Genetic testing for inherited disease susceptibility is a powerful tool that helps to reduce morbidity and mortality associated with familial cancer syndromes. The identification of individuals with germ-line mutations in BRCA1 and BRCA2 (associated with hereditary breast and ovarian cancers); MLH1, MSH2, MSH6, and PMS2 (associated with Lynch syndrome); and RET (associated with familial medullary thyroid cancer), among many others, has led to targeted screening and interventions that can lead to reductions in morbidity, and in some cases, mortality.1,2 Although it is generally assumed that familial cancer syndromes are largely identifiable through a thorough assessment of the family history, there are overlaps in cancer susceptibility syndromes (e.g., ovarian cancer can be due to either a BRCA1/2 or a Lynch syndrome–associated mutation); moreover, family history can be limited by adoption, small family size, and poor communication within families. Genetic evaluation usually has proceeded in a serial fashion, starting with the most likely genes and then, if testing for these is negative, consideration of additional testing. However, this process can lead to “testing fatigue” in both providers and patients, incomplete evaluation when patients do not return to complete the recommended testing, and considerable lengthening of the time frame required for testing. Against this background, advances in technology have allowed development of multiplex gene panels in which many genes (from 6 to 110) can be assessed simultaneously by massively parallel sequencing. The potential advantage of this approach is an efficient and timely evaluation that may be only marginally more expensive than standard-of-care genetic testing, or in the case of serial testing, potentially less expensive. However, little is known to date about key aspects of such testing, including the likelihood of a positive result, the rates of variants of unknown significance, and the clinical utility of testing using multiplex panels.

In this issue of Genetics in Medicine, LaDuca et al.3 describe their initial results obtained from tests using four multiplex panels performed in a Clinical Laboratory Improvement Amendments–certified commercial laboratory (Ambry Genetics) on a large series of individuals. These panels, similar to those offered by other commercial laboratories (Invitae, GeneDx, Fulgent Diagnostics, University of Washington, and Myriad Genetics), contain a mix of high-penetrance and presumed moderate-penetrance genes. In general, guidelines for clinical management are available for individuals with mutations in high-penetrance, but not in moderate-penetrance, genes. It is worth noting that BRCA1 and BRCA2 were not initially included in these versions of the Ambry panels due to patent concerns (although they are currently included). Questions and concerns regarding the use of these panels have been previously published4 and include (i) finding unanticipated high-penetrance mutations (e.g., detecting a TP53 mutation in a family without features consistent with Li–Fraumeni syndrome); (ii) managing high rates of variants of uncertain significance (VUSs); (iii) determining the clinical utility of mutations in moderate-penetrance susceptibility genes; and (iv) the rationale for the selection of genes on the panels because mutations in some have not been definitively associated with susceptibility either for cancer at all (e.g., XRCC2—not included in the Ambry Genetics panel but part of other commercially available panels) or the cancer for which the panel is intended (e.g., MUTYH in a breast cancer panel).

The study by LaDuca et al.3 provides critical data to begin to answer these important questions. Of 2,079 patients, 8.3% had deleterious mutations in the genes tested (determined by the proprietary Ambry Variant Analyzer according to the guidelines of the American College of Medical Genetics and Genomics), with frequencies ranging from 7.2% of those tested with the ovarian cancer panel to 9.6% with the “cancer” panel. Of the total, importantly, only 71 patients (3.4%) had mutations in genes for which clinical guidelines are available (TP53, PTEN, MLH1, MSH2, MSH6, PMS2, SMAD4, APC, biallelic MYH, CDH1, and STK11). The majority (47/74 (64%)) of these mutations were found in the colon cancer panel and had corresponding clinical guidelines. Of the patients tested with the colon cancer panel, 8% (46/557) had a positive clearly clinically actionable result, as compared with <1% (7/874) with such a result detected using the breast cancer panel. This result is not surprising because of the number of genes in which mutations are known to confer a greatly increased risk of colorectal cancer and thus illustrates a much higher clinical utility for panel testing in this context. However, the utility of identifying mutations in moderate-penetrance genes even on the colon cancer panel remains unclear.

Unanticipated findings in high-penetrance genes can be viewed as both an advantage and a disadvantage of multiplex testing. As an example, the authors report that 30% of patients with mutations in the colon panel would not meet the criteria for clinical testing. However, this result must be interpreted with caution because neither all the family history nor all the patient information may have been reported; moreover, the particular mutations that were detected were not specified. The counterargument is that the detection of unanticipated high-penetrance mutations associated with specific syndromes—such as mutations in (i) CDH1 (associated with hereditary diffuse gastric cancer), for which the standard recommendation would be prophylactic gastrectomy, and (ii) TP53 (linked to Li–Fraumeni syndrome), which is associated with a particularly high morbidity and mortality—are challenging to manage in the absence of a supportive family history. Interestingly, no STK11 (Peutz–Jehgers syndrome) mutations were identified using any panel, suggesting that it is recognized only clinically. In this cohort, 10 TP53 and 2 CDH1 mutations were detected. Three of the TP53 mutation carriers were diagnosed with breast cancer before the age of 35 years, so genetic testing would have been warranted clinically. Pretest counseling for panel mutation testing must include a discussion of the low, but real, likelihood of an unanticipated finding that could have major implications for medical management. On the basis of previous work in presymptomatic testing for TP53 mutations in family members of those with known mutations, 45% of individuals decline testing.5 Counseling models for multiplex testing must incorporate this consideration.

Most of the deleterious mutations found in this series were in moderate-penetrance genes for which there are no clinical management guidelines and thus no clear clinical utility. Even for genes about which much is known (such as CHEK2), how to use mutational information for clinical care is uncertain. In addition, the discovery of a mutation in such a moderate-penetrance gene may not provide a compelling explanation for the family history that may have prompted testing in the first place. Cancer penetrance associated with CHEK2 may be influenced by specific mutations (e.g., CHEK2 1100delC versus I157T),6 as well as the associated family history.7 Risks of other cancers, such as colon cancer, appear to be modestly elevated, but whether this should alter screening recommendations (including age at which to begin or frequency of screening) is unknown. For many of the other genes (such as NBN, BRIP1, and MRE11), even less is known about their true association with cancer susceptibility. Clinical genetic testing should be considered in the context of its use as a tool to improve the medical management of patients. Thus, because the identification of deleterious mutations in moderate-penetrance genes currently does not influence medical management, their inclusion in commercially available genetic testing panels should be carefully considered going forward.

As anticipated, VUSs were frequently observed, with rates of 15–26% (approximately double the deleterious mutation rate). Critical for ordering providers is the need to prepare patients for the possibility of this type of result. Clinical management remains based on the family history in this setting. Providers and testing laboratories also need to establish methods to reexamine these results periodically, particularly VUSs in high-penetrance genes. In addition, programs that offer tracking of VUSs in families with disease, without burdening the patient with additional cost, are of great value in providing additional information to determine pathogenicity. Although the high rate of VUSs is of concern, it is anticipated that with additional testing, rates will decline as they have with BRCA1/2 testing. However, it may be more difficult to determine the significance of VUSs in moderate-penetrance genes, adding to complexity and potential confusion for both patient and provider.

Multiplex panel testing for cancer susceptibility holds promise, and the study by LaDuca et al.3 has provided us with important initial information. The VUS rate is reasonable for this early stage of panel use. The yield of mutations in genes with associated clinical management guidelines, particularly in breast cancer patients, is low but is appreciable in those evaluated for colon cancer. Panel testing for inherited mutations may equip non–cancer geneticists with a mechanism with which they can identify individuals who carry a mutation associated with a cancer susceptibility syndrome so that they can be appropriately referred to cancer genetics centers, which may be particularly important for diseases with high inherited mutation rates, such as pheochromocytoma.8 However, the inclusion of moderate-penetrance genes in these panels remains a concern because currently the identification of deleterious mutations in these genes does not (and should not, without proper evidence) lead to changes in medical management. The results of this study emphasize the need for international collaborative efforts to study rare moderate-penetrance mutations and to develop standard-of-care guidelines based on accurate estimates of cancer risk for the benefit of both patients and their providers.


The authors declare no conflict of interest.