Draft Genetic Test Review

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Cystic Fibrosis
Clinical Validity

(436KB)

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CLINICAL VALIDITY

Question 18: How often is the test positive when the disorder is present?
Question 19: How often is the test negative when the disorder is not present?
Question 20: Are there methods to resolve clinical false positive results in a timely manner?
Question 21: What is the prevalence of the disorder in this setting?
Question 22: Has the test been adequately validated on all populations to which it may be offered?
Question 23: What are the positive and negative predictive values?
Question 24: What are the genotype/phenotype relationships?
Question 25: What are the genetic, environmental or other modifiers?

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CLINICAL VALIDITY

Question 21: What is the birth prevalence of cystic fibrosis in the prenatal setting?

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Cystic fibrosis prevalence: Newborn screening trials

Cohorts identified as part of newborn screening trials (utilizing measurements of immunoreactive trypsinogen and/or DNA testing) can also be used to estimate the prevalence of cystic fibrosis. In many of these trials, extensive efforts were made to identify false negative results, in order to quantify screening performance. The following section presents cystic fibrosis prevalence estimates for non-Hispanic Caucasians, based on results from 17 newborn screening trials. Some published trials are excluded, because they either did not attempt complete ascertainment or did not sufficiently document the diagnostic criteria or population studied. Others were not published in English or were presented in proceedings of a meeting that are difficult to obtain.

Table 3-20 contains a brief description of the 17 selected studies, carried out in four geographical areas: the United Kingdom (UK), Australia/New Zealand, the United States, and Europe (not including the UK). For each study, the inclusive dates, the location, and the ethnic make-up of the study population are provided. In addition, three possible sources of bias in ascertainment are listed. The first two summarize the compliance with follow-up screening tests (i.e., the percentage of newborns with initially high immunoreactive trypsinogen [IRT] measurements from whom a second requested sample could be obtained) and with diagnostic tests (i.e., the percentage of newborns who received a sweat test, when indicated). The last column indicates whether or not the study was likely to under-ascertain newborn cases of cystic fibrosis, due to some of the affected pregnancies being identified prenatally and selectively terminated.

Taking non-compliance into account
In a two-step IRT screening protocol, the first measure of compliance is the proportion of those with screen positive results on the first sample who submit a second sample. The second measure of compliance is the proportion of those remaining screen positive who then receive diagnostic sweat testing. In 14 of the 17 studies in Table 3-20, compliance with the protocol was high. In the other three studies, compliance at the diagnostic or follow-up screening level was between 74 and 81 percent. If the risk of cystic fibrosis were the same among newborns whose parents did not comply with the complete testing protocol as among those who were compliant, then that factor could be taken into account. If IRT levels were lower among newborns who were not fully tested, however, the risk for cystic fibrosis in that group might well be lower. Although IRT levels were not provided on these studies for the non-compliant group, all three studies attempted to identify all cases, regardless of screening status and compliance. These efforts at complete ascertainment are considered to compensate for the non-compliance. For this reason, none of these studies are corrected for compliance.

Table 3-20. Demographic and Study-Related Characteristics of Newborn Cystic Fibrosis Screening Trials

Study Number	Location	Years	Race/Ethnicity	Possible Biases
				Compliance		Prenatal Diagnosis
				Screen	Diag
United Kingdom
1	Leeds	1975-94	NHC	NR	NR	Yes
2	East Anglia	1980-89	NHC	NR	NR	No
3	N. Ireland	1983-89	NHC	NR	NR	No
4	Wales/Midlands	1985-90	NHC	NR	100%	Yes
5	Trent	1989-94	NHC	99%	99%	Yes
Australia/New Zealand
6	New South Wales	1981-93	NHC	98%	NR	Yes
7	New Zealand	1983-86	NHC	NR	NR	No
8	Victoria	1987-93	NHC	NA	100%	Yes
9	Adelaide	1989-93	NHC	NA	100%	Yes
United States
10	Colorado	1982-87	Mixed	77%	100%	No
11	Wisconsin	1985-94	Mixed	NA	81/100%	Yes
12	W. Pennsylvania	1987-91	Mixed	NR	NR	Yes
Europe (w/o the United Kingdom)
13	Normandy, France	1980-82	NHC	NA	NR	No
14	Vienna, Austria	1988-91	NHC	NA	74%	Yes
15	Northeastern Italy	1988-91	NHC	100%	NR	Yes
16	France	1989-90	NHC	93%	NR	Yes
17	Brittany, France	1993-99	NHC	NA	100%	Yes

NHC = non-Hispanic Caucasians; Mixed = NHC and at least one other racial/ethnic group; NR = not reported; NA = not applicable; Diag = diagnosis

Taking mixed racial/ethnic populations into account All but the three studies from the United States appear to have been conducted in populations that were nearly exclusively composed of non-Hispanic Caucasians. Since other races (along with Hispanic Caucasians) are reported to have lower prevalences of cystic fibrosis, it is important to take this factor into account. In two of the U.S. studies (Hammond et al., 1991; Gregg et al., 1993), cystic fibrosis prevalence was reported for both the entire population and for Caucasians alone. However, neither study analyzed prevalences separately in the Hispanic or Asian populations. These two groups are estimated to have much lower prevalences (about 1:9000 and 1:30,000, respectively). The third study reported that both Blacks and other racial/ethnic groups were included but provided no separate estimates. None of the three studies provided racial/ethnic breakdowns for the affected and unaffected populations. The present analysis deals with this issue by deriving estimates of the racial/ethnic distribution for each study from the National Center for Health Statistics (National Center for Health Statistics 1989; 1993), using a one year time period near the middle of the study. In this analysis, the total number of non-Hispanic Caucasians is derived by multiplying the total number of study subjects by the proportion estimated to be non-Hispanic Caucasian. Then, the number of newborns with cystic fibrosis is reduced by the number of cases expected in other racial/ethnic groups. This is estimated by computing the number of newborns for the other racial/ethnic groups (Black, Hispanic Caucasian and Asian) and then multiplying that number by the appropriate prevalences.

Taking the impact of prenatal diagnosis into account Of the 17 newborn screening trials summarized in Table 3-21, 12 (71 percent) were active during the time period when prenatal diagnosis was possible (1988 or later). The effect of prenatal diagnosis on the birth prevalence of cystic fibrosis depends on the percentage of pregnancies terminated after identification of a fetus with two mutations. Of these 12 studies, three from Australia/New Zealand (Ranieri et al., 1994; Balnaves et al., 1995; Wilcken et al., 1995) and one from France (Scotet et al., 2000) estimated the impact of prenatal diagnosis on the birth prevalence. One study provided the rate by year from 1987 through 1993 (Balnaves et al., 1995). Overall, these trials identified 80 prenatal diagnoses and selective terminations of fetuses with cystic fibrosis in the cohort who would otherwise have been tested as newborns. Three of these trials were active only after 1988; the fourth (Wilcken et al., 1995) screened roughly half of the newborns after 1988. Dividing the total number screened in these four trials by the number of prenatal diagnoses can provide a rough approximation of the impact of prenatal diagnosis for those studies not providing such data. Based on this, approximately one prenatal diagnosis can be expected for every 16,800 newborns tested (1,258,381 / 80). Studies reporting results since 1988 that did not account for prenatal diagnosis are corrected by this factor. Under this assumption, a study that tested 168,000 newborns since 1988 would have identified 10 additional newborns with cystic fibrosis, had prenatal diagnosis not been available. If studies were active both before and after 1988, the number of newborns tested after 1988 is estimated assuming a uniform recruitment rate (unless yearly recruitment was provided). It is possible that prenatal diagnosis is less common in the United States and this correction will over estimate the prevalence.

Table 3-21 shows the total number of newborns screened for each of the studies (numbers assigned in Table 3-20), along with the total number of cases identified. The last four columns contain the corrected numbers tested and cases identified, along with an estimate of the prevalence and the corresponding 95% confidence intervals. Corrections for individual studies are described in text following the table. Below each of the four geographical groups is a summary prevalence, computed using a random effects model (Berlin et al., 1989). At the bottom of the table is the summary prevalence for all studies. Figure 3-8 shows the same data in graphical form. The overall estimate for prevalence in non-Hispanic Caucasians is 1:2509, but there is considerable heterogeneity between studies (χ² = 35.0, p< 0.001). This is greatly reduced when the results are stratified by geographic region. Even then, however, heterogeneity exists in the prevalence estimates from the United Kingdom and Europe.

Table 3-21. Reported and Adjusted Cystic Fibrosis Prevalence in Non-Hispanic Caucasians, Derived from Newborn Screening Studies

Study Number	Reported		After Adjustment
	Tested	Cases (% MI)1	Tested	Cases	Prevalence	95% CI
United Kindom
1	81,778	37 (14)	81,778	37	1:2210	1604-3139
2	211,344	98 (18)	211,344	98	1:2157	1792-2706
3	108,422	70 (17)	108,422	70	1:1549	1226-1987
4	227,183	78 (8)	227,183	78+14	1:2469	2041-3125
5	437,959	170 (14)	437,959	170+26	1:2234	1956-2561
Subtotal	χ2=6.3, p=0.01		1,066,696		1:2123	1838-2451
Australia/New Zealand
6	1,204,000	451 (20)	1,204,000	451+24	1:2535	2323-2789
7	210,751	78 (24)	210,751	78	1:2702	2165-3418
8	309,873	112 (16)	309,873	112+32	1:2152	1844-2533
9	88,752	29 (24)	88,752	29+6	1:2536	1823-3640
Subtotal	χ2=2.7, p=0.1		1,813,376		1:2465	2259-2689
United States
10	279,399	73 (16)	181,552	73-7	1:2751	2162-3556
11	325,173	82 (21)	285,502	82-3+17	1:2974	2467-3743
12	105,734	20 (NR)	86,702	20-1+5	1:3613	2428-5637
Subtotal	χ2=0.8, p=0.5		553,753		1:2963	2480-3540
Europe (w/o United Kingdom)
13	78,800	23 (13)	78,800	23	1:3426	2283-5405
14	19,882	12 (NR)	19,882	12+1	1:1538	909-2888
15	157,992	42 (NR)	157,992	42+9	1:3098	2356-4160
16	513,440	122 (12)	513,440	122+31	1:3356	2889-4003
17	343,756	118 (14)	343,756	118+18	1:2528	2157-3052
Subtotal	χ2=7.7, p=0.01		1,113,870		1:2875	2359-3503
Total	χ2=35.0, p<0.001		4,547,695		1:2509	2286-2754

Figure 3-8. Individual and Combined Estimates of Cystic Fibrosis Prevalence in non-Hispanic Caucasians, Derived from Newborn Screening Trials

The following explains how the numbers were derived for each study listed in the table.

United Kingdom

1 Leeds – Newborn screening was offered between 1975 and 1994; the more recent time period utilized IRT measurements (Littlewood et al., 1995). No adjustments have been made to the numbers.

2 East Anglia Newborn screening was offered between 1981 and 1990 using IRT measurements (Green et al., 1992). No adjustments have been made to the numbers.

3 Northern Ireland – Newborn screening was offered for four years using IRT measurements (Roberts et al., 1988). No adjustments have been made to the numbers.

4 Wales and the Midlands – Newborn screening for cystic fibrosis was performed on alternative weeks, using a two-step IRT protocol (Ryley et al., 1988). The rate of meconium ileus (8 percent 6/78) was significantly lower than in the corresponding control population (28 percent 19/68), and in the other newborn studies reported (Table 3-21). The authors point out that a non-participating hospital was often used as a referral center for infants with meconium ileus. Thus, the allocation of 19 patients with meconium ileus was not random. The present analysis takes this into account by allocating six additional cystic fibrosis infants with meconium ileus to the study group. During four of the six years of the study, prenatal diagnosis could have reduced the number of cases identified. An additional nine cases (4/6 * 227,183 / 16,800) are estimated to have been prenatally diagnosed in this population.

5 Trent – Over about six years, newborn screening for cystic fibrosis was offered, using a two-step IRT protocol and, later, a three stage protocol, using IRT and mutation analysis (Pollitt et al., 1997). The entire study was performed during a time when prenatal diagnosis for cystic fibrosis was available. Overall, 26 (437,859/16,800) prenatally diagnosed cases would be expected in this population.

Australia/New Zealand

6 New South Wales – Nearly all of these newborns were screened using a two-step IRT protocol. Subsequently, 200,000 were tested using IRT and mutation analysis (Wilcken et al., 1995). During the time when newborn screening was offered, 24 cases were known to be prenatally diagnosed and terminated, bringing the total cases identified during the 14 years of study to 475. No other corrections are performed.

7 New Zealand – Newborns were screened for cystic fibrosis using a two-step IRT protocol over a seven year time period (Wesley et al., 1989). No modifications are made to the reported numbers. It was assumed that the population was mainly northern European Caucasians.

8 Adelaide – Newborns were screened during one year, using a two-step IRT protocol, beginning in 1989 (Ranieri et al., 1994). For the remaining four years, IRT and mutation analysis was used. During the study, there were 32 prenatal diagnoses and terminations for cystic fibrosis leading to a total of 144 cases identified. No other modifications are made to the reported numbers.

9 Victoria – During the four years of this study, all newborns were tested, using a combination of IRT and mutation analysis (Balnaves et al., 1995). Six prenatal diagnoses were made during that time, leading to a total of 35 cases of cystic fibrosis being identified in the cohort. No other modifications are made to the reported numbers.

United States

10 Colorado – A pilot study was conducted as part of the statewide newborn screening program (Hammond et al., 1991). Screening was by two-step IRT, with sweat testing as the diagnostic test. Compliance and the mixed racial/ethnic nature of the population tested complicate the analysis of prevalence in this study. Only about three-quarters of those with an initially elevated IRT measurement submitted a second sample. In reporting the results of their study, the authors corrected for this non-compliance. However, the study design included follow-up methodology that was designed to identify all missed cases, and three cases of cystic fibrosis were identified in the group with initially elevated IRT measurements that were not re-tested. Because of the study's follow-up, the adjustment (as performed by the authors) appears not to be appropriate and was not performed in our analysis. The study reported that 70 of the 73 cases were Caucasian and that the associated prevalence was 1:3073 (70:215,110), but it did not distinguish Hispanic from non-Hispanic. Based on 1987 vital statistics, 84.4 percent of the Caucasian population in Colorado was non-Hispanic. Taking this into account reduces the denominator to 181,552 (215,110 * .844). Among Hispanic Caucasians, cystic fibrosis is estimated to have occurred in four newborns. Thus, the corrected non-Hispanic Caucasian prevalence of cystic fibrosis is 1:2750 (66:181,552). The study was performed prior to the availability of prenatal diagnosis.

11 Wisconsin - A pilot study was conducted as part of newborn screening in Wisconsin (Gregg et al., 1993; Gregg et al., 1997). Screening was by two-step IRT, with DNA testing incorporated in the later years. A significant proportion of those with elevated screening results did not have the sweat testing completed, but the study's extensive follow-up procedures would have detected any cases occurring in this group. The study reported a rate for Caucasians (1:3431) in the first report (Gregg et al., 1993) but did not stratify by race/ethnicity in the second. Based on the 1989 vital statistics for Wisconsin, 87.8 percent of the population is non-Hispanic Caucasian, 2.2 percent Hispanic, and 10.0 percent Black. Thus, the corrected denominator for non-Hispanic Caucasians is 285,502 (325,173*.878). Three of the observed cases are estimated to have occurred in other racial/ethnic groups. An additional 17 cases (285,502/16,800) would have been detected by newborn screening, had prenatal diagnosis not been available and the present analysis also takes this into account.

12 Western Pennsylvania – A pilot newborn screening trial was conducted in several hospitals in the Pittsburgh area (Spence et al., 1993). Screening was by two-step IRT, with DNA testing incorporated in the later years. Even though the population was of mixed race/ethnicity, the study did not take this into account. Based on the 1989 vital statistics for Pittsburgh (percentage of Caucasian and Black) and Pennsylvania (percentage of Hispanic Caucasian), 82.0 percent of the screened population is estimated to be non-Hispanic Caucasian, 2.5 percent Hispanic, 14.5 percent Black and 1 percent Asian. Thus, the corrected denominator for non-Hispanic Caucasians is 86,702, and the corrected number of cases is 19 (one case expected in other racial/ethnic groups has been removed). The study was performed during a time when prenatal diagnosis for cystic fibrosis was widely available, yet this was not taken into account. An additional five cases (86,702/16,800) would have been detected by newborn screening, had prenatal diagnosis not been available and these cases have been included in our estimates.

Europe (w/o United Kingdom)

13 Normandy, France – Over a two year time period, newborns were tested using a one-step IRT protocol (Tavert and Duhamel, 1983). Twenty-three infants with cystic fibrosis were identified. No changes to the reported numbers have been made.

14 Vienna, Austria – Over a three year time period newborns were screened, using a single IRT measurement (Larsen et al., 1994). Twelve cases of cystic fibrosis were confirmed by sweat testing. The study was performed during a time when prenatal diagnosis was available. One prenatal diagnosis might have occurred in this group (19,992/16,800) and our estimate of prevalence includes this case.

15 Northeastern Italy – Over a three year time period, IRT measurements were used, along with meconium lactase activity on selected samples (Pederzini et al., 1995). During the study, prenatal diagnosis for cystic fibrosis was available, and an estimated nine cases were diagnosed (157,992/16,800). A total of 51 cases of cystic fibrosis are expected in this population.

16 France, Collaborative Study – Over two years, newborn screening was performed in eleven laboratories using a two-step IRT protocol (Dhondt et al., 1993). A total of 122 cases of cystic fibrosis was identified. Another 31 cases are added in the present analysis (513,440/16,800) to account for possible prenatal diagnoses.

17 Brittany, France – Over 10 years, this ongoing screening program identified 118 newborns with cystic fibrosis (Ferec et al., 1995; Scotet et al., 2000). At the beginning, the program used a two-step IRT protocol. In 1993, the program replaced the follow-up IRT measurement with DNA measurements. Prenatal diagnosis identified 18 cases. Thus, 136 cases of cystic fibrosis occurred during the time period covered by the project.

References

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Dhondt JL, Farriaux JP, Briard ML, Boschetti R, Frezal J. 1993. Results of pilot screening activities in the French neonatal screening program – cystic fibrosis, congenital adrenal hyperplasia and sickle cell disease. Screening 2:87-97.

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