B7 Coreceptor Molecules as Clinically-Relevant Surrogate Biomarkers in the Hyper IgD Syndrome (HIDS) form of Mevalonate Kinase Deficiency (MKD)

Cholesterol and isoprenoids are critical in membrane biogenesis, embryonic
development, intracellular signal transduction, vesicular trafficking, and cell
cycle progression (Goldstein et al 2006; Gong et al 2006a,b).
HIDS/MKD is an autoinflammatory disease characterized by systemic inflammation
without an apparent infectious etiology. Mevalonate kinase deficiency (MKD) is
a human enzyme deficiency with diverse phenotypes, including severe mevalonic
aciduria (MA) and HIDS (Drenth, 1999; Simon, 2004). Clinically, MA and HIDS are
distinguished based upon the extent of neurologic involvement, which is
prominent in MA and predominantly absent in HIDS (Prietsch, 2003; Hoffmann,
1993). Both individuals with MA and HIDS may present with hepatosplenomegaly,
lymphadenopathy, anemia, increased erythrocyte sedimentation rates and levels
of C-reactive protein, leukocytosis, and increased urinary leukotriene
excretion (Prietsch, 2003). People with HIDS suffer recurrent febrile crises
during childhood characterized by elevated serum IgD and IgA1 levels, skin
rash, arthralgia and myalgia (Rigante, 2009; Steichen, 2009; Touitou, 2008; van
der Hilst, 2008; Sornsakrin, 2008), yet studies by Ammouri and coworkers
(Ammouri, 2007) suggest a poor correlation between elevated IgD in serum and
MKD. With regard to pathogenic MVK (the gene responsible for the phenotypic
spectrum of MKD) mutations, the c.1129G>A (p.V377I) is pathognomonic for the
HIDS form of MKD, and this allele confers temperature sensitivity to the MVK
protein (Houten, 2002).

MVK and inflammation: Patients with MKD exhibit dysregulation of IL-1*, a major
inflammatory cytokine. Stimulated peripheral blood mononuclear cells (PBMCs)
cultured from individuals with MKD hypersecrete IL-1*, and this is exacerbated
by addition of lovastatin (Drenth, 1996; Frenkel, 2002). Hypersecretion of
IL-1* was recently shown to be linked to a shortage of geranylgeranylated
proteins (Houten, 2003a and 2003b; Mandey, 2006b). More recently, Marcuzzi and
coworkers (Marcuzzi, 2008) recapitulated the inflammation observed in MKD by
administering aminobisphosphonates (which inhibit the mevalonate pathway) to
BALB/c mice. Intervention with exogenous isoprenoids (geraniol, farnesol and
geranylgeraniol) effectively reversed the aminobisphosphonate-induced
inflammation. We speculate that a deficiency in dolichol production, linked to
loss of MVK activity since dolichol is made from isoprenoid precursors, could
lead to aberrant protein glycosylation and shedding or secretion of IgD which
is normally present as membrane-bound immunoglobulin in B lymphocytes. We
further hypothesize that shortage of dolichol results in aberrant expression of
costimulatory B7-glycoprotein expression on various cell types (see below).
Finally, if oxysterol production is altered in MKD, this could alter immune
function (Hannedouche, 2011).
Costimulatory B7-glycoproteins in the MKD mouse model of HIDS: We determined
the activation status and proliferative capacities of splenic lymphocyte
populations from Mvk+/- mice, a phenocopy of HIDS. Mvk-/- mice are embryonic
lethal, while Mvk+/- mice demonstrate increased serum levels of IgD, IgA1, and
TNF*, temperature dysregulation, hematological abnormalities, and splenomegaly.
Flow cytometry analysis of cell surface activation markers on T and B
lymphocytes, and macrophage populations, demonstrated aberrant expression of B7
glycoproteins in all splenic cell types studied. In Mvk+/- CD4 and CD8 T cells,
alterations in expression of CD25, CD80, CD152, and CD28 were observed (Fig. 2,
upper left and right quadrants). Mvk+/- splenic macrophages expressed altered
levels of CD80, CD86, CD40, and CD11c (Fig. 2, lower left quadrant), while
Mvk+/- B lymphocytes had differential expression of CD40, CD80, and CD86 (Fig.
2, lower right quadrant).
We postulate that imbalances in the expression of cell surface proteins
necessary for activation, proliferation, and regulation of the intensity and
duration of an immune response may result in defective T cell activation,
proliferation, and effector functions in the murine HIDS model, and potentially
in human HIDS. We will examine this hypothesis in HIDS patients, and will
further determine if these B7 glycoprotein molecules correlate with clinical
severity, other known biomarkers of HIDS, as well as isoprenoid metabolites
reflecting overall cholesterol pathway function. Our studies hold promise for
identifying surrogate biomarkers specific for human HIDS, and will
simultaneously expand our limited understanding of the pathophysiology of this
rare disorder.

For figures and references: see study protocol.

The objective of this study is to assess potentially new and unique biomarkers
that will be specific to patients with HIDS as surrogate outcomes for eventual
larger, cohort-controlled clinical studies. Our longitudinal design in a small
pilot group will also identify new mechanisms of pathophysiology in HIDS
patients. The primary hypothesis is that the costimulatory B7 glycoprotein
abnormalities identified in the murine MKD model will be recapitulated in sera
obtained from human HIDS patients, either before, during, or after febrile
episodes. The secondary hypothesis posits that B7 glycoprotein molecule levels
will correlate with clinical symptomatic severity score, other known biomarkers
of HIDS (IgD, IgA and IL-6), markers of inflammation (CRP/ESR and leukocyte
count), and/or markers of isoprenoid metabolism (lathosterol as a measure of
cholesterol synthesis, cholesterol and oxysterols, urine mevalonic acid (MVA),
farnesyl glucuronide, dolichol and ubiquinone, among others).

Observational prospective pilot study.

To begin, at least 14 days following any fever or symptoms from the last
febrile episode, research participants will begin a 24 hour urine collection
and obtain a blood draw at the end of this 24 hour period. Body temperature and
clinical symptom monitoring will begin at this time. Once a temperature of
*100.4*F (*38.0*C) as well as a combined clinical score of * 20 is noted,
participants will be considered to be in a febrile episode. At this time, a
second 24-hour urine collection period will begin, followed by a second blood
draw at the end this second 24-hr urine collection period. After 72 hours from
the beginning of the febrile episode, participants will begijn a third 24-hour
urine collection, followed by a third blood draw. As soon as the participants
body temperature returns to <100.4*F (<38.0*C) as well as a combined clinical
score <20, participants will begin a final 24 hour urine collection period,
followed by a final blood draw. A follow-up visit or phone call will be
conducted one month after the final specimen collection to record any adverse
events.
During the entire study, participants are instructed to follow their usual
diet. As far as medications, use of abortive or symptomatic medications,
including anakinra or steroids, as well as medications used on demand (e.g.
antipyretics/analgesics) will be prohibited for this single febrile episode.

The study design is summarized in Fig. 3, page 13, of the protocol.

- Venipuncture: The vein in which the needle has been inserted to draw blood
may become sore and red. A temporary *black and blue mark* may develop, and
rarely fainting may occur. Blood draws will cause some minor pain and carry a
small risk of bleeding and/or infection at the puncture site. A blood clot
could develop and go to the lungs. Such problems are exceedingly rare.
- Skin Biopsy: The risk of a skin biopsy includes scarring, bleeding, or
infection.
- Being off medication: Risk includes increased temperature, and exacerbation
of usual HIDS symptoms during a febrile episode. This is a burden to the
patient because of fever, pain and general malaise. Without treatment, a HIDS
attack will end spontaneously in about 5-6 days.