was to come, however. A hastily conducted post-hoc analysis of the trial was
done, which reported at the HIV Vaccines Trials Network Conference in Seattle
in November 2007, and was then published in The
Lancet in 2008.1 This
uncovered a more alarming possibility: the vaccine, though it could not
possibly have caused HIV infection in itself, may have made some participants
more vulnerable to HIV.
the higher rate of HIV infection in vaccine recipients was not statistically
significant. But further analysis suggested that the rate in a subgroup of male
volunteers were more likely to be real than random.
was only one infection in female volunteers during the trial, though there were
twelve (six in each arm) during the two-year follow-up period. While this looks
like a startling finding at first, it was not highlighted in the analysis, and
reflects three facts; firstly, although 40% of all trial participants were
women, they lagged behind men in trial recruitment and did not have so much
time to become infected with HIV; secondly, HIV prevalence amongst heterosexual
male contacts of female trial volunteers was considerably lower than it was in
the partners of gay male volunteers; thirdly, anal sex transmits HIV more
easily. The HIV infection figures below are for men only.
apparently slight excess of HIV infection in those receiving vaccine started
looking more significant when the results were stratified according to the
level of pre-existing immunity volunteers had to the Ad5 virus.
people with high levels of pre-existing immunity to the Ad5 adenovirus had been
excluded from the STEP trial, but, as we said above, a decision to double the
trial’s size had been taken in August 2005 when reasonable immune responses
were observed in this group. The trial had reached its target of 3000
volunteers by March 2007, and the analyses that follow are based on 1503 male
volunteers who received at least one shot of vaccine or placebo and 1363 male
volunteers who received at least two shots (the full schedule was three
injections at zero, one and six months).
were stratified into four groups according to Ad5 antibody levels in their
- In those with the
lowest antibody levels, 4% of volunteers caught HIV regardless of whether they
had vaccine or placebo.
- In those with
medium-low levels, 4.4% of those given vaccine caught HIV but only 2.2% given
placebo. In those with medium-high levels, 6.1% given vaccine caught HIV and
3.0% given placebo, so twice as many given the vaccine caught HIV in both of
- In those with the
highest levels of Ad5 antibodies, 4.4% given vaccine versus 1.2% given placebo
became infected. This amounts to 3.5 times as many, although in this case numbers
were very small (seven on vaccine and two on placebo).
cases the confidence intervals overlapped, and no result in any category was
near statistical significance. But the direction of change was consistent.
classical definition of ‘statistical significance’ is that there is a less than
one-in-20 probability that the effects observed could be due to chance. The
round table heard that there was a one-in-17 probability that the higher rates
of HIV infection seen in male volunteers who had high levels of pre-existing
Ad5 immunity could have been a chance effect, as opposed to an effect of the
possibility to be considered, however, is that high Ad5 immunity conferred some
kind of pre-existing resistance to HIV infection in the placebo group – which
was undermined by the vaccine. It could mean that there is some characteristic
in both the vaccine and the placebo
groups that may have to be taken into account, and this would then create what
is called a “double sided” possibility – which changes the probability that
observed effects are due to chance.
would mean that there was one chance in 35 that the difference in infection
rates observed in male volunteers with high Ad5 immunity was a random effect –
well within the boundary of statistical significance.
differences were there between the volunteer groups? In line with this being a
randomised double-blind trial, there were no behavioural differences between
those receiving placebo and those receiving vaccine.
were significant differences between volunteers with low and high levels of Ad5
immunity. People with higher levels of Ad5 immunity, recruited in the second
wave of enrolment, were twice as likely to be female, nearly half as likely to
be white and gay, and somewhat younger. We know there is already a gender bias
in the infections, but was this due to gender, because more women were black,
or for some other reason?
with high pre-existing immunity to Ad5 were also half as likely to be
circumcised. This was a potentially significant finding. Nearly two-thirds of
the men in the trial (64%) were circumcised. Excluding men with uncertain
circumcision status, there were 26 infections each in circumcised vaccine and
placebo recipients, but in uncircumcised recipients only six receiving placebo
caught HIV versus 26 receiving vaccine – the latter figure representing a
fourfold greater risk of HIV acquisition for vaccine over placebo.
information on the STEP trial emerged at the CROI conference in February 2008.3
Further data were presented from the whole group suggesting that the Ad5
vaccine specifically increased the vulnerability of uncircumcised men to
infection through insertive anal sex.
whole trial group, the difference in infection rates between vaccine and
placebo recipients was marginally statistically significant, with a p-value of
0.044 if the effect observed was caused by the vaccine alone (so-called
‘one-tailed’ probability), and a p-value of 0.077 if the effect was caused by
some combination of a direct vaccine effect and some kind of protective
characteristic of the placebo group that was removed (‘two-tailed’
group that had high levels of adenovirus immunity, however, there were 21
infections in vaccine recipients and only nine in placebo recipients, and this
was statistically significant (p = 0.02 one-tailed, 0.029 two-tailed).
Mark Robertson, presenting preliminary data on immune responses in the trial,4
showed that volunteers with high Ad5 immunity levels had higher levels of
generalised CD4 cell activation. Susan Buchbinder of UCSF said that the
incidence of HIV infection, at about 4 to 5% a year, was the same in vaccine
recipients regardless of their Ad5 immunity level, but the incidence of
infection was lower in placebo
recipients who had higher levels of Ad5 immunity, ranging from 4% a year in
those with the lowest level to 1.2% a year in those with the highest level. The
vaccine may therefore somehow have removed a protective effect of Ad5 immunity.
a univariate model and in a multivariate model that progressively eliminated
other possible confounding factors, high Ad5 immunity conferred a threefold
higher risk of infection in vaccine recipients than in placebo recipients,
compared to no increased risk in people with low Ad5 immunity when compared to
the placebo group.
Buchbinder added, the real reason for the additional risk in vaccine recipients
may have had little to do with Ad5 immunity, or it may only have had an
was one more important risk factor for HIV acquisition in vaccine recipients
when compared to the placebo group: circumcision, or rather, the lack of it.
vaccine recipients were, in univariate and multivariate analyses, four times
more likely to become infected with HIV if they received vaccine than if they
received placebo. In contrast there was no difference at all in infection rates
between circumcised vaccine versus placebo recipients.
questions after the presentation, Buchbinder commented that the risk to
uncircumcised men was greater than the risk to men with high Ad5 immunity, and
that the latter may be a passive marker for the former. This was because the
men with high Ad5 immunity, who were largely recruited in the second wave, also
tended to come from countries and communities with lower rates of circumcision;
they were younger and more likely to come from Latin America, for instance.
commented that preliminary data hinted that the enhancement of infection risk
was specifically seen in uncircumcised gay men who largely or exclusively had
insertive anal sex. This would imply that the vaccine was undermining some
immune mechanism that normally protected uncircumcised ‘tops’ from infection
through the mucosa of the foreskin.
this would not explain in itself why placebo recipients were less likely to get
HIV if they had high Ad5 immunity, so the two effects may be synergistic.
concern arising from the STEP trial results was that the vaccine might have
stimulated some recipients’ immune systems in a way that sensitised, rather
than immunised, people to HIV. If that were the case, it could mean that
recipients of the V520 vaccine could be unusually vulnerable to HIV for a long
time – possibly for the rest of their lives. Immune responses to the vaccine
did appear to be long-lasting and had faded only slightly in a group of early
vaccinees two years after injection.
by October 2009, when more follow-up data were presented at the AIDS Vaccine
2009 conference in Paris, it was beginning to seem that the excess risk to
adenovirus-experienced male recipients was fading over time.5
Susan Buchbinder told the conference that an excess risk was still evident in
Ad5-seropositive men, but it was no longer trending towards significance. The
risk was still of borderline statistical significance in uncircumcised men,
though it had declined too.
told the conference that 60 new infections had occurred in study participants
between unblinding in October 2007 and January 2009. Twelve occurred in women
(six in each group) and 48 in men (26 in the vaccine group and 22 in the
are still many questions left unanswered by the STEP trial. Buchbinder said
that the trial participants would need to continue to be followed to find out
if the apparent extra vulnerability to HIV conferred by the vaccine was
long-lasting, and Mark Robertson outlined a whole range of other factors that
would be analysed, including other immune activation markers and CD4 subsets, markers
of genetic vulnerability to infection such as HLA genotypes, herpes (HSV-2)
status, seminal immune-cell subsets and seminal HIV viral load in
seroconverters, and so on.