# VRBPAC January 31, 2001

LYMErix Lyme Disease Vaccine, safety update

Introduction

LYMErix, Lyme Disease Vaccine, manufactured by SmithKline Beecham
Pharmaceuticals, is indicated for the prevention of Lyme disase in
subjects 15-70 years of age. The vaccine is composed of 30 mcg
recombinant lipidated OspA with alum. OspA is the major outer surface
protein of the spirochete, *Borrelia burgdorferi*, the etiologic agent
of Lyme disease.

In 1993, the Sponsor began their initial studies of recombinant OspA as
a human antigen in Europe. In 1994, a US IND was initiated and the
sponsor began a dose ranging safety and immunogenicity study. In June
1994, VRBPAC met to discuss the clinical design of Phase 3 studies
intended to evaluate the safety and efficacy of Lyme disease vaccines.
The sponsor initiated the Phase 3 efficacy study in January 1995.
Results of this study were submitted to the FDA as part of a Product
License Application on September 15, 1997, and presented to VRBPAC on
May 26, 1998. FDA licensed LYMErix on December 21, 1998.

The safety and efficacy data that formed the basis for the approval of
LYMErix are summarized in the Summary Basis of Approval; a copy of this
document is provided with the briefing package.

Prior to licensure of the LYMErix recombinant OspA vaccine, several
research reports and epidemiological studies suggested that either
anti-OspA antibodies or OspA-specific T cells, particularly in people
who have DR4 major histocompatability (HLA) alleles, were associated
with the development of treatment resistant Lyme arthritis (TRLA). TRLA
is a rare syndrome in which as many as 10% of persons who experience
arthritis as a late clinical manifestation of Lyme disease have
persistent symptoms for months or even several years after antibiotic
treatment (Steere *et al*., Arthritis and Rheumatism 37: 878-888, 1994).
Research into the pathogenesis of TRLA is ongoing, and invokes concepts
about the fundamental nature of antigen recognition by T lymphocytes
that are still incompletely understood. In 1998, one research group
hypothesized that TRLA was a consequence of T cell recognition of a
particular peptide of the *B. burgdorferi* OspA via presentation by
particular HLA alleles. The HLA types in question included the so-called
rheumatoid arthritis-associated or "RA" alleles, exemplified by
DRβ1\*0401; many of these share a sequence in the third hypervariable
region of the DRβ1 chain. Further, the authors hypothesized that the
human cell surface adhesion molecule Lymphocyte Function Antigen 1
(LFA-1), which is expressed on activated T cells, contained a homologous
peptide that could continue to stimulate T cell activation after
elimination of the spirochete. In this hypothesis, the exact mechanism
by which chronic T cell stimulation and (Th1) cytokine secretion would
actually lead to immunopathology and frank arthritis was not specified.

Data presented in support of this hypothesis (Gross *et al.*, Science
281: 703-706, 1998) included the demonstration that T cells from
synovial fluid for 14 of 16 TRLA patients (but 5 of 16 patients with
treatment responsive Lyme arthritis, or 3 of 9 control arthritis
patients) produced interferon γ, and/or proliferated, in response to
whole OspA. Synovial T cells also responded to a particular
immunodominant peptide of OspA (amino acids 164-175), and a recently
developed binding algorithm predicted binding of this OspA~164-175~
peptide to the HLA DRβ1 0401 molecule. Further, a peptide from LFA-1 had
a 6 amino acid homology with the OspA peptide as well as a high score in
the binding algorithm; direct binding of the LFA-1 peptide to DRβ1 0401
was demonstrated *in vitro*, implying the potential for autoimmune
cross-reactivity between OspA and LFA-1. Data was also presented
indicating that synovial T cells from two TRLA patients produced
increased interferon γ in response to OspA, the OspA peptide, human
LFA-1, and the LFA-1 peptide.

On the other hand, other data presented in this and subsequent
publications do not necessarily support this hypothesis. For example,
cells from several TRLA subjects studied did not appear to recognize
either OspA, the OspA peptide, LFA-1, or the proposed cross-reactive
LFA-1 peptide (despite having one of the relevant RA alleles), and some
responses were quite low or spontaneous cytokine release without
stimulation quite high (Gross *et al.*, Science 281: 703-706, 1998). It
should also be noted that relatively few subjects have been studied to
date, and that the studies reported have not been confirmed by other
investigators. In follow up studies from the same group, only T cells
from synovial joint fluid were reactive, while peripheral blood T cells
from the same patients were not (Chen *et al.*, Arthritis Rheum. 42:
1813-1822, 1999). When T cells specific for the OspA~164-175~ peptide
were cloned from the synovial fluid of 3 TRLA patients, only a small
fraction of these clones bound an MHC DRβ1 0401 tetramer loaded with the
LFA peptide (Meyer *et al.*, PNAS 97: 11433-38, 2000), and only 11%
proliferated in response to human LFA-1 protein (SKB CMI report; see
below). The latter findings highlight the uncertainty of the LFA-1
cross-reaction at this time.

Recent studies from other investigators also suggest alternate
interpretations of the data concerning T cell responsiveness in TRLA
patients. Investigations using a combinatorial peptide library to screen
a *B. burgdorferi*-specific T cell clone derived from a patient with
chronic neuroborreliosis found not only several *Borrelia* antigens that
could stimulate proliferation, but also several other potential
autoantigens that had minimal (or no) obvious sequence homology with the
putative bacterial antigens (Hemmer *et al*., Nature Med. 5: 1375-1382,
1999). Such results imply that structural homology may be neither
necessary nor informative, and that T cell recognition may have
flexibility and plasticity that is different in different environments.
Similarly, another group used peptide spot synthesis to perform a
complete peptide substitution analysis of antigens stimulating T cell
hybridomas derived from OspA-immunized DR4 transgenic mice, including
hybridomas responsive to the OspA~164-175~ peptide. Binding motifs
identified by these analyses were used to screen databases for matching
human or mouse peptides, and these candidate autoantigens were again
used to test stimulation of the original clones. Several dozen mammalian
proteins with apparent cross-reactivity were readily identified by this
approach, suggesting that T cell cross-reactivity may actually be quite
common (Maier *et al.*, Eur. J. Immunol. 30: 448-457, 2000); if so, the
existence of such cross-reactions alone would not imply a molecular
mimicry outcome, as autoimmunity is obviously uncommon.

The hypothesis discussed above concerns the role of OspA and
OspA-responsive T cells in the pathogenesis of TRLA, which follows
infection with *Borrelia burgdorferi.* It is unclear whether there is
any relationship between this complication of Lyme disease itself (as
initiated by infection with a living, motile pathogenic bacterium), and
vaccination with purified OspA protein. Nonetheless, the clinical trials
investigating safety and efficacy of LYMErix were prospectively
designed to examine whether vaccinees experienced higher incidences of
rheumatological or neurological symptoms than placebo recipients. No
increased incidence was observed. Of note, it likely that about 30% of
the individuals enrolled in the clinical efficacy trial carried a DR4
allele, and approximately 5% an RA allele.

SmithKline Beecham also incorporated an exploratory research study of
cell mediated immune (CMI) responses to OspA in LYMErix vaccinees in
the pivotal efficacy trial, and licensure of LYMErix included a
commitment to provide a final report of this study to FDA. This report
has been received. In this study, peripheral blood lymphocytes (PBLs)
obtained from about 50 vaccinee and 50 placebo subjects were HLA typed
and studied for proliferation and cytokine secretion following
stimulation with OspA or the OspA~164-175~ peptide. PBLs from 13/41
(32%) vaccinees proliferated in response to whole (unlipidated) OspA,
and 9 of the 13 had a "positive" proliferative response to the
OspA~164-175~ peptide. However, responses to the peptide were very low,
generally much lower than the response to whole OspA protein. Not
surprisingly, cells from only 1/44 of the placebo subjects responded.
Cytokine (IFN γ and IL4) responses to either whole OspA or any of the
peptides were uniformly very low, with only a few sporadic marginal
positives. Although 30% of the vaccinees had peripheral blood CMI
responses to OspA, these responses were low, and the responses to the
OspA~164-175~ peptide almost undetectable. These results were also
analyzed by stratifying vaccinees into 3 groups according the presence
of DRβ1\*0401 /\*0404 ((RA alleles), 6 subjects; DRβ1\*11 (no increased
risk of TRLA), 10 subjects; or other DR alleles (mixed, possibly some
risk with some alleles but only low resolution typing performed), 25
subjects. No trends were apparent, especially as responses were low and
numbers of subjects small. There were also no obvious correlations
between adverse events in any of these subjects and either OspA
responsiveness or HLA type. Finally, reactivity to LFA-1 or the LFA-1
peptide was not studied; thus these studies neither support nor refute
the hypothesis concerning OspA and LFA-1 cross reactivity.

Also of note, with possible implications for the safety of LYMErix^^,
are recent reports from a number of laboratories suggesting that long
recognized mitogenic properties of OspA may be due to the ability of
OspA (and other bacterial lipoproteins) to initiate signal transduction,
lymphocyte activation, and inflammatory responses by engaging the newly
described family of cell surface receptors known as Toll-like receptors
(TLR). *In vitro* induction of IL-12 secretion by macrophages
(Brightbill *et al.*, Science 285: 732-736, 1999), activation of NFκB
(Hirshfeld *et al.*, J. Immunol. 163: 2382-2386, 1999; Lorenz *et al*.,
Infect. Immun. 68: 6398-6401, 2000), and stimulation of TNF α production
(Lien *et al.*, J. Biol. Chem. 274: 33419 - 33425, 1999) have been
reported to date. At the present time, the relevance of these *in vitro*
studies to vaccination, and any implications for the dose of OspA used,
are not clear.

At the time of licensure, the original PLA submission contained safety
data regarding the administration of 18,047 doses of the of OspA Lyme
disease vaccine (30 mcg) to 6,476 subjects who were ≥ 15 years of age.
For details of the safety analyses refer to the SBA. While exposure to
LYMErix was associated with significant increases in transient
post-injection symptoms, no increase was seen in the incidence of any
chronic musculoskeletal events, and in the incidence of any conditions
associated with chronic Lyme disease when vaccinees were compared to
control. It was noted that subjects with a prior history of
musculoskeletal conditions and those with a self-reported history of
Lyme disease were more likely to experience musculoskeletal events than
subjects without such prior history, but there were no differences in
incidence of musculoskeletel events when placebo and vaccine-recipients
were compared within any of these groups.

However, because of theoretical concerns about the association of
anti-OspA antibodies and limitations of prelicensure studies in
detecting rare adverse events, the committee recommended that additional
post-licensure studies be performed to further evaluate the safety
profile of LYMErix. At the time of approval, SmithKline Beecham agreed
to conduct a Phase 4 post-marketing study in 25,000 vaccinated
individuals (a synopsis of the study is provided in the briefing
document), to establish a pregnancy registry to specifically monitor
adverse events in pregnant women, to complete a preclinical reproductive
toxicity study, and to complete a cellular immunity study.

Since licensure of LYMErix, reports of adverse events occurring
following receipt of the vaccine have circulated in the media. The
purpose of today's meeting is to review the safety database, which
supported the licensure of LYMErix, to update the committee on the
status of post-marketing studies and adverse events reported to the
VAERS system. (A summary of the VAERS data is provided with the briefing
document.)