Comparing tumour heterogeneity in primary tumour, circulating tumour cells and its metastases

Therapeutic decisions in lung cancer are increasingly dependent on adequate
tumour tissue biopsies. However, amongst others, tumour heterogeneity and
technical issues with the handling of tissue allow adequate diagnosis in only
part of patients(1,2). We are interested in the tumour heterogeneity, as it
provides crucial information regarding the metastatic processes of tumours and
the response to treatment.
To study the tumour heterogeneity adequately, we plan to obtain material from
all accessible metastatic tumour sites and the primary tumour from patients
with metastasized lung cancer ([N]SCLC) after they have passed away, by an
autopsy.
As CTC*s are an important prognostic factor, and play an important role in the
metastatic process, all subjects that participate in this study will have 15mL
blood drawn at a regular outpatient visit, in order to obtain CTC*s. These
CTC*s will be studied for their genetical characteristics and will be compared
to both the original tumour and its metastases. We think it is important to
incorporate CTC*s as liquid biopsies in the project as they may help to bypass
the problems of normal biopsies: CTCs do not have the issue of contamination
with normal cells and DNA/RNA from leukocytes that come with this technique can
be harvested in the same run(3).
Therefore, CTCs may replace current tumour biopsy practices when an adequate
number of tumour cells can be detected, while also giving the option for
further mutation analysis(4,5). But while specific mutations have been
assessed, it is unknown whether CTC*s accurately reflect the heterogeneity of
the malignancies.
In this study we can assess whether the CTC*s do reflect the heterogeneity by
comparing them to the original tumour. At the same time we are able to assess
the differences between original tumour and metastases and the differences in
the heterogeneity by comparing SNP, mutations and DNA instability. Our
hypothesis is twofold: Firstly, that the metastatic processes will have less
heterogenicity than the primary tumour. Secondly, that the CTC*s will mimic
original tumour heterogeneity, but are more closely related to the metastases
and that this is visible in the mutations and genetic profile.

- Comparing the heterogeneity of the original tumour and its metastases.
- Comparing the mutations in CTC*s to those of both the original tumour and the
metastatic processes.
- Assessing whether certain mutations precede the others (so called trunk and
branch mutations).
- Analyzing whether mutations in metastases all originated from the primary
tumour.

Terminal patiënts with proven lung cancer (NSCLC or SCLC, cytological or
pathological proven) will be asked to participate. The treating physician will
assess wether he can bring this research up with the patient. If he deems the
patient can't discuss it, he will abstain from doing so, but otherwise he will
give the patient an introduction and the written information.
If the patient approves, we will draw some blood from which we can obtain CTCs.
After the participants death, the family will be asked to consent with an
obduction, which will be recorded in the patients file according to common
practice. A warm obduction will be performed during which biopsies will be
taken. These biopsies will be processed by the pathological department of the
UMCG as patient material. The biopsies will be used for genetic testing on
heterogeneity and mutational load and specific mutations. The different
biopsies will be compared to one another using paired statistics

While the risk associated with the study is inherently low (it mostly takes
places after the participant has deceased), we do believe there could be an
emotional strain considering the nature of participating in this study. The
question of autopsy is not always easy for a patient, but we believe that the
insights we could gain by this research could be important in the future.
Ofcourse it goes without saying that physicians will bring up the subject only
with patients they deem capable of handling this topic. And they will bring the
subject as tactfully as possible.