DISCUSSION:
We report IRIDA in fraternal twins who presented with severe microcytic
anaemia, hypoferremia, hypoferritinemia and low transferrin saturation
states. Two missense mutations (p. G442R; p.E522K) in the TMPRSS6gene presenting in a compound heterozygous form were detected using
massively parallel gene sequencing. The mutations sequenced in our
Australian twins have been previously identified in several other IRIDA
patients [4,7,12]. To date, only one other case report of a
French-Canadian kindred has identified the precise compound heterozygous
missense mutations of TMPRSS6 seen in our patients [7]. In
parallel with the findings reported by Khuong-Quang et al. (2013)
[7], our patients presented with severe hypochromic microcytic
anaemia, hypoferremia and negligible transferrin saturation. Similar
blood count and serum iron abnormalities have been seen in other IRIDA
patients with either the p.G442R or the p.E522K missense mutation
[4,12]. However, unlike the French-Canadian siblings [7], our
twins showed borderline normal to low serum ferritin levels and failed
to respond to simple oral iron replacement therapies. It was only after
the administration of parenteral iron infusions that profound and
sustained improvements in the haemoglobin and serum ferritin of our
patients were observed.
Reasons for these clinical differences despite a similar genotype has
been a contentious issue within the IRIDA arena. Genotype-phenotype
studies of a large European cohort of IRIDA patients across multiple
unrelated families have proposed that missense mutations affecting theTMPRSS6 mutation through a compound heterozygous phenomenon
typically present with less severe anaemia and milder hypoferremia
[6]. In addition, they usually respond to oral iron therapy in
comparison to patients with single homozygous mutations or other lesion
types (e.g. frameshift, nonsense). However, this appears to be at
odds with our patients. The clinical heterogeneity of IRIDA been
attributed to diverse TMPRSS6 mutations affecting variable
domains within the encoded MT-2 product, leading to different degrees of
dysfunction [6,12]. The p.G442R mutation of the TMPRSS6 gene
is known to target and alter the activity of the second C1r/C1s, urchin
embryonic growth factor and bone morphogenetic protein 1 (CUB) domain of
MT-2, which is involved in its autoactivation through autocatalytic
cleavage [4,6,12]. In contrast, the p.E522K mutation disrupts the
second class A low-density lipoprotein receptor (LDLRA) domain further
downstream [6,12]. This domain has been proposed to assist with the
conformational folding of MT-2 to achieve autocatalytic cleavage, whilst
also facilitating interactions between hemojuvelin and MT-2. Silvestri
and co-workers (2009) [12] showed through a series of separately
transfected HeLa cell lines with multiple missense mutations targeting
the TMPRSS6 gene, including p.G442R and p.E522K, that the latter
was more mutagenic. LDLRA domain alterations propagated through the
p.E522K lesion showed reduced expression of MT-2 on the cell membrane of
transfected cells and was unable to interfere with hemojuvelin [12].
In contrast, the p.G442R mutation and subsequent aberrations in the CUB
domain did not alter the expression of the MT-2 protein on the cell
membrane [12]. Moreover, CUB mutations still retained partial,
albeit lowered hemojuvelin cleavage capabilities [12]. A compound
heterozygous inheritance mechanism for both missense mutations may
therefore cause a severe IRIDA phenotype that does not respond to oral
iron therapy through complete dysfunction of the mutated MT-2 encoded
product.
Of interest, we observed that despite a reasonable trial of oral iron
replacement therapy, neither child showed any clinical improvement in
red cell indices or serum iron levels. Previously published reports of
IRIDA patients with missense mutations affecting the CUB and LDLRA
domains, including the French-Canadian kindred, have successfully
treated their patients with oral iron supplements [6,7].
Interestingly, the French-Canadian siblings only showed clinical
improvements after one year of oral iron therapy in combination with
ascorbic acid [7]. The shorter time course and exclusive use of oral
iron supplementation without ascorbic acid may account for the lack of
response seen in our patients. Nevertheless, parenteral iron infusions
are reported to yield clinical benefits. It has been previously shown
that recovery of red cell indices and ferritin is often slow and
prolonged with an ongoing need for parenteral iron administration
[2,6]. Surprisingly, we observed marked and sustained improvements
in red cells indices and ferritin levels in our patients after a single
iron infusion.
In summary, we have reported IRIDA in paediatric, fraternal twins with
compound heterozygous missense mutations in the TMPRSS6 gene. We
show that despite knowledge of these TMPRSS6 mutations
[4,6,7,12], they confer extensive clinical heterogeneity. Therapy
should therefore be directed on a case-by-case basis. Currently, the
frequency of parenteral iron administration in IRIDA cases refractory to
oral iron replacement therapy is not established. Future prospective
cohort studies will hopefully elicit appropriate therapeutic guidelines
for IRIDA.