Health Policy Advisory Committee on Technology

Health Policy Advisory Committee on Technology Technology Overview Blood and stool biomarker testing for colorectal cancer screening July 2015

State of Queensland (Queensland Department of Health) 2015 This work is licensed under a Creative Commons Attribution Non-Commercial No Derivatives 3.0 Australia licence. In essence, you are free to copy and communicate the work in its current form for non-commercial purposes, as long as you attribute the authors and abide by the licence terms. You may not alter or adapt the work in any way. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc-nd/3.0/au/deed.en. For further information, contact the HealthPACT Secretariat at: HealthPACT Secretariat c/o Clinical Access and Redesign Unit, Health Service and Clinical Innovation Division Department of Health, Queensland Level 2, 15 Butterfield St HERSTON QLD 4029 Postal Address: GPO Box 48, Brisbane QLD 4001 Email: HealthPACT@health.qld.gov.au Telephone: +61 7 3328 9180 For permissions beyond the scope of this licence contact: Intellectual Property Officer, Department of Health, GPO Box 48, Brisbane QLD 4001, email ip_officer@health.qld.gov.au, phone (07) 3328 9824. Electronic copies can be obtained from: http://www.health.qld.gov.au/healthpact DISCLAIMER: This Brief is published with the intention of providing information of interest. It is based on information available at the time of research and cannot be expected to cover any developments arising from subsequent improvements to health technologies. This Brief is based on a limited literature search and is not a definitive statement on the safety, effectiveness or costeffectiveness of the health technology covered. The State of Queensland acting through Queensland Health ( Queensland Health ) does not guarantee the accuracy, currency or completeness of the information in this Brief. Information may contain or summarise the views of others, and not necessarily reflect the views of Queensland Health. This Brief is not intended to be used as medical advice and it is not intended to be used to diagnose, treat, cure or prevent any disease, nor should it be used for therapeutic purposes or as a substitute for a health professional's advice. It must not be relied upon without verification from authoritative sources. Queensland Health does not accept any liability, including for any injury, loss or damage, incurred by use of or reliance on the information. This Brief was commissioned by Queensland Health, in its role as the Secretariat of the Health Policy Advisory Committee on Technology (HealthPACT). The production of this Brief was overseen by HealthPACT. HealthPACT comprises representatives from health departments in all States and Territories, the Australian and New Zealand governments and MSAC. It is a sub-committee of the Australian Health Ministers Advisory Council (AHMAC), reporting to AHMAC s Hospitals Principal Committee (HPC). AHMAC supports HealthPACT through funding. This Brief was prepared by Benjamin Ellery, Jacqueline Parsons and A/Prof Tracy Merlin from Adelaide Health Technology Assessment (AHTA), School of Population Health, University of Adelaide and Linda Mundy from the HealthPACT Secretariat..

Summary of findings At this stage, the stool-based DNA test, ColoGuard, appears to be a sensitive method for the detection of CRC and precancerous lesions / adenomas in an asymptomatic screening population, with a considerable trade-off in specificity compared to FOBT. This trade-off has implications for patients (due to resultant increase in unnecessary colonoscopies) and the healthcare system (additional costs). The acceptability of stool-based DNA tests compared to FOBT, and potential impacts on population screening programs, remains an area for further research. A cost-effectiveness analysis using the data from the study by Imperiale et al 51 would be beneficial to inform decision making on whether or not stool DNA testing has a place in screening for the prevention of CRC in Australia. Based on the available studies, the evidence on the accuracy of blood-based biomarker tests for CRC is incomplete and inconclusive due to inconsistent study results. One study on the blood-based test, Epi procolon, recruited a study population which was representative of the Australian FOBT screening population, but did not provide a direct comparison with FOBT and reported only sensitivity data (not specificity) for pre-crc pathology 41. Another study 35 on Epi procolon compared diagnostic accuracy results with FOBT, but recruited a much smaller study sample enriched with CRC-positive patients. Such a study design renders the results as non-applicable to the Australian screening setting. In Australia, the use of a blood-based test for CRC, ColoVantage Plasma, is being piloted following an agreement between the manufacturer and a participating health insurer. Diagnostic accuracy data on ColoVantage Plasma are anticipated with the publication of a trial including more than 2,000 Australian and Dutch participants. HealthPACT advice This assessment identified a number of new blood-based biomarker and stool-based tests for the detection of colorectal cancer that are currently being developed and undergoing testing both nationally and internationally including clinical trials. In comparison to the faecal occult blood test currently in use in the Australian National Bowel Cancer Screening Program, these tests are expensive and as such would be prohibitive for population screening. Although the stool-based DNA test, ColoGuard, appears to be a sensitive method for the detection of CRC and precancerous lesions/adenomas in an asymptomatic screening population; there is a reduction in specificity compared to the ifobt, which would result in an increased number of unnecessary colonoscopies being performed. This would not only have implications for the patient who would experience an unnecessary invasive and stressful procedure, but would also impact on existing colonoscopy waiting lists. In addition, there is a lack of evidence supporting the use of these tests to detect early adenomas. Blood and stool biomarker testing for CRC screening: July 2015 i

This assessment highlighted that Australian and international evidence supports that the ifobt is currently the most sensitive screening test for use in population screening programs for CRC. At this time, there is insufficient evidence to support the introduction of blood-based biomarker tests for the use in population screening in Australia; however; due to the rapidly evolving evidence base around a number of these tests, HealthPACT recommends that this technology be reassessed in 24 months. Blood and stool biomarker testing for CRC screening: July 2015 ii

Contents Summary of findings... i HealthPACT assessment... i Patient indication... 1 Description of the technology... 1 Company or developer... 2 Reason for assessment... 2 Licensing, reimbursement and other approval... 3 Disease description and associated mortality and morbidity... 5 Current technology... 9 International utilisation... 12 Diffusion of technology in Australia... 11 Cost infrastructure and economic consequences... 12 Ethical, cultural or religious considerations... 13 Evidence and Policy... 14 Safety and effectiveness... 14 Stool-based tests... 17 Blood-based tests... 23 Economic evaluation... 26 Ongoing research... 28 Other issues... 28 Appendix A... 30 References... 31 Blood and stool biomarker testing for CRC screening: July 2015 iii

Technology, Company and Licensing Register ID Technology name Patient indication Description of the technology WP151 Blood and stool biomarker tests for colorectal cancer screening People at average risk of colorectal cancer Bowel cancer, which includes cancers of the colon, recto-sigmoid junction and rectum, is a major cause of morbidity and mortality in Australia. In 2006, based on the evidence of several randomised controlled trials (RCTs), which demonstrated that bowel cancer mortality could be reduced by 15 33 per cent by regular bowel screening using the guaiac FOBT, Australia introduced a population-based bowel cancer screening program. 1 This program uses an automated immunochemical faecal occult blood test (ifobt) a 4 th generation FOBT - to detect minute traces of haemoglobin in stool samples. 2 Since the introduction of this screening program, several new colorectal cancer (CRC) diagnostic tests have been brought to market (Table 1). The aim of this Brief is to provide a preliminary assessment of the safety and diagnostic accuracy of these new blood and stool-based biomarker tests in comparison to the established FOBT, and in particular how they perform in the detection of early colorectal cancer and adenomas. It is not a systematic appraisal of all available evidence, nor is it placed to definitively report on the effectiveness of these new technologies in clinical and / or population screening settings. Blood and stool-based tests for CRC utilise in vitro diagnostic (IVD) test kits which target one or more biological markers (biomarkers) known to be involved in the development of cancer. Most of these tests are based on modifications of DNA sequences that are present only at low levels in normal cells, but at increased levels when cancerous cells a are present in the colon and rectum. Other tests are designed to target modified RNA sequences. 3, 4 Blood or stool samples are processed according to in-house laboratory protocols developed by private biotechnology companies. Occasionally, the development of these tests occurs in collaboration with universities and / or government agencies and non-government / not-forprofit organisations. For example, ColoVantage Plasma was co-developed by Clinical Genomics and the CSIRO b and it has undergone clinical evaluation by Flinders Centre for Innovation in Cancer, Flinders University of South Australia. 5 As yet, the results of this evaluation have not been published in a peer-reviewed journal. It is thought that DNA methylation occurs in the early stages of tumour development and can serve as a marker for precancerous lesions, including adenomas. 6 Several tests have been developed based on the detection of hyper-methylated DNA, with increased levels a To some extent, the expression of these DNA sequences is increased in pre-cancerous cells also. b CSIRO = Commonwealth Scientific and Industrial Research Organisation Blood and stool biomarker testing for CRC screening: July 2015 1

being detected in blood samples from individuals with colorectal polyps or CRC, with corresponding low levels (undetectable) in pooled plasma from individuals without colorectal neoplasia. 7 The majority of blood tests currently on the market for use in colorectal cancer detection, test for aberrant DNA methylation levels of the SEPT9 gene, which is involved in cytokinesis and cell cycle control. 6 These tests include the ColoVantage test c, Epi procolon and ms9 (see Table 1). ColoVantage Plasma test is designed to detect aberrant DNA methylation in two genes, BCAT1 and IKZF1, where hyper-methylation has frequently been observed in benign colorectal polyps and malignant cancers (colorectal neoplasia). 7 Company or developer Various companies have developed blood and stool tests to screen for CRC (see Table 1). Besides Clinical Genomics and the CSIRO producing the Australian DNA-based test (ColoVantage Plasma), international companies providing blood tests for CRC include Quest Diagnostics (USA), Epigenomics (Germany / USA), ARUP Laboratories (USA), Enzo Clinical Labs (USA) and Warnex Medical Laboratories (Canada). Exact Sciences Corporation (USA) offers stool-based DNA biomarker testing for CRC. Reason for assessment Colonoscopy is an invasive procedure that requires a bowel preparation regimen prior to the procedure with most jurisdictions having waiting lists for patient to access to semiurgent and routine colonoscopy in the public health system. 8 Colonoscopy is not used as a first-line CRC screening tool in Australia, which has an established CRC population-based screening program using the ifobt, with only those individuals testing ifobt positive being referred on to colonoscopy. The reduction in morbidity and mortality due to CRC FOBT screening is well-recognised; however, only 35 per cent of the 930,000 Australians invited to partake in FOBT in the 2011-2012 financial year participated. 9 Although it has been suggested that the need for stool sampling remains a barrier to the uptake of CRC screening using FOBT in the general population 10, 11, there is also evidence to suggest that faecal aversion becomes a non-issue once eligible individuals have participated in an FOBT screening program, as evidenced by continued compliance with such programs. 12 However, of concern is that blood-based tests for CRC are currently being marketed to consumers in Australia as a more convenient d and acceptable alternative to FOBT without a rigorous assessment of the safety and effectiveness of these tests in a non-symptomatic population. c Differs from the Australian developed ColoVantage Plasma test d For example, a blood sample for CRC testing may be requested at the same time a pathology form is completed for other blood work when a patient visits a general practitioner. Blood and stool biomarker testing for CRC screening: July 2015 2

Stage of development in Australia ColoVantage Plasma is the only test currently available to a limited population of Australians within a defined location (see Diffusion of technology in Australia). Development of an updated version of the test has begun e, and it is conceivable that more tests will become available over the next few years. Yet to emerge Experimental Investigational Nearly established Established Established but changed indication or modification of technique Should be taken out of use Licensing, reimbursement and other approval In general, IVDs and related instrumentation are pathology tests which are intended to assist in clinical diagnosis or making decisions concerning clinical management. The Therapeutic Goods Administration (TGA) regulatory framework for IVDs changed in July 2010, and in-house laboratory tests now receive a similar level of regulatory scrutiny as commercial kits. As the ColoVantage Plasma test is a DNA test and is currently provided only as an in-house IVD, it would be classified as a Class 3 in-house IVD. 13 Laboratories that manufacture in-house Class 3 IVDs are required to notify the TGA of the types of IVDs manufactured in each laboratory for inclusion on a register. These laboratories must be accredited by the National Association of Testing Authorities (NATA). In addition, they must demonstrate compliance with the suite of standards for the validation of in-house IVDs as published by the National Pathology Accreditation Advisory Council (NPAAC), for each test manufactured. 13 Several IVDs that target CRC specific DNA/RNA biomarkers were identified during the literature search as shown in Table 1. Where possible, details of the regulatory status for these IVDs in in the relevant jurisdictions have been provided. e Personal communication (email), Flinders Centre for Innovation in Cancer, 28 July 2014. Blood and stool biomarker testing for CRC screening: July 2015 3

Table 1 Summary of identified in vitro diagnostic medical tests for DNA/RNA based colorectal cancer screening IVD Developer Licensed provider/s of test (if different from developer) Description/stage of development Biomarker/s used Sample source Regulatory status ColonSentry GeneNews Ltd, Ontario, Canada Enzo Clinical Labs, Enzo Biochem Inc, NY, USA Available in US as an in-house IVD; US licence exclusive to Enzo Clinical Labs; established in Canada, Malaysia and China mrna; ANXA3, CLEC4D, LMNB1, PRRG4, TNFAIP6, and VN1 (over-expressed in CRC), and IL2RB (under-expressed in CRC relative to bowel tissue) Blood Approved by New York State Department of Health ColoGuard Exact Sciences Corporation, Wisconsin, USA Laboratory Corporation of America (LabCorp), North Carolina, USA Ready for commercialisation in North America DNA + immunochemical; combines assays for KRAS mutations, aberrant NDRG4 and BMP3 methylation, and β-actin with a haemoglobin immunoassay Stool Granted PMA by FDA in August 2014 a ColoVantage Quest Diagnostics, New Jersey, USA This version of the test for the USA market developed under licence from Clinical Genomics (see entry below) Commercialised in North America DNA; aberrant methylation of SEPT9 Blood No regulatory information on this test found ColoVantage Plasma Clinical Genomics Pty Ltd, NSW, Australia NA Limited use in Australia commenced in July 2014 with a pilot in cooperation with local GPs and health insurer; inhouse testing by Clinical Genomics DNA; aberrant methylation of BCAT1, IKZF1 Blood Can be used under TGA regulations governing the marketing and use of Class 3 IVDs Epi procolon Epigenomics AG, Berlin, Germany ARUP Laboratories, Utah, USA; Warnex Laboratories, Quebec, Canada (since merged with DIAGNOS Inc, Quebec, Canada) Nearing commercialisation in the USA (ARUP), Canada (Warnex/DIAGNOS Inc) and China (BioChain Institute Inc); established in Europe DNA; aberrant methylation of SEPT9 Blood CE marked since 2012; FDA has not granted PMA on grounds that Epigenomics have not demonstrated increased screening compliance in the target population ms9 Abbott Molecular, Illinois, USA Worldwide non-exclusive licensing Established Europe and Asia- Pacific DNA; aberrant methylation of SEPT9 Blood CE marked DNA, deoxyribonucleic acid; NA, not applicable; IVD, in vitro diagnostic medical device; mrna, messenger RNA (ribonucleic acid); TGA, Therapeutic Goods Administration;, FDA, US Food and Drug Administration; PMA, premarket approval a In the USA, CMS (Centers for Medicare and Medicaid Services) are currently considering whether to cover testing using Cologuard once every three years for Medicare beneficiaries between the ages of 50 and 85, and who are asymptomatic for colorectal disease and at average risk of CRC. Blood and stool biomarker testing for CRC screening: March 2015 4

Australian Therapeutic Goods Administration approval Yes No Not applicable Technology type Technology use ARTG number not applicable (see Error! Reference source not found. above) Diagnostic (triage test) Diagnostic / screening Disease description and associated mortality and morbidity In normal body tissues, cells constantly multiply and die in a regulated way. However, the cell s regulatory mechanisms do not always function correctly, and abnormal cells which multiply and spread in an uncontrolled manner can result in one of the many diseases broadly referred to as cancer. Cancers differ by the specific type of cell involved and the specific organ or body in which the disease began. 14 Colorectal cancer (CRC) develops via a multi-stage process in which a series of cellular mutations occur in the epithelial cells that line the large intestine (i.e. the colon and rectum). Most commonly, CRC develops over time from benign adenomas, which can vary in size from tiny nodules to polyps many cm across, but can also arise from de novo lesions. Given the relatively slow disease progression, the early detection and removal of small cancers, and polyps that may become cancerous, is recognised as an effective strategy to 15, 16 prevent morbidity and mortality due to CRC. Epidemiological data indicate that CRC is the second most frequently occurring cancer in Australia. For the year 2010, the age standardised incidence of CRC was 61.8 per 100,000 f for both sexes combined. In the same year, 73.7 males per 100,000 were diagnosed with CRC, compared with 51.1 females per 100,000 (Figure 1). While the number of cases of CRC diagnosed over the last decade has increased for both males and females (Figure 2), this is a reflection of Australia s ageing population. It is well recognised that the risk of CRC increases with age, as indicated by the differences observed when age-specific incidence and mortality are compared to the age-standardised rates (compare Figure 1 with Figure 2 and Figure 3). Compared to known deaths from other cancer types, CRC remains the second most common cause of death due to cancer in Australia (16.3 deaths per 100,000 in 2011). More men die from CRC compared to women; in 2011, there were 19.7 and 12.7 deaths per 100,000 g among Australian males and females, respectively (Figure 3). 17 f Adjusted to the Australian 2001 population. g Rates adjusted to the Australian 2001 population. Blood and stool biomarker testing for CRC screening: March 2015 5

Figure 1 Colorectal cancer incidence and mortality by year, expressed as rate per 100,000 and agestandardised to Australian 2001 population 17 Figure 2 Age-specific colorectal cancer incidence rate by age group in Australia, expressed as cases per 100,000 for the year 2011 17 Blood and stool biomarker testing for CRC screening: March 2015

Figure 3 Age-specific colorectal cancer mortality rate per 100,000 by age group in Australia, 2011 17 In New Zealand, published data for CRC incidence and mortality are available up until 2010 18, and unless otherwise specified, all rates described here are for that year. The incidence of CRC, based on the number of registered cases, was 45 per 100,000 when standardised to the WHO h world population. Similar to the rates observed in Australia, more cases of CRC are diagnosed in New Zealand men than in women (49.4 versus 41.1 cases per 100,000), although the difference observed by sex is less than in Australia. CRC is the second most common cancer registered in New Zealand and the second most common cause of death from cancer, accounting for 14.1 per cent of cancer deaths. Unlike Australia, the agestandardised incidence of CRC in New Zealand declined between 2000 and 2010; however, the WHO world standard population differs markedly from the Australian 2001 standard population, and this is likely to account for the major part of this difference. There are notable differences in CRC registration and mortality rates between Māori and non-māori New Zealanders. In fact, CRC is one of few cancers for which registration and mortality rates among Māori have been historically lower than the rates observed among non-māori. In 2010, CRC was the fourth most commonly registered cancer and fourth most common cause of death from cancer for Māori. For non-māori, CRC was the most commonly registered cancer and the second most common cause of death from cancer. Selected registration and mortality data for CRC by sex and ethnicity (i.e. Māori versus non- Māori) are shown in Table 2. 18 h World Health Organization Blood and stool biomarker testing for CRC screening: March 2015

Table 2 Age-standardised registration and mortality rates for colorectal cancer in New Zealand, by sex and ethnicity, 2010 18 Registrations (per 100,000) Deaths (per 100,000) Males Maori Non-Maori Total 35.9 50.2 49.4 16.7 19.7 19.6 Females Maori Non-Maori Total 34.4 41.7 41.1 10.8 15.2 15.0 Rates shown are standardised to the World Health Organization world population and expressed as the number of registrations/deaths per 100,000 Speciality Technology setting Various i Primary care and community care (for testing); general hospitals and private consulting rooms for specialist followup where necessary, i.e. when further investigation (colonoscopy) is indicated after a positive screening result Impact Alternative and/or complementary technology Blood-based tests for CRC are investigational and are not yet at a stage of development to be considered for population screening or to replace screening with FOBT. Blood-based tests are being marketed as an alternative for individuals who may consider faecal sampling to be an unacceptable screening modality; however access to blood-based tests is limited at present and would require that patients pay for the full cost of testing. The appropriateness of blood-based tests as a complementary technology alongside FOBT in a population screening program will depend on factors such as evidence supporting the applicability of blood-based tests to the population eligible for FOBT screening, cost, accessibility and acceptability to patients. However, this Brief is not intended to be a systematic review of comparative effectiveness of different tests for CRC in a population screening setting, but rather to provide a preliminary indication on the diagnostic accuracy of blood-based and stool-based biomarkers for CRC, compared to FOBT where possible. Relevant details on cost, level of technology diffusion in Australia, and ethical implications are also briefly discussed. i A general practitioner would order the test and collection would most likely be through a standard pathology service. The blood sample would then be transported for processing to an in house laboratory. Individuals with positive results would then be referred for further investigation, usually by colonoscopy. The specialists of relevance in the management of patients with polyps or CRC would include gastroenterologists, colorectal surgeons, radiologists and radiation oncologists. Blood and stool biomarker testing for CRC screening: March 2015

Current technology In 2006 the Australian Government introduced the National Bowel Cancer Screening Program (NBCSP) which offered ifobt to individuals turning 55 and 65 years of age The program was expanded to include 50 year-olds in July 2008 and 60 year olds in July 2013. 1 From January 2015, the NBCSP is inviting Australians turning 50, 55, 60, 65, 70 and 74 years of age and the program is being incrementally expanded to include all Australians aged 50 to 74 years by 2020 who will be invited to participate in a biennial FOBT screen. 19 The most recent data collected on the NBCSP indicates of the 963,518 invitations issued resulted in an overall participation rate of 33.4 per cent for the 12-month period between June 2012 and June 2013. A number of individuals opted out of the program (39,525) giving an overall response rate of 37.5 per cent (50, 55 and 65 year olds only). The low participation rate may be due to a number of factors including: lack of awareness about the screening program, the FOBT or about the seriousness or consequences of CRC; remoteness, with higher rates of participation in major cities and regional areas compared with lower rates in remote and very remote areas; age: participation rates increased with age and therefore participation rates are expected to increase with the expanded program including Australians aged 50 to 74 years; sex: females were more likely to participate; 20 and a significant number of individuals opting for screening outside of the NBCSP by FOBT, e.g. the Rotary Bowelscan Program or by colonoscopy, usually, but not always, for appropriate clinical reasons. 21 Definitive diagnosis following a positive FOBT result requires subsequent investigation, usually by colonoscopy, the gold standard for detection of pre-cancerous adenomas and CRC. Of those tests completed and returned during the 2012-2013 period, 7.5 per cent were positive, and approximately 70 per cent of these individuals are known to have undergone colonoscopy. Of these, one in 32 were diagnosed with a confirmed or suspected cancer and 19, 20 at least one in 17 were diagnosed with advanced adenoma. NBCSP data obtained from Queensland (2006-2010) indicates that 182,434 participants returned a valid FOBT kit, of which 14,005 individuals (7.7%) had a positive FOBT result. 1 Complete histology data were obtained from the 4,817 (34.4%) individuals who tested positive and who underwent a colonoscopy in the public hospital system. Of these, 3,072 (63.8%) underwent a polypectomy, 2,311 (48.0%) were diagnosed with an adenoma and 209 (4.3%) were diagnosed with CRC. Definitive diagnostic accuracy measures of the FOBT performance in the NBCSP cannot be calculated due to the missing data describing the remaining 9,188 individuals who tested FOBT positive and did not undergo a colonoscopy in the public health system. Blood and stool biomarker testing for CRC screening: March 2015

The purpose of bowel screening using FOBT is to detect low grade bleeding from CRCs and adenomas. The ifobt has a positive predictive value (PPV) of 3.6 per cent for bowel cancer, that is, of those people with a positive ifobt result, 3.6 per cent tested FOBT positive and were actually diagnosed with bowel cancer. It is important to note that the PPV reflects test sensitivity and the low prevalence of bowel cancer in the population being screened. ifobt sensitivity and specificity for bowel cancer are 83.4 per cent and 92.6 per cent respectively. However of greater importance in a screening test, the negative predictive value (NPV) of the ifobt for bowel cancer is 99.9 per cent, giving reassurance that those individuals who are FOBT negative truly do not have bowel cancer. FOBT performance results are summarised in Table 3. It was not possible for the analysis of the NBCSP to report a PPV or NPV for adenomas. Table 3 Performance of Magstream HemSp, the ifobt in the NBCSP 2006-08 1 Screening result Cancer diagnosed Cancer not diagnosed Total Positive FOBT 887 3.6% PPV 23,899 24,786 Negative FOBT 176 0.06% false negatives 297,378 99.9% NPV 297,554 Total 1,063 83.4% sensitivity PPV = positive predictive value, NPV = negative predictive value 321,277 92.6% specificity 322,340 A more in-depth analysis of the results of the NBCSP was conducted for the 2-year period 2006-08. Of the 1,418,068 eligible invitees, 42 per cent (595,705) returned the FOBT. A positive FOBT was reported in a total of 43,764 (7.3%) of these individuals. The outcomes of those testing positive and the 92.7 per cent of individuals who tested negative are summarised in Table 4. Table 4 Outcomes of the NBCSP 2006-08 1 Screen detected outcomes: 43,764 (7.3%) individuals tested positive Cancer 2 years Cancer >2 years Advanced adenomas Other adenomas No cancer or adenoma 1,518 (3.5%) 57 (0.1%) 3,282 (7.5%) 1,928 (4.4%) 36,979 (84.5%) Non-positive screen and follow-up: 551,941 (92.7%) Cancer 2 years Cancer >2 years No cancer 265 (0.05%) 316 (0.06%) 551,360 (99.9%) Blood and stool biomarker testing for CRC screening: March 2015

A further analysis of data from a study population obtained from this screening cohort indicated that of those people with a diagnosis of bowel cancer who were invited to participate in the NBCSP, 11.4 per cent died. Of those people never-invited to participate in the NBCSP who had a diagnosis of bowel cancer, 19.6 per cent died, translating to a 68 per cent higher risk of death when compared to NBCSP invitees. After correction for lead-time bias there was still a statistically significant 15 per cent higher risk of death in the neverinvited group. This reduced mortality is thought to be due to the earlier detection of bowel cancer due to the screening program, allowing for better treatment options and prognosis, compared to non-invitees who present with advanced bowel cancer. In New Zealand, a four year pilot screening program commenced in 2011 to determine if a national CRC screening program is warranted; however implementation on the national level will not occur prior to a full evaluation of all collected pilot program data. 22 Diffusion of technology in Australia In late July 2014, a pilot of the ColoVantage Plasma test commenced in the Blue Mountains region of New South Wales. The Blue Mountains region was chosen because: (a) there is access to a supportive local health insurer (see Cost infrastructure and economic consequences); (b) pathology services are readily accessible, and; (c) the area is known to have a strong network of local primary healthcare providers. j This allowed residents of the Blue Mountains to have access to blood-based CRC testing through their general practitioner (GP) who arranged for patients to have a blood sample taken at a pathology collection centre. Patients could then return to their GP at a later date for the results of the test. The stated aim of the pilot was to work through the logistics of collecting samples, collate feedback from local GPs, and monitor the participation of Blue Mountains residents. k Based on this information, it would appear that the data being collected is for internal purposes, rather than generating data for an evidenced based comparison of ColoVantage Plasma with FOBT, the established screening modality in Australia. A trial, which is expected to provide relevant diagnostic accuracy data for ColoVantage Plasma test is underway, but results are yet to be published l. j http://www.clinicalgenomics.com/media/media-releases. k Personal communication (email) with Palin Communications, public relations consultancy on behalf of Clinical Genomics, 28 July 2014. l Personal communication (email) with lead study investigator, 28 July 2014. Blood and stool biomarker testing for CRC screening: March 2015

International utilisation Country Level of use of CRC biomarker-based screening Trials underway or completed Limited use Widely diffused Canada Germany USA Malaysia China Spain Japan Cost infrastructure and economic consequences The cost of the ColoVantage Plasma test to consumers in the Blue Mountains trial is $125 per test. Clinical Genomics have an arrangement with private health insurer Westfund Health. Westfund Health has agreed to cover the test for their members, but the test will cost $125 for participants who are not insured by Westfund. l The costs of other tests developed/available in the US are summarised in Table 5. A recent online news report noted that the developer of Cologuard, Exact Sciences, has proposed a discounted price of US$502 per test to the Centers for Medicare and Medicaid Services (CMS), in the event that a decision is made to cover Medicare beneficiaries between the ages of 50 and 85 years. According to Exact Sciences, the proposed pricing for Cologuard is based on Medicare pricing for tests for KRAS gene mutations, faecal haemoglobin testing, and RAR-α (retinoic acid receptor alpha) genetic testing for leukaemia. 23 Table 5 Cost comparison of selected biomarker-based colorectal cancer screening tests and FOBT24, 25 ColonSentry Epi procolon Cologuard FOBT Sample source Blood Blood Stool Stool Targeted marker(s) mrna DNA; aberrant methylation of SEPT9 DNA + immunochemical; combines assays for KRAS mutations, aberrant NDRG4 and BMP3 methylation, and β-actin with a haemoglobin immunoassay Haemoglobin detected by immunoassay Price to payer per test (USD) $795 $150 $599 $10-$40 DNA, deoxyribonucleic acid; FOBT, faecal occult blood testing; mrna, messenger RNA (ribonucleic acid); USD, US dollars Blood and stool biomarker testing for CRC screening: March 2015

It is apparent that even the least expensive of these biomarker-based tests are many times more costly than an FOBT (cost range A$10-$40 depending on the test used m ). 26 In the event that the diagnostic accuracy of a CRC biomarker test outperforms FOBT, any potential downstream health benefits of a funded screening program incorporating CRC biomarker testing will be achieved only at a substantially higher cost outlay. Cost-effectiveness will also be predicated on the uptake of these tests 27, i.e. if blood-based CRC tests are adopted as a replacement for FOBT screening, then significant increases in screening compliance will need to occur, leading to significant additional health gains, in order to offset the increase in costs. Costs of biomarker-based tests might reduce if used widely. Ethical, cultural or religious considerations Companies marketing IVDs for CRC screening using genetic material have an ethical imperative to provide transparency about the limitations as well as the benefits of their tests. Comparisons with FOBT are therefore required before biomarker-based CRC screening can be marketed as a replacement or alternative screening modality (see section on Safety and effectiveness). A positive result will still require clinical management which is no different from that occurring after a positive FOBT result, i.e. colonoscopy is indicated. However, if the biomarker-tests are not as accurate at triaging patients at risk of CRC than FOBT, then given the screening population is healthy false positive test results will lead to unnecessary colonoscopies and biopsies (leading to physical and psychological impacts on the patient and opportunity costs for the health system n ), while false negative test results will mean that patients forgo early identification of a potentially fatal disease. If any government funded screening program for CRC using genetic material is to occur in the future, the public invited or seeking CRC screening will need to be informed of the same limitations (and advantages) of the test as is currently provided for FOBT screening, i.e. the likelihood of a false positive or false negative result and the consequences of a positive test. It is worth noting that there are differences in the type of tests used in screening and predictive genetic testing. The main difference is that predictive tests seek to determine risk of future disease development for hereditary disorders, whereas DNA screening tests are more aligned in purpose to conventional medical diagnostic tests which define a current condition. DNA screening tests, unlike predictive genetic testing, provides a result which leads to the diagnosis or exclusion of disease by a confirmatory testing method. Although this may have implications for the future, the main utility lies in the information it provides about the current health state. In the context of CRC biomarker testing, asymptomatic persons are screened for changes consistent with the presence of early CRC or pre-crc pathology. 28 Results are dichotomous m http://www.inwmml.org.au/_uploads/_ckpg/files/cspii_inwmml_fobt_infosheet_jan_13.pdf n That is, in terms of how the funding that has been consumed for unnecessary colonoscopies might have been used for other health services. Blood and stool biomarker testing for CRC screening: March 2015

(either positive or negative) for current disease status and are intended to inform clinical management. Therefore, the management of the ethical implications unique to predictive testing for heritable CRC risk (e.g. the need for genetic counselling) is not applicable to screening for CRC. The uptake of stool-based DNA tests in clinical practice may be affected by the same issues affecting the FOBT, i.e. the preference to avoid faecal sampling among some groups in the screening population. This may vary across different ethnic, cultural and religious groups. As indicated previously, once uptake has occurred, it is known that compliance in an FOBT program, with biennial screening, is likely to be ongoing. No specific information regarding the implications of stool-based or blood-based DNA tests for CRC among different cultural or religious groups was located at the time of preparing this Brief. Evidence and Policy Safety and effectiveness Studies describing a variety of candidate biomarkers for CRC screening tests are abundant in the scientific and medical literature. Many of these studies report on prototype assays without clinical results (i.e. experimental studies). 29-33 These studies are not discussed further. Recent relevant studies reporting biomarker test accuracy against a relevant reference standard (colonoscopy and / or surgical or histological findings) were selected for the discussion of safety and effectiveness. The main study characteristics and results are summarised in Table 6 (stool-based tests) and Table 8 (blood-based tests). A glossary of screening related terms is provided in Appendix A. Of the 11 clinical studies that were identified, nine were diagnostic case-control studies (level III-3 evidence), with cases being selected from among populations either scheduled to undergo or already having undergone colonoscopy or surgery. All 11 studies (see Table 6 and Table 8) reported diagnostic accuracy results of the biomarker test relative to a reference standard of colonoscopy or histopathology, and five studies 34-38 also reported the results of various FOBTs against the reference standard. Two studies reported that the markers under investigation were poor predictors for the presence of CRC 36, 39, i.e. they were found/known a priori to be associated with other cancer types in addition to CRC o 37, 40, or the association with other cancer types remains uncertain. Additionally, the biomarkers selected by Tao and colleagues 37 yielded a sensitivity to detect CRC and advanced adenomas which was much lower than the sensitivity observed with o Sheng and colleagues investigated the potential of transferrin as a biomarker for CRC. Blood and stool biomarker testing for CRC screening: March 2015

ifobts, limiting the use of the chosen markers as alternatives to traditional CRC screening. p Further study of these investigational biomarkers appears to have been largely discontinued due to the lack of promising results. The results of these studies are therefore only of limited relevance to this Brief and are not discussed further. In contrast to the findings described above, a number of studies 34, 35, 38, 41-43 (Table 6 and Table 8) have reported diagnostic accuracy of IVDs for CRC which is similar to the diagnostic accuracy observed for an ifobt, using biomarkers that are considered to be highly specific to CRC. Proprietary biomarkers including SEPT9, NDRG4, BMP3, BCAT1, and IKZF1 have been prominent in the literature during recent years and there has been progressive development of tests employing these markers. As indicated in Table 1, several of these testing protocols are ready for commercial use, or are soon to receive regulatory approval overseas. Three studies involving such tests are discussed in more detail below. p Tao et al selected a range of inflammatory peptides / proteins including C-reactive protein, complement C3a anaphylatoxin and tissue inhibitor metalloproteinases. As noted above, the role of these proteins in cancers other than CRC cannot be ruled out. Blood and stool biomarker testing for CRC screening: March 2015

Table 6 Studies of stool-based biomarkers identified as potential CRC screening test alternatives Study design and level of evidence Location, population and study sample size Biomarker(s) CRC specific? Comparator test Reference standard Accuracy outcomes a *Imperiale et al 2014 34 Multi-centre study of test accuracy, with blinded comparison against a valid reference standard among consecutive patients Level II evidence USA / Canada N=9,989 Population: asymptomatic persons 50-84 years considered to be at average risk for colorectal cancer and who were scheduled to undergo screening colonoscopy. DNA + immunochemical; combines assays for KRAS mutations, aberrant NDRG4 and BMP3 methylation, and β-actin with a haemoglobin immunoassay (Cologuard) Other cancers, particularly organs connected to the digestive tract (i.e., pancreas and liver), may shed markers which are targeted in the Cologuard test Immunochemical FOBT (OC FIT- CHEK, Polymedco) Colonoscopy The sensitivity of Cologuard and FOBT in the detection of CRC was 92.3% and 73.8%, respectively (p=0.002) In the detection of advanced precancerous lesions the sensitivities were 42.4% and 23.8%, respectively, for Cologuard and FOBT (p<0.001) Cologuard and FOBT yielded specificities of 89.8% and 96.6%, respectively, among participants with a negative result on colonoscopy (p<0.001) Sheng et al 2009 36 Diagnostic casecontrol study Consecutive patients Level III-3 evidence China N=110 Population: Asymptomatic, low-risk patients (n=34), mean age 61 years (range 35-82 years) Pre-malignant (n=36), mean age 60 years (range 60-84 years) CRC (n=40), mean age 63 years (range 38-81 years) Protein; transferrin No Immunochemical FOBT (Hemosure ifob Test, WHPM, Inc, USA) Colonoscopy with histological confirmation of findings The transferrin assay detected 76.4% of CRCs and premalignant lesions compared to 69.0% by FOBT 95%CIs NR Note: The markers under investigation were poor predictors for the presence of CRC, i.e. they were found/known a priori to be associated with other cancer types in addition to CRC. These results are not discussed further. CRC, colorectal cancer; DNA, deoxyribonucleic acid; FOBT, faecal occult blood test; NR, not reported, a Overall values for CRC and premalignant lesions combined, unless otherwise specified * Denotes a study selected for specific discussion of results based on the following: most recently published, highest level of evidence available, and identified in the methods a test intended for commercial use as opposed to studies involving prototype assays or studies less representative of clinical practice, by comparison. Also, the results based on tests using markers known to be, or potentially associated with cancers other than CRC have not been discussed. Blood and stool biomarker testing for CRC screening: March 2015 16

Stool-based tests Imperiale et al 2014 A large multicentre study (N=12,776) 34 recruited persons aged 50 to 84 years from across 90 sites q in the USA and Canada (level II diagnostic evidence). The aim of the study was to assess the diagnostic performance of a multi-target stool-based CRC screening test, which is being marketed in the USA as Cologuard (Exact Sciences Corporation, Madison, WI, USA), compared to FOBT (OC FIT-CHEK, Polymedco). Eligibility for inclusion required that participants were asymptomatic and at average risk for CRC (e.g. no previous positive FOBT result within 6 months, rectal bleeding or history of CRC or bowel disease), suggesting the included study population was similar to the population eligible for FOBT screening in Australia. Study participants were also scheduled to undergo colonoscopy within 90 days from the date of providing informed consent. Cologuard testing involved collection of stool samples which were received by a single centre for distribution for processing r among three laboratories in the USA. Each laboratory received a similar distribution of stool specimens on the basis of colonoscopy findings and testing was performed in a blinded manner, i.e. results of colonoscopy and FOBT (described below) were not revealed to laboratory personnel. FOBT (OC FIT-CHEK) was performed as per instructions provided by the manufacturer (Polymedco, Cortlandt Manor, NY) and used the same sample provided for the Cologuard test. Refrigerated samples were sent in batches to a single laboratory for blinded analysis. A positive FOBT result was indicated by the presence of haemoglobin exceeding 100 ng s per millilitre of buffer solution. Colonoscopy involved a routine bowel preparation for each patient following stool collection; however, no dietary or medication restrictions were required. Colonoscopists were required to describe the extent of examination (whether the endoscope was advanced to the caecum t ), rate the quality of bowel preparation (using a modified Aronchick scale 44 ) and record the size and location of lesions. Biopsy and surgical specimens were sent to the histopathology laboratory typically used for each study site. High grade findings were sent to a central site for a second assessment by a gastrointestinal pathologist. Disagreements were resolved by consensus of a least two central pathologists. The analysis of findings was based on data from all participants who had valid results for Cologuard, FOBT and colonoscopy (n=9,989) u. Results from the main analysis are provided in Table 7. q Private practice and academic settings. r Buffered stool samples were homogenised, separated in to aliquots and frozen at -80 C on receipt. s Nanograms. t The caecum is considered to be the beginning of the large intestine. u Of the original 12,776 participants recruited, 464 withdrew consent, 1,168 did not undergo colonoscopy and 128 did not provide a stool sample. Of the 11,016 participants remaining, 689 did not have a valid stool DNA Blood and stool biomarker testing for CRC screening: March 2015 17

The main strength of this study was the design, which included a valid reference standard, and recruited a large study population of persons aged over 50 at average risk for CRC. This study population is mostly applicable to the eligible screening population in Australia, especially given the National Bowel Cancer Screening Program will be offered to average risk individuals ranging from 50 to 74 years of age by 2020. The data tabulated in Table 7 indicate that Cologuard is more sensitive than the FOBT used in this study for the detection CRC and advanced adenoma. However, the FOBT has a significantly higher specificity, which as previously discussed, is of importance in a screening test. The higher sensitivity of the Cologuard test comes with the associated trade-off of low specificity, meaning that in a population screening setting, a higher false positive rate would be expected, along with a higher number of unnecessary colonoscopies. The balance of sensitivity and specificity of the test and the resultant impact on individuals and the health system would need to be considered along with other criteria should the test be assessed for suitability for population screening. It should be emphasised that the stool-based DNA test described by Imperiale et al is not an exclusively genetic assay; it does contain an immunochemical component to detect the presence of haemoglobin in the stool and the results should be interpreted accordingly. The authors reported that the haemoglobin component in isolation yielded diagnostic accuracy that was not significantly different to the diagnostic accuracy of FOBT. Imperiale et al reported no safety issues associated with the use of Cologuard. The study received financial support from Exact Sciences Corporation. test result, 304 did not have a valid colonoscopy result and 34 did not have a valid FOBT result. Thus 9,989 participants were included in the final analysis. Blood and stool biomarker testing for CRC screening: March 2015 18