STEP:
Study of Tumor Educated Platelets for early detection of breast cancer in blood in BRCA1/2 mutation carriers

Women with a genetic predisposition for breast cancer (BrCa) due to a
BRCA1/BRCA2 mutation have a strongly increased lifetime risk of developing
BrCa. As treatment and survival chances of BrCa strongly depend on disease
stage, it is paramount that BrCa is detected in the earliest stage possible.
Therefore, these women are monitored regularly from age 25 to 70 (clinical
examination, X-ray/ MRI scans). Current screening strategies have suboptimal
sensitivity and specificity, however. Moreover, these screening modalities are
unpleasant and not performed during pregnancy.
In this study, we will validate a test for early detection of BrCa in blood.
Initially, within STEP we aimed to evaluate the ThromboSeq test, which is
based on analysis of tumour RNA biomarkers in Tumour Educated Platelets (TEPs).
However, recent insights have shown that this test is less accurate in breast
cancer detection, moreover optimalisation of the protocol is necessary.
Therefore, within STEP, anothe liquid biospy test will be investigated, i.e.
the meD-seq assay. Proof-of-concept studies are highly promising. Thus, in the
current study we aim to evaluate the MeD-seq test for detection of breast
cancer in the specific clinical setting of women with a BRCA1/BRCA2 mutation
who are being screened for BrCa. If BrCa can be detected by the MeD-seq test in
early stage, screening with MRI/X-ray can potentially be replaced (partly) or
complemented with the MeD-seq test. If the MeD-seq test can detect BrCa earlier
in time, potentially a less intensive treatment is needed.

We will also collect an additional blood sample at each blood withdrawal for
future studies of cell free tumor DNA (ctDNA) or Tumor educated Platelets for
early detection of cancer in blood.

- To study sensitivity and specificity of the MeD-seq test for detection of
BrCa in female BRCA1/2 mutation carriers in an early stage.
- To determine if the MeD-seq test can detect BrCa earlier in time than current
surveillance tools (MRI/mammography) in female BRCA1/2 mutation carriers.
- To set-up a biobank with blood samples of BRCA1/2 mutation carriers
undergoing regular breast screening, for future research aimed at early
detection of cancer in blood using the ThromboSeq test and ctDNA or newly
developed liquid biopsy methods.

A nested case-control study will be conducted. About 1400-1700 female BRCA1/2
mutation carriers will be approached for participation, of which we expect 50%
to participate.
We will collect half-yearly blood draws (2 blisters of 10 ml) of female BRCA1/2
mutation carriers in an initial period of 3 years, 6 blood withdrawals per
participant. A subset of partcipants will be asked (after the 5th blood
withdrawal) if the want to partcipate for a longer period - i.e. 2 extra yeras,
4 extra blood withdrawals. The blood withdrawals will take place just before
the regular screening moment and, depending on the personal screening program,
also in between screening moments. Blood withdrawals will be arranged by the
participants themselves, in a licenced blood withdrawal location. This location
will be registered by the researcher. All necessities for blood withdrawal and
shipping to Vumc will be sent to the home adress of participants. Participants
receive extensive intsruction about the study and blood withdrawal bij
phonecall and written.
A total of about 700-875 participants are aimed to be included, depending on
the number of participants that will partcipate in the extended follow up after
the first 3 years. An estimated 5250 blood samples for TEP analysis and 5250
blood samples for ctDNA studies will be collected from 875 BRCA1/2 mutation
carriers.

The MeD-seq test will be conducted on the blood samples of the participants who
developed breast cancer (cases) and matched controls, derived from the same
cohort.
Controls will meet the following criteria:
- the control was not diagnosed with BrCa by regular screening at the moment
that this blood sample was taken, and no diagnosis of breast cancer in at least
1 year of follup up
- the mutation status is the same as in the case (either BRCA1 or BRCA2
mutation)
- the age of the control may differ with a maximum of 5 years from the age of
the case at the moment of BrCa diagnosis

Subsequently, the sensitivity, the specificity, and the positive predictive
value of the new method will be calculated, as well as the proportion of women
with a positive MeD-seq test at a point earlier in time than the date of
detecting abnormalities during regular screening.

The study protocol consists of a questionnaire study and a 6 or 10 blood
withdrawals within three years or five years. Neither of these methods causes
significant risks for the participants of the study. The blood withdrawal and
sending of the samples will be arranged by the participants themselves. This
will demand time and effort.
There are no direct benefits for women participating in this study. If the
MeD-seq test proves to detect BrCa earlier and with less false positives and
false negatives, this test might complement or replace the current surveillance
tools in the future, thereby reducing the burden associated with surveillance
for the group of BRCA1/2 mutation carriers as a whole.