Letters to the Editor 1235

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8 Niiya M, Uemura M, Zheng XW, Pollak E, Dockal M, Scheiinger F,

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Late onset thrombosis in a case of severe protein S deficiency

due to compound heterozygosity for PROS1 mutations
M . J . H E E B , * S . G A N D R I L L E , J . A . F E R N A N D E Z , * J . H . G R I F F I N * and P . F . F E D U L L O
*Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA, USA; INSERM Unite de Recherche 765,
Paris; AP-HP Hopital Europeen Georges Pompidou, Service dHematologie Biologique A, Paris; Universite Paris Descartes, Unite de Formation et
de Recherche des Sciences Pharmaceutiques et Biologiques, Paris, France; and Pulmonary and Critical Care Medicine, University of California at
San Diego Medical Center, San Diego, CA, USA

To cite this article: Heeb MJ, Gandrille S, Fernandez JA, Griffin JH, Fedullo PF. Late onset thrombosis in a case of severe protein S deficiency due to
compound heterozygosity for PROS1 mutations. J Thromb Haemost 2008; 6: 12357.

Plasma protein S is an anticoagulant whose heterozygous

genetic deciency is associated with increased risk of venous
thrombosis and juvenile stroke [13]. Several families have been
described with individuals who were homozygous or compound heterozygous for protein S defects and severely protein
S-decient. These families were identied from Thailand,
Japan, China and Turkey, and they include two AfricanAmerican children. Nine individuals presented as neonates
with life-threatening thrombotic purpura fulminans, and
required life-long treatment with a source of protein S and/or
anticoagulants (heparin or warfarin) [48]. Yet remarkably,
several other severely protein S-decient individuals did not
experience a serious thrombotic event until ages of 1020 years
Correspondence: Mary Jo Heeb, Department of Molecular and
Experimental Medicine MEM180, The Scripps Research Institute,
10550 N. Torrey Pines Rd., La Jolla, CA 92037, USA.
Tel.: +1 858 784 2185; fax: +1 858 784 2113.
E-mail: heeb@scripps.edu
DOI: 10.1111/j.1538-7836.2008.02994.x
Received 1 April 2008, accepted 16 April 2008
 2008 International Society on Thrombosis and Haemostasis

old, and one individual with 18% total protein S antigen was
asymptomatic at age 23 years [2,9,10]. Thrombosis in these
young adults was recurrent, including deep vein thrombosis,
pulmonary embolism, mesenteric vein thrombosis, sagittal
sinus thrombosis, and ankle ulcerations. Here we describe
another severely decient individual in an attempt to advance
understanding of the variable phenotypes.
The propositus, a 23-year-old Filipino male in California,
experienced multiple venous thrombosis episodes, beginning
with lower limb thrombosis at age 10. Record of duration of
anticoagulant treatment following that event was unavailable.
At age 20, he underwent ileal resection and ileostomy as a result
of mesenteric vein thrombosis. He was treated sequentially with
heparin, warfarin, and then subcutaneous heparin. At age 21,
he experienced a small left fronto-parietal subcortical hemorrhage following head trauma while on 2000 units of heparin
subcutaneously daily. He recovered without signicant neurologic residua. Physical examination at age 23 was remarkable
only for scars from his ileal surgery and for chronic bilateral
punctuate erythematous lesions on the dorsum of each foot.
Subcutaneous heparin (7500 units every 12 h) was resumed
until a physician could be located near his home city to
administer warfarin under close supervision.

1236 Letters to the Editor

Table 1 Characteristics of the protein S-deficient family

I-1*
I-2
II-1
II-2
II-3
II-4
II-5

Sex/age

Cofactor
activity

% PS total

M/23
F/37
M/12
M/11
F/5
M/3
F/1.2

<7
28
42
38
128
32
26

10
52
67
70
125
47
70

17
8
8
2
4
8

2
10
11
8
19
4
7

% PS free
0.2
5.4
12
16
>78
13
9

0
1
2
9
1
2

% PS free**

PS-direct

%
C4BP

APC
resistance

2
16
27
27
109
15
18

1.12
1.24
NT
1.30
1.73
NT
NT

74
95
127
82
117
64
140

1.06
1.20
0.82
1.10
1.01
NT
0.96

*Propositus. Not tested. Staclot S kit, Diagnostica Stago. Elisa of plasma [13]. Elisa of polyethylene glycol supernatant of plasma [13].
**Asserachrom free protein S antigen Elisa, Diagnostica Stago. Ratio of factor Xa-1-stage clotting times determined without/with anti-protein S
antibodies [13]; normal range, 1.31.9. APC (activated protein C) resistance: normalized ratio of APTT determined in the presence/absence of
APC for test plasma diluted in factor V-decient plasma; normal range, 0.81.2 [14].

The patient and his sister reported that several family

members residing in the Philippines died of cardiovascular
events, including their mother at age 50, a sister at age 35, and a
brother at age 20. Two sisters, ages 31 and 29 years at the time
of referral, had experienced recurrent venous thrombosis and
were receiving warfarin therapy. Another sister, age 38, and a
brother, age 25, residing in the Philippines, had no history of
thromboembolic disease. A sister in California was asymptomatic at the time of the study (at age 37), but she later
suffered recurrent venous thrombosis and is now receiving
warfarin.
Before heparin treatment was resumed for the propositus,
blood samples anticoagulated with trisodium citrate were
taken from him, his sister in California, and this sisters ve
children. Informed consent was obtained. The patients
brinogen level was 177, hemoglobin/hematocrit 14.9/45.1,
sedimentation rate 1, white blood count 7.4, platelet count
280 000, prothrombin time 12.1/11.6 s, APTT 27.9/26.1 s,
factor X 115%, antithrombin 92%, and protein C 108%. The
patients sister had factor X and protein C levels of 99% and
95%, respectively.
Multiple protein S-related parameters were obtained
(Table 1). The propositus (I-1) had barely detectable levels
of free protein S or of anticoagulant cofactor activity for
activated protein C, while his sister (I-2) and four of her ve
children had low levels of each of these. Total protein S was
10% of normal for the propositus, while his sister and two of
her children had low total protein S, and two of her children
had low-normal levels (normal range, 70130%). Protein S
anticoagulant activity in the absence of activated protein C
(termed PS-direct) was consistent with other protein S
parameters. The APC/protein C activity ratio was twice the
normal level only for the propositus, possibly indicating
above normal levels of thrombin generation and protein C
activation, consistent with a mildly elevated level of thrombin-antithrombin complexes (5.6 lg L)1). The normalized
APC resistance ratio was low-normal for subject II-1, for
reasons that were not apparent.
To identify mutations in the PROS1 gene, DNA was
sequenced from all subjects except II-5, for all exons and exon/
intron junctions and for part of the promoter region [11]. A
novel mutation was identied in one allele of all subjects who

had low free protein S, namely a deletion of 14 base pairs in

intervening sequence L, )10 to )24 upstream of exon 13.
This mutation probably impairs splicing of PROS1 premRNA, which usually requires a cis-acting element 1030
bases upstream from the acceptor site. The mutation is in the
putative branch point of intron L.
The propositus, but not the other subjects studied, carried a
second novel mutation, namely a CT mutation at )168 in the
promoter region, in an Sp1 binding site. This mutation is in a
promoter region that is highly conserved among species, and is
within an indispensable Sp1 transcription regulation site
located at nt )178 to )165 [12]. Presumably this compound
heterozygosity for PROS1 mutations caused the observed
severe deciency and recurrent thrombosis in the propositus.
The propositus in this study is one of only a few known
individuals with severe protein S deciency who remained free
of thrombosis until age 10 or later and who survived into
adulthood. He may carry a genetic polymorphism(s) that
partly compensates for the increased thrombotic risk caused by
low protein S levels.
Acknowledgements
We gratefully acknowledge A. Gruber, Y. Montejano, M.-L.
Aubry and L. Tonnu for technical assistance, the NIH for
support from grants HL21544 and HL070002, and the Stein
Endowment Fund.
Disclosure of Conflict of Interests
The authors state that they have no conict of interest.
References
1 Schwarz HP, Fischer M, Hopmeier P, Batard MA, Grin JH. Plasma
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Hamostaseparameter bei 55 Patienten mit venosen und/oder arteriellen Thromboembolien. Schweiz Med Wochenschr 1989; 119: 4939.

 2008 International Society on Thrombosis and Haemostasis

Letters to the Editor 1237

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Severe protein S deficiency resulting from two novel mutations

in PROS1 presenting with a relatively mild clinical phenotype
I . S . R . C A R T E R , * J . H E W I T T , * C . H . P U , * J . K . W U , C . J . C A R T E R and R . T . A . M A C G I L L I V R A Y *
*Centre for Blood Research and Department of Biochemistry & Molecular Biology; Centre for Blood Research and Department of Pediatrics; and
Centre for Blood Research and Department of Pathology & Laboratory Medicine, University of British Columbia, Vancouver, BC, Canada

To cite this article: Carter ISR, Hewitt J, Pu CH, Wu JK, Carter CJ, MacGillivray RTA. Severe protein S deficiency resulting from two novel mutations
in PROS1 presenting with a relatively mild clinical phenotype. J Thromb Haemost 2008; 6: 12379.

Protein S (PS) is a vitamin K-dependent plasma glycoprotein that functions as a non-enzymatic cofactor for activated
protein C (APC) in the degradation of coagulation factors Va and VIIIa, thereby eliminating the prothrombinase
and tenase complexes, respectively [1,2]. In plasma, approximately 30% of PS circulates in a free form, and the
remainder is found complexed with C4b-binding protein;
only free PS functions as an anticoagulant [3]. The human
PS gene, PROS1, is located on chromosome 3p11.1-q11.2
[4], where it spans 80 kb of genomic DNA and contains 15
exons and 14 introns. Molecular analysis of PROS1 is
complicated by the presence of a homologous pseudogene
(PROS2) at 3p21-cen [4]. Hereditary PS deciency (PSD) is
an autosomal dominant disorder associated with a risk of
Correspondence: Ross T. A. MacGillivray, Centre for Blood Research,
University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
Tel.: + 1 604 822 3027; fax: + 1 604 822 4364.
E-mail: macg@interchange.ubc.ca
DOI: 10.1111/j.1538-7836.2008.03012.x
Received 11 April 2008, accepted 21 April 2008
 2008 International Society on Thrombosis and Haemostasis

recurrent thrombosis due to a failure to attenuate the

coagulation cascade [5]. Compound heterozygosity is rare
for PS, as such a genotype is usually incompatible with
adult life without treatment [6]. PS is a modular protein
composed of a c-carboxyglutamic acid domain, a thrombinsensitive linking region, four epidermal growth factor-like
domains, and a large domain similar to sex hormonebinding globulin (SHBG) at residues 243635 [4].
Proposita A (individual II-1 in Fig. 1A) is of Filipino
descent, and suffered from a superior and inferior sagittal
cerebral venous sinus thrombosis at age 14 months after
dehydration resulting from a viral syndrome. At that time, a
functional PS activity of 0.2 U mL)1 was measured using the
Staclot PS assay (Diagnostica Stago, Parsippany, NJ, USA),
with a resulting diagnosis of PSD. The patients twin sister,
father and paternal grandfather were also found to have PSD
(Fig. 1A). Since this early thrombosis in proposita A, neither
twin has experienced a further thrombotic event in the past
20 years. Proposita B (individual II-3 in Fig. 1B) is a Caucasian with Downs syndrome who was diagnosed with PSD (PS
functional activity of < 0.2 units mL)1) after she developed
an iliofemoral deep vein thrombosis following orthopedic
surgery at 13 years of age (for a family pedigree, see Fig. 1B).