J Clin Immunol (2015) 35:147157

DOI 10.1007/s10875-015-0135-7

ORIGINAL RESEARCH

Variable Phenotype of Severe Immunodeficiencies Associated

with RMRP Gene Mutations
Winnie Ip & H. Bobby Gaspar & Robert Kleta & Estelle Chanudet &
Chiara Bacchelli & Alison Pitts & Zohreh Nademi & E. Graham Davies &
Mary A. Slatter & Persis Amrolia & Kanchan Rao & Paul Veys &
Andrew R. Gennery & Waseem Qasim

Received: 8 September 2014 / Accepted: 23 January 2015 / Published online: 8 February 2015
# Springer Science+Business Media New York 2015

Abstract
Purpose Mutations in RMRP primarily give rise to Cartilage
Hair Hypoplasia (CHH), a highly diverse skeletal disorder
which can be associated with severe immunodeficiency. Increased availability of RMRP mutation screening has uncovered a number of infants with significant immunodeficiency
W. Ip : H. B. Gaspar : E. G. Davies : W. Qasim
Molecular and Cellular Immunology Section, Institute of Child
Health, University College of London, London, UK
W. Ip : H. B. Gaspar : E. G. Davies : W. Qasim
Department of Clinical Immunology, Great Ormond Street Hospital,
London, UK
R. Kleta
Centre for Nephrology University College London, Royal Free
Hospital, London, UK
E. Chanudet : C. Bacchelli
Centre for Translational Omics-GOSgene, Institute of Child Health,
University College, London, UK
A. Pitts : Z. Nademi : M. A. Slatter : A. R. Gennery
Institute of Cellular Medicine, Newcastle University, Newcastle upon
Tyne, UK
Z. Nademi : M. A. Slatter : A. R. Gennery
Bone Marrow Transplant Unit, Royal Victoria Infirmary, Great North
Childrens Hospital, Newcastle upon Tyne, UK
P. Amrolia : K. Rao : P. Veys
Bone Marrow Transplantation Department, Great Ormond Street
Hospital, London, UK
W. Ip (*)
Molecular and Cellular Immunology Section, UCL Institute of Child
Health, 30 Guilford Street, London WC1N 1EH, UK
e-mail: w.ip@ucl.ac.uk

but only mild or absent skeletal features. We surveyed the

clinical and immunological phenotype of children who have
undergone allogeneic haematopoietic stem cell transplantation
for this condition in the UK.
Methods Thirteen patients with confirmed RMRP mutations
underwent allogeneic stem cell transplantation (SCT) at two
nationally commissioned centres using a variety of donors and
conditioning regimens. Records were retrospectively
reviewed.
Results Median time from clinical presentation to diagnosis
was 12 months (range 1 to 276 months), with three infants
diagnosed with severe combined immunodeficiency (SCID)
without radiographical manifestations of CHH. A total of 17
allogeneic procedures were performed on 13 patients including two stem-cell top-ups. The median age at transplant was
32.4 months (range 1.5 to 125 months). Of the eleven surviving patients, median follow-up was 50 months (range 21.6 to
168 months).
Conclusions RMRP mutations can cause short stature and significant immunodeficiency which can be corrected by allogeneic SCT and the diagnosis should be considered even in the
absence of skeletal manifestations.

Keywords Allogeneic stem cell transplantation . cartilage hair

hypoplasia . metaphyseal chondrodysplasia .
immunodeficiencies . variable phenotype

Abbreviations
CHH
Cartilage hair hypoplasia
RMRP Ribonuclease mitochondrial RNA processing
SCT
Stem cell transplantation
SCID
Severe combined immunodeficiency

148

Introduction
Cartilage Hair Hypoplasia (CHH) was first described in
Amish populations as an autosomal recessive form of
metaphyseal chondrodysplasia with short-limbed dwarfism
and light-coloured hypoplastic hair [1] and affects 1.5/1000
births in this North American community compared to an
incidence of 1 in 20,000 Finnish births, the most affected
European population [2]. It has also been reported in patients
from other parts of Europe, South America, and Asia [35].
Characteristic radiological findings include short tubular
bones with widened, scalloped and sclerotic metaphyses most
notable in knees and ankles [6]. Other findings include coneshaped epiphyses, short metacarpals and phalanges, lumbar
lordosis, and femoral bowing [7, 8]. A variety of extraskeletal features have been recognised ranging from
Hirschsprungs disease to variable degrees of immunodeficiency, bone marrow dysplasia, granulomatous inflammation
and predisposition to malignancies [912]. The genetic basis
of CHH maps to chromosome 9p, a region encoding an untranslated RNA component of the mitochondrial RNAprocessing (RMRP) complex [13]. Mutations in RMRP cause
disruption of ribosomal processing and disruption of cell cycle
progression in rapidly dividing cells such as lymphocytes and
chondrocytes [14]. Over 70 different mutations have been
described [15] with a common mutation (70 A>G) accounting
for 92 % of Finnish CHH patients and 48 % of non-Finnish
patients [3]. Clinical heterogeneity and variable degree of immunodeficiency have been previously reported [9, 16, 17]. In
subjects with the most severe immunodeficiency, marrow insufficiency or haematological malignancy, allogeneic
haematopoietic stem cell transplant (SCT) offers the possibility of life saving therapy. Transplant outcomes have previously been published in a European collaborative study [18] with
overall survival of 63 % in cohort of 16 children treated between the years 19912006. Here we survey the UK experience of allo-SCT infants with RMRP mutations and uncover a
highly variable clinical phenotype, including children presenting with SCID but without skeletal manifestation of CHH.

Patients and Methods

Thirteen patients with confirmed RMRP mutations (P1-P13)
and short stature underwent allo-SCT at Great Ormond Street
Hospital in London (n=8) and Great North Childrens Hospital in Newcastle upon Tyne (n=5) during a 14-year period
from 1998 to 2013.
Three siblings (P3, P4 and P5) are from a consanguineous
South Asian family. P9, P12 and P13 have previously been
reported in [18];P10 was reported in [12]. Median time from
presentation to diagnosis was 12 months (range 1 to
276 months). Eleven of 13 patients were alive at last follow-

J Clin Immunol (2015) 35:147157

up with a median follow-up period of 50 months from transplantation (range 21.6 to 168 months).
RMRP Gene Mutation Analysis
P1, P6 P13 underwent sequence analysis of the RMRP gene
by Regional Molecular Genetics Lab (Great Ormond Street
NHS Trust, London). RMRP gene mutation screening for P2
was performed at Centre for Pediatrics and Adolescent Medicine Freiburg University Hospital. P3-5 had gene mutation
analyses (linkage analyses, custom DNA capture and Next
Generation Sequencing) performed by UCL Genomics and
GOSgene (Centre for Translational Omics, UCL Institute of
Child Health).
Immunological Evaluation
Lymphocyte populations were enumerated using flow cytometry. Whole blood was labelled with combinations of monoclonal antibodies conjugated with fluorescein isothiocyanate
(FITC), phycoerythrin (PE), allophycocyanin (APC),
peridinin chlorophyll protein (PerCP), or fluorochrome combinations with cyanines (PerCP-Cy5.5, APC-Cy7, and PECy7) (BD Biosciences). Lymphocyte subsets were detected
with reagent containing CD3 FITC, CD16+56 PE, CD45
PerCP-Cy5.5, CD19 APC, CD4 PE-Cy7, and CD8-APCCy7. Nave, effector, and memory T cell populations were
detected with CD45RA FITC, CD27 PE, CD45 PerCP, and
CD4 or CD8 APC.
T-cell repertoire was evaluated either by flow analysis of
TCRV expression as percentage of total CD3+ T population;
or by CDR3 TCR molecular spectratyping. For the latter, template cDNA corresponding to rearranged transcripts with different CDR3 lengths from specific TCR variable region genes
was amplified by PCR and the products run on AB3130 Genetic Analyzer (Applied Biosystems) and analyzed with SpA
Web-based software [19]. T-cell functionality was assessed by
measuring T-cell proliferation in response to mitogen (PHA)
as described previously [20].
Thymopoiesis was quantified by nave T-cell numbers (as
above) and levels of T-cell receptor rearrangement excision
circles (TRECs) measured by real-time qPCR as described
previously [21].
B-cell immunity was evaluated by immunoglobulin levels
(IgG, A, M) measured with a Dade Behring (Milton Keynes)
nephelometer (BN II), according to the manufacturers instructions; and by measuring vaccine responses against childhood immunisations.
Analysis of Chimerism
Engraftment studies were performed by short tandem repeat
variability on peripheral blood samples using PowerPlex 16

J Clin Immunol (2015) 35:147157

system (Promega, Southampton, United Kingdom) for PCR

amplification; followed by analysis on AB3130 Genetic
Analyser with Gene Mapper v4.0 software (Life Technologies, Carlsbad, Calif). If indicated, lineage-specific chimerism
analysis was performed after selection of CD3/CD15/CD19+
cells using the magnetically activated cell sorting (MACS)
system (Miltenyi Biotech, Surrey, United Kingdom).

Results
Clinical Features (Table 1)

The main clinical features for the 13 patients are

summarised in Table 1. Typical features of short stature, skeletal abnormalities and fine silky hair were seen in 4 (P2, P9,
P12, P13) patients. All presented with short stature at presentation with one having disproportionate growth; however only
7 had radiological changes typical of CHH. Of the three affected siblings, two had skeletal features that were noted after
a genetic diagnosis of CHH was made. The other sibling (P4)
had normal radiographs at the age of 9 years. The eldest (P3)
presented at 5 months of age with clinical features of SCID
and received SCT from a matched family donor. The younger
2 siblings (P4 and P5) were diagnosed at birth with undefined
SCID based on family history and underwent SCT at 3 years
and 1.5 year, respectively. Here, a molecular diagnosis of
RMRP mutation was confirmed after SCT (see below).
All but one patient (P1) presented either in infancy or early
childhood with immunodeficiency . One patient (P2) had features of Omenns syndrome (skin rash, hepatosplenomegaly,
lymphopenia, and eosinophilia). Atypically, P1 presented at
9 years of age with longstanding history of enteropathy, recurrent chest infections and growth failure. Previously radiographs taken of spine and legs at age of 18 months had shown
mild flaring of metaphyses and bowing of femora. Repeat
radiographs at 3 years showed lytic areas in the metaphyses
of femur and tibia and cone-shaped epiphyses suggestive of
CHH. Immunological investigations showed combined immunodeficiency; patient underwent SCT and had genetic confirmation of CHH.
Only 3 patients (P2, P3, P5) had typical skeletal changes
seen in CHH; 2 of whom had changes detected after SCT.
Laboratory Findings
The results of immunological evaluations at presentation are
summarised in Table 2. All thirteen patients had lymphopenia
and/or CD4 lymphopenia. CD8 lymphopenia was present in 9
of 13 patients. B cell numbers were low in 5 patients and 2 had
pan hypogammaglobulinemia. The majority (9 out of 11) had
absent or impaired proliferative response to PHA. Three of 4

149

patients had abnormal T cell repertoire. P2 (Omenns syndrome) had abnormal spectratype with oligoclonal expansions. P6 and P7 had reduced percentage of T cells expressing
a V marker without oligoclonal expansions. TRECs were
measured in 9 out of 12 patients and they were low or absent
in all.
Thirteen different mutations in the RMRP gene were identified (Table 3) including 3 new mutations that have not been
reported previously. Five patients (P1, P2, P6, P10, P12) were
compound heterozygotes for either duplication or insertion in
the promoter region and a single nucleotide substitution in the
transcribed region. P1 had a 17-nucleotide insertion at position 20 located between the TATA box and the transcription
initiation site on one allele and a 748C>T point mutation in the
5 region of the other allele. P2 had on one allele a 15-bp
duplication at position 25 in the promoter region followed
by a 4C>T point mutation in the second allele. P6 (of mixed
race African and Caucasian) was compound heterozygote for
the common 70A>G mutation, and a 7-nucleotide insertion at
position 20 in the promoter region on another allele. P10 had
monoalleic 13 bp duplication in the promoter (not previously
described) along with previously reported 4C>T point mutation on the other allele (described in McCann LJ et al. 2014).
P12 inherited from one parent a 28-nucleotide duplication at
position 28 (not previously described), and a 154 G>C point
mutation from another parent. P7 and P8 were compound
heterozygous for single base pair substitutions only. P7 had
a 151 G>A point mutation in one allele which is newly identified but position 151 is known to be highly conserved between species. This patient carried the common 70 A>G mutation in the other allele. P8 was compound heterozygous for
two previously reported mutations.
Homozygous mutations were identified in six patients, two
of whom carried the common 70 A>G mutation (P9 and P13).
Three siblings (P3, P4 and P5) who had originally undergone
SCT for undefined SCID were homozygous for 35 C>A nucleotide substitutions. This molecular diagnosis was uncovered by linkage analyses performed on all members of the
family which had identified a 46 Mb region containing around
426 genes. Sanger sequencing confirmed the homozygous
mutations within the RMRP gene. Compound heterozygous
mutations have previously been reported at the same position
of 35 in CHH in association with a 70A>G mutation. The
affected P3 domain is critical for RMRP binding to protein
subunits and nucleolar localisation [22]. Interestingly homozygous 35 C>A mutations in our kindred have resulted in
different skeletal phenotypes, with only P3 and P5 having
radiological evidence of skeletal dysplasia. More recently, a
newborn member of the extended family was found to encode
the same mutations but has completely normal immunological
and growth parameters at 1 year of age. There are perhaps yet
unknown modifier genes at play that determine the manifestations of genetic mutations in the RMRP gene.

Malabsorption, diarrhoea, Failure to thrive

Siblings

17 m

120 m

108 m

ND c

+0.4

M
0m

P 5*

17 m

+
-

+
+ ADV gut

-c

+<<0.4

F
3m

P6

Degree of metaphyseal dysplasia

disproportionate short stature present at birth

3m

Neutropenia
(auto immune)

ND

+ <0.4

M
8m

P7

276 m

Sclerosing
cholangitis

+
-

++

+<<<0.4

M
24 m

P8

12 m

+
+ EBV,
ADV,
HHV6
+f
+g
-

+ <0.4

F
0m

P 9^

+
+ EBV associated
B cell lymphoma
Neutropenia,
Granulomatous
skin lesions
1m

+
+ EBV

+ <0.4

F
32 m

P10#

6m

+
-

+d

+ <0.4

M
19 m

P 11

1m

+ EBV
LPD

+
+e

+ <0.4

F
29 m

P 12^

4m

+
+ EBV,
HHV6

++

+ <0.4

M
11 m

P 13^

pure red cell aplasia secondary to EBV

ADV and HHV6 enteritis

recurrent chest infections and bronchiectasis

recurrent ear infections

these three siblings did not undergo skeletal surveys until after the diagnosis of CHH was made. P3 had typical features of CHH (degree of metaphyseal dysplasia ++) P4 had normal X-ray. P5 had mild
metaphyseal and epiphyseal changes without shortening of long bones

P9-P13 were treated at Great North Childrens Hospital, Newcastle upon Tyne

P1-P8 were treated at Great Ormond Street Hospital, London

# Previously described in McCann et al. 2014

^ Previously described in Bordon et.al 2010

++ bowed, shortened long bones, marked metaphyseal cupping, cone-shaped metaphyses in phalanges

+ shortening of long bones and metaphyseal changes

- shortening of long bones but no clear signs of metaphyseal dysplasia

5m

24 m

Time to diagnosis
(from presentation)

+
Omenns

+
+
-

Other Features

GI symptoms
Anaemia
Malignancy

+
-

+ severe
varicella

+ EBV

ND c

ND c

+
+ RSV, Para flu,
CMV, Candida,
polio stool
+
-

+0.4

+0.4

+a<<<0.4

+<<<0.4

F
0m

P 4*

F
5m

P 3*

F
5m

P2

M
108 m

P1

Clinical features at presentation in 13 patients with cartilage hair hypoplasia

General features
Sex
Age at Presentation
Clinical features
Short Statue
(height %ile) at
presentation
Radiological skeletal
changes b
Fine silky hair
Recurrent Infections

Table 1

150
J Clin Immunol (2015) 35:147157

P1

P2

P9

Patient on Rituximab at time of evaluation

26 months
951 (14003700)
251 (7002200)
76 (4901300)
0a (3901400)
179 (130720)
ND
ND
ND
ND

1885 (14003700)
459 (7002200)
714 (4901300)
1064 (3901400)
615 (130720)
ND
Absent
Absent
IgG
+IgM

390 (25005500)
160 (16004000)
140 (5601700)
430 (3002000)
1060 (1701100)
ND
Absent
Absent
IgA

32 months

P 11

620 (25005500)
200 (16004000)
400 (5601700)
370 (3002000)
160 (1701100)
ND
ND
Impaired
IgA, IgM

1280 (25005600)
210 (18004000)
820 (5901600)
320 (4303000)
660 (170830)
ND
V low
Absent
Normal

0 months

P5

570 (14003700)
190 (7002200)
9 (4901300)
290 (3901400)
ND (130720)
ND
Absent
Absent
IgG, IgA, IgM

2 years

P 12

210 (25005600)
110 (18004000)
30 (5901600)
670 (4303000)
680 (170830)
36 % V expression
V low
Normal
IgG,A,M

3 months

P6

ND not done

* T-cell repertoire was evaluated either by analysis of TCRV expression as a percentage of the total CD3+ T population; or by CDR3 TCR spectratyping

24 months
12 months
Lymphocyte subsets (cells/L) (normal range of values for age)
300 (14003700)
723 (21006200)
170 (7002200)
389 (13003400)
420 (4901300)
194 (6202000)
470 (3901400)
311 (7202600)
660 (100480)
914 (160950)
ND
ND
ND
ND
Impaired
Low
Normal IgG, IgA, IgM
Normal IgG, IgM IgA

P8

0 months

P4

5 months

P3

P 10

Pre-transplant immunology in 13 patients with Cartilage Hair Hypoplasia

Age at Evaluation
9 years
5 months
Lymphocyte subsets (cells/L) (normal range of values for age)
CD3
270 (12002600)
1290 (25005600)
CD4
130 (6501500)
1250 (18004000)
CD8
70 (3701100)
20 (5901600)
CD19
50 (270860)
210 (4303000)
CD56
230 (100480)
300 (170830)
T cell repertoire*
Normal
Oligoclonal expansion
TRECs
Absent
Absent
Proliferative response to PHA
Absent
Impaired
IgG, IgA, IgM
IgG
IgG, IgA

Table 2

915 (21006200)
417 (13003400)
221 (6202000)
334 (7202600)
212 (180920)
ND
V low
ND
Normal

23 months

P 13

510 (19005900)
440 (14004300)
20 (5001700)
510 (6102600)
820 (160950)
48 % V expression
Low
Impaired
IgG,A,M

10 months

P7

J Clin Immunol (2015) 35:147157

151

152
Table 3

J Clin Immunol (2015) 35:147157

RMRP mutations and phenotype in 13 patients with Cartilage Hair Hypoplasia (CHH)

Patient

Ethnicity

Allele 1

Allele 2

Immunological
phenotype

Skeletal phenotypeb

Mutation references
(allele 1; allele 2)

Caucasian

20_-4 dup

4 C>T

CID

Mild features of CHH

at 3 years

Ridanpaa et al. 2001 [13];

Ridanpaa et al. 2002 [3]

Caucasian

25_-11 dup

4 C>T

CID+OS

Mild features of CHH

Ridanpaa et al. 2001 [13];

Ridanpaa et al. 2002 [3]

3
4
5
6

South Asian
South Asian
South Asian
African

35 C>A a
35 C>A a
35 C>A a
20_-14 dup

35 C>A a
35 C>A a
35 C>A a
70 A>G

CID
SCID
SCID
SCID

Mild features of CHH

Normal
CHH
Normal

Caucasian

151 G>Aa

70 A>G

CID

Skeletal survey not done

Ridanpaa et al. 2001 [13]

Caucasian

146 G>A

242 A>G

CID

Mild features of CHH

Kavadas et al. 2008 [16];

Bonafe et al. 2005 [22]

Caucasian

70 A>G

70 A>G

CID

CHH

Ridanpaa et al. 2001 [13]

10
11

Caucasian
South Asian

26_-14 dup
30 A>G

4 C>T
30 A>G

CID
CID

Atypical
Atypical

McCann et al. 2014 [12]

12
13

South Asian
Caucasian

28_-1dup
70 A>G

154 G>C
70 A>G

CID
CID

Atypical
CHH

Ridanpaa et al. 2001 [13]

Ridanpaa et al. 2002 [3];

Ridanpaa et al. 2001 [13]

Bold are new mutations

CID Combined Immunodeficiency, OS Omenns syndrome
a

Mutations not previously reported but nucleotides 35 and 151 are highly conserved between species

This refers to phenotype recorded at last follow-up

P11 carried homozygous nucleotide substitution 30A>G in

the coding region, a mutation that has not yet been described
but lies within a block of conserved sequence in the P3
domain.
Hematopoietic Stem Cell Transplantation (Table 4)

(P3, P4, P5, P6, P7 and P8) were conditioned with

fludarabine, melphalan and serotherapy with Alemtuzumab
(or ATG in P8) except for P5 who received matched sibling
bone marrow graft. Three patients (P2, P10, P11) were conditioned with combination of Treosulfan fludarabine and
Alemtuzumab.
Chimerism & Immune Reconstitution (Table 5)

A total of 17 allogeneic procedures were performed on 13

patients. The median age at transplant was 32.4 months (range
1.5 to 125 months). The sources of stem cells were bone
marrow (n=9), peripheral blood stem cells (PBSC) (n=4)
and cord blood (n=2). Two patients received second procedures after poor T-cell immune reconstitution following unconditioned cord blood infusion from a matched sibling donor
(P5); and after failure of engraftment following cord blood
transplant with low cell dose (P6). Two patients (P9 and
P11) had second CD34+ stem cell selected procedures for
red cell aplasia. Donor grafts were derived from 1, 2, or 3
antigen-mismatched unrelated donor (n=7), fully matched unrelated donor (n=3), fully matched family donor (n=1) and
matched sibling donor (n=4). Conditioning regimen differed
slightly between the two centres. Three patients (P9, P12,
P13) received conditioning with Cyclophosphamide in combination with Busulfan or Treosulfan but none of the other
patients had full myeloablative conditioning. Six patients

Eleven patients were alive at median follow-up of

50 months (range 21.6 to 168 months), all of whom achieved
stable full or high level donor chimerism. Two patients (P2
and P5) were mixed chimeras with high levels of T-cell donor
chimerism (79 and 73 %, respectively) but low levels of myeloid (0 and 17 %, respectively) and B-cell (40 and 24 %,
respectively) chimerism. P8 is also a mixed chimera but split
chimerism is not available.
All of the surviving patients have recovered normal numbers for age range of lymphocyte, CD3, CD4, CD8, CD19 and
CD16/56 counts post SCT; as well as normal numbers and
proportions of nave T cells. Levels of TRECs were normal
in 4 patients. T cell repertoire as assessed by spectratyping was
performed on 5 out of 10 patients and all had normal or mostly
normal Gaussian distribution of all V-beta families in T cells.
T-cell function measured by PHA response was tested in 10

CSA/MTX
BM
100 %

A&W
30 m

P9

48 m 72 m (top-up)
7/10 MMUD
B/Cy
CSA
BM
100 %

A&W
168 m

GvHD prophylaxis
Stem cell source
Chimerism

Outcome
Last follow-up from tx

P8

125 m
10/10 MUD
F/M/A
CSA/MP
BM
53 %WB

A&W
71 m

A&W
142 m

CSA/MMF
BM
100 %

F/M/C

5m
10/10 MFD

P3

A&W
90 m

Unconditioned (1st)
F/M (2nd)
CSA/MMF(2nd)
Cord (1st) BM (2nd)
27 % WB 70 % CD3
18 % CD15 25 %
CD19
A&W
21.6 m

1.5 m (1st) 20 m (2nd)

MSD (1st) MSD (2nd)

P5

A&W
48 m

28.8 m 38.4 m (top-up)

9/10 MMUD
T/F/C
CSA/MMF
BM
100 %

P 11

CSA/MMF
PBSC
100 %

F/M/C

36 m
9/10 MMUD

P4

Died 1 m post

42 m
9/10 MMUD
T/Cy/C
CSA/MMF
BM
100 %

P 12

A&W
24 m

CSA/MMF
Cord (1st) PBSC (2nd)
100 %

8 m (1st) 23 m (2nd)
8/10 MMUD (1st) 9/10
MMUD (2nd)
T/F (1st) F/M/C (2nd)

P6

32.4 m
MSD
B/Cy/C
CSA
BM
100 % CD3 91 %
CD15 100 % CD19
A&W
108 m

P 13

Died 9 m post

CSA/MMF
PBSC
100 % WB 100 %
CD3 100 % CD15

F/M/C

16.8 m
10/10 MUD

P7

Cyclophosphamide 200 mg/kg

ATG 15 mg/kg

Campath 1 mg/kg

Melphalan 140 mg/m2

Treosulfan 42 g/m2

Fludarabine 160 mg/m2

Busulfan 16 mg/kg

Doses of conditioning regimen (total dose)

MUD Matched unrelated donor, MSD Matched sibling donor, MMUD Mismatched unrelated donor, MFD Matched family donor, A Anti-thymocyte globulin, F Fludarabine, M Melphalan, C Campath, B
Busulfan, T Treosulfan, Cy Cyclophosphamide, CSA Cyclosporin, MTX Methotrexate, MMF Mycophenolate Mofetil, MP Methylprednisolone, BM Bone marrow, PBSC Peripheral blood stem cell, WB
whole blood, A&W alive and well

36 m
10/10 MUD
T/F/C
CSA/MMF
PBSC
99 % CD3 87 %
CD15 95 % CD19
A&W
24 m

P 10

CSA/MMF
BM
22 % WB 79 %
CD3 0 % CD15
40 % CD19
A&W
50 m

T/F/C

B/F

Conditioning

8m
9/10 MMUD

P2

133 m
MSD

P1

Haematopoietic stem cell transplant characteristics in 13 patients with Cartilage Hair Hypoplasia

Age at transplant
Donor

Table 4

J Clin Immunol (2015) 35:147157

153

ND not done

50 m
2190
1280
730
520
150
Normal
Low at 7 m post tx
Normal
Normal
Normal
+ ND

1250
740
440
330
370
Normal
Normal
Normal
Normal
Borderline low IgG, A, M

+ ND

P2

30 m

P1

Post transplant immunological evaluation in surviving patients

Time post BMT

Lymphocyte subsets (cells/L)
CD3
CD4
CD8
CD19
CD56
T cell repertoire (spectratype)
TRECs
Nave T cells
Proliferative response to PHA
IgG, A, M
Vaccine response to
-Tetanus
-Hib

Table 5

+
+

3200
1390
1430
340
550
Near Normal
Low 2y9m post tx
Normal
Normal
Normal

142 m

P3

+
+

3630
1590
1820
500
270
Normal
Normal
Normal
Normal
Normal

90 m

P4

3660
2000
1120
780
390
Normal
Normal
Normal
Normal
Normal

21.6 m

P5

ND
ND

2350
1840
430
110
230
ND
Low at 6 m post tx
Normal
ND
Normal

24 m

P6

+
+

860
430
300
120
200
ND
Normal
ND
Normal
Normal

71 m

P8

+
+

848
475
354
226
57
ND
Low
ND
Normal
Low IgA

168 m

P9

ND
ND

1861
1296
459
411
920
ND
Low
Normal
Normal
Normal

24 m

P 10

+
+

3898
1974
1791
543
187
ND
Normal
Normal
Normal
Normal

48 m

P 11

+
+

1540
768
680
212
146
ND
Low
Normal
Normal
Normal

108 m

P 13

154
J Clin Immunol (2015) 35:147157

J Clin Immunol (2015) 35:147157

patients and was normal in all. The majority of patients

regained normal B cell immunity with 9 of the 11 surviving
patients having normal levels of serum immunoglobulins and
off replacement therapy. P1 had borderline low levels of IgG,
IgA, and IgM but was able to demonstrate vaccine response
with good levels of tetanus antibodies. All 9 patients who had
vaccine responses tested all demonstrated protective levels of
antibodies to tetanus and/or Haemophilus influenzae type B.
Complications Post HSCT
Viral infections and GVHD were the most common transplant
related complications. 1 patient (P3) had resistant CMV reactivation in the blood and was treated with anti-viral drugs as
well as CMV-specific CTLs. This patient also developed
blood sepsis with Streptococcus oralis and Enterococcus
faecalis and required intensive care. 1 patient (P4) had both
EBV and Adenovirus reactivation, along with Grade 3 gut
GVHD associated with pneumatosis intestinalis and was treated with steroids. P5 also developed EBV reactivation and was
treated with Rituximab. P6 had chronic skin GVHD grade 2.
P7 developed adenoviraemia on day +18 and was treated with
Adenovirus-specific CTLs with good effect. However he developed encephalopathy on day +41, a novel strain of human
Astrovirus was identified on a brain biopsy manuscript in
press, [34]. The patient died 9 months post-transplant from
Pseudomonas pneumonia. P12 developed pulmonary haemorrhage and multiorgan failure and died on day +32.

Discussion
This cohort of patients with RMRP mutations demonstrates
that although CHH may typically present with distinctive phenotypic features of skeletal dysplasia, disproportionate short
stature, hypoplastic hair and associated immunological deficiency, in the immunology clinic the phenotype can be highly
variable and may include severe immune deficiency with or
without associated short stature. [9, 16, 2224]. Importantly,
characteristic metaphyseal changes seen in CHH may not be
present at birth and often manifest after 2 years of age [8,
2527]. In this cohort, 5 subjects were transplanted in infancy
before the onset of radiological changes and only 7 out of 13
patients had evidence of characteristic radiological features
such as metaphyseal dysplasia and bowing of femora. Remarkable clinical heterogeneity may even occur within
sibships [9, 16], and we found that of three siblings in our
cohort with identical homozygous mutations in the RMRP
gene, only two had clinical and radiological skeletal manifestations of CHH. Interestingly newborn screening of a cousin
from this consanguineous kindred uncovered a further subject
with identical homozygous mutations but no evidence of either immune deficiency or skeletal dysplasia.

155

The degree of immunodeficiency reported in CHH patients

is also highly variable but predominantly affects cell-mediated
immunity, whilst humoral immunity is often intact [9, 10, 23,
28], although in this cohort 6 patients had B cell lymphopenia
and 2 were panhypogammaglobulinaemic.. The most severely
affected infants may present with features of SCID with characteristic TB+/NK+ immunological phenotype which may
present as Omenns syndrome whereby leaky autoreactive
T cell clones mediate skin and gut inflammatory complications. This degree of immunodeficiency can certainly be detected by SCID newborn screening by measuring levels of Tcell receptor excision circles (TREC). Indeed, over a 2 year
period in California where 993,724 infants were screened, 15
were diagnosed with SCID of whom 1 had RMRP mutation
[29].
The RMRP locus is non-protein coding but in vitro testing
of RNase MRP mRNA and rRNA cleavage suggests that reduction in mRNA cleavage is associated with milder skeletal
phenotypes but more pronounced immunodeficiency, haematological abnormalities, as well as hair hypoplasia; in contrast
RMRP mutations that decrease rRNA cleavage in ribosomal
assembly are strongly correlated with the degree of bone dysplasia [15, 30].
In this cohort we report 83 % survival with broad normalisation of humoral and cell-mediated immunity following successful transplantation. All (eventually) received conditioning, including reduced intensity regimens with serotherapy.
In exceptional circumstances, faced with a severely unwell
infant and the availability of a fully matched sibling donor, it
may be possible to infuse bone marrow into patients with
RMRP mutations, but with the expectation that a second, definitive procedure will be needed. Given the T-/lowB+NK+ phenotype, pre-conditioning is generally warranted due to the
likelihood of residual host immunity and importance of ensuring full donor chimerism. In this cohort, all survivors have
stopped receiving immunoglobulin replacement therapy and
have been vaccinated and are living unrestricted lifestyles.
Longer term follow up will be required to confirm full reconstitution of immunity.
One particular concern is the 7 fold risk of malignancy
compared to an age-adjusted expected incidence reported in
(non-transplanted) Finnish CHH patients. Non-Hodgkins
Lymphoma and squamous cell carcinoma, are most common
with the majority of patients being diagnosed between the
ages of 1544. [31, 32]. One patient transplanted in the UK
not included in this current cohort as molecular diagnosis was
not confirmed, but previously reported [18] had clinical features of CHH and received allo-SCT after developing EBVassociated diffuse large B-cell Non-Hodgkins lymphoma at
the age of 11 years. The extent to which allo-SCT mitigates
against the risk of haematological malignancy is unknown,
but in the case of full donor chimeras, any intrinsic transformational risk of haematological cells is presumably abrogated.

156

A number of subjects have mixed myeloid or B cell chimerism, and here it may be that donor derived T cell immune
surveillance is sufficient to control transformation events. Experience from other disorders, including Fanconi anaemia [33]
suggests that the risk of malignancy in other organ systems,
including the skin, may persist.
In summary we report the UK transplant experience for
CHH and include a group of subjects who presented with
short stature and immunodeficiency as predominant features
rather than typical CHH skeletal changes. We advocate that
RMRP mutation screening is considered in any child with
undefined combined immunodeficiency even in the absence
of skeletal features.
Acknowledgments The authors would like to thank Kerra Pearce and
Mike Hubank (UCL Genomics) for their assistance with microarrays and
NGS work.
Funding Source This work was undertaken at GOSH/UCL Institute of
Child Health which received funding from the Department of Healths
NIHR Biomedical Research Centres funding scheme and GOSH charity
special trustees. The UK service for primary immunodeficiency received
support from the Department of Health via the national commissioning
group (NCG).
Financial Disclosure Winnie Ip has no financial relationships relevant
to this article. All the other authors also have no financial disclosures
relevant to this article.
Conflict of Interest Winnie Ip has no conflict of interest to disclose. All
the other authors also have no conflicts of interest relevant to this article to
disclose.

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