Introduction
The intention to carry out radiology and pathology reviews was noted in the original FH01 protocol, although their design and conduct was not specified. In this chapter, we summarise the protocols of the reviews, report progress on the reviews and give some preliminary results. The reviews are substantial pieces of work, involving the collation from multiple centres of radiological images and biological material. Consequently, they are still ongoing, but there are already some interesting observations, notably for the radiology review.
Radiology review
Radiology review is standard practice in major screening studies.61,62 This radiology review has two components: a rereading of mammograms of cancers and selected non-cancer cases for radiological features and their correlation with pathological and biological features; and a case–control study of mammographic density. The first component of the review includes:
Determination of the observed radiological features of the cancers on mammography, to identify diagnostic features with verified poor outlook either on histology, biological features or outcome (in the long term, survival and disease-free survival).
Radiological audit to identify those tumours which could have been detected at a screen previous to the diagnosis, (i.e. potential false-negatives), with a review of the reasons for failure of mammographic diagnosis. This is particularly relevant to the 26% interval cancers.
Comparison with other age groups or risk profiles, including:
The review includes the films of the cancers including their previous mammograms, as in the radiology reviews of the previous breast screening studies such as the Age Trial and the Breast Screening Frequency Trial.61,62 The review differs from that of previous studies in three important respects. First, it includes mammograms from subjects who never developed cancer during the study, two per cancer matched for age, centre and date of screening. Second, the X-rays are digitised (where not already digital) so that reviewers can view the mammograms without either readers or films having to travel. Third, the review includes the density study mentioned above.
Digitisation of the analogue mammograms was by Array Corporation's 2905 X-ray film digitiser (Array Corporation USA, Hampton, NH, USA), which gives a pixel size of 3600 by 4800 and DICOM resolution of 1 mm = 20 pixels, equivalent to 12 bit. The DICOM images were converted to bitmap images as this format is the most suitable for uploading to the web. This conversion reduces the resolution to 8 bit. Digital mammograms were anonymised, assigned a unique study number, then converted to bitmap (8 bit) format and uploaded to the web.
The formatted and anonymised images were uploaded into Image-box, version 1 (University of Southampton, Southampton, UK), where there is further compression of the images to 550 by 900 pixels. We used the image database developed for the Prospective study of Outcomes in Sporadic and Hereditary breast cancer (POSH) radiological review.64 This is web based and was developed by Kevin Wheeler of Southampton University, who is employed by Professor James Batchelor, in the Department of Computer Science.
Features of the POSH database include:
It is web based and password controlled.
It incorporates anonymised scanned mammographic and ultrasound images.
Recording sheets appear online to match images and screening events under examination.
It enables a greater number of radiologists to participate and can therefore be opened up to volunteers from the centres.
It allows for two readers per study.
A third reader arbitrates on any differences (we have a limited list of final decision arbitrators). For each field the final observation is adopted if both initial readers agree or two out of three after arbitration agree. If all three differ, we take as the final observation the decision of the third reader.
The database uses Breast Imaging-Reporting and Data System (BI-RADS) terms so that it is suitable for publication in American journals.
Studies are reviewed starting with the latest screening episode and progressing backwards through the series so that any information on the site of the cancers can be used to look for subtle earlier signs (in the same way as interval cancer audit is generally done).
A major outcome of the first component of the radiology review is radiological audit of cancers arising in FH01 – could these have been picked up at previous screens, and what are the key radiological signs of this? The inclusion of the non-cancers will give us information about specificity of earlier signs of malignancy, yielding an estimate of the likely effect on false-positives of a change in practice or training. Other outcomes include a quality assessment of both radiology and radiography in FH01. It should be noted, however, that with the advent of digital mammography, accuracy will improve in any case for this young population with relatively dense breast tissue.
The second component of the radiology review is a case–control study of breast density, again using at least two age-matched controls per cancer case. The major aim is to determine whether or not breast density is a risk factor in this population at enhanced familial risk, as it is in the general population.65 Secondary outcomes will be the determination of whether per cent density or absolute dense area is the better predictor of breast cancer risk in this population, and how far in advance of diagnosis does density predict risk. Density was measured by the Cumulus interactive threshold computer program version 4 (University of Toronto, Toronto, Canada),66 operated by a single radiologist (Ruth Warren) with extensive experience in reading mammograms for density, both visually and using Cumulus. The program yields measures of dense area and total breast area. From these, the per cent dense area and the non-dense area can be calculated.
Density was read on digitised mammograms as described above. We have digitised mammograms for 103 cancer cases and 231 disease-free controls. The multiple readings for earlier mammographic signs of malignancy, quality assessment and radiology/pathology/biology correlation is ongoing. Density readings were available for 101 cases and 228 controls. Some results of the density study are already available. shows the dense area, total breast area and per cent density for cases and controls. The cases have slightly higher values than the controls for all three measures, but especially so for absolute dense area. As expected, per cent density was negatively correlated with age, although this was of borderline significance (correlation coefficient −0.10; p = 0.06).
Mean and SD of dense area, total area and per cent density on mammograms of 101 cases and 228 controls
Formal analysis was by conditional logistic regression, taking into account the individual matching of cases and controls.
There was a significant increase in risk of cancer with absolute breast density after adjustment for menopausal status (p = 0.03), with an 8% increase per 10 cm2 of dense tissue [odds ratio (OR) = 1.08, 95% CI 1.01 to 1.19]. The difference was more marked in premenopausal women (p = 0.008), defined as having had a menstrual period within the last 6 months. There was a 12% increase in risk per 10 cm2 of dense tissue (OR = 1.12, 95% CI 1.02 to 1.22). This remained significant after adjusting for HRT, age at menarche, parity and age at first birth (OR = 1.15, 95% CI 1.02 to 1.29; p = 0.01). A non-significant decrease in risk was observed in postmenopausal women; however, only 25 (8%) women were postmenopausal.
Per cent breast density did not have a significant effect on risk, regardless of menopausal status, unless adjusted for total breast area. In terms of both significance (p = 0.008 vs p = 0.03) and the magnitude of the standardised regression coefficient (0.41 vs 0.34), absolute dense area was a stronger predictor of risk than total area-adjusted per cent density.
Further analysis will focus on time between the mammographic examination from which density was estimated and diagnosis of cancer, on association of density with other breast cancer risk factors, and on the combined effects of density and other factors on breast cancer risk. In the meantime, the conclusions from the density component of the radiology review are that absolute density is a stronger predictor of breast cancer risk than per cent density in this population, and that absolute density increases risk in addition to the effect of other breast cancer risk factors. There is suggestive evidence that the effect is stronger in premenopausal women.
Pathology review
This will be a standardised review of conventional histopathological features (grade, type, size, etc.), which will be compared with the original pathology laboratory determination. In addition, this gives an opportunity to record other morphological features that are increasingly recognised as important characteristics of specific tumour subtypes, such as central scar formation, lymphocytic response, pushing or infiltrative tumour margin and degree of stromal response.67 In addition, features of ‘background’ non-involved breast tissue will be documented, which will be of relevance to the linked radiology review (see Radiology review above). It is suggested, therefore, that all haematoxylin- and eosin-stained slides for each case are requested for review. The pathology review form is shown in .
This is similar to the pathology reviews carried out in the various UK breast screening trials. However, in addition to the standard pathological variables, we also propose to stain the tumour samples for the recently discovered molecular subtypes of tumour, to determine the underlying aggressiveness of cancers occurring in the FH01 risk group. An ‘intrinsic gene set’ identified by Perou et al.68 and validated by Sortie et al.69 has led to the recognition of five ‘molecular’ subgroups: luminal (Lum) A, Lum B + C, human epidermal growth factor receptor-2 (HER-2) positive, basal and ‘normal-like’. These molecular subgroups of breast cancers have been shown to differ in their clinical behaviour, with HER-2-positive and basal groups exhibiting the poorest prognosis. BRCA1-associated breast cancers frequently exhibit a basal phenotype,70 but basal tumours are also more common in non-BRCA-associated cancers arising in young women,71 and may be associated with loss of BRCA function through other mechanisms such as gene methylation.72 There is, however, growing evidence that the basal subtype of breast cancer may not be a single entity,73,74 and different subsets may differ in their clinical behaviour and in potential therapeutic targets. Recent research identifying molecules that are highly effective at targeting BRCA-null tumours (and therefore potentially all or a subgroup of basal tumours) underlines the importance of accurately establishing the molecular phenotype of breast cancers.75 This study will employ a wide panel of markers that should better identify biologically important tumour subsets. In addition, whole sections will be analysed for BRCA1-methylation status (methodology already optimised).
As a comparison group, we will interrogate the POSH database64 to identify a cohort of age-matched non-family history cases on which the same analysis will be performed as part of the POSH study.
The tumour samples are still being collated, which is proving a demanding job in terms of administration and governance, with material transfer agreements being required for > 70 sites. For one centre with seven cancers, the tumour material has been transferred to Barts Health and the rereading of invasive status and grade is 100% in agreement with that in the original pathology laboratory, and correlation of tumour size with that of the original laboratory is 0.9997.
