acute in vivo HIV-1 infection was investigated using hu-spl-PBMC-NSG mice, NOD-SCID-IL2rγ-/- mice intrasplenically injected with human PBMCs, which develop productive in vivo infection after intrasplenic inoculation with HIV-1. IL-15 superagonist treatment potently inhibited acute HIV-1 infection in hu-spl-PBMC-NSG mice even when delayed until three days after intrasplenic HIV-1 inoculation. Removal of NK cells from the human PBMCs prior to intrasplenic injection into NSG mice completely abrogated IL-15 superagonist-mediated suppression of in vivo HIV-1 infection. Thus, the in vivo activation of NK cells, integral mediators of the innate immune response, by treatment with an IL-15 superagonist increases their anti-HIV activity and enables them to potently suppress acute in vivo HIV-1 infection. These results indicate that in vivo activation of NK cells may represent a new immunotherapeutic approach to suppress acute HIV-1 infection.
April 09, 2015 in Cure research, Immune-based therapies, Innate immunity, Latency reversing agents, Natural killer cells | Permalink | Comments (0)


Every industry has its issues, which may be a significant sampling block for success and achievement of any goals. With the advanced technology of today, it will be interesting to see how designers cope with the pace. Technology keeps on changing which require a change in approach from designers from security to interfaces, which appeal to a specific audience. Web design London offer best practices for solving most of the expected and unexpected challenges to designers. Flexible and versatility are the primary drivers of today's designers who want to remain useful and relevant in the market. However, with almost every organization and even individuals today owning a website, challenges have become a regular business because many of those who hold such sites are not tech savvy and lack the necessary skills to utilize on this phenomenal opportunity.
· Security is one the major concerns for websites today. Designers and users are all at the risk of hackers and malware. To encrypt a site in a way that will keep it free from such vices has been for a long time proved a challenge.
· Ensuring consistency of experience that users have with sites is a significant concern for designers' today. In pursuit of better skills. Designers are required to keep reviewing their sites regularly and this will mean spending enormous time on the site.
· Another challenge facing designers is being able to create a responsive site, which can be supported by multiple gadgets. Everyday electronic devise manufactures are improving version of applications on their devices, and for a designer, this requires constant review of the site to ensure compatibility with such new changes.
· Designers of today face a challenge in keeping the ever-changing customer of today coming back to the site often. With many other places to choose from, retaining customers in a website is one of the significant issues of concern, which designers have to address.
Survival in this industry depends on how best a designer can address these challenges and emerging issues. However, web design London gives lifeline on most problems facing the designer of today.


text:
France and Spain turned away Nave Andromeda oil tanker carrying stowaways
Will Humphries, Southwest Correspondent | Isambard Wilkinson, Madrid | Lucy Fisher
Tuesday October 27 2020, 12.00pm, The Times
The Nave Andromeda was stormed by British special forces on Sunday after a mayday call from the crew
HENRY NICHOLLS/REUTERS
Spanish and French authorities refused to take stowaways from a Liberian oil tanker just days before it was involved in a suspected hijacking in the English Channel, The Times has learnt.
The Nave Andromeda, which was stormed by British special forces on Sunday night after issuing a mayday call, may have tried to set down the stowaways at a French port in the days leading up to the incident, but the request was rejected, according to the shipping journal Lloyd's List.
Attempts by the ship's Greek captain to disembark the stowaways in the Canary Islands on October 15 and in Saint Nazaire on October 20 were both refused by the Spanish and French authorities.
The Liberian-flagged ship then travelled towards Southampton but when

redpajama_set_name: RedPajamaCommonCrawl


text:
Pennyroyal
Pennyroyal refers to two plants in the mint family, Lamiaceae
The mints, taxonomically known as Lamiaceae or Labiatae, are a family of flowering plants. They have traditionally been considered closely related to Verbenaceae, but in the 1990s, phylogenetic studies suggested that many genera classified in Verbenaceae belong instead in Lamiaceae...
. For the American species, see American pennyroyal. The European pennyroyal, Mentha pulegium, (also called Squaw Mint, Mosquito Plant, and Pudding Grass), is a plant
in the mint
Mentha is a genus of flowering plants in the family Lamiaceae . The species are not clearly distinct and estimates of the number of species varies from 13 to 18. Hybridization between some of the species occurs naturally...
In biology, a genus is a low-level taxonomic rank used in the biological classification of living and fossil organisms, which is an example of definition by genus and differentia...
, within the family Lamiaceae
. Crushed Pennyroyal leaves exhibit a very strong fragrance similar to spearmint
Mentha spicata syn. M. cordifolia is a species of mint native to much of Europe and southwest Asia, though its exact natural range is uncertain due to extensive early cultivation. It grows in wet soils...
. Pennyroyal is a traditional culinary herb, folk remedy, and abortifacient
Abortifacient
An abortifacient is a substance that induces abortion. Abortifacients for animals that have mated undesirably are known as mismating shots....
. The essential oil of pennyroyal is used in aromatherapy
Aromatherapy is a form of alternative medicine that uses volatile plant materials, known as essential oils, and other aromatic compounds for the purpose of altering a person's mind, mood, cognitive function or health....
, and is also high in pulegone
Pulegone
Pulegone is a naturally occurring organic compound obtained from the essential oils of a variety of plants such as Nepeta cataria , Mentha piperita, and pennyroyal. It is classified as a monoterpene....
, a highly toxic volatile organic compound
Volatile organic compound
Volatile organic compounds are organic chemicals that have a high vapor pressure at ordinary, room-temperature conditions. Their high vapor pressure results from a low boiling point, which causes large numbers of molecules to evaporate or sublimate from the liquid or solid form of the compound and...
affecting liver and uterine function.
Culinary and medicinal uses
Pennyroyal was commonly used as a cooking herb by the Greeks and Romans. The ancient Greeks often flavored their wine with pennyroyal. A large number of the recipes in the Roman cookbook of Apicius
Apicius is the title of a collection of Roman cookery recipes, usually thought to have been compiled in the late 4th or early 5th century AD and written in a language that is in many ways closer to Vulgar than to Classical Latin....
call for the use of pennyroyal, often along with such herbs as lovage
Lovage is a tall perennial plant, the sole species in the genus Levisticum, in the family Apiaceae, subfamily Apioideae, tribe Apieae.-Distribution:...
, oregano
Oregano – scientifically named Origanum vulgare by Carolus Linnaeus – is a common species of Origanum, a genus of the mint family . It is native to warm-temperate western and southwestern Eurasia and the Mediterranean region.Oregano is a perennial herb, growing from 20–80 cm tall,...
and coriander
Coriander is an annual herb in the family Apiaceae. Coriander is native to southern Europe and North Africa to southwestern Asia. It is a soft, hairless plant growing to tall. The leaves are variable in shape, broadly lobed at the base of the plant, and slender and feathery higher on the...
. Although still commonly used for cooking in the Middle Ages, it gradually fell out of use as a culinary herb and is seldom used so today.
Even though pennyroyal oil is extremely poisonous, people have relied on the fresh and dried herb for centuries. Early settlers in colonial Virginia used dried pennyroyal to eradicate pests. Pennyroyal was such a popular herb that the Royal Society published an article on its use against rattlesnakes in the first volume of its Philosophical Transactions in 1665.
Pennyroyal tea
A herbal tea, tisane, or ptisan is a herbal or plant infusion and usually not made from the leaves of the tea bush . Typically, herbal tea is simply the combination of boiling water and dried fruits, flowers or herbs. Herbal tea has been imbibed for nearly as long as written history extends...
is the use of an infusion
An infusion is the outcome of steeping plants with desired chemical compounds or flavors in water or oil.-History:The first recorded use of essential oils was in the 10th or 11th century by the Persian polymath Avicenna, possibly in The Canon of Medicine.-Preparation techniques:An infusion is very...
made from the herb. The infusion is widely reputed as safe to ingest in restricted quantities. It has been traditionally employed and reportedly successful as an emmenagogue
Emmenagogue
Emmenagogues are herbs which stimulate blood flow in the pelvic area and uterus; some stimulate menstruation. Women have used plants such as mugwort, parsley and ginger to prevent or terminate early pregnancy...
(menstrual flow stimulant) or as an abortifacient
. Pennyroyal is also used to settle an upset stomach and to relieve flatulence. The fresh or dried leaves of pennyroyal have also been used when treating colds, influenza, abdominal cramps, and to induce sweating, as well as in the treatment of diseases such as smallpox and tuberculosis, and in promoting latent menstruation. Pennyroyal leaves, both fresh and dried, are especially noted for repelling insects. However, when treating infestations such as fleas, using the plant's essential oil should be avoided due to its toxicity to both humans and animals, even at extremely low levels.
Pennyroyal essential oil
An essential oil is a concentrated hydrophobic liquid containing volatile aroma compounds from plants. Essential oils are also known as volatile oils, ethereal oils or aetherolea, or simply as the "oil of" the plant from which they were extracted, such as oil of clove...
is extremely concentrated. It should never be taken internally because it is highly toxic; even in small doses, consumption of the oil can result in death.Macht, David I. (1913). "The Action of So-Called Emmenagogue Oils on the Isolated Uterus with a Report of a Case of Pennyroyal Poisoning." Journal of the American Medical Association LXI(2):105-107. The metabolite menthofuran is thought to be the major toxic agent. Complications have been reported from attempts to use the oil for self-induced abortion
Self-induced abortion
A self-induced abortion is an abortion performed by the pregnant woman herself outside the recognized medical system. Although the term can include abortions induced through legal, over-the-counter medication, it also refers to efforts to terminate a pregnancy through alternative, often more...
. For example, in 1978 an eighteen year-old pregnant woman from Denver, Colorado died within one week after consuming one ounce of concentrated Pennyroyal oil in an attempt to self-induce abortion. There are numerous studies that show the toxicity of pennyroyal oil to both humans and animals.
Since the U.S. Congress
The United States Congress is the bicameral legislature of the federal government of the United States, consisting of the Senate and the House of Representatives. The Congress meets in the United States Capitol in Washington, D.C....
passed the Dietary Supplement Health and Education Act in October 1994, all manufactured forms of pennyroyal in the United States
The United States of America is a federal constitutional republic comprising fifty states and a federal district...
have carried a warning label against its use by pregnant women. This substance is not regulated by the U.S. Food and Drug Administration.
Attributed and possible contributions to death
1897- A twenty-three year old British woman died eight days after swallowing a tablespoon of pennyroyal in order to induce menstruation.
circa 1909- The Supreme Court of Indiana convicted a Mr. Carter of prescribing and administering pennyroyal pills to a pregnant woman who died two months after her miscarriage.
August 1912- A sixteen year-old girl from Maryland consumed thirty-six pennyroyal pills to induce abortion. An autopsy revealed that the herbal abortion was only partially successful, and was later, "...completed by mechanical means..." before the girl's death.
November 1978- An eighteen year-old pregnant woman from Denver, Colorado died after ingesting one ounce of concentrated pennyroyal oil in an effort to abort her fetus. Prior to her death, the victim had reported using the dried leaves of the plant to induce menstruation with no ill effects.
July 1994- Kris Humphrey, a twenty-four year-old woman, died in California. At the time of her death, Humphrey unknowingly had an ectopic pregnancy, and drank a tea made with pennyroyal extract. According to one news report, there is disagreement over whether Humphrey died as a result of the ectopic pregnancy or from pennyroyal poisoning.
Historical and literary mentions
In the Homeric Hymn of Demeter
In Greek mythology, Demeter is the goddess of the harvest, who presided over grains, the fertility of the earth, and the seasons . Her common surnames are Sito as the giver of food or corn/grain and Thesmophoros as a mark of the civilized existence of agricultural society...
, Demeter in the guise of an old woman as she searches for the abducted Persephone
In Greek mythology, Persephone , also called Kore , is the daughter of Zeus and the harvest-goddess Demeter, and queen of the underworld; she was abducted by Hades, the god-king of the underworld....
refuses red wine but accepts a drink of barley, water and pennyroyal called kykeon
Kykeon
Kykeon was an Ancient Greek drink made mainly of water, barley and naturally occurring substances. It was used at the climax of the Eleusinian Mysteries to break a sacred fast, but it was also a favourite drink of Greek peasants.Kykeon is mentioned in Homeric texts: the Iliad describes it as...
Aristophanes , son of Philippus, of the deme Cydathenaus, was a comic playwright of ancient Athens. Eleven of his forty plays survive virtually complete...
made reference to pennyroyal as abortifacient
in Lysistrata
Lysistrata is one of eleven surviving plays written by Aristophanes. Originally performed in classical Athens in 411 BC, it is a comic account of one woman's extraordinary mission to end The Peloponnesian War...
and Peace
Peace (play)
Peace is an Athenian Old Comedy written and produced by the Greek playwright Aristophanes. It won second prize at the City Dionysia where it was staged just a few days before the ratification of the Peace of Nicias , which promised to end the ten year old Peloponnesian War...
. Mari Sandoz
Mari Sandoz
Mari Susette Sandoz was a novelist, biographer, lecturer, and teacher. She was one of Nebraska's foremost writers, and wrote extensively about pioneer life and the Plains Indians, and has been occasionally referred to as Mari S...
writes in her book, Slogum House, "She was the fifth of twelve children in the river-bottom family, with a mother who laid the cards and brewed tansy, pennyroyal and like concoctions for luckless girls who were in need."
The grunge
Grunge is a subgenre of alternative rock that emerged during the mid-1980s in the American state of Washington, particularly in the Seattle area. Inspired by hardcore punk, heavy metal, and indie rock, grunge is generally characterized by heavily distorted electric guitars, contrasting song...
band Nirvana
Nirvana (band)
Nirvana was an American rock band that was formed by singer/guitarist Kurt Cobain and bassist Krist Novoselic in Aberdeen, Washington in 1987...
recorded a song named "Pennyroyal Tea."
"Pennyroyal Tea" is a song by the American grunge band Nirvana. Featured on the band's third and final studio album, In Utero , it was initially scheduled to be released as the third single in April 1994...
In the book Jitterbug Perfume
Jitterbug Perfume
Jitterbug Perfume is Tom Robbins' fourth novel, published in 1984. The major themes of the book include the striving for immortality, the meaning behind the sense of smell, individual expression, self-reliance, sex, love, and religion. Beets and the god Pan figure prominently...
by Tom Robbins
Thomas Eugene "Tom" Robbins Thomas Eugene "Tom" Robbins Thomas Eugene "Tom" Robbins (born July 22, 1936 is an American author. His best-selling novels are serio-comic, often wildly poetic stories with a strong social and philosophical undercurrent, an irreverent bent, and scenes extrapolated from...
, pennyroyal is used by Kudra to keep from becoming pregnant.
William Carlos Williams was an American poet closely associated with modernism and Imagism. He was also a pediatrician and general practitioner of medicine, having graduated from the University of Pennsylvania...
opens his "Kora in Hell: improvisations" city lights books with
"Fools have big wombs. For the rest? - here is pennyroyal if one knows to use it. But time is only another liar so go along the wall a little further......"
In the Mortal Engines
Mortal Engines is the first of four novels in Philip Reeve's quartet of the same name, which is also known as the Hungry City Chronicles in the United States...
by Philip Reeve
Philip Reeve is a British author and illustrator. He presently lives on Dartmoor with his wife Sarah and their son Samuel.-Biography:...
, Professor Pennyroyal is a bogus historian who leads the protagonists of the story to near death.
In the Science Fiction book "The Technician" by Neal Asher, Penny Royal is the name of rogue black AI.
The musician Jesca Hoop mentions "Pennyroyal wine" in her 2008 song, Enemy.
ARKive - images and movies of the Pennyroyal (Mentha pulegium)


text:
Theory of Change in Rwanda
Home/News/Save the Children supported Literacy Boost Project handed over to the Government for sustainability.
Save the Children supported Literacy Boost Project handed over to the Government for sustainability.
By Elysee Niyigena
"I am always happy to bring my child to a school with well-trained teacher, a school that also engages children in fun learning activities", said an excited Jeannette, a mother of Teresa, 7-year-old.
"My wife was advised by the Head of our Village about the good education at this Primary School", stated another happy husband, Lambert. Him and other parents at the Kinjonjo Primary School enjoy to bring their children to school because their children like it.
Kinjonjo Primary School is one of the 103 schools that benefited from Save the Children supported literacy boost project for the last five years, and officially handed over to the government of Rwanda in December 2017. Through training, supply of age-appropriate reading materials, the project reached 101,377 Children, 1431Teachers, 130 Head Teachers/Education leaders, 43,456 Parents, 2,790 Local Leaders, and 241 Book Sector Actors.
"Thanks to Save the Children that we have now well trained teachers that support children's learning. We have well stocked reading corners in every class of primary one to three. Parents are happy to send children here and the number of students is increasing", stated Daphrosa Mukashema, the School Head Teacher.
The Government of Rwanda with support from Save the Children contributed to the creation of a conducive environment for all children to learn, and strengthen alliances with the local community to help ensure learning, protection and wellbeing of their children.
"Before Save the Children coming to Gicumbi, there was a high rate of children dropping out of school, and there was no conducive learning environment for children. We are happy that children like it here now. I am positive that this will go a long way in improving the quality of education for our children" concluded the Head teacher.
"We grateful that the project has contributed to the reduction of school dropouts. The project is phasing out today, but we are not leaving Gicumbi as an organization. We have other projects contributing to the education of children in this district and around the country", Philippe Adapoe, Country Director, Save the Children Rwanda.
"We are thankful to Save the Children for their contribution. We will work hard so that what has been achieved through Literacy Boost is sustained. We believe in long lasting and sustainable actions. Literacy improving interventions are important in advancing the vision of our country. We all want to see a Rwanda where every child can read and write in our language perfectly", noted David Nkurunziza, Director of Good Governance, Northern Province, and represented the government of Rwanda during the handover ceremony.
Save the Children International is a company limited by guarantee, registered in England and Wales with company number 3732267 and a charity registered in England and Wales with charity number 1076822. Its registered office is St Vincent House, 30 Orange Street, London, WC2H 7HH.
© 2019 Save the Children International | Legal information
Global Campaign


text:
package org.springframework.cloud.launcher.dataflow;

import org.junit.Ignore;
import org.junit.Test;
import org.junit.runner.RunWith;

import org.springframework.boot.test.context.SpringBootTest;
import org.springframework.test.context.junit4.SpringRunner;

@RunWith(SpringRunner.class)
@SpringBootTest({ "spring.cloud.bus.enabled=false", "eureka.client.enabled=false" })
public class DeployerApplicationTests {

	@Test
	public void contextLoads() {
	}

}

redpajama_set_name: RedPajamaGithub


Welcome to JazzGuitarLessons.net! I'm honoured to have you on board, ready to learn how to play jazz guitar! This is a huge topic and don't worry, you're not alone!
I believe every guitarist has the potential to learn jazz guitar playing convincingly, and I'm here to help on the journey with some of the best guitar lessons you'll find online.
This website's mission is to make the above statement a reality through free online jazz guitar lessons. In essence, the entire website has one goal: to help you with free jazz instruction. You'll get free training with lessons about chords, improvisation, guitar scales and songs. And the website may also help simply by broadening your listening horizons or changing the way your organize your practice time.
JazzGuitarLessons.net has been at home on the web since 2009, so as you can imagine I've built up hundreds of lessons, articles, videos and podcasts on everything from jazz guitar standards to which jazz guitar players (legends) to listen to. You can read about the creator of these lessons here.
If it's your first time, I know all of the information can be overwhelming, so I've created this page, with some of the very best jazz guitar resources, to get you started on how to play jazz guitar. Every journey starts with the first step!
Some of your heroes are: Wes Montgomery, Pat Metheny, George Benson, John Scofield and other great jazz guitarists.
Sounds familiar? Even if you only said yes to TWO of the above points, you've come to the right place. The lessons here will help you progress.
Your ultimate goal is to play well on jazz tunes (aka standards).
This means executing the melody, accompanying (with chords), improvising (single-note solos) and perhaps add a touch of chord melody when possible on the jazz tune you're working on.
All you have to remember is play on standards! Play jazz tunes and/or jazz blues you enjoy listening to. Learn the melody, learn the chords and learn to solo. That's it!
How to Play Jazz Guitar … in 4 "Easy" Steps!
Dedicate time to learning jazz tunes (standard songs) each day. Picks new songs often and learn the melody first, then the chords. Build yourself a solid list of 10 tunes.
First, learn basic chords to accompany on your repertoire. Then improve upon your comping bag of tricks constantly.
Improvise with basic tools at first, then keep improving your jazz guitar soloing as your tastes, ears and skills evolve. You'll have to solo lots and lots to finetune your ideas and discover your true musical voice.
As an intermediate or advanced player, look into chord melody guitar lessons to create better, more refined performances on standards.
Then rinse and repeat! That's how all great jazzers learn how to play jazz guitar and improve your jazz guitar playing. Simple, Uh?! Seems almost too simple in a way.
To get you started, do yourself a favor and check out the most frequently visited and shared free lessons and learn jazz guitar on JazzGuitarLessons.net below. You'll find plenty of ideas on exactly what to do with the four steps above. You can also use the right sidebar to navigate the most frequently visited lessons.
You can also find JazzGuitarLessons.net on Facebook, Twitter and YouTube here.


The biennial CRA Conference at Snowbird is the flagship invitation-only conference for the leadership of the North American computing research community. The upcoming conference will be held July 16-18, 2018 in Snowbird, Utah. The New Chairs Workshop will take place on July 16, 2018.
The upcoming conference dates are revised somewhat from past schedules with the intent of making the conference more family friendly by avoiding weekends. Because of the long lead time for reserving space at Cliff Lodge, this is being implemented gradually.
To view the agenda and presentations from the 2016 conference visit: https://cra.org/events/snowbird-2016/#agenda. Names and sessions highlighted in blue have an associated presentation.


text:
#ifndef _IAS_SM_Mon_ProcessLockFile_H_
#define _IAS_SM_Mon_ProcessLockFile_H_

#include <commonlib/commonlib.h>
#include <commonlib/sys/fs/FileLock.h>
#include "sm/cfg/dataobjects.h"

namespace IAS {
namespace SM {
namespace Mon {

/*************************************************************************/
/** The ProcessLockFile class.
 *
 */
class ProcessLockFile : public ::IAS::SYS::FS::FileLock {
public:

	enum State{
		PS_ENDED,
		PS_STARTED
	};

	virtual ~ProcessLockFile() throw();

	void setProcessPidAndState(unsigned int iPid,State iState);
	void getProcessPidAndState(unsigned int& iPid,State& iState);

protected:
	ProcessLockFile(const String& strLockDir,
					const ::org::invenireaude::sm::cfg::Service* pService,
			   	    int iInstanceIdx);

	String getFileName(const ::org::invenireaude::sm::cfg::Service* pService,
			   	      int iInstanceIdx);

	friend class ::IAS::Factory<ProcessLockFile>;
};

/*************************************************************************/
}
}
}

#endif /* _IAS_SM_Mon_ProcessLockFile_H_ */


text:
Q: Char array seems to be overriding other data So I have another piece of code here which utilize the printchar function I wrote earlier:
void prints(char input[],int length){
 for (int i=1;i<=length;i++){
       printchar(input[i]);
  }
}

And it printed the boot sector of my os instead of input.
So I tried to create an array:
char test[]="TEST";

and print the individual letter with printchar():
and as expected, it printed the boot sector again. If I replace the array with regular char, it print the value of char.
Also, I'm pretty sure my printchar() function is working properly, because if I give it a letter to print, it print the letter, just as expected.
Here is my printchar function:
void printc(char in){
    asm volatile("mov ah,0x0e\n"
         "mov al,%0\n"
         "int 0x10\n"
         "mov al,0\n"
    ::"r"(in)
    :"eax","ax"
    );
}

A: This is a C problem, not an assembly problem. Arrays start at 0, not 1, and use semicolons, not commas, so for (int i=1,i<=length,i++){ should be for (int i=0;i<length;i++){.

A: As you said you are working with boot environment, are you sure your stack is set up correctly(after jumping to 32-bit) ?
Just to be sure, check all segment registers are set up correctly.
Also if you are in 32-bit mode, you can't(shouldn't) print using the bios int.
And how do you know it printed the bootsector exactly?

redpajama_set_name: RedPajamaStackExchange