Assignee: 3i Research Exploitation Limited (London, GB2)
Appl. No.: 478045
Filed: June 7, 1995
Assistant Examiner: Grun; James L.
Attorney, Agent or Firm: Nixon & Vanderhye
United States Patent 5,605,843 February 25, 1997
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Riverbend DS Assocation Home Page » Resources » Patents » Prenatal Screening » Kit for Prenatal Screening for Down's Syndrome Kit for Prenatal Screening for Down's Syndrome |
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Inventors: Canick; Jacob A. (Newton, MA); Wald; Nicholas J. (London, GB); Cuckle; Howard S. (London, GB); Haddow; James E. (Sebago Lake, ME) Assignee: 3i Research Exploitation Limited (London, GB2) Appl. No.: 478045 Filed: June 7, 1995 |
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Primary Examiner: Green; Lora M. Assistant Examiner: Grun; James L. Attorney, Agent or Firm: Nixon & Vanderhye United States Patent 5,605,843 February 25, 1997 |
Parent Case Text
This is a division of application Ser. No. 07/908,875, filed Jul. 7, 1992 now U.S. Pat. No. 5,506,150, which is a continuation of 07/457,687, filed as PCT/GB00557 on Jul. 11, 1988, now abandoned.
Claims
We claim:
1. An assay kit for a sample of serum from a pregnant
woman at a stage before the beginning of the third trimester of pregnancy to
determine if a pregnant woman is carrying a fetus having an increased risk of
Down's syndrome, said kit consisting of:
an immunoassay means to
determine a level of unconjugated oestriol in said sample,
an
immunoassay means to determine a level of alpha-fetoprotein in said sample, and
an immunoassay means to determine a level of human chorionic
gonadotrophin in said sample.
2. An assay kit for a sample of serum from
a pregnant woman at a stage before the beginning of the third trimester of
pregnancy to determine if a pregnant woman is carrying a fetus having an
increased risk of Down's syndrome, said kit consisting of:
an
immunoassay means to determine a level of unconjugated oestriol in said sample,
an immunoassay means to determine a level of dehydroepiandrosterone
sulfate in said sample,
an immunoassay means to determine a level of
alpha-fetoprotein in said sample, and
an immunoassay means to determine
a level of human chorionic gonadotrophin in said sample.
3. An assay kit
for a sample of serum from a pregnant woman at a stage before the beginning of
the third trimester of pregnancy to determine if a pregnant woman is carrying a
fetus having an increased risk of Down's syndrome, said kit consisting of:
an immunoassay means to determine a level of dehydroepiandrosterone
sulfate in said sample,
an immunoassay means to determine a level of
alpha-fetoprotein in said sample, and optionally
an immunoassay means to
determine a level of human chorionic gonadotrophin in said sample.
Description
BACKGROUND OF THE INVENTION
This invention relates to antenatal
screening for risk of Down's Syndrome in a foetus.
Risk of Down's
syndrome in a foetus is known to increase with the age of the mother and it is
this fact which forms the basis for selection of mothers for further
investigation. Further testing involves sampling of the amniotic fluid by
amniocentesis, a procedure which is not completely free from risk to the mother
or foetus, induction of miscarriage being a recognised hazard.
The
amniotic fluid test for Down's syndrome (the recognition of an extra chromosome
21 in foetal cells) is diagnostic.
There is a need for a screening
method which will identify pregnancies most at risk so as to justify
amniocentesis with its attendant risks.
An object of the present
invention is to provide an improved screening procedure.
[...]
DESCRIPTION OF THE INVENTION
According to the present invention
there is provided a method of screening for Down's syndrome in a foetus,
comprising assaying a sample of maternal serum, taken before the beginning of
the third trimester of pregnancy, for at least one serum constituent selected
from;
unconjugated oestriol;
progesterone;
16-alpha-hydroxy-dehydroepiandrosterone sulphate
(16-alpha-hydroxy-DHEAS);
dehydroepiandrosterone sulphate (DHEAS), and
precursors or metabolites thereof,
the results of said assay being
indicative of increased risk of foetal Down's Syndrome.
The method may
also include an additional assay for alpha-fetoprotein.
It is preferred
that the method include a further assay for human chorionic gonadotrophin (hCG).
In a particularly preferred method three assays are carried out for
alpha-fetoprotein (AFP), unconjugated oestriol and human chorionic
gonadotrophin.
In another preferred embodiment the method includes
additional assays for either or both of
16-alpha-hydroxy-dehydroepiandrosterone sulphate
(16-alpha-hydroxy-DHEAS); and,
dehydroepiandrosterone sulphate (DHEAS)
In deriving the risk indicator the assay results may be interpreted in
combination with maternal age.
The present invention also provides an
assay kit comprising means for conducting the assays for metabolites identified
above and means for producing from the said assay results an indication of
increased risk of foetal Down's syndrome.
This invention is based on the
fact that serum AFP, and unconjugated oestriol (uE.sub.3) in maternal blood in
early pregnancy are significantly lower in affected pregnancies than unaffected
pregnancies, and therefore their precursors including 16-alpha-hydroxy DHEAS and
DHEAS are also affected, and serum progesterone and human chorionic
gonadotrophin are both greater in affected pregnancies than in unaffected
pregnancies.
The abnormal unconjugated oestriol, progesterone and AFP
levels in Down's syndrome pregnancies reflect abnormal synthesis metabolism in
the foetus and/or placenta. FIG. 1 of the accompanying drawing sets forth the
metabolic pathways relating to the biosynthesis of oestriol (and progesterone)
during pregnancy, the oestriol biosynthesis route being shown in bold type. The
abbreviations DHEAS means dehydroepiandrosterone sulphate and 16-OH DHEAS means
its 16-hydroxy-derivative.
AFP is manufactured by the yolk sac and the
foetal liver.
Thus, as shown in FIG. 1, oestriol, derived from
cholesterol of maternal origin, is converted, via pregnenolone and DHEAS to
16-hydroxy-DHEAS which is then converted by placental activity to oestriol which
returns, in unconjugated form, into maternal serum where, subsequently, it is
converted to oestriol sulphates and glucuronides.
Progesterone arises by
placenta activity which converts cholesterol to pregnenolone and hence to
progesterone, a portion of the pregnenolone being passed to the foetus, for
conversion to oestriol by the route described above, and the remainder returned
to maternal serum.
The median maternal serum unconjugated oestriol level
at 13 to 27 weeks gestation in seventy-seven pregnancies associated with Down's
syndrome was 0.73 multiples of the median (MoM) value for 385 unaffected control
pregnancies matched for maternal age, gestational age, and duration of storage
of the serum sample. This result was statistically highly significant. Serum
unconjugated oestriol is a better discriminator between Down's syndrome and
unaffected pregnancies than maternal age or serum AFP. The extent of the
discrimination was also independent of maternal age and largely independent of
serum AFP. When used in combination, the three variables, age, serum AFP and
serum unconjugated oestriol, were more effective than any one by itself or two
in combination at identifying Down's syndrome. The inclusion of serum
progesterone leads to significant increase of the effectiveness of the
detection.
The median maternal serum progesterone level at 13 to 27
weeks gestation in 77 pregnancies associated with Down's syndrome was 1.19
multiples of the median value (MoM) for 385 unaffected control pregnancies of
the same gestational age. The increase in values was statistically significant
(X.sub.1.sup.2 =14, based on analysis of variance of the ranks of matched sets).
There was more overlap in the distribution of progesterone between affected and
unaffected pregnancies than was the case for unconjugated oestriol.
In
normal pregnancy the foetal adrenal cortex produces DHEAS which enters the
foetal circulation and passes to the foetal liver where most of it undergoes
16-alpha-hydroxylation. The newly formed 16-alpha-hydroxy-DHEAS reaches the
placenta where a portion of it is converted in four enzymatic steps to oestriol.
Unconverted 16-alpha-hydroxy-DHEAS is also likely to pass into the mother's
circulation. It is possible that in maternal serum almost all of the
16-alpha-hydroxy-DHEAS is derived from the foetus rather than from the mother,
and therefore measurement of this substance is a specific marker of foetal
development.
Down's syndrome foetuses are less mature than unaffected
foetuses and so pregnancies associated with foetal Down's syndrome would be
expected to have lower 16-alpha-hydroxy-DHEAS levels than unaffected
pregnancies. It is by similar reasoning that lower levels of DHEAS will be found
in the maternal blood of women carrying foetuses with Down's syndrome than in
those carrying unaffected foetuses.
Therefore, either or both of DHEAS
and 16-alpha-hydroxy-DHEAS provide useful markers of Down's syndrome in early
pregnancy, particularly when the results are viewed in combination with the
other indicators referred to herein.
The method of the present invention
increases the efficiency of screening of women for Down's syndrome by
identifying a greater proportion of women with affected pregnancies as eligible
for amniocentesis (i.e. increasing the proportion of true positives) without
also increasing the proportion of women with unaffected pregnancies who, by some
criteria, would be eligible for amniocentesis (i.e. without increasing the
proportion of false positives).
[...]
A way of combining information on median serum AFP and median
serum unconjugated oestriol levels which gives equal weight to both tests is to
regard a screening result as positive if either level is less than or equal to
the same percentlie of normal. Using this approach, for a given number of
affected pregnancies with positive results there were fewer unaffected
pregnancies with positive results when both tests were used than when each test
was considered alone (Table V).
TABLE V
______________________________________
False-positive rate (%) using
Downs Age,
Syndrome
Age AFP uE.sub.3
Age & Age & AFP
Detectn.
alone alone alone AFP uE.sub.3
and uE.sub.3
(%) (a) (b) (c) (d) (e) (f)
______________________________________
60 31 33 28 20 14 12
55 25 28 19 16 11 9.1
50 19 24 16 12 8.8 7.0
45 15 20 13 9.8 6.7 5.3
40 11 17 11 7.3 5.0 3.9
35 7.5 14 8.5 5.3 3.7 2.8
30 5.1 11 6.6 3.6 2.5 1.9
25 3.3 8.2 5.0 2.2 1.6 1.2
20 1.9 6.0 3.5 1.3 0.9 0.7
______________________________________
Note:
A result is positive if (a) age is advanced, (b) AFP is low, (c) uE.sub.3
is low or, for (d), (e) and (f), the risk of Downs Syndrome (at term) is
high.
TABLE VIII
______________________________________
Down False-positive rate (%) using
Syndrome age with:
Detection AFP AFP uE.sub.3
Rate and and and AFP, uE.sub.3
(%) AFP uE.sub.3
hCG uE.sub.3
hCG hCG and hCG
______________________________________
80 44 34 27 29 20 20 16
75 37 27 21 23 15 15 12
70 30 22 16 18 12 11 8.6
65 25 18 12 15 8.8 8.1 6.4
60 20 14 9.5 12 6.7 6.0 4.7
55 16 11 7.2 9.1 5.0 4.4 3.4
50 12 8.8 5.4 7.0 3.7 3.2 2.5
45 9.7 6.7 3.9 5.3 2.7 2.3 1.7
40 7.3 5.0 2.8 3.9 1.9 1.6 1.2
35 5.3 3.7 1.9 2.8 1.3 1.0 0.8
30 3.6 2.5 1.2 1.9 0.8 0.7 0.5
25 2.2 1.6 0.8 1.2 0.5 0.4 0.3
20 1.3 0.9 0.4 0.7 0.3 0.2 0.2
______________________________________
TABLE IX
______________________________________
Without DHEAS With DHEAS
Meas d Risk % Det PPV d Risk % Det PPV
______________________________________
AEHP 1.771 232 63.6 62 1.795
232 64.3 61
AEH 1.680 227 60.6 65 1.700
228 61.3 64
EHP 1.601 226 58.1 68 1.632
226 59.1 67
AEP 1.585 225 57.5 68 1.619
226 58.7 67
EH 1.539 225 56.1 70 1.565
225 56.9 69
AHP 1.517 225 55.4 71 1.546
225 56.3 70
AH 1.493 225 54.6 72 1.519
225 55.5 71
EP 1.390 227 51.4 76 1.434
226 52.8 75
HP 1.272 231 47.9 82 1.312
229 49.1 80
H 1.268 231 47.8 82 1.305
229 48.8 80
AE 1.159 237 44.7 88 1.193
235 45.6 86
AP 1.080 243 42.6 92 1.133
239 44.0 89
E 1.006 249 40.7 96 1.051
246 41.9 94
A 0.703 285 34.7 113 0.767
277 35.8 110
P 0.656 291 34.0 116 0.753
278 35.5 111
DHEAS -- -- -- -- 0.333
321 30.7 128
______________________________________
d = distance, in standard deviation units, between affected and unaffecte
groups.
Risk is quoted as a ratio of 1: the figure in the Table.
PPV = positive predictive value and is an indicator of the odds of the
actual occurrence of Down's Syndrome in the foetus. It is quoted as a
ratio of 1: the figure in the Table.
[...]
| Source: http://www.uspto.gov/patft/ | |
| Revised: February 14, 2001. | |