Key words tumor cell dissemination - circulating tumor cell - disseminated tumor cell - prognostic
factor - breast cancer
Schlüsselwörter Tumorzelldissemination - zirkulierende Tumorzellen - disseminierte Tumorzelle - Prognosefaktor
- Mammakarzinom
Introduction
Dissemination of single cancer cells via blood stream is currently considered a crucial
step during metastatic cascade. Historically, the first report on tumor cells in peripheral
blood from a patient with a solid tumor has been published 150 years ago by Thomas
Ashworth, a Melbourne-based surgeon ([Fig. 1 ]) [1 ]. Since then, an extensive body of evidence has been accumulated on the relevance
and the biological meaning of hematogenous cancer spread. Isolated tumor cells are
generally referred to as circulating tumor cells (CTCs) in the peripheral blood or
disseminated tumor cells (DTCs) when encountered in bone marrow or other organs. In
recent years, studies have shown with level 1 evidence that both CTCs and DTCs predict
worse clinical outcome in early and metastatic setting [2 ], [3 ], [4 ], [5 ].
Fig. 1 First page of the publication “A case of cancer in which cells similar to those in
the Tumours were seen in the blood after death” including the first description of
circulating tumor cells in 1869 [1 ].
Currently, numerous translational research projects are investigating this issue in
breast cancer. Among 58 clinical trials registered at ClinicalTrials.gov and EudraCT
which are recruiting or planned, the vast majority (53) focuses on CTCs in the blood,
with only five trials designed to explore DTCs in the bone marrow. 41% of the trials
are being conducted in the United States of America, followed by China (12%) and Germany
(10%). A smaller number of studies has been initiated in France (9%) and Sweden (5%)
([Fig. 2 ]). Only four trials are phase III studies: two of them are based in Germany (DETECT
III and DETECT V), one in India and one in China.
Fig. 2 Circulating and disseminated tumor cell – current study landscape with an overview
of open or planned clinical trials registered at ClinicalTrials.gov and EudtraCT.
Beyond prognostication and therapy monitoring, CTC/DTC-guided treatment decisions
are considered the most exciting potential of tumor cell dissemination. In 2018, results
from the first positive trial on therapy based on CTC status were presented at the
San Antonio Breast Cancer Symposium [6 ]. These and data from other studies have been recently thoroughly discussed at the
yearly Consensus Conference on Tumor Cell Dissemination during the Annual Meeting
of the German Society of Senology. In this review, we comment on the current data
on hematogenous tumor cell dissemination and its possible clinical applications in
the near future.
Early Breast Cancer
Two large meta-analyses confirmed that presence of CTCs or DTCs in patients with non-metastatic
breast cancer (BC) is associated with poor survival [2 ], [4 ]. Similarly, patients with persistent CTCs/DTCs after completion of (neo)adjuvant
chemotherapy are more at risk for developing a relapse [7 ], [8 ]. Recently, two studies have investigated whether a blood-test performed during routine
follow-up in asymptomatic patients may identify those with particularly poor prognosis.
The first analysis was conducted by Sparano et al. on a large group of HER2-negative
BC patients participating in the ECOG-ACRIN trial E5103 [9 ], [10 ]. This trial was originally designed to explore the use of bevacizumab in combination
with adjuvant chemotherapy. 546 patients presented for blood sampling five years after
diagnosis. In 4.8% of cases at least one CTC was detected per 7.5 ml blood by using
the CellSearch assay. In the subgroup of 353 patients with hormone receptor (HR-)positive
disease, the recurrence rates per person-year of follow-up in the CTC-positive and
CTC-negative groups were 21.4 and 2.0%, respectively. In the multivariate analysis,
a positive CTC result was associated with a 13.1-fold higher risk of recurrence.
Similar results were reported by the German SUCCESS-A trial [11 ]. In this phase III trial patients with high-risk BC were first randomized to 3 cycles
of epirubicin-fluorouracil-cyclophosphamide followed by either 3 cycles of docetaxel
or 3 cycles of gemcitabine-docetaxel, followed by a second randomization to 2 vs.
5 years of zoledronate treatment. Presence of CTCs five years after chemotherapy was
assessed using the CellSearch system. At least one CTC was detected in 7.8% of patients.
In HR-positive patients, CTC status was a significant prognostic factor for recurrence-free
survival (hazard ratio 5.14 in the univariate and 5.95 in the multivariate analysis,
respectively).
These two trials showed for the first time that the remaining relapse risk after BC
treatment can be assessed using CTC detection. Future studies will clarify which strategies,
e.g. intensified follow-up or extended adjuvant endocrine therapy, should be offered
to patients with CTC persistence.
Next to therapy monitoring and estimating prognosis, CTC detection might be used to
predict therapy benefit. Retrospective analyses of NSABP-B31 and NCCTG-N9831 have
suggested that some patients might benefit from HER2-directed treatment even though
they have HER2-negative disease [12 ], [13 ]. Using CTC detection, the phase II Treat-CTC trial aimed to identify these HER2-negative
early BC patients that might respond to additional trastuzumab therapy. Patients with
centrally confirmed HER2 nonamplified primary tumors and at least one CTC per 15 ml
blood (CellSearch assay) following surgery and (neo)adjuvant chemotherapy were randomized
(1 : 1) to receive either 6 cycles of trastuzumab or 18 weeks of observation. Primary
endpoint was the CTC detection rate (≥ 1 CTC/15 ml blood vs. no detectable CTCs) 18
weeks after randomization [14 ]. Secondary endpoints were invasive disease-free survival (iDFS) and cardiac safety.
Of 1317 patients screened 95 (7.2%) were CTC-positive. Sixty-three of these patients
were randomized to trastuzumab (31 patients) and observation (32 patients). At week
18, 5 out of 31 patients that had received trastuzumab and 4 out of 32 in the observation
arm were still CTC-positive (p = 0.765). The study was stopped due to the recommendations
of an independent data committee after a median follow-up of 13 months. One-year iDFS
was 93.8% (95% CI 77.3 – 98.4) in the observation vs. 84.8% (95% CI 63.4 – 94.2) in
the trastuzumab arm. Several limitations might explain the negative results of Treat-CTC.
First, only few patients were randomized and the CTC detection rate at week 18 was
low. Second, results were biased by the use of endocrine treatment in hormone receptor-positive
patients and third, CTCs were merely enumerated but not further characterized. As
results of the NSABP-B47 trial recently demonstrated that there is no benefit for
adjuvant trastuzumab in patients with HER2-negative tumors [15 ], determination of HER2 amplification in CTCs might help to more precisely guide
HER2-targeted treatment. This concept is currently addressed by the German DETECT
studies in the metastatic setting (see below).
A major limitation of using CTCs to estimate prognosis or predict treatment efficacy
is their low detection rate in early breast cancer. Detection of single disseminated
tumor cells (DTCs) in the bone marrow is an alternative approach that is feasible
especially at the time-point of primary surgery: after bone marrow aspiration from
the iliac crest, DTCs are generally detected by isolation of mononuclear cells, immunocytochemistry
(pan-cytokeratin staining) and cytomorphologic criteria [16 ]. To confirm their prognostic relevance, a large pooled analysis (PADDY) was presented
at the 2018 San Antonio Breast Cancer Symposium [17 ]. Individual patient data from 10 307 patients, derived from 11 centers across Europe
and the U. S., was analyzed. Of all patients included, 2814 (27.3%) had at least one
detectable DTC. The proportion of DTC-positive patients was higher in patients with
larger tumor burden (tumor size, lymph node involvement, p < 0.001) or more aggressive
tumor (ER/PR negativity, HER2 positivity or higher grading, p < 0.001). DTC detection
was significantly associated with an impaired prognosis: after a median follow-up
of 7.6 years the multivariate hazard ratios (HR; 95% CI) were 1.23 (1.06 – 1.43; p < 0.006)
for overall survival (OS) and 1.30 (1.12 – 1.52, p < 0.001) for disease-free survival
(DFS). There was a statistically significant interaction between biological tumor
subtype and DTC-positivity (p = 0.014) and the prognostic impact was especially pronounced
in Luminal B (defined as ER/PR-positive, HER2-negative and G3) patients. Additionally,
it was shown previously that DTCs are already detectable in patients with pre-invasive
breast cancer (i.e. ductal carcinoma in situ, DCIS) [18 ], [19 ]. These results emphasize the hypothesis of parallel tumor progression where metastatic
disease derives from single cells that migrate and disseminate from very early breast
cancer lesions [20 ], [21 ]. Future trials should now investigate whether DTC detection is useful to tailor
adjuvant treatment. For example the HER2 expression of DTCs differs from that of the
primary tumor and patients with HER2-positive DTC have an increased risk of metastatic
relapse and might therefore benefit from trastuzumab treatment [22 ], [23 ]. Moreover, DTC detection seems to predict efficacy of bone-targeted therapy. The
prospective MRD-1 trial found that zoledronic acid might eliminate DTCs from the bone
marrow and a retrospective analysis of a large data-set revealed that bisphosphonates
are more effective for the prevention of distant metastasis in DTC-positive patients
[24 ], [25 ], [26 ]. Currently there are prospective trials ongoing to evaluate the effect of denosumab
in DTC-positive patients (NCT01545648, NCT02682693).
Metastatic Breast Cancer
In the last two decades, numerous studies have confirmed presence of CTCs in patients
with metastatic BC as a strong independent prognostic factor [3 ], [27 ], [28 ]. Due to large numbers of CTCs encountered in patients with advanced disease, the
cutoff of 5 CTCs per 7.5 ml blood is usually used in case of CellSearch-based detection
(in contrast to early BC, where patients with at least one CTC are referred to as
CTC-positive) [29 ]. Recently, Cristofanilli et al. evaluated CTC samples from 2436 women with metastatic
BC from 17 European centers participating in the European Pooled Analysis Consortium
and a single large U. S. institution, the MD Anderson Cancer Center in a pooled analysis
[30 ]. Patients with < 5 CTCs per 7.5 ml blood had significantly longer median OS than
those with ≥ 5 CTCs (36.3 vs. 16.0 months, p < 0.0001). This association was independent
of the tumor subtype, prior treatment and disease location.
In most patients, CTC counts decline rapidly after start of systemic therapy. However,
patients with persistently high CTC numbers are significantly more at risk for relapse
[31 ]. Martin et al. measured CTCs in patients receiving palliative chemotherapy and reported
that CTC numbers after the first cycle of treatment were associated with response
to therapy and survival in the multivariate analysis [32 ]. Thus, CTC dynamics are able to predict whether the patients will benefit from therapy
sooner than conventional radiological imaging. However, it remains unclear which therapeutic
strategy should be offered to non-responders. In the SWOG 0500 trial, blood samples
were evaluated before and during first-line chemotherapy in 595 patients [31 ]. Out of these, 123 patients had increased CTC counts both prior to therapy and after
the first cycle and were randomized either to continue treatment or switch to another
chemotherapy regimen ([Fig. 3 ]). PFS and OS were similar in both arms. Possibly, CTC persistence identifies patients
resistant to conventional chemotherapy, independent of the cytotoxic drug used, who
might rather benefit from targeted or experimental treatment approach.
Fig. 3 Study design of the SWOG 0500 trial (A Randomized Phase III Trial to Test the Strategy
of Changing Therapy Versus Maintaining Therapy for Metastatic Breast Cancer Patients
who Have Elevated Circulating Tumor Cell Levels at First Follow-Up Assessment).
CTC-guided Therapy Decisions in Metastatic Setting: Ready for Prime Time?
CTC-guided Therapy Decisions in Metastatic Setting: Ready for Prime Time?
In recent years, several studies have aimed at exploring the potential of CTCs to
guide treatment in metastatic BC. Two large studies have been initiated in France
(Institut Curie) and Germany (DETECT study group), respectively. While the DETECT
trials are still recruiting, the results of the French STIC CTC trial were presented
at the San Antonio Breast Cancer Symposium 2018 [6 ]. This study was designed to investigate whether CTC counts may select optimal therapy
for patients with HR-positive HER2-negative metastatic disease ([Fig. 4 ]). 778 patients were randomized before start of first-line therapy: in the standard
arm patientʼs oncologist decided whether the patient was suitable for endocrine therapy
or chemotherapy; in the experimental arm the treatment was based on the CTC numbers.
In about half of the patient population, a concordant low risk estimate was obtained
(i.e. clinically low risk and < 5 CTCs per 7.5 ml blood). These patients received
endocrine therapy in both the control arm and the CTC arm. At the other side of the
spectrum were 13.5% of patients whose clinical risk estimation and the blood test
both showed high risk situation (i.e. clinically high risk and ≥ 5 CTCs); these patients
received chemotherapy in both arms. In between, in about 40% of the population, the
risk estimation (clinical vs. blood-based) was discordant and patients received either
endocrine therapy or chemotherapy according to their risk profiles and randomization
arm.
Fig. 4 Design of the STIC CTC trial – the first positive study on CTC-based treatment interventions
(Randomized Trial to Evaluate the Medico-economic Interest of Taking Into Account
Circulating Tumor Cells to Determine the Kind of First Line Treatment for Metastatic,
Hormone-receptors Positive, Breast Cancers).
The progression-free and overall survival were identical in both arms, showing that
standardized CTC-based risk estimation was not inferior to the – often poorly reproducible
– clinical assessment. Interestingly, in patients with discordant risk estimation
(i.e. clinically low risk but ≥ 5 CTCs or clinically high risk but < 5 CTCs), the
PFS and OS were significantly longer if patients received chemotherapy. Admittedly,
the interpretation of these results is not easy: the STIC CTC trial was designed before
the approval of CDK4/6 inhibitors and the majority of patients with HR-positive HER2-negative
metastatic BC is now treated with endocrine-based combination treatment [33 ]. Having said that, the evidence provided by this trial demonstrates that assessment
of CTC numbers is able to identify patients in need of a treatment more effective
than endocrine monotherapy.
In contrast, the DETECT study program aims at investigating the influence of CTC characteristics
– and not CTC counts – on response to therapy. Currently, the choice of systemic therapy
in the metastatic setting is based on the properties of the primary tumor or metastasis.
However, numerous studies have shown that expression profiles of CTCs may differ from
both the primary tumor and distant metastasis [21 ], [34 ], [35 ]. The first phase III trial within this large translational project is the DETECT
III study ([Fig. 5 ]). In this trial, patients with HER2-negative metastatic BC are first screened for
HER2-positive CTCs and, in case such cells are detected, randomized to standard therapy
± HER2-targeted treatment with lapatinib. Patients with HER2-negative CTCs are offered
several study options within the DETECT IV trials. Women with HER2-positive tumors
may participate in the DETECT V trial and receive dual anti-HER2 blockade in combination
with either chemotherapy or endocrine therapy plus CDK4/6 inhibitor ribociclib.
Fig. 5 Study design of the DETECT trials – the largest study program on CTC-guided therapies
worldwide [36 ].
Conclusions
In conclusion, the detection and characterization of CTCs and DTCs holds great promise
to more precisely monitor and target the source of metastatic spread. For this purpose
it is important to better understand the biological principles of tumor evolution.
In the future, combining molecular characteristics (e.g. single cell sequencing) of
CTCs and DTCs with clinical data, immunological markers and other forms of liquid
biopsy (e.g. circulating tumor DNA or miRNA) might help to identify biomarkers or
drugable targets to more precisely monitor and/or tailor systemic treatment.
