Clinical Trial Details
— Status: Active, not recruiting
Administrative data
| NCT number |
NCT04868396 |
| Other study ID # |
Glioma stem cell organoids |
| Secondary ID |
|
| Status |
Active, not recruiting |
| Phase |
|
| First received |
|
| Last updated |
|
| Start date |
April 10, 2021 |
| Est. completion date |
September 1, 2024 |
Study information
| Verified date |
August 2023 |
| Source |
Maastricht Radiation Oncology |
| Contact |
n/a |
| Is FDA regulated |
No |
| Health authority |
|
| Study type |
Observational
|
Clinical Trial Summary
Rationale: Glioblastoma (GM) is the most frequent incurable adult brain tumor with median
survival of 15 months after diagnosis, despite extensive treatment with surgery, radiation
therapy and chemotherapy. Tumor recurrence is inevitable after which life prolonging
therapies are no longer available. The development of new treatments for GM is being hampered
by inter-and intratumoral heterogeneity of tumors and their microenvironment, which currently
cannot be predicted accurately with current diagnostics.
Objective: To establish primary patient derived organoid cultures from GM to study mechanisms
that contribute to aggressive tumor growth and treatment resistance in primary and recurrent
GM.
Study design: Preclinical study, using patient derived glioblastoma tissue. Study population:
Patients 18 years or older, with newly diagnosed glioblastoma.
Main study parameters/endpoints: Intra-and inter organoid genetic and epigenetic
heterogeneity that is representative for GM. Nature and extent of the burden and risks
associated with participation, benefit and group relatedness: Minimal burden, since the
biopsies are part of a regular neurosurgical procedure (debulking); which intends to
eradicate the macroscopical tumorload in order to optimize survival benefit. The tissue
(biopsy) that will be used for this trial is part of the tumor tissue that is resected during
the standard debulking.
Benefit: no benefit for the patient.
Description:
Patients with glioblastoma (GM) have a median overall survival of approximately 15
months.Standard therapy for GM encompasses maximum surgical resection followed by radiation
and chemotherapy using temozolomide (TMZ) (1). Regardless of initial tumor response, tumor
recurrence is inevitable, after which survival drops to less than 6 months. GM tailored
approaches targeting oncogenes that might drive the growth of the bulk of primary tumors,
have been unsuccessful so far in clinical trials(2), creating a large unmet need warranting
new approaches to overcome intrinsic and acquired resistance to current treatment schedules.
The objective of this research is to establish primary patient derived organoid cultures from
GM to study mechanisms that contribute to aggressive tumor growth and treatment resistance in
primary and recurrent GM.
1. Inter-and intratumoral heterogeneity in glioblastoma. Tumor tailored approaches for GM
are being hampered by inter-and intratumoral heterogeneity of both microenvironment and
genomic alterations in GM cells. It has been shown that tumors are composed of multiple
clones harboring distinct genetic alterations (3-7). The clonal evolution model posits
that tumor formation is initiated in a cell of origin and is followed by the subsequent
accumulation of multiple genetic and epigenetic alterations, leading to tumor cell
survival and growth advantage (8). Divergent genetic alterations in early transformed
cells give rise to a variety of clones under the selective pressure of the tumor
microenvironment (3-7). An important microenvironmental stressor is intratumoral
hypoxia, which is frequent in GM and a negative prognostic and predictive factor
associated with reduced survival (9,10).
Emerging evidence implicates a subpopulation of tumor cells with characteristics of
normal stem cells so-called glioma stem cells (GSC) in intrinsic and acquired treatment
resistance. GSC are endowed with specific properties including high tumor initiating
ability, unlimited self-renewal potential and capacity for multipotent differentiation,
generating a diverse progeny(11). GSC are marked by common stem cell markers including
CD133+, SOX2, Olig1, and have been shown to reside in the perivascular region as well as
in hypoxic areas. GSC are expanded under hypoxia12depend on glycolysis (13,14).
Combined with their low proliferation, increased DNA repair, high anti-oxidant activity
and among others, make GCS more resistant to conventional treatments (radiation and
temozolomide) then non-GSC (15,16) {Jamal, 2012 #52}.
This implies that GSCs form an important driver of GM recurrence after chemoradiation.
There are currently no effective treatments to eliminate glioma stem cells. Blocking
hypoxia signalling in tumors (inhibiting self-renewal and survival of GSC cells)(12,17)
and blocking the NOTCH stem cell pathway (rendering GSCs sensitive to radiation(18) and
TMZ(19-21)), seem to promising but drugs interfering with these pathways have not passed
beyond early phase clinical trials yet(22).
Current standard of care of newly diagnosed glioblastoma is multimodal and consists of
surgery, radiation therapy and TMZ, an alkylating agent modifying purine bases of DNA
(O6-guanine; N7-guanine and N3-adenine). The addition of TMZ to radiotherapy has
increased the overall survival of GM patients significantly, but only up to 14.6
months(1). Intratumoral hypoxia has been shown to decrease therapeutic efficacy of RT
and chemotherapy(23). Hypoxic GM cells are genetically unstable and show increased MGMT
expression and thereby resistance to alkylating TMZ chemotherapy(24).
In non-GSCs, MGMT promoter methylation is a predictive marker for response to TMZ
treatment(25,26). However the interpretation of the MGMT methylation assay is complex,
since the extent of MGMT promoter methylation in GM is heterogeneous and the level of
heterogeneity is underestimated, since only GM biopsies or fragmented GM tissue is being
analysed(27,28). Importantly, MGMT is also expressed in the normal brain endothelial
cells and in immune cells including tumor infiltrating cells(29). Thus depending on the
extent of normal tissue contamination in GM biopsies the level MGMT methylation may also
differ.
Since intra-tumoral heterogeneity in MGMT expression cannot be objectivated using
currently available diagnostics and since undertreatment of patients should be prevented
at all times, at present most patients diagnosed with ´de novo´ GM, receive TMZ;
although there seems to be little benefit of TMZ in GMs with MGMT unmethylated glioma
cells.
Designing new treatment protocols for newly diagnosed glioblastoma is quite complex,
since diagnostic tools, predicting inter-and intra-GM heterogeneity of MGMT promoter
methylation status (or level of MGMT expression), are lacking. Additionally, the level
of MGMT expression, needed for significant TMZ response and improvement of clinical
outcome, is not known. Thus since most GMs are defined by both MGMT methylated and MGMT
unmethylated tumor clones, combining TMZ with drugs that target non-methylated glioma
cells/glioma stem cells and/or microenvironment might be necessary.
2. Glioblastoma recurrence Even if a macroscopically complete resection of a GM can be
achieved, tumor cells remain at the resection site. It has been shown that GM cells have
a high capacity for dissemination. Invading tumor cells escape at the periphery of the
tumor mass and diffusely infiltrate the normal brain parenchyma. Deeply infiltrated
tumor cells are more likely to escape surgery and what is not known is whether
infiltration is a property of a more resilient cell population that initiates and drives
tumor recurrence. Even after postoperative chemotherapy and radiation outside of the
field of surgery to reduce infiltrative tumor cells, almost all GM recur, mostly around
the resection cavity. If a gross total resection cannot be performed; primary radiation
therapy and chemotherapy are also able to reduce clonal diversity, but are thus
insufficient to prevent recurrence.
This implies that tumor cells resistant to multiple therapies persist in the brain parenchyma
around the tumor cavity after gross total resection or in the remaining tumor after
chemoradiation which are responsible for tumor repopulation, making them a critical target to
overcome tumor recurrence. Genomic analysis of GM has shown that dominant clones in recurrent
tumors are composed of clones representative of the primary tumor as well as new clones with
little resemblance to the original tumor(3,30,31,32,33). Consequently targets identified
based on the analysis of the primary tumor may not be informative to identify the best
molecular targets to prevent recurrence(5).
After failure of first line GM treatment (radiation, TMZ), recurrence seems to be accompanied
with a phenotypic shift of GSCs to the mesenchymal (MES) subtype (loss of CD133; gain of
BMI1, SOX2 and CD44)(34-38). Such cells are more aggressive, invasive and angiogenic than
primary tumor derived GSCs, mostly the proneural (PN) subtype (CD133+, CD15+)(39-41). MES GSC
also show a higher expression of the inflammasome genes, such as IL6, IL8, IL1B1C and CXCL2,
reinforcing the notion that interplay with the microenvironment and plays an important role
in recurrence and progression(42,43). GM specific clinical trials are being developed, using
immune modulators. It has been shown that MSH (mutS homologue) mutations correlate with TMZ
resistance, as they are neither found in pre-treatment GM nor radiotherapy post-treatment GM,
but were detected in approximately half of recurrent GM patients treated with TMZ and
radiotherapy. This strongly indicates that MSH6 alterations are associated with resistance to
alkylating agent therapy. Thus GM patients who initially respond to TMZ may acquire MMR
defective hypermutator phenotype(44-46). It is well established that an intact mismatch
repair and base excision repair (BER) contribute to effective TMZ cytotoxicity.
Pharmacological inhibition of BER using PARP inhibitors either alone or in combination with
TMZ has shown promise in clinical trials. However, acquired resistance to PARP inhibitors is
observed through up-regulation of base excision repair and homologous recombination repair to
compensate for diminished BER (47,48).
In conclusion, understanding the therapy induced genetic and epigenetic alterations of the
remaining tumor cells and GSCs and also the impact of standard of care to the
microenvironment /niche, in which recurrence occurs, will guide the development of new
treatments.
In this project we will use patient derived glioma stem cell organoids(49), mimicking de novo
GM and its intratumoral heterogeneity. Also these patient-derived organoids (PDO) organoids
will be used to study acquired temozolomide resistance and thus can be used for identifying
novel targeted agents.
Tumor cell heterogeneity in MGMT expression and its relevance for TMZ response will also be
addressed using these organoids, using single cell proteomics and IHC for MGMT, GSC markers
and exome sequencing (for common driver mutations) to identify those populations that
clonally expand under TMZ (RT) selection and those that disappear.
Another interesting feature we will explore using PDO is the possibility to analyze the
supernatants for circulating tumor DNA (ctDNA) or exosomes (secreted vesicles which contain
RNA, small non-coding RNA, proteins as well as DNA) before during after acquired TMZ
resistance which might lead to the identification of pharmacological response and predictive
biomarkers. Such biomarkers 'liquid biopsy' may become useful to measure treatment response
in blood or lumbar fluid of GM patients and enable dose-modification (escalation or
deintensification or termination).
The development of the PDOs (costs of materials) will be funded (607061 PI M. Vooijs, MAASTRO
and KWF grant Alpe D'Huzes PI M.Vooijs, MAASTRO. There are no additional costs associated
with obtaining the tumor tissue.
