doi:10.1111/codi.12600
Narrative review
Genetic testing in inherited polyposis syndromes – how and why? G. H. Lee*†, S. J. Payne‡, A. Melville‡ and S. K. Clark*† *The Polyposis Registry, St Mark’s Hospital, Harrow, UK, †Department of Surgery and Cancer, Imperial College London, London, UK and ‡Northwest Thames Regional Genetics Service, Kennedy Galton Centre, St Mark’s Hospital, Harrow, UK Received 9 October 2013; accepted 5 December 2013; Accepted Article online 24 February 2014
Abstract There have been recent advances in genetic testing enabling accurate diagnosis of polyposis syndromes by identifying causative gene mutations, which is essential in the management of individuals with polyposis syndrome and predictive genetic testing of their extended families. There are some similarities in clinical presentation of various polyposis syndromes, which may pose a challenge to diagnosis. In this review, we discuss the clinical presentation of the main polyposis syndromes and the process
of genetic testing, including the latest advancement and future of genetic testing. We aim to reiterate the importance of genetic testing in the management of polyposis syndromes, potential pitfalls associated with genetic testing and recommendations for healthcare professionals involved with the care of polyposis patients.
Introduction
The polyposis syndromes
Recent advances in molecular genetic testing have allowed an accurate diagnosis of various polyposis syndromes by identifying causative gene mutations. Since the discovery of the adenomatous polyposis coli (APC) gene on chromosome 5q in 1991 [1–3] responsible for familial adenomatous polyposis (FAP), genetic mapping techniques have identified genes mutated in other inherited polyposis syndromes such as MUTYH-associ ated polyposis (MAP), Peutz–Jeghers syndrome (PJS) and juvenile polyposis syndrome (JPS). Except for MAP which is genetically recessive, the rest are inherited in an autosomal dominant manner. Although there are some similarities between these polyposis syndromes, genetic testing enables clinicians to diagnose, treat and manage patients and their families specifically according to the inherited disease. In this review, we discuss the clinical presentation of the main polyposis syndromes, the process of genetic testing, cost involved, role in management of disease, pitfalls and recommendations for clinicians involved with the care of polyposis patients.
Correspondence to: Mr Gui Han Lee, The Polyposis Registry, St Mark’s Hospital, Watford Road, Harrow HA1 3UJ, UK. E-mail: [email protected]
Keywords Polyposis syndromes, genetic testing, management
Familial adenomatous polyposis
FAP is an autosomal dominantly inherited disease characterized by the development of hundreds to thousands of adenomatous polyps in the colon and rectum. In almost all cases, an individual with FAP will develop colorectal cancer at a mean age of 40–50 years without surgical intervention [4]. Adenomatous polyps also occur in the duodenum, which in 5% progress into cancer if left untreated [5]. Other cancers associated with FAP include thyroid, hepatoblastoma and brain. The incidence of FAP is 1 in 8324 births [6]. The clinical diagnosis of FAP is based on the presence of more than 100 adenomatous polyps in the colon and rectum. Other characteristic extra-intestinal features of FAP include congenital hypertrophy of the retinal pigment epithelium, epidermoid cysts, osteomas, desmoids, supernumerary teeth and adrenal gland adenoma. The clinical presentation of FAP in the past has often been with colorectal cancer. However, the introduction of screening programmes and polyposis registries has significantly reduced the number of deaths from colorectal cancer by early identification and surveillance of at-risk individuals and provision of timely prophylactic surgery [7]. Attenuated familial adenomatous polyposis (AFAP) is a milder form of FAP characterized by fewer colorectal
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Clinically affected patient (>10 adenomatous polyps)
Genetic counselling
Genetic testing (blood test)
Mutation detected
No mutation detected
Diagnosis confirmed
Clinical diagnosis only
Individualised care Predictive testing available for relative
Manage patient and relatives with routine colonoscopies
Enter into study to identify other causative genes
Preimplantation genetic testing can be offered Enhanced understanding of phenotype and penetrance
Figure 1 Flow diagram of the management of a suspected polyposis patient.
polyps and later age of onset [8]. It may be challenging for clinicians to diagnose AFAP and attempts to define diagnostic criteria have been made. However, it has been generally agreed that a positive family history of adenomatous polyposis inherited in an autosomal dominant manner with multiple colorectal adenomas (but fewer than 100) is diagnostic of AFAP [9,10]. MUTYH-associated polyposis
MAP is an autosomal recessively inherited condition where patients present with multiple adenomatous polyps due to biallelic mutation in the MUTYH gene. The
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clinical presentation of MAP can be difficult to differentiate from AFAP due to lower numbers of colorectal polyps (15–200 adenomas) and presentation at a later age compared with FAP [11,12]. MAP patients have a lower chance of developing upper gastrointestinal disease [13]. However, the risk of developing colorectal cancer is estimated to approach 100% at 60 years in individuals with biallelic mutation in the MUTYH gene, although this may be an overestimate due to ascertainment bias [14]. There is also a modestly increased risk in monoallelic mutation carriers but this is not considered sufficiently high to warrant screening [15].
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Peutz–Jeghers syndrome
PJS is an autosomal dominantly inherited condition where patients develop multiple characteristic polyps in the gastrointestinal tract, mainly in the small intestine. These polyps histologically show interdigitating smooth muscle bundles in the mucosa, which appear to be ‘frond-like’. Another important clinical feature of PJS is the characteristic mucocutaneous hyperpigmentation presenting as macules most commonly on the vermillion border of the lips, hands, buccal mucosa and feet, due to increased deposits of melanocytes and melanin in the epidermal–dermal junction of the skin. The incidence of PJS is estimated to be between 1 in 50 000 and 1 in 200 000 [16]. Clinical presentation of PJS is mainly associated with the characteristic polyps, which may lead to bowel obstruction, acute or chronic anaemia associated with gastrointestinal bleeding, prolapse of a polyp and, in the case of children, small bowel intussuception. Such presentation normally requires endoscopic investigation and surgical intervention. Diagnostic criteria for PJS include two or more histologically confirmed Peutz–Jeghers polyps, any number of Peutz–Jeghers polyps detected in one individual with a family history of PJS in a close relative, characteristic mucocutaneous pigmentation in an individual who has a family history of PJS in a close relative, or any number of Peutz–Jeghers polyps in an individual who also has characteristic mucocutaneous pigmentation [17,18]. Although PJS polyps are benign, there is a significantly increased risk of gastrointestinal cancer. Various studies show high rates of other cancers including breast, pancreatic and gynaecological cancers. A recent systematic review by Beggs et al.[19] summarizes recommendations for surveillance and follow-up for PJSassociated cancers.
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was 39%, with significantly elevated relative risk compared with the general population [21]. The incidence of gastric cancer associated with gastric polyps is around 21% [22]. It is generally agreed that the diagnostic criteria for JPS involve having one of the following presentations – five or more juvenile polyps in the large intestine or any number of juvenile polyps with a positive family history [23].
Genetic testing in polyposis Familial adenomatous polyposis
Mutation of the APC gene located in the long arm of chromosome 5 (5q 22.2) is responsible for FAP. A mutation is detected in up to 90% of patients with a clinical diagnosis of FAP [24]. In about 15–20% of FAP cases, the APC gene mutation is de novo [25]. It is important for clinicians first to identify the characteristic presentation of FAP prior to genetic testing. However, once the causative mutation has been identified, at-risk relatives of FAP patients can undergo predictive genetic testing, which is usually recommended in late childhood or adolescence [26], a paradigm that is also applicable to other inherited polyposis syndromes. MUTYH-associated polyposis
Mutation in the mutY human homologue (MUTYH) gene located on chromosome 1p34.1 is responsible for MAP [27]. MAP should be considered first in patients with a phenotype similar to FAP or AFAP and a pedigree suggesting autosomal recessive inheritance, and also in all other cases with more than 20 adenomas when APC testing is negative.
Juvenile polyposis syndrome
Attenuated familial adenomatous polyposis
JPS is the development of characteristic benign hamartomatous polyps along the gastrointestinal tract. The term ‘juvenile’ is used to describe the histological characteristics of having dense stroma with inflammatory infiltrate and mucus-filled cystic glands in the lamina propria. The incidence of JPS is estimated at 1 in 100 000 [20]. The number of polyps varies in JPS patients (ranging from 5 to over 100 polyps in a lifetime) and the main clinical presentation is gastrointestinal bleed or anaemia associated with juvenile polyps. However, there is an increased risk of developing gastrointestinal cancer. Specifically, the incidence of colorectal cancer in one study
Presentation of AFAP can be due to mutation in either the APC or MUTYH gene. A study by Nielson et al. showed that up to 72% of families with a clinical diagnosis of AFAP have such a mutation [9]. It is recommended that genetic testing should be guided by the inheritance pattern in the pedigree (i.e. if apparently autosomal dominant, test for APC mutation first) [9,10]. Peutz–Jeghers syndrome
Germline mutation in the STK11 (LKB1) gene in chromosome 19p13.3 is responsible for PJS [28,29]. It has
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been reported that the rate of mutation detection is over 94% in PJS provided that the diagnostic criteria are met [18]. Predictive testing is recommended in earlier childhood to allow appropriate management of any episodes of abdominal pain [19]. Juvenile polyposis syndrome
Germline mutations in two genes have been identified as causing JPS, namely BMPR1A and SMAD4. Approximately 20% of patients have mutations in BMPR1A and approximately 21% have mutations in SMAD4 [30]. The overall mutation detection rate of BMPR1A and SMAD4 is approximately 45% [31]. However, the detection rate varies depending on the type and method of genetic testing. Hereditary haemorrhagic telangectasia (HHT) is an autosomal dominant disease leading to multiple arteriovenous malformations (AVMs) ranging from telangectasia (small AVMs) to major pulmonary or cerebral AVMs. A combined syndrome of JPS and HHT has been specifically connected to mutation of the SMAD4 and endoglin (ENG) genes. Recent figures from Cleveland Clinic show up to 81% correlation of SMAD4 mutation to HHT in JPS patients [32]. Predictive testing is usually undertaken in late childhood or earlier if symptomatic.
Process of genetic testing Once clinical and histopathological features raise strong suspicion of a polyposis syndrome, patients can be referred for genetic testing. The geneticist or genetic counsellor will take and assess a detailed medical and
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family history and, if appropriate, take a blood sample for genetic testing. Genetic counselling
There are significant psychosocial implications of a diagnosis of a hereditary polyposis syndrome, for both the patient and the immediate family members, especially in young adults [33–35]. Therefore, during a genetic counselling consultation, patients receive advice on practical, emotional and financial issues surrounding genetic testing. It is essential that patients understand what genetic testing involves, including the implications of a positive or a negative test result. Patients can experience complex emotions and appropriate support is available. For example, to help patients with the possible burden of sharing a genetic diagnosis with their relatives, a letter explaining the family history and genetic information can be given to patients to pass on to their at-risk family members. Cost of genetic testing
There are two main types of genetic testing in polyposis syndromes: mutation characterization in an index case and predictive testing for at-risk relatives. For mutation detection, various techniques are available for identifying specific common mutations, gene sequencing and chromosome analysis, with current costs in the region of £500 in the UK using current techniques [36], which is likely to decrease significantly in the near future with the introduction of next generation sequencers. Once a mutation is identified, confirming
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Figure 2 Example of DNA sequence analysis to identify APC mutation. DNA sequence analysis to show the recurrent APC deletion c.3183_3187delACAAA, p.Gln1062*. The top panel is the normal APC sequence. The bottom panel is APC from a patient carrying the deletion of five bases (underlined in the top panel) on one allele. The arrow marks the start of the mixed sequence where the normal and deleted alleles overlap.
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the presence or absence of that mutation in at-risk family members is straightforward and inexpensive.
surveillance in those confirmed not to have inherited the condition.
Financial and confidentiality issues
Predictive testing
Clinicians emphasize the importance of identifying potentially inherited colorectal cancers in patients due to the ever-increasing accuracy of genetic testing and preventive management available for patients and their extended families [37]. However, some patients may be concerned about the potential financial implications and confidentiality issues surrounding a positive genetic test. According to the Association of British Insurers, insurance companies can ask for family history and diagnostic genetic test results prior to taking up a policy. However, there is a moratorium until 2017 stating that insurers cannot ask for predictive genetic test results nor force an individual to have any genetic testing after approval of an insurance policy [38]. Therefore it is advisable for patients to arrange insurance policies prior to taking genetic testing. Clinicians must ensure that adequate consent is obtained from an individual with a polyposis syndrome before disclosing any information to other medical professionals involved in management of associated conditions, or to other family members for the purposes of predictive testing. Exceptionally, a decision to breach confidentiality may be made in cases where there is potential harm to a relative of not being informed of their risk. The potential harm to the relative of not knowing their risk should be considered to outweigh the potential harm to the patient from confidentiality being broken. Clinicians are advised to provide adequate support for patients in disclosing information to their relatives in receiving predictive genetic testing, without the need to breach confidentiality. Genetics departments offer support and assistance in these aspects. The regulations concerning these aspects vary internationally and liaison with local clinical genetics departments is strongly recommended.
Once a genetic mutation is confirmed, predictive testing of relatives at high risk can be carried out. Predictive testing is particularly useful due to its ease and availability in genetics centres. It is less invasive (DNA is extracted from a simple peripheral blood sample) and more cost effective than colonoscopies. Predictive genetic testing currently costs around £150 in the UK compared with around £600 for colonoscopy [39]. Risks and costs of colonoscopy are higher in children due to the potential need for general anaesthetic and need for specialist endoscopists. Once a negative predictive test is established, there is no need for subsequent clinical follow-up.
Advantages of genetic testing Confirmation of the diagnosis
It can be valuable to have a confirmed genetic diagnosis, particularly in cases where phenotype can be confusing (e.g. AFAP). This also allows us to improve our understanding of penetrance and disease features, which can to some extent be biased if based on clinical diagnosis alone. A definitive diagnosis may also give significant peace of mind to those diagnosed and their relatives, and prevents unnecessary
Family planning
Apart from predictive testing, prenatal diagnosis can be offered for mutation carriers, to allow them to have children free of polyposis. Pre-implantation genetic diagnosis is also possible, although services are restricted to a few specialist centres. Although these tests are not often requested, they may provide important options for patients prior to starting a family. Individual management
There is undoubted evidence of genotype–phenotype correlation in FAP, with particular respect to the number and age of onset of colorectal adenomas. Gayther et al.[40] suggested that mutations in codon 1309 and beyond were associated with more severe and earlier onset of disease. A recent report from Germany confirmed that patients with mutation in codon 1309 developed bowel symptoms more than 10 years earlier than patients with other mutations [41]. A recent review by Nieuwenhuis and Vasen classified FAP into profuse, intermediate and attenuated phenotype and grouped the associated mutations [42]. The profuse phenotype was common in mutations from codons 1250 to 1464. These findings can identify patients with a severe form of polyposis who may require early surgical intervention with restorative proctocolectomy with ileal pouch-anal anastomosis rather than colectomy with ileorectal anastomosis [43]. Therefore, such knowledge of an individual’s mutation can directly influence the choice and timing of surgery. Desmoid tumours have been linked to mutations at the 30 end of the APC gene, beyond codon 1399, and
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there is evidence that delaying surgery in female patients at high risk with such a mutation may lead to a reduction in incidence of desmoid tumour [44–46]. Having confirmation of STK11 gene mutation is invaluable in the management of young children at risk of PJS. It allows a rapid and appropriate clinical response to the common and vague symptom of abdominal pain, which in a child with PJS may represent developing intussusception, and prevents overreaction in children who have not inherited the mutation. Detection of mutation in SMAD4 in a patient with JPS allows appropriate screening for HHT to be undertaken, particularly in individuals undergoing procedures under general anaesthetic who may have increased prevalence of arteriovenous malformation [47].
detectable, however, in bowel mucosa biopsies taken between polyps. In some instances, siblings have been reported with FAP resulting from a parent with no evidence of the condition, who actually has a mutation in a gonad but not in the gastrointestinal tract. Genetic testing to identify mosaicism can be challenging and laborious, involving detailed mutation analysis. In some instances, DNA analysis of colonic biopsies may be required to identify the mutation responsible. Clinical management of patients with mosaicism provides an additional challenge to clinicians and may account for a significant proportion of AFAP. Due to wide variation in the phenotype, it is particularly difficult to predict the severity of disease and manage at-risk relatives [51].
Difficult situations Other genes Known genes
The greatest challenge arises when patients exhibit clinical and histopathological signs of a polyposis syndrome with negative mutation detection. This may arise for a number of reasons. First, there may be technical difficulty in identifying mutations in the known genes. However, the detection rates have risen as techniques have evolved. In addition, a better understanding of the role of ‘non-coding’ and promoter regions has recently led to the identification of pathogenic mutations in these areas. For example, an APC gene promoter deletion was identified following demonstration of imbalanced RNA expression in an individual with previous negative APC and MUTYH gene sequencing [48]. At another centre, promoter gene deletions have been identified using direct sequencing followed by multiplex ligation-dependent probe amplification of genomic DNA [49]. Mosaicism
Around 15–20% of FAP cases arise de novo and it is estimated that up to one-fifth of these cases are associated with mosaicism [50]. Mosaicism is the presence of two or more genetically different cell lines in an individual. This occurs during embryogenesis when a mutation arises in a single cell and is therefore present in all resulting daughter cells. The mutation will thus be present in tissues derived from the original mutated cell but not in others. In the context of a polyposis syndrome, mosaicism may result in a mutation being present in the bowel (resulting in polyposis phenotype) but in too low a concentration in blood to detect. The mutation should be
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Since the first description of the MUTYH gene mutation in a Welsh family with APC gene negative polyposis in 2002, subsequent research has led to the characterization of MAP [27,52]. However, in MAP there is initial difficulty in clinically differentiating from AFAP. Without adequate family history, genetic testing may be the only way in which to distinguish between these polyposis syndromes. Although there are common mutations that account for over 80% of MAP in Caucasians, other specific mutations have been found in individuals of Asian origin [53,54]. Recent advances have led to the identification of other genes responsible for polyposis syndromes. The latest discovery has been constitutional inherited (germline) mutations POLE and POLD1 in adenomatous polyposis, which resulted from the use of whole-genome sequencing in patients within a large cohort of colorectal polyposis patients recruited to the CORGI (Colorectal Cancer Gene Identification) study [55]. This study showed that heterozygous POLE and POLD1 constitutional inherited variants had high penetrance and may be further associated with endometrial cancer. Improvements in genetic testing with introduction of next generation sequencing and RNA analysis will continue to enhance the detection of mutations in polyposis syndromes. Furthermore, next generation sequencing has been shown to be cost effective and highly reproducible in genetic testing [56].
Conclusion The introduction of genetic testing has brought about significant changes in the management of inherited
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polyposis. Clinicians are now able to diagnose and provide individualized care depending on the genetic defect of the patient group, and predictive testing enables early diagnosis of asymptomatic family members. Genetic testing gives opportunity for family planning and asymptomatic relatives can be reassured with negative predictive genetic testing. Formation of polyposis registries, early diagnosis and adequate endoscopic and surgical management have significantly improved outcomes in polyposis with genetic testing playing a vital role in the patient care pathway [47].
Author contributions G. H. Lee: literature search and drafted article. S. J. Payne: critically revised drafted article. A. Melville: critically revised drafted article. S. K. Clark: conception and design of article, final approval of the version to be published.
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Colorectal Disease ª 2014 The Association of Coloproctology of Great Britain and Ireland. 16, 595–602
Narrative review
Genetic testing in inherited polyposis syndromes – how and why? G. H. Lee*†, S. J. Payne‡, A. Melville‡ and S. K. Clark*† *The Polyposis Registry, St Mark’s Hospital, Harrow, UK, †Department of Surgery and Cancer, Imperial College London, London, UK and ‡Northwest Thames Regional Genetics Service, Kennedy Galton Centre, St Mark’s Hospital, Harrow, UK Received 9 October 2013; accepted 5 December 2013; Accepted Article online 24 February 2014
Abstract There have been recent advances in genetic testing enabling accurate diagnosis of polyposis syndromes by identifying causative gene mutations, which is essential in the management of individuals with polyposis syndrome and predictive genetic testing of their extended families. There are some similarities in clinical presentation of various polyposis syndromes, which may pose a challenge to diagnosis. In this review, we discuss the clinical presentation of the main polyposis syndromes and the process
of genetic testing, including the latest advancement and future of genetic testing. We aim to reiterate the importance of genetic testing in the management of polyposis syndromes, potential pitfalls associated with genetic testing and recommendations for healthcare professionals involved with the care of polyposis patients.
Introduction
The polyposis syndromes
Recent advances in molecular genetic testing have allowed an accurate diagnosis of various polyposis syndromes by identifying causative gene mutations. Since the discovery of the adenomatous polyposis coli (APC) gene on chromosome 5q in 1991 [1–3] responsible for familial adenomatous polyposis (FAP), genetic mapping techniques have identified genes mutated in other inherited polyposis syndromes such as MUTYH-associ ated polyposis (MAP), Peutz–Jeghers syndrome (PJS) and juvenile polyposis syndrome (JPS). Except for MAP which is genetically recessive, the rest are inherited in an autosomal dominant manner. Although there are some similarities between these polyposis syndromes, genetic testing enables clinicians to diagnose, treat and manage patients and their families specifically according to the inherited disease. In this review, we discuss the clinical presentation of the main polyposis syndromes, the process of genetic testing, cost involved, role in management of disease, pitfalls and recommendations for clinicians involved with the care of polyposis patients.
Correspondence to: Mr Gui Han Lee, The Polyposis Registry, St Mark’s Hospital, Watford Road, Harrow HA1 3UJ, UK. E-mail: [email protected]
Keywords Polyposis syndromes, genetic testing, management
Familial adenomatous polyposis
FAP is an autosomal dominantly inherited disease characterized by the development of hundreds to thousands of adenomatous polyps in the colon and rectum. In almost all cases, an individual with FAP will develop colorectal cancer at a mean age of 40–50 years without surgical intervention [4]. Adenomatous polyps also occur in the duodenum, which in 5% progress into cancer if left untreated [5]. Other cancers associated with FAP include thyroid, hepatoblastoma and brain. The incidence of FAP is 1 in 8324 births [6]. The clinical diagnosis of FAP is based on the presence of more than 100 adenomatous polyps in the colon and rectum. Other characteristic extra-intestinal features of FAP include congenital hypertrophy of the retinal pigment epithelium, epidermoid cysts, osteomas, desmoids, supernumerary teeth and adrenal gland adenoma. The clinical presentation of FAP in the past has often been with colorectal cancer. However, the introduction of screening programmes and polyposis registries has significantly reduced the number of deaths from colorectal cancer by early identification and surveillance of at-risk individuals and provision of timely prophylactic surgery [7]. Attenuated familial adenomatous polyposis (AFAP) is a milder form of FAP characterized by fewer colorectal
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Clinically affected patient (>10 adenomatous polyps)
Genetic counselling
Genetic testing (blood test)
Mutation detected
No mutation detected
Diagnosis confirmed
Clinical diagnosis only
Individualised care Predictive testing available for relative
Manage patient and relatives with routine colonoscopies
Enter into study to identify other causative genes
Preimplantation genetic testing can be offered Enhanced understanding of phenotype and penetrance
Figure 1 Flow diagram of the management of a suspected polyposis patient.
polyps and later age of onset [8]. It may be challenging for clinicians to diagnose AFAP and attempts to define diagnostic criteria have been made. However, it has been generally agreed that a positive family history of adenomatous polyposis inherited in an autosomal dominant manner with multiple colorectal adenomas (but fewer than 100) is diagnostic of AFAP [9,10]. MUTYH-associated polyposis
MAP is an autosomal recessively inherited condition where patients present with multiple adenomatous polyps due to biallelic mutation in the MUTYH gene. The
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clinical presentation of MAP can be difficult to differentiate from AFAP due to lower numbers of colorectal polyps (15–200 adenomas) and presentation at a later age compared with FAP [11,12]. MAP patients have a lower chance of developing upper gastrointestinal disease [13]. However, the risk of developing colorectal cancer is estimated to approach 100% at 60 years in individuals with biallelic mutation in the MUTYH gene, although this may be an overestimate due to ascertainment bias [14]. There is also a modestly increased risk in monoallelic mutation carriers but this is not considered sufficiently high to warrant screening [15].
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Peutz–Jeghers syndrome
PJS is an autosomal dominantly inherited condition where patients develop multiple characteristic polyps in the gastrointestinal tract, mainly in the small intestine. These polyps histologically show interdigitating smooth muscle bundles in the mucosa, which appear to be ‘frond-like’. Another important clinical feature of PJS is the characteristic mucocutaneous hyperpigmentation presenting as macules most commonly on the vermillion border of the lips, hands, buccal mucosa and feet, due to increased deposits of melanocytes and melanin in the epidermal–dermal junction of the skin. The incidence of PJS is estimated to be between 1 in 50 000 and 1 in 200 000 [16]. Clinical presentation of PJS is mainly associated with the characteristic polyps, which may lead to bowel obstruction, acute or chronic anaemia associated with gastrointestinal bleeding, prolapse of a polyp and, in the case of children, small bowel intussuception. Such presentation normally requires endoscopic investigation and surgical intervention. Diagnostic criteria for PJS include two or more histologically confirmed Peutz–Jeghers polyps, any number of Peutz–Jeghers polyps detected in one individual with a family history of PJS in a close relative, characteristic mucocutaneous pigmentation in an individual who has a family history of PJS in a close relative, or any number of Peutz–Jeghers polyps in an individual who also has characteristic mucocutaneous pigmentation [17,18]. Although PJS polyps are benign, there is a significantly increased risk of gastrointestinal cancer. Various studies show high rates of other cancers including breast, pancreatic and gynaecological cancers. A recent systematic review by Beggs et al.[19] summarizes recommendations for surveillance and follow-up for PJSassociated cancers.
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was 39%, with significantly elevated relative risk compared with the general population [21]. The incidence of gastric cancer associated with gastric polyps is around 21% [22]. It is generally agreed that the diagnostic criteria for JPS involve having one of the following presentations – five or more juvenile polyps in the large intestine or any number of juvenile polyps with a positive family history [23].
Genetic testing in polyposis Familial adenomatous polyposis
Mutation of the APC gene located in the long arm of chromosome 5 (5q 22.2) is responsible for FAP. A mutation is detected in up to 90% of patients with a clinical diagnosis of FAP [24]. In about 15–20% of FAP cases, the APC gene mutation is de novo [25]. It is important for clinicians first to identify the characteristic presentation of FAP prior to genetic testing. However, once the causative mutation has been identified, at-risk relatives of FAP patients can undergo predictive genetic testing, which is usually recommended in late childhood or adolescence [26], a paradigm that is also applicable to other inherited polyposis syndromes. MUTYH-associated polyposis
Mutation in the mutY human homologue (MUTYH) gene located on chromosome 1p34.1 is responsible for MAP [27]. MAP should be considered first in patients with a phenotype similar to FAP or AFAP and a pedigree suggesting autosomal recessive inheritance, and also in all other cases with more than 20 adenomas when APC testing is negative.
Juvenile polyposis syndrome
Attenuated familial adenomatous polyposis
JPS is the development of characteristic benign hamartomatous polyps along the gastrointestinal tract. The term ‘juvenile’ is used to describe the histological characteristics of having dense stroma with inflammatory infiltrate and mucus-filled cystic glands in the lamina propria. The incidence of JPS is estimated at 1 in 100 000 [20]. The number of polyps varies in JPS patients (ranging from 5 to over 100 polyps in a lifetime) and the main clinical presentation is gastrointestinal bleed or anaemia associated with juvenile polyps. However, there is an increased risk of developing gastrointestinal cancer. Specifically, the incidence of colorectal cancer in one study
Presentation of AFAP can be due to mutation in either the APC or MUTYH gene. A study by Nielson et al. showed that up to 72% of families with a clinical diagnosis of AFAP have such a mutation [9]. It is recommended that genetic testing should be guided by the inheritance pattern in the pedigree (i.e. if apparently autosomal dominant, test for APC mutation first) [9,10]. Peutz–Jeghers syndrome
Germline mutation in the STK11 (LKB1) gene in chromosome 19p13.3 is responsible for PJS [28,29]. It has
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been reported that the rate of mutation detection is over 94% in PJS provided that the diagnostic criteria are met [18]. Predictive testing is recommended in earlier childhood to allow appropriate management of any episodes of abdominal pain [19]. Juvenile polyposis syndrome
Germline mutations in two genes have been identified as causing JPS, namely BMPR1A and SMAD4. Approximately 20% of patients have mutations in BMPR1A and approximately 21% have mutations in SMAD4 [30]. The overall mutation detection rate of BMPR1A and SMAD4 is approximately 45% [31]. However, the detection rate varies depending on the type and method of genetic testing. Hereditary haemorrhagic telangectasia (HHT) is an autosomal dominant disease leading to multiple arteriovenous malformations (AVMs) ranging from telangectasia (small AVMs) to major pulmonary or cerebral AVMs. A combined syndrome of JPS and HHT has been specifically connected to mutation of the SMAD4 and endoglin (ENG) genes. Recent figures from Cleveland Clinic show up to 81% correlation of SMAD4 mutation to HHT in JPS patients [32]. Predictive testing is usually undertaken in late childhood or earlier if symptomatic.
Process of genetic testing Once clinical and histopathological features raise strong suspicion of a polyposis syndrome, patients can be referred for genetic testing. The geneticist or genetic counsellor will take and assess a detailed medical and
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family history and, if appropriate, take a blood sample for genetic testing. Genetic counselling
There are significant psychosocial implications of a diagnosis of a hereditary polyposis syndrome, for both the patient and the immediate family members, especially in young adults [33–35]. Therefore, during a genetic counselling consultation, patients receive advice on practical, emotional and financial issues surrounding genetic testing. It is essential that patients understand what genetic testing involves, including the implications of a positive or a negative test result. Patients can experience complex emotions and appropriate support is available. For example, to help patients with the possible burden of sharing a genetic diagnosis with their relatives, a letter explaining the family history and genetic information can be given to patients to pass on to their at-risk family members. Cost of genetic testing
There are two main types of genetic testing in polyposis syndromes: mutation characterization in an index case and predictive testing for at-risk relatives. For mutation detection, various techniques are available for identifying specific common mutations, gene sequencing and chromosome analysis, with current costs in the region of £500 in the UK using current techniques [36], which is likely to decrease significantly in the near future with the introduction of next generation sequencers. Once a mutation is identified, confirming
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Figure 2 Example of DNA sequence analysis to identify APC mutation. DNA sequence analysis to show the recurrent APC deletion c.3183_3187delACAAA, p.Gln1062*. The top panel is the normal APC sequence. The bottom panel is APC from a patient carrying the deletion of five bases (underlined in the top panel) on one allele. The arrow marks the start of the mixed sequence where the normal and deleted alleles overlap.
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the presence or absence of that mutation in at-risk family members is straightforward and inexpensive.
surveillance in those confirmed not to have inherited the condition.
Financial and confidentiality issues
Predictive testing
Clinicians emphasize the importance of identifying potentially inherited colorectal cancers in patients due to the ever-increasing accuracy of genetic testing and preventive management available for patients and their extended families [37]. However, some patients may be concerned about the potential financial implications and confidentiality issues surrounding a positive genetic test. According to the Association of British Insurers, insurance companies can ask for family history and diagnostic genetic test results prior to taking up a policy. However, there is a moratorium until 2017 stating that insurers cannot ask for predictive genetic test results nor force an individual to have any genetic testing after approval of an insurance policy [38]. Therefore it is advisable for patients to arrange insurance policies prior to taking genetic testing. Clinicians must ensure that adequate consent is obtained from an individual with a polyposis syndrome before disclosing any information to other medical professionals involved in management of associated conditions, or to other family members for the purposes of predictive testing. Exceptionally, a decision to breach confidentiality may be made in cases where there is potential harm to a relative of not being informed of their risk. The potential harm to the relative of not knowing their risk should be considered to outweigh the potential harm to the patient from confidentiality being broken. Clinicians are advised to provide adequate support for patients in disclosing information to their relatives in receiving predictive genetic testing, without the need to breach confidentiality. Genetics departments offer support and assistance in these aspects. The regulations concerning these aspects vary internationally and liaison with local clinical genetics departments is strongly recommended.
Once a genetic mutation is confirmed, predictive testing of relatives at high risk can be carried out. Predictive testing is particularly useful due to its ease and availability in genetics centres. It is less invasive (DNA is extracted from a simple peripheral blood sample) and more cost effective than colonoscopies. Predictive genetic testing currently costs around £150 in the UK compared with around £600 for colonoscopy [39]. Risks and costs of colonoscopy are higher in children due to the potential need for general anaesthetic and need for specialist endoscopists. Once a negative predictive test is established, there is no need for subsequent clinical follow-up.
Advantages of genetic testing Confirmation of the diagnosis
It can be valuable to have a confirmed genetic diagnosis, particularly in cases where phenotype can be confusing (e.g. AFAP). This also allows us to improve our understanding of penetrance and disease features, which can to some extent be biased if based on clinical diagnosis alone. A definitive diagnosis may also give significant peace of mind to those diagnosed and their relatives, and prevents unnecessary
Family planning
Apart from predictive testing, prenatal diagnosis can be offered for mutation carriers, to allow them to have children free of polyposis. Pre-implantation genetic diagnosis is also possible, although services are restricted to a few specialist centres. Although these tests are not often requested, they may provide important options for patients prior to starting a family. Individual management
There is undoubted evidence of genotype–phenotype correlation in FAP, with particular respect to the number and age of onset of colorectal adenomas. Gayther et al.[40] suggested that mutations in codon 1309 and beyond were associated with more severe and earlier onset of disease. A recent report from Germany confirmed that patients with mutation in codon 1309 developed bowel symptoms more than 10 years earlier than patients with other mutations [41]. A recent review by Nieuwenhuis and Vasen classified FAP into profuse, intermediate and attenuated phenotype and grouped the associated mutations [42]. The profuse phenotype was common in mutations from codons 1250 to 1464. These findings can identify patients with a severe form of polyposis who may require early surgical intervention with restorative proctocolectomy with ileal pouch-anal anastomosis rather than colectomy with ileorectal anastomosis [43]. Therefore, such knowledge of an individual’s mutation can directly influence the choice and timing of surgery. Desmoid tumours have been linked to mutations at the 30 end of the APC gene, beyond codon 1399, and
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there is evidence that delaying surgery in female patients at high risk with such a mutation may lead to a reduction in incidence of desmoid tumour [44–46]. Having confirmation of STK11 gene mutation is invaluable in the management of young children at risk of PJS. It allows a rapid and appropriate clinical response to the common and vague symptom of abdominal pain, which in a child with PJS may represent developing intussusception, and prevents overreaction in children who have not inherited the mutation. Detection of mutation in SMAD4 in a patient with JPS allows appropriate screening for HHT to be undertaken, particularly in individuals undergoing procedures under general anaesthetic who may have increased prevalence of arteriovenous malformation [47].
detectable, however, in bowel mucosa biopsies taken between polyps. In some instances, siblings have been reported with FAP resulting from a parent with no evidence of the condition, who actually has a mutation in a gonad but not in the gastrointestinal tract. Genetic testing to identify mosaicism can be challenging and laborious, involving detailed mutation analysis. In some instances, DNA analysis of colonic biopsies may be required to identify the mutation responsible. Clinical management of patients with mosaicism provides an additional challenge to clinicians and may account for a significant proportion of AFAP. Due to wide variation in the phenotype, it is particularly difficult to predict the severity of disease and manage at-risk relatives [51].
Difficult situations Other genes Known genes
The greatest challenge arises when patients exhibit clinical and histopathological signs of a polyposis syndrome with negative mutation detection. This may arise for a number of reasons. First, there may be technical difficulty in identifying mutations in the known genes. However, the detection rates have risen as techniques have evolved. In addition, a better understanding of the role of ‘non-coding’ and promoter regions has recently led to the identification of pathogenic mutations in these areas. For example, an APC gene promoter deletion was identified following demonstration of imbalanced RNA expression in an individual with previous negative APC and MUTYH gene sequencing [48]. At another centre, promoter gene deletions have been identified using direct sequencing followed by multiplex ligation-dependent probe amplification of genomic DNA [49]. Mosaicism
Around 15–20% of FAP cases arise de novo and it is estimated that up to one-fifth of these cases are associated with mosaicism [50]. Mosaicism is the presence of two or more genetically different cell lines in an individual. This occurs during embryogenesis when a mutation arises in a single cell and is therefore present in all resulting daughter cells. The mutation will thus be present in tissues derived from the original mutated cell but not in others. In the context of a polyposis syndrome, mosaicism may result in a mutation being present in the bowel (resulting in polyposis phenotype) but in too low a concentration in blood to detect. The mutation should be
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Since the first description of the MUTYH gene mutation in a Welsh family with APC gene negative polyposis in 2002, subsequent research has led to the characterization of MAP [27,52]. However, in MAP there is initial difficulty in clinically differentiating from AFAP. Without adequate family history, genetic testing may be the only way in which to distinguish between these polyposis syndromes. Although there are common mutations that account for over 80% of MAP in Caucasians, other specific mutations have been found in individuals of Asian origin [53,54]. Recent advances have led to the identification of other genes responsible for polyposis syndromes. The latest discovery has been constitutional inherited (germline) mutations POLE and POLD1 in adenomatous polyposis, which resulted from the use of whole-genome sequencing in patients within a large cohort of colorectal polyposis patients recruited to the CORGI (Colorectal Cancer Gene Identification) study [55]. This study showed that heterozygous POLE and POLD1 constitutional inherited variants had high penetrance and may be further associated with endometrial cancer. Improvements in genetic testing with introduction of next generation sequencing and RNA analysis will continue to enhance the detection of mutations in polyposis syndromes. Furthermore, next generation sequencing has been shown to be cost effective and highly reproducible in genetic testing [56].
Conclusion The introduction of genetic testing has brought about significant changes in the management of inherited
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polyposis. Clinicians are now able to diagnose and provide individualized care depending on the genetic defect of the patient group, and predictive testing enables early diagnosis of asymptomatic family members. Genetic testing gives opportunity for family planning and asymptomatic relatives can be reassured with negative predictive genetic testing. Formation of polyposis registries, early diagnosis and adequate endoscopic and surgical management have significantly improved outcomes in polyposis with genetic testing playing a vital role in the patient care pathway [47].
Author contributions G. H. Lee: literature search and drafted article. S. J. Payne: critically revised drafted article. A. Melville: critically revised drafted article. S. K. Clark: conception and design of article, final approval of the version to be published.
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