A pilot study about culturing laryngeal tumor tissue organoids to facilitate new treatment strategies for laryngeal cancer in the future

Despite intensive treatment, prognosis of laryngeal cancer (LC) remains poor.
Five years overall survival is 60% and an accurate treatment is of paramount
significance to improve overall survival. Most patients with LC will receive
larynx preserving (chemo)radiation without knowing the sensitivity of the LC.
Selecting LC with low (chemo)radiosensitivity could prevent unnecessary
(chemo)radiation.
Organoids which are tumor-derived three-dimensional cancer stem cells that
mimic in vivo tumor characteristics were explored and efficacy has been tested
in our well-established collaboration with the UMCG departments of Biomedical
Sciences of Cells & Systems/Radiation Oncology, Medical Oncology, Ear Nose
Throat/Head and Neck Surgery, Pathology and Maxillofacial Surgery. Recently,
the optimized organoids culture methodology for squamous esophageal cancers
resulted in the parallel development of a culture methodology for organoids of
head and neck squamous cell which was shown to be successful in six out of
fourteen tumors. In this study we would like to develop and evaluate the
efficacy of a solid ex vivo LC tumor model (= LC organoids) of patient derived
LC tumor material by whole genome DNA sequencing. With solid LC organoids we
would be able to test the effects of standard (chemo)radiation on self-renewal
and regrowth potential of the LC stem cell derived organoids in future. Solid
organoids predicting the patients (chemo)radiation response could lead to an
improvement of LC treatment, by allowing selection of patients who will benefit
from surgical treatment bypassing (chemo)radiation and as such improving
survival and reducing side effects thereby increasing post-treatment quality of
life.
With this pilot study, based on the methods described by Tanaka and Nagle (7,
8), we aim:
- to develop and evaluate solidity of patients* specific ex vivo LC organoids
in which at least 50% of the obtained tumor samples are viable and
proliferating at week 2 and 6
- to analyse difference of solidity of ex vivo model in primary and recurrent
LC tissue
The findings of this pilot project will guide the design of early phase
clinical trials in LC patients with the ultimate goal to develop less toxic and
more effective treatment strategies.

Primary Objective: We want to further develop and evaluate our patients*
specific ex vivo LC organoids in which at least 50% of the obtained tumor
samples are viable and proliferating at week 2 and 6.

Secondary Objective: We will analyse differences in solidity of ex vivo LC
organoids between primary and recurrent LC tissue.

In this prospective study, fresh LC tissue will be collected to test
feasibility of the patients specific ex vivo LC organoids.
Durations: Study period of 42 months
Setting: Biopsies will be taken at the departments of Otolaryngology / Head and
Neck Surgery or Maxillofacial Surgery of the University Medical Center
Groningen. During routine biopsy or during/after resection under general
anaesthesia, three to five additional biopsies of the tumor (maximum 0.5 cm3)
will be removed by using an endoscope. The subjects will not undergo extra
procedures in the course of the research: only routinely procedures are
performed (i.e. endoscopic examination in the outpatient clinic or under
general anaesthesia for tumor staging, resection of tumor by neck dissection,
total laryngectomy). Also, 2ml blood will be withdrawn for DNA sequencing.
Organoids of the ex vivo model will be developed and tested in the Laboratory
of Medical Oncology and Laboratory of Cell Biology / Radiation Oncology,
University Medical Center Groningen.

For the reliable translational use of these organoids we need to confirm their
origin, define their genomic alterations and test genomic (in)stability over
time. Therefore, we will analyze the original tumor and its organoids
derivative after 2 weeks and after 6 weeks of culturing by whole genome DNA
sequencing (~30x coverage (~100 Gb)). The 6 weeks period has been chosen
because of the clinical translation of this model: organoids based treatment
strategies can only be tested during a maximum of six weeks staging period
between tissue biopsy and start of treatment. After collection of the original
tumor specimen for organoids culturing we will store part of this tumor tissue
for DNA-analysis at -20 degree Celsius. The other part will be used for
organoids culturing. After a maximum of three months culturing, the organoid
will be destroyed.

We will analyse differences in solidity of ex vivo LC organoids between primary
and recurrent LC tissue by using a Chi-square test with SPSS.

We expect no significant additional burden or risks associated with
participation. During routine biopsy or during/after resection under general
anaesthesia, three to five additional biopsies of the tumor (maximum 0.5 cm3)
will be removed by using an endoscope.