Posts Tagged ‘South Africa’

HPV vaccines for South Africa: coming to a school near you!

19 May, 2013

From The Independent Online:

HPV and cervical cancer: courtesy Russell Kightley Media

HPV and cervical cancer: courtesy Russell Kightley Media

“Cape Town – Government will start administering cervical cancer vaccines in schools from February next year, Health Minister Aaron Motsoaledi has announced.

Speaking during the health budget vote debate in the National Assembly on Wednesday, Motsoaledi said government hoped to negotiate lower prices for the vaccine, which treats the Human Papilloma Virus (HPV) – the major cause of cervical cancer among women.

Quoting experts, he said cervical cancer affected 6000 South African women a year, 80 percent of them black. More than half the women affected died of the disease.

While the HPV vaccine presented an opportunity to prevent women from contracting cancer, there were still obstacles to overcome.”

This is a really, really big deal for South Africans – and pity is, the vaccine will not be given to boys, or universally to girls.

Seriously: all the science says that giving it to boys as well limits spread of the viruses far better; not making it universally available will mean all sorts of recriminations around unequal access (read: to less privileged kids ONLY as part of the government programme at first).

But a big step in the right direction!

ViroBlogy: 2012 in review

1 February, 2013

So: thank you, anyone who clicked in, and regular visitors.  You make it worthwhile!!

The WordPress.com stats helper monkeys prepared a 2012 annual report for this blog.

Here’s an excerpt:

4,329 films were submitted to the 2012 Cannes Film Festival. This blog had 33,000 views in 2012. If each view were a film, this blog would power 8 Film Festivals

Click here to see the complete report.

CCHFV in South Africa

1 February, 2013

I am indebted to the National Institute for Communicable Diseases (NICD) in Johannesburg for their very informative newsletter, from which I culled this.

I would also like to very sincerely congratulate Professor Barry Schoub, a long-time former Director of the NICD, on his  African Society for Laboratory Medicine (ASLM) Lifetime Achievement Award!  Very well deserved.

Crimean-Congo haemorrhagic fever

Two cases of Crimean-Congo haemorrhagic fever (CCHF)  acquired in South Africa have been laboratory confirmed  in January 2013.

On 1 January 2013, a 31-year-old male working as a  game warden on private game ranch near Jagersfontein  (Free State Province) presented with clinical features  suggestive of CCHF. The patient did not report any tick  bites or direct exposure to unprocessed meat or  slaughtering of animals. The Centre for Emerging and  Zoonotic Diseases of the NICD/NHLS confirmed infection  with CCHF virus by PCR and serology testing.

A second case of CCHF was laboratory confirmed on 12  January 2013 in a 44-year-old male hospitalised in  Bloemfontein, Free State Province. He had been on a  farm in Pomfret, North West Province (situated ±5 km  from the border with Botswana), where he was bitten by  a tick. Three days later he developed symptoms, and  presented with fever, rash, conjunctivitis and pharyngitis.  No laboratory-confirmed cases were identified in 2011- 2012.

Human CCHF cases have been reported annually  from South Africa since 1981, when it was first  recognised in the country; between 0 and 20 cases of  CCHF are diagnosed each year. Through nearly thirty  years of passive surveillance, a total of 187 cases has  been laboratory confirmed. Although cases have been  reported from all of the nine provinces, more than half of  the cases originate from the semi-arid areas of Northern  Cape Province (31.5% of cases) and Free State Province  (23% of cases).

CCHF infection is generally asymptomatic in many species  of wildlife (including antelope) and livestock animals  (including cattle, sheep, goats, hares and ostriches).  Humans  become  infected  sporadically  by  ticks,  particularly  Hyalomma ticks, which are both reservoirs  and vectors for CCHF virus. Other modes of transmission  include direct contact with blood/tissues of infected  animals, and in the case of healthcare workers, through  direct contact with the blood/tissue of infected patients;  nosocomial outbreaks are well described and have been  associated with high mortality rates. Disease may be  severe in people, with case-fatality rates reported as 3 –  30% across various studies.

Detailed information for healthcare workers regarding  CCHF can be found on the NICD website  http:// http://www.nicd.ac.za/ (see General Public FAQ, or Health Workers FAQs here).

Setting up a platform for plant-based influenza virus vaccine production in South Africa

5 May, 2012

A virus-like particle formed by influenza virus haemagglutinin budding out of plant cells. By Russell Kightley Media

See it also on Scoop.itVirology News

Our (very) recently-published article on plant-made flu vaccines in BMC Biotechnology:

Setting up a platform for plant-based influenza virus vaccine production in South Africa

Elizabeth Mortimer, James M Maclean, Sandiswa Mbewana, Amelia Buys, Anna-Lise Williamson, Inga I Hitzeroth and Edward P Rybicki

Background
During a global influenza pandemic, the vaccine requirements of developing countries can surpass their supply capabilities, if these exist at all, compelling them to rely on developed countries for stocks that may not be available in time. There is thus a need for developing countries in general to produce their own pandemic and possibly seasonal influenza vaccines. Here we describe the development of a plant-based platform for producing influenza vaccines locally, in South Africa. Plant-produced influenza vaccine candidates are quicker to develop and potentially cheaper than egg-produced influenza vaccines, and their production can be rapidly upscaled. In this study, we investigated the feasibility of producing a vaccine to the highly pathogenic avian influenza A subtype H5N1 virus, the most generally virulent influenza virus identified to date. Two variants of the haemagglutinin (HA) surface glycoprotein gene were synthesised for optimum expression in plants: these were the full-length HA gene (H5) and a truncated form lacking the transmembrane domain (H5tr). The genes were cloned into a panel of Agrobacterium tumefaciens binary plant expression vectors in order to test HA accumulation in different cell compartments. The constructs were transiently expressed in tobacco by means of agroinfiltration. Stable transgenic tobacco plants were also generated to provide seed for stable storage of the material as a pre-pandemic strategy.

Results
For both transient and transgenic expression systems the highest accumulation of full-length H5 protein occurred in the apoplastic spaces, while the highest accumulation of H5tr was in the endoplasmic reticulum. The H5 proteins were produced at relatively high concentrations in both systems. Following partial purification, haemagglutination and haemagglutination inhibition tests indicated that the conformation of the plant-produced HA variants was correct and the proteins were functional. The immunisation of chickens and mice with the candidate vaccines elicited HA-specific antibody responses.

Conclusions
We managed, after synthesis of two versions of a single gene, to produce by transient and transgenic expression in plants, two variants of a highly pathogenic avian influenza virus HA protein which could have vaccine potential. This is a proof of principle of the potential of plant-produced influenza vaccines as a feasible pandemic response strategy for South Africa and other developing countries.”

I have mentioned time and again that going green is the sensible thing to do: here is a concrete example of how my research group is trying to go about it.  This is a very sensible technology for rapid-response vaccine production, and especially for emerging or orphan or pandemic virus threats.  We got really good expresion levels of H5N1 HA protein via transient expression in plants, and have already started on pandemic H1N1 HA expression.  Let’s hope some governmental types in SA take some notice!

I thank Russell Kightley Media for the specially-commissioned graphic of budded HA-only VLPs.

 

HIV Vaccines From Bangkok – 4, and final….

22 September, 2011

Thursday morning started with three parallel oral sessions – and I chose Symposium 07, Characterization of Breakthrough Viruses.  The second talk – by Morgane Rolland, in the US Military HIV Research Program – detailed a study of the sieve analysis of breakthrough viruses in the RV144 Thai trial.  They wished to see whether or not the vaccine could block infection of specific variants, and thought they might see that viruses in vaccinees were evolutionarily distant from the insert in the vaccine, relative to the placebo arm.

HIV and its life cycle

The saw no differences in virus diversity over 10 sequences per person, in 121 people,  71 of whom were in the placebo arm.  They did note, however, that linked transmissions showed less diversity in the env gene than normal – 1 vs 10%.  Over 75% of cases had a single founder virus, in both placebo and vaccine arms.  There was no significant divergence from the vaccine sequence in either group in anything but the Pro aa sequence – with some non-significant evidence for Env variation.

When they looked for Env sites under selection in gp120, they saw 4 in the placebo group at positions 181, 208, 327 and 359 – with less variation in vaccine than placebo recipients.  Rolland speculated that this could be to do with entry being more restrictive in vaccinees?  4 different sites in the vaccine group were under selection: they found that for MHC I epitopes there was a greater distance for vaccine than placebo groups, with a result that was not significant for MHC II epitopes.

There was a trend toward longer Env V2 loop sequences in vaccinees at later times, and a reduced number of cysteines in Env among vaccinees – this was seen also in the VAX004 trial.

Phil Berman – formerly of VaxGen, which made the gp120 for RV144 and earlier trials – mentioned that there was lower variance in Env than in the unsuccessful VAX 003 trial.  Jerome Kim noted that men seroconverting had a much higher incidence of HCV infection – which could be associated with undeclared IV drug use.

Katharine Barr of Univ Alabama spoke next, on the increased incidence of multiple variant transmission of HIV in VAX003 injection drug users.  She noted that this efficacy trial was of gp120 in IV drug users, while VAX004  was in MSM and high-risk women: they speculated that differences if any could be due to transmission route – as in, IV route vs sexual.  She further noted that in RV144, the best (non-significant) effect was in low-risk heterosexuals.

Something that was a little disturbing to me, given HIV transmission in our part of the world is overwhelmingly by heterosexual sex, was that the IV route is responsible for 10% of world infections.  They had looked at transmitted founder viruses – the ones going in and replicating in recipients.  They predicted that consensus of a low diversity lineage is the sequence of the founder virus – and that several founders would give multiple low variance lineages.

She noted that 80% of heterosexual infections are established by single viruses, so there exists a window of opportunity of viral vulnerability when vaccine-induced immunity could block infection.  However, with MSM, the multiple infection goes up to 40%; while injection drug users (IDUs) are less studied, multiplicity goes up  60% in one study and 31% in another….

Looking at Vax003 results, they had asked how high a barrier there had been for placebo infections, and whether in vaccinees there were more or fewer founder viruses?  While they had found that there was an 44% incidence of multiple variant transmission in the  placebo arm, and  22% in the vaccinees, this was unfortunately not significant, given the low numbers.  There was a median of 1.8 viruses per transmission vs 1.3, but this too was not significant.  However, it could mean there is a higher bar for vaccine protection among IDUs, which has important implications for which groups to use in vaccine trials.

Katherine incidentally gave the best answer yet heard to a long and detailed question: “I think that’s a really good question but I have zero data to address it…” = I don’t know.

Which prompted thoughts of new conference drinking games: take a shot every time you hear a speaker say “I would like to thank the organisers for inviting me…”, or “Our hypothesis [generally pronounced hy-PAH-the-sis] was…”, or a question which starts with either “…really good talk / great data” or “So – ummmm – when you/we did…”.

Paul Edlefsen (Fred Hutchinson Cancer Res Ctr) described a sieve analysis of RV144 [and started: “So...umm...” = another shot!].  He repeated the finding that observed correlates of risk generated two hypotheses; namely, that high IgG response to Env protected from HIV infection while a high IgA response interfered with protection.  Additionally, analysis of the antibody response using scaffold V region showed that a high V2 response correlated with a lower infection rate.  He noted that the STEP trial results showed a distinct difference in Gag between vaccine and placebo groups.  He noted further that were only 110 usable subjects in RV144, so they could only detect large sieve effects in their study of CTL and Ab epitope responses.

MUCH MIND-NUMBINGLY BORING STATISTICAL METHODOLOGY FOLLOWED…sorry, Paul!

There were 2 sites of evidence for sieving – aa positions 169 and 181 in the Env V2 loop, in the middle of a region identified by Ab binding array data.  There was also some evidence of covariation among pairs of aa residues in the V2 loop for vaccinees only.

After a long and complicated structural question, he gave the second-best answer of the conference: “I could say that I do, or that I don’t – but I have so little expertise in this area…(laughter)”.  And after long rambling statement: – “I’m sorry, was there a question in there?”

Brandon Keele (National Cancer Inst, MD) described work on NHPs which they had extended to studying human transmission of HIV, on transmitted/founder viruses.  NHP studies show multiple founders because doses are high generally, in order to get 100% infection rates.  One study using very low dose multiple intrarectal exposures to see if one can immunise macaques showed that one virus could do it.  Animals followed up from early times stayed with one evolving variant.

He noted that the consensus sequences in humans posited to have had one transmitted variant are average in  neutralisation susceptibility.  These viruses are all functional in vitro and in vivo and one can get full length viral clones ex NHPs which recap original founder viral load and pathogenicity.  All such viruses use the CCR5 coreceptor.  All HIV clones replicate in CD4 T-cells but not in  macrophages.  The transmission signature is to increase Env processing and infectivity.

They now mix cloned viruses with tags so can follow them in NHP challenge experiments, as most challenge studies have used virus with <1% diversity, which represents a clone in any one epitope – which he felt to be non-reflective of the real world .

The closing plenary session was opened by IAVI‘s Wayne Koff, who remarked that he had heard someone say “The  airport….”, in answer to the session name “Where do we go next?”….

Jeffrey Boyington (Vaccine Res Ctr) described some very impressive work on using structure of Env for rational immunogen design, specifically to target the CD4 binding site as a good target for broadly neutralising Ab.  They used crytallographic data to make proteins best mimicking the struc and then used them as immunogens.  They had used stabilised resurfaced gp120 with mutations around the binding site and isolated dozens of Abs with them from several infected subjects.  Part of the process involved stabilising flexible regions by bolstering cysteine content, removing glycans from the site of interest and adding them to immunodominant sites, and using Chikungunya virus VLPs to multimerise spike proteins for maximal immunogenicity.  Boyington noted that there were 80 native trimers on the surface of the VLPs, and that one can put the Outer Domain of gp120 on the tip of each monomer.  They get good Ab back for gp120 and get CD4 binding site Ab in rabbits.  In rhesus monkeys primed with gp140 trimers they got good boosting and better Abs to the CD4 BS.

Altogether a very impressive account – and one which advances to possibility of other opportunities for the design of other good broad-binding vaccine epitopes.

Rick King of IAVI followed, with an account of the current status and future directions of vector-based HIV vaccines.  He stated that most HIV vaccines now involve vectors – so there is a wealth of data that can be efficacious, so how to use it?  He thinks that we want the next generation of vectored vaccines to block infection and control virus load – meaning a combination of Ab and cellular responses.He noted that in NHPs, SIV protection is possible, and that it requires Env in the vaccine – and that the mechanism of protection is under intense investigation right now.

He further noted that in a DNA prime MVA boost vaccine regime, protection is associated with the avidity of the Abs.  Thus, a major goal is to improve the response to Env, by identifying the nature of the protective response, and enhancing and using native Envs to do it.  He stated in this context that there were only two vaccine regimens using native spike protein – and that one of them is the SA AIDS Vaccine Initiative (SAAVI) vaccine.

It was possible to engineer Env to bind a broader array of broadly neutralising Ab and to incorporate it into vesicular stomatitis virus (VSV) instead of the native G protein spike, or into canine distemper virus (CDV, a measles relative), which replicates in lymphoid tissue.  One could also bias processing of Env in CDV to get better cleavage and presentation.  The rCDV could be put into ferrets and shown to replicate.

He said that while the RV144 vaccine did not control viral load, vaccines can control SIV replication, so we need to have those components in HIV vaccines.  For instance, recombinant live cytomegalovirus (CMV) expressing the whole proteome of SIV could control the virus, this was associated with CD8 effector memory T-cells.

He thought we need to capitalise on information on mechanisms of control, and to increase immunity by use of replicating vectors and heterologous prime/boost combos, and deal with diversity by broadening the response.  The reason for replicating vectors was because live attenuated virus works for SIV – preventing infection and controlling replication.  Possibilities were vaccinia, measles, VSV, Sendai, CMV, AdV, CDV and VSV-HIV chimaeras.  As for diversity, one could increase the number of epitopes by using mosaics, and direct responses using conserved epitopes, as Tomas Hanke has demonstrated in IAVI-funded trials using chimpanzee Ad as prime then MVA as a boost with his HIVCONS Ag.

Finally, there was what I consider to have been the best talk of the conference – simply because it was much wider in scope than the rest: Steven Reed of the Infectious Disease Res Inst, Seattle, described new generation adjuvants for use with HIV.  He started by noting that adjuvants were necessary for lots of things; eg: for T-cell vaccines for TB and leishmania; for Ab response-broadening (Cervarix, HPV vaccine); Ag dose sparing (eg flu); to combat immune sensescence, and for vaccine therapy.

They had focused on a toll-like receptor (TLR4) agonist as an adjuvant, following work that showed that the well-known MPL was a TLR4 agonist ,and vaccines including TLR agonists had been used unknowingly since 1885.

He thinks the ideal adjuvant should have no effect on lymphocytes, no systemic effects, no non-specific B or T cell responses, should elicit potent long-lived responses, should redirect ongoing immune responses, and should be safe and effective in all age groups.  They had accordingly designed GLA – based on lipid A – to bind TLR4: this was purely synthetic, and induces Th1 CD4 helper cells and a broad humoral immunity.  They used a hexaacyl chain length that was preferred by human TLR4, which is restricted to macrophages and dendritic cells, has transient local effects, and reduces inflammation so as to get better central memory.

They can also formulate it differently for different vaccines and can get very different effects thereby.  For example, emulsion alone stimulates Th2 responses while GLA stimulates Th1 even in combo with an emulsion, which helps in leishmania and TB vaccines.

He noted that alum-based adjuvant stimulated mainly a Th2 response, while adding GLA gives a Th1 response with the same antigen.  They get good Ab diversity with GLA and expansion of it with the malaria vaccine – and Ab diversity leads to better neutralisation (eg transl med 2011).

GLA increases and broadens the haemagglutination-inhibtion (HAI) Ab response to the influenza vaccine Fluzone, which contains lots of inactivated virions.  He noted one gets a better protective response against “drifted” viruses – which have evolved away from the vaccine strains – with GLA.  Baculovirus-made H5N1 vaccine requires 30x less vaccine to get the same response with GLA.

It is also possible to get mucosal immunity by IM vaccination with HIV gp140, according to Robin Shattock’s results.

Reed noted that intradermal adjuvants are very rare – and that this looks good with flu vaccines delivered this way.  They were in the process of optimising the adjuvant formulation for intradermal delivery to increase vaccine potency, get mucosal immunity, and CD8+ T-cell responses.  Dermal dendritic cells have a wider range of TLRs than Langerhans cells – so Sanofi target them with ID delivery, and GLA works well to amplify the response.  It was impressive that they could protect ferrets with a single ID vaccine shot of flu vaccine.  It was also interesting that they are working with Medicago Inc., who have one of the most successful plant-produced influenza virus vaccine candidates, presently in human trial.

Thereafter, closing remarks from the conference organiser were as one would expect; people were honoured for their present and long-term contributions – notably Jose Esparza – and the venue of the next conference was announced to be Boston, with Dan Barouch as Organising Chair.

It was a good conference, with all of the high-intensity interactions and presentations one would expect from such a loaded topic.  However, it possibly suffered from over-emphasis of the “RV144 results” – which weren’t that impressive, in my opinion – as part of an effort to keep up perceived momentum from announcement of the RV144 success (small as it was) from the previous meeting.  For me, the highlights were the envelope antigen design talks, and what I managed to catch of the actual virology, and especially analysis of diversity by massively parallel sequencing.

We still don’t have an effective HIV vaccine – but we’re getting closer. 

HIV Vaccines From Bangkok – 2

16 September, 2011

Big News Day: HIV Vaccine Conference, Tuesday 13th September
The first plenary session of the conference had as its theme “Novel approaches in clinical evaluation through global collaboration” – and it was graced by the presence of no fewer than three scientists in full military dress uniform complete with medals, from the USA and Thailand (Nelson Michael and Jerome Kim and Punnee Pitisuttithum), reflecting the significant involvement of both countries’ military in the RV144 efficacy trial.

It was probably fitting, however, that it was led off by Pontiano Kaleebu of the MRC/UVRI Research Unit on AIDS from Uganda, on Africa’s contributions to AIDS vaccines.  He said that Africa had been crucial to the endeavour for a number of good scientific and societal reasons, but principally because most infections are there:  some countries are up to 15% of total population being HIV+ and sub-Saharan Africa contributed 17% of global infections in 2010.  Factors influencing vaccine efficacy that were unique were the great diversity of viruses, the mainly heterosexual transmission of viruses, diverse HLA alleles and significant preexisting vector immunity.

Small Buddha used for offerings, temple of the Reclining Buddha

South African scientists – largely drawn from the University of Cape Town, he says, modestly – had been responsible for the only vaccines designed in Africa, which were now in clinical trial.  Africa had been part of much work on epidemiology and variation of HIV-1.  Africa and Africans had contributed to understanding transmission events, mechanisms of early viral control and immune escape, and had helped in the addition of new broadly neutralising MAb derived from patients in African cohorts.

The first vaccine trial in Africa was done in 2000 in Uganda, and there have been many since:  30 trials or 17% of all trials have been done in Africa, mainly by  IAVI (13)  and the HVTN (11).

He noted that financing had declined and that the  reduced vaccine pipeline was a challenge, as many well-established sites have no vaccine to trial.  Another challenge was that new prevention successes means lower viral incidence, so trials have to be bigger – and may be impossible in certain cohorts.  There was also a challenge in the up and downstream HIV research imbalance in Africa – where there was no research infrastructure in many centres so sampes got shipped out, while clinical trials were large and well serviced.

His conclusion was that Africa had made significant contributions to vaccine research and development, but that challenges such as those mentioned could threaten further work.

Dr Punnee Pittisutithum of Mahidol University in Bangkok described how Thailand had a national plan established as early as 1993, and revised in 2006, to transfer technology, and to collaborate with a variety of institutions and countries, in HIV vaccine research and development and prevention efforts.  A collaboration in 1997 with Japan had used recombinant BCG as a subtype E vaccine prime, boosted with live vaccinia virus.  There had since been14 preventive trials including 2 efficacy trials.  In 1997 they had established a plan to monitor circulating virus – and now 1765 Thai viruses had been sequenced, and they had a very good idea of variation and currently circulating viruses.
They had an impressive infrastructure to set up and monitor clinical trials, which accounted for the success of the Thai trials over the years.  The partially successful RV144 efficacy trial had resulted in a study of correlates of protection involving 35 investigators from many institutes, including in Thailand.  She also made a point – as many others did subsequently of thanking the 16 402 Thai men and women who participated in the RV144trials.

Jerome Kim of the Walter Reed Army Institute of Research and Deputy Director of the US Military HIV Research Program spoke next, on correlates, sieve analysis and clinical development of the RV144 trial.

His first news was that there was a correlation of high Ab concentration to Env in vaccinees with a low risk of infection – resulting from 4000-odd samples analysed in the last two years for correlation. The work was the result of 35 investigators from 25 institutions collaborating on samples gathered during the trial.

A unique finding was that the gp120 and ALVAC vaccines were novel immunogens – the gp 120 from Vaxgen (also used in previous trials) used a N-terminal 11 aa replacement from a HSV gD glycoprotein epitope, which may have affected presention of Env HIV epitopes: its presence changed the binding of mAbs directed against gp120 up to 10-fold better for conformation-specific epitopes in the V2 and V3 loop epitopes, but not for linear epitopes.
There had been a good reaction to V2 peptides by intracellular cytokine staining (ICS) assays, with the response by CD4+ T-cells mainly.  One unfortunate finding was that gp120 Ab binding dropped 1.5 logs from 12 to 24 weeks post the last vaccination.

Sieve analysis – a new term to me, but denoting analysis of breakthough HIV or other infections in vaccine trials for selection pressure by the vaccine – looked at the gp120 V2 loop sequence in placebo and vaccine arm infectees – and saw selection.

It was left to the ununiformed Barton Haynes – Duke Human Vaccine Institute – to actually break the big news on correlates in the RV144 case control study.  Bart gave a lesson in the careful exposition of a complex topic of potentially huge significance in the HIV vaccine world, by starting with an explanation of what they had discovered – correlates of risk rather than of protection – and exactly what it meant.

He explained that a correlate of risk may be causal, or that it could be a surrogate marker.  Their team had looked at 41 infected and  205 uninfected vaccinees and 40 placebo recipients,  followed for 3 yrs.  There were only 41 cases, so the statistical study was only powered to pick up strong correlates and could miss weak ones.

Pilot work noted that most Ab responses were directed to the gp120 V2 loop.  In brief, their work found that there were two correlations associated with infection rate: the first was that serum IgG binding to a scaffolded V2 loop correlated inversely with infection rate.  The second was that Env binding of serum IgA correlated positively with infection rate: the statistical analysis showed a 43% reduction in infection rate associated with high serum IgG to the V2 region and a 54% increase in infection rate with high serum IgA binding.  Their hypothesis to explain this result was that high V2 IgG Ab levels were protective and low plasma IgA was associated with protection.

Ongoing analysis wase focused on looking at 9 PBMC-produced cytokines – and medium to high cytokine levels seemed to be correlated with protection.  Epitope mapping of IgA binding find C1 peptide response was correlated with infection, so maybe plasma monomeric IgA can block antibody dependent cytotoxicity (ADCC) caused by IgG binding?  He noted that only monomeric plasma IgA was collected in this trial, and mucosal dimeric IgA was to be collected next.

Haynes thought the way forward is to see if correlates of risk are causal correlations, by new clinical trials, or by having the trial Abs tested in non-human primates (NHPs) for passive protection.  They were presently testing the IgA association with risk by looking at mAbs in macaques binding all sorts things in Env, to see if IgA inhibited other Ab binding.

While the result was undoubtedly interesting – and unexpected – I was not convinced that it was as significant as it was made out to be, for a number of reasons – and a number of people I spoke to at the conference agreed.  Thus, in my slightly jaundiced opinion, the relatively weak correlations with risk applied for just one region in Env for this one vaccine combination with a unique monomeric gp120 product, for one subtype of HIV-1, in one population, for a trial in which the efficacy was only 30-odd percent, and which even then was only just significant.  They also tested only plasma antibodies, which may not give the full picture, and got next to no cellular response, which could correlate with the total absence of virus control seen in both placebo or vaccine recipients who became infected.

Accordingly, I think the massive obsession with analysing the results of the trial may turn out in retrospect to have been a bit of a waste, compared to testing new products which have markedly better responses in preclinical trials.  Another view can be seen here.

Giuseppe Pantaleo of the Centre Hospitalier Universitaire Vaudois in Lausanne closed out the session, with a presentation on poxvirus vector-based vaccines beyond the RV144 trial.    He pointed out that modified vaccinia Ankara (MVA) was the result of 571 passages in chick embryo fibroblast (CEF) cells; that the ALVAC canarypox vaccine resulted from 200 such passages, and vaccinia Copenhagen had had the deliberate deletion of 18 ORFS to result in the NYVAC vaccine.  The two former vaccines did not replicate in humans; however, replication competent poxviruses give appropriate innate immune cytokine responses and CD4 help.  To this end, NYVAC is known to grow in human primary keratinocytes, is highly attenuated, has no effect on dendritic cell (DC) maturation, and one gets higher levels and longer persistence of expressed antigens, cross presentation of Ag by MHC I and II receptors and stimulation of memory T-cell responses.

They tested replication competent and a replication deficient NYVAC and DNA expressing the same Ag and compared them, with  a boost of type C gp120.  They compared the effect of DNA priming or not, and scarification or intramuscular injection for NYVAC, with DNA and NYVAC both expressing Env and a Gag/Pol polyprotein.  The gag gene in constructs makes particles and a trimeric secreted gp140.  Pantaleo noted that the DNA plus regime elicited much more cellular immunity and a predominantly Gag/Pol response, while NYVAC alone gives 70-80% response to Env.  In the DNA+ group there was a balanced CD4 and CD8 T-cell response.  High Elispot results get long term and durable response.  There was no difference between scarification and im immunisaiton, and no increase of immune response with protein boost.  There was also no difference between rep and non rep NYVAC.  In the no DNA group the rep virus was lots better.

Most HIV-1 neutralising Ab response was in the DNA- groups and SHIV neutralisation was restricted to the DNA- group.

In ADCC assays for the DNA+ group there was no advantage in boosting with protein, and response decayed later with some animals being negative; in the DNA- group responses were considerably higher, there was an advantage to boosting,and all animals were positive.  In cross-type titring assays there was good cross-binding of IgG, with the DNA- groups being better.

The lesson from this was that a greater magnitude of T-cell responses do not necessarily correlate with neutralising (NAb) responses.

For plasma IgA responses they see the same distribution as for IgG.  In the DNA- groups they get very little response up to 3 months, then good responses 8-9 months, which then wane after 12.  Their response would be to boost with poxvirus plus Env at 12 months.  Pantaleo thinks we need compressed regimens to reduce the time of reduced protection, that we should try Env-only regimens, and that we should tailor vectors for optimal Ab responses.

My opinion on this is that one should try for Env-specific Ab responses AND Gag- and other protein-specific T-cell responses, elicited at the same time by immunisation in different limbs.

There were 6 Tuesday afternoon sessions, in two sets of three, so some judicious choices were needed.  I went with Oral Sessions 3 and 5, entitled Novel Immunogens and Inserts, and Acute Infection/Viral Diversity, respectively.

Oral 03: Novel Immunogens and Inserts
Two stand-out talks for me were one by A Flamar, and another by M Zhou – with a third on my favourite virus-like particles, by L Yang.

The Flamar talk reported targetting to CD40 receptors of five 19-32 aa peptides containing a string of known highly conserved CD4 and CD8 T-cell epitopes from Gag, Nef and Pol covalently linked to a lipid tail for antigen presenting cell (APC) uptake.  These have been tested and found to be therapeutic already.  The targetting is done using a MAb targetting CD40 with the HIV5 pep attached to the heavy chain C-terminus. The epitopes are 2 from Gag, 2 from Nef and one from pol.  The MAb is a humanised one with mouse Vh and Vkappa portions, which binds monocytes and APCs specifically.  The immunogen expands HIV peptide-specific CD4 and 8 T-cells from HIV+ patients.  They get broad peptide-specific responses and CD4 and CD8 polyfunctional responses.  The latter are CTL-characteristic and can kill target cells as well as suppressing HIV replication in vitro.

They are presently humanising the V region for clinical manufacture and testing in mice and NHP.

The Zhou talk discussed the use of mimotopes – peptide sequences mimicking native epitopes – displayed via phage surfaces, which mimic a membrane-proximal or MPER gp41 epitope, and bind Ab from an elite controller of virus load.  The M13 display library is made by env-specific PCR and fragmentation followed by cloning, and is bound by immobilised IgG.  “Panned” phage is eluted and amplified.  They get epitopes localised to gp41, inclusing the MPER region, using EC26-2a4 Ab.  The core epitope overlaps the known binding site of the broadly-neutralising MAb 2F5 but is distinct: the sequence is  NEQELLELDK.  They used this as an immunogen after a env DNA prime as phage plus adjuvant x3 – and got neutralising Ab back.  This is a genuinely exciting result, as it builds on much speculation regarding just how good Abs directed against this region are at neutralising a wide rtange of HIV variants – and answers some of the questions about how difficult they are to make.  These two sorts of immunogen – one aimed at T-cell responses, the other at neutralising Ab response – may yet be a valuable adjunct to other vaccines containing more conventional ingredients.

The third talk by Yang was very useful in that it demonstrated the possibility of using insect cells – which can be cultured very reliably at large scale, and are already used to make a major human vaccine (GSK’s anti-HPV Cervarix) – to make genuine HIV virus-like particles, with a Gag shell inside a membrane, studded with processed Env spikes.  I was especially interested as we have already used the same technology to make Gag-only VLPs, which are a real possibility as a subunit vaccine.  However, routine production of VLPs is complicated by the fact thats VLP are usually produced in low quantity, there is poor cleavage of Env, there is often recombinant baculovirus contamination, and poor batch consistency.

They used VLPs Drosophila S2 cells that had been stably transfected with plasmids encoding HIV-1 Gag and Env.  S2 cells are good as they grow up to high density and can easily be cultured in suspension – in a WAVE bioreactor in this case, which is scalable up to hundreds of litres.  They get glycosylated Env which undergoes appropriate cleavage and ends up as spike protein on budded Gag-containing VLPs.  They get 23 million cells per ml, and 8 mg of gp120 / litre – which is an excellent yield for VLPs.  Appropriate Mabs bind the spikes, indicating correct conformation.  Upon immunisation with a DNA prime and VLP plus CpG boost, they get a good Ab response which is weakly neutralising.  An ADCC test was also positive.  The T-cell response was a relatively poor CD4 but good CD8 cell.  The result is not entirely new – there were two posters at the conference describing the same thing, and our group has used stably transfected insect cells to make baculovirus-free VLPs – but they have investigated production at scale, and have shown appreciable and appropriate immunogenicity for what may be a valuable future component of heterologous prime-boost regimens.

Oral 5: Acute Infection/Viral Diversity
While I probably should have been more interested in acute infections, and there were several most worthy talks on this, I am inexorably drawn as a result of my history in virology and with HIV, to studies of virus diversity and especially of virus evolution over time and between individuals.  So there were really only two talks in it….

L Yin presented a fascinating account of the use of deep pyrosequencing to look at the evolution of viral diversity in peripheral blood cells in single individuals over time.  The study used pyrosequencing of cell-associated virus – which of course, reflects the whole history of the individual’s infection as integrated DNA – to look at diversity and both real and inferred longitudinal variation, given multiple blood samples from the individuals over time.  They used samples from children infected at birth, for time spans of 18 months to 6 years.  In six children, 4 of the 6 showed big differences in virus populations, while 2 did not not.  The biggest diversity was for R5 coreceptor binding sequences, illustrating immune selection of viruses.

Their conclusion was that deep sequencing was a robust method for evaluation of complexity and population structure and for evaluation of the virus historical record in an individual.  It was also easily possible to compare cell-associated and plasma-isolated virus, as DNA and cDNA respectively.

V Novitsky from the Essex lab presented on the dynamics of changes in Gag sequences in the global epidemic; how they change over time and the probable age of HIV subtype C in particular.

They sampled databases for sequences of 500 bases and up for gag, and found only 1800 -odd suitable sequences: these were mostly from South Africa, and Zambia, Malawi and Botswana.  The sequences were reduced for dating purposes to 433 by criteria such as <10 per year per country, while  966 were used for diversity.  They arbitrarily defined 9 groups of about 150 viruses over 20 years from 1983.  Interestingly, there was no clustering by year of sampling, or extinguishing of lineages.  SA had profound founder effects for 2 groups of viruses;  for diversity of Gag over time, one could see significant increase over time.  The p17 C terminus had the highest changes for this protein, p24 less so and spread throughout the sequence, with the most changes in the rest of Gag (p15).  Only 20 aa positions over 500 show consistent selection pressure changes, meaning the consensus sequence of Gag fom Subtype C HIV-1 remains pretty much same over more than 25 years.  They estimated the time of viral diversification from other subtypes to have been around 1959 – with a hefty uncertainty.

While the results may not seem exciting – and indeed, some people said “What’s new?” – the fact that Gag consensus sequences have essentially been stable over a protracted period is interesting; so too is the fact that lineages do not seem to have disappeared as one sees with influenza viruses with immune selection.  A very interesting virus, HIV-1….

A recycled virus to protect against TB?

25 August, 2011

News from the University of Cape Town site:

“UCT is taking part in the Phase IIb proof-of-concept efficacy trial of a candidate tuberculosis vaccine, a study that will involve people living with the human immunodeficiency virus (HIV).

Researchers from the Institute of Infectious Disease and Molecular Medicine will screen and test patients living in Khayelitsha, using the vaccine known as MVA85A. The patients are HIV positive but have not been infected with TB.

This is the first proof-of-concept efficacy trial in people infected with HIV using MVA85A, which is being developed by the Oxford-Emergent Tuberculosis Consortium (OETC), a joint venture between the University of Oxford and Emergent BioSolutions, and Aeras, a non-profit partnership focusing on TB vaccine regimens.

The MVA85A vaccine candidate is intended to boost the response of immune-essential T-cells already stimulated by the Bacille Calmette-Guerin (BCG) vaccine, also used against tuberculosis.”

So – fantastic, and it involves the alma mater, but what does it have to do with viruses??  Note the throwaway “…using MVA85A…”: while this could be an adjuvant, or some kind of carrier, it is in fact a live virus.  Modified Vaccinia Ankara, in fact, meaning it is a variant of the tried-and-true smallpox virus vaccines that have been with us since Edward Jenner did his thing on the 14th of  May, 1796.  Poxviruses, and especially vaccinia and fowlpox viruses, can also be genetically engineered to express foreign proteins, because they have large genomes and can tolerate even quite large insertions without it affecting the virus much.  There is a useful recent paper on the subject in PLoS One; inevitably, Wikipedia  has an article on it too.  Not a very good one, however!

It does have this, though:

Modified Vaccinia Ankara (MVA) virus, is a highly attenuated strain of vaccinia virus that was developed towards the end of the campaign for the eradication of smallpox by Professor Anton Mayr in Germany. Produced by hundreds of passages of vaccinia virus in chicken cells, MVA has lost about 10% of the vaccinia genome and with it the ability to replicate efficiently in primate cells”.

So, two important features:

  1. the virus replicates in chicken cells and in chicken eggs, meaning it can be cultivated at large scale
  2. it does not replicate in primate, and in fact not in most mammal, cells

It does, however, undergo a significant portion of the life cycle in mammalian cells – only virion maturation does not occur.  This means that genes inserted into the MVA virus genome with appropriate poxvirus promoters may be expressed in cells containing virus particles even if the virus does not multiply.  The other Wikipedia page mentioning it – the Vaccinia page – has this:

“Modified vaccinia Ankara: a highly attenuated (not virulent) strain created by passaging vaccinia virus several hundred times in chicken embryo fibroblasts. Unlike some other vaccinia strains it does not make immunodeficient mice sick and therefore may be safer to use in humans who have weaker immune systems due to being very young, very old, having HIV/AIDS, etc.”

And THAT’S why MVA as a TB vaccine vector in what amounts to a high-risk environment for HIV infection in South Africa: because the vaccine won’t cause complications in immune-suppressed individuals.

As I have previously discussed here, MVA has also been used as a vector for a component of the South African HIV-1 vaccine developed at UCT that is currently in Phase I clinical trial in SA and in the USA: there the MVA was engineered to express both a gp150 Env and a polygenic fusion protein GRTTN (Gag-RT-Tat-Nef), and was the boost component to a dual-component DNA vaccine expressing both singly.

It is encouraging that technology that has been touted for many years is finally seeing the mainstream: a large clinical trial combining immunogenicity with efficacy.  Malaria antigens are also being delivered by MVA in clinical trials; HIV Env antigens were delivered using avian poxviruses in the only HIV vaccine efficacy trial that showed any positive effects at all – so the promise is finally being fulfilled.

A sword turned into a ploughshare.  We need to see more of them!

…and the virus marches on….

1 September, 2009

From News24 this evening:

SA’s H1N1 deaths now 27

2009-09-01 17:03

Cape Town – The number of swine flu deaths in South Africa has risen to 27, and confirmed infections to 5 841, the National Institute for Communicable Diseases said on Tuesday.

“There is also ongoing and widespread community transmission,” it said in a statement.

Of the 27 fatal cases, 12 were pregnant women, five of whom had no identified underlying conditions.

The institute repeated its standard warning that people with depressed immunity, asthma, diabetes, or chronic lung, kidney and heart problems, or who were pregnant should seek early treatment with antivirals.

and on 26th August:

SA wants own H1N1 flu vaccine

2009-08-26 22:29

Cape Town – South Africa has no choice but to develop its own H1N1 flu vaccine, Health Minister Aaron Motsoaledi said on Wednesday, citing concerns treatment will not be available to poorer nations.

“South Africa has arrived at a situation where we have no option but to start developing our own vaccine capacity, not only for H1N1, but generally,” Motsoaledi told parliament.

“The disturbing feature about today’s world… has been expressed by the minister of health for Cambodia… who noted that the developed world, after producing the vaccine, may want to cover their own population first before thinking about the developing world,” Motsoaledi said.

Anyone remember reading that before, anywhere?  Watch this space….

First African-developed HIV vaccine goes to trial

21 July, 2009

Finally, finally, a product of our 10-odd-year-old South African HIV vaccine development programme goes into Phase I human trial, in South Africa!

I say “our” because I was part of the overall team; however, the two vaccines which comprise the SA AIDS Vaccine Initiative (SAAVI) / HIV Vaccine Trials Network (HVTN) trial – designated SAAVI 102/HVTN 073 – were designed and developed by others.

The vaccines consist of a DNA component, consisting of an artificial “polygene” dubbed Grttn (for Gag-RT-Tat-Nef) and a truncated Env (gp150) cloned separately into vector plasmids, and the same genes recombined under the control of different promoters into the genome of a poxvirus (Modified Vaccinia Ankara, MVA).  These and their testing in mice and non-human primates have been described in published work: see here and here for relevant journal articles on the MVA and DNA components.

From the July 20th SAAVI press release:

The test vaccines – called SAAVI MVA-C and SAAVI DNA-C2 – have shown promising results in animal testing. The SAAVI DNA-C2 vaccine was constructed in South Africa using a plasmid backbone provided by the Dale and Betty Bumpers Vaccine Research Center (VRC) of NIAID, while the MVA vaccine was designed by the team at UCT and constructed and manufactured in the USA.

“Reaching this important milestone of translating our discoveries in the laboratory to testing in humans would not have been possible without the support of a large team of people from the University of Cape Town, together with national and international collaborations.  An effective vaccine against HIV/AIDS remains a top global health priority and it is our hope that the evaluation of these vaccines in clinical trial will provide some important answers that will bring us closer towards this goal,” says Prof. Anna-Lise Williamson, leader of the vaccine development team and joint staff member of UCT’s Institute for Infectious Disease and Molecular Medicine, and the National Health Laboratory Services (NHLS).

The SAAVI DNA-C2 was constructed in South Africa and manufactured in the US by Althea Technologies. The MVA vaccine was manufactured by Therion Biologics, USA. The vaccines will be tested in a prime-boost approach where the SAAVI DNA-C2 vaccine will be given to prime the immune response and the SAAVI MVA-C vaccine to boost or enhance the immune response.

National and international press got hold of the story in a big way – unsurprisingly, given as there is the 5th IAS Conference on AIDS Pathogenesis going in in Cape Town at the same time, which incidentally has its own live blog feed.

The University of Cape Town is obviously pleased with the press release (see here); however, the launch had its fair share of controversy: Associated Press reporter Michelle Faul posted a story yesterday entitled “South Africa begins AIDS vaccine trial, cuts funds“, which has been taken up by a wide spectrum of especially foreign media.  According to Faul:

“South Africa launched a high-profile trial of an AIDS vaccine created by its own researchers Monday, a proud moment in a nation where government denial, neglect and unscientific responses have helped fuel the world’s worst AIDS crisis.

After a government official lauded the project at a ceremony at Cape Town’s Crossroads shantytown, the scientist leading the research said state funding had been halted.

The contrast between Monday’s hopeful vaccine launch and the revelation of funding cuts raised questions about whether the government was backsliding on its pledge to combat AIDS.

Anna-Lise Williamson, an AIDS researcher at the University of Cape Town, told The Associated Press the clinical trial would continue with U.S. money. But she said South Africa’s Department of Science and Technology had pulled its funding in March, while the project’s other sponsor, the state electricity utility Eskom, did not renew its contract when it expired last year.

Neither government spokesmen nor Eskom immediately returned calls seeking comment about funding cuts.”

In the midst of light, there is darkness…frequently, thanks to ESKOM

I have blogged on my personal view elsewhere; suffice it to say that bad decisions were made, and  9 years worth of momentum has effectively been lost – along with a number of very experienced personnel, and many years worth of accumulated and very relevant experience.

For an illustration of the product pipeline which existed behind the current trial offerings – and which may now never be developed – click here  for published descriptions of our plasmid- and BCG-vectored and virus-like particle (VLP) subunit vaccines.

But who knows, this current trial may even show promise – and then it will all have been worth it.  Let’s live in hope!

H1N1: coming to a South African home near you, soon

14 July, 2009

And after a very pleasant holiday, I come back to work to find…85 cases of confirmed pandemic H1N1 in South Africa!

Yes, it is true – at least, as far as the National Institute for Communicable Diseases (NICD) Director, Prof Barry Schoub, is concerned: he was featured yesterday on eTV News explaining how it was all going.  And it is “mild” according to him: it looks the same as standard flu, although most cases so far are due to people bringing it into the country, without much community spread…yet

In an article just published by the Independent Online (IOL), Kanina Foss says:

Swine flu cases will probably spike when schools reopen next week. Health officials will monitor schools, but are still advising that mild cases should be treated no differently from seasonal flu.

Only patients with serious symptoms – such as high fever, persistent vomiting, pain in the chest, or shortness of breath – should seek medical assistance. These are symptoms that people would seek medical assistance for anyway, says National Institute for Communicable Diseases (NICD) deputy director Lucille Blumberg.

The number of confirmed swine flu cases in South Africa is 75 [since modified, see above]. Once this number reaches 100, the NICD will stop counting. It will focus instead on severe cases and those at high risk because of compromised immune systems, such as HIV-positive people.  The institute will also monitor schools.  Counting cases was resource intensive, said the NICD on Monday, and served no more purpose than counting cases of seasonal flu. The overwhelming majority of cases worldwide had been mild, and had required no special treatment.

The World Health Organisation (WHO) has said the spread of the H1N1 virus is inevitable, and the NICD is expecting many more South Africans will be infected. It is unsure how the country’s high HIV prevalence will affect the severity of infections.

“It’s something we need to monitor very carefully,” said Blumberg.

The highest number of confirmed cases are in Gauteng (39), followed by the Eastern Cape (nine), and Western Cape (five).

Oh, and in my other persona, a post on AIDS denialism….


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