Transcript Glioma heterogeneity and the LAT-1

Glioma heterogeneity and the L-Amino
acid transporter-1 (LAT1):
A first step to stratified BPA-based BNCT?
D. Ngoga1; C. L. Schütz1; A. Detta1; S. Green2;
G. Cruickshank1
1University
of Birmingham, School of Cancer Sciences,
Queen Elizabeth Hospital, Department of Neurosurgery
2University
of Birmingham, Queen Elizabeth Hospital,
Department of Medical Physics
ICNCT-16, June 2014, Helsinki
Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Glioma Genetics
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To date, glioblastoma subject to the most extensive genomic
profiling of any cancer (Dunn, et al Genes Dev. 2012 26: 756-784)
The Cancer Genome Atlas Research Network (Parsons et al.
2008) and other studies have enabled us to systematically and
comprehensively define the genomic landscape of
glioblastoma.
Malignant gliomas are characterized by genetic instability and
complex alterations in:
 Chromosome structure
 Copy number.
 Variation in post translational modification.
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Glioma Genetics
Patterns of gene expression have been collated to identify
molecular subgroups with putative prognostic or predictive
significance.
Verhaak, et al. 2010
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Isocitrate Dehydrogenase – 1 mutation
Perhaps the most important of the discoveries arising from the
TCGA studies.
IDH-1 cytosolic component of the glycolytic pathway (Bleeker, et
al. 2009)
A spontaneous mutation in the gene for IDH1R132 identified in
 12% of patients GBM.
 12%–17% of AML (Mardis et al. 2009)
 Majority of central and periosteal cartilaginous tumors
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(Amary et al. 2011)
23% of cholangiocarcinomas (Borger et al. 2012)
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Isocitrate Dehydrogenase – 1 mutation
Found in majority of LGG
Confers a significantly improved prognosis.
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Methylation and treatment response
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The DNA repair enzyme 0-6-methylguanine-DNA
methyltransferase (MGMT) is responsible for reversing the
damage caused by temozolomide.
Methylation of the promoter region encoding MGMT has a
critical role in patients response to Temozolomide.
Only ~ 50% of patients with GBM have methylated MGMT.
(Esteller, et al. 2000)
The remainder of patients with unmethylate, IDH WT tumours
continue to have have poor prognosis.
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
The challenge for BNCT
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Understand how BNCT fit in this new framework of
understanding of GBM biology?
Understand and explain differences in treatment response to
BNCT.
Understand the impact of glioma heterogeneity on Boron
uptake and therefore the effectiveness of BNCT.
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
BPA based BNCT
4-dihydroxyboryl-L-phenylalanine (BPA)
 no selectivity/specificity, but increased AA need ( LAT1)
 low toxicity, resistance in tumor cells, low background
 coupling to fructose / mannitol to increase solubility
 uptake mostly regulated via amino acid transporter 4F2hc/LAT1
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
4F2hc/LAT1 - Structure
SLC3A2 has been shown to interact
with SLC7A7 (Pfeiffer, R;et al 1999)
heavy chain
(also called CD98)
light chain
The monomers alone do not
function as transporter!
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Study aims
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To understand role of LAT1 genetics in glioma
To determine the influence of LAT1 gene expression of patient
survival.
Understand the interaction between LAT1 gene expression and
know mutations relevant to GBM (IDH1 mutation)
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Study Method 1
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Interrogate publically available gene expression databases
(REMBRANDT)
Genes encoding the constituent proteins of the LAT1
transporter (SLC3A2 and SLC7A5) as well as related transporter
SLC7A7 were analysed.
REMBRANDT - Repository for Molecular Brain Neoplasia Data
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 1 – SLC3A2 (Heavy chain)
Gene Expression Plot (SLC3A2)
ALL GLIOMA (454)
ASTROCYTOMA (148)
GBM (228)
MIXED: (11)
NON_TUMOR: (28)
OLIGODENDROGLIOMA: (67)
UNKNOWN: (67)
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
Kaplan-Meier Survival Plot for Samples
with Differential SLC3A2 Gene
Expression (All Glioma)
Number of samples in group
Up-Regulated:
96
Intermediate:
247
Log-rank p-value:
Up-Regulated vs. Intermediate: 9.322837E-4
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 1 – SLC3A2 (Heavy chain)
Kaplan-Meier Survival Plot for Samples with
Differential SLC3A2 Gene Expression
(Oligodendroglioma)
Number of samples in
group:
Up-Regulated:
14
Intermediate:
36
Log-rank p-value
Up-Regulated vs.
Intermediate: p=0.0062
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 1 – SLC7A5 (Light chain)
Gene Expression Plot (SLC7A5)
ALL GLIOMA (454)
ASTROCYTOMA (148)
GBM (228)
MIXED: (11)
NON_TUMOR: (28)
OLIGODENDROGLIOMA: (67)
UNKNOWN: (67)
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
Kaplan-Meier Survival Plot for Samples
with Differential SLC7A5 Gene
Expression (All Glioma)
Number of samples in group:
Up-Regulated:
8
Down-Regulated: 30
Intermediate:
305
Log-rank p-value
Up-Regulated vs. Down-Regulated: 0.073
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 1 – SLC7A7 (y+LAT1)
Gene Expression Plot (SLC7A7)
ALL GLIOMA (454)
ASTROCYTOMA (148)
GBM (228)
MIXED: (11)
NON_TUMOR: (28)
OLIGODENDROGLIOMA: (67)
UNKNOWN: (67)
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
Kaplan-Meier Survival Plot for Samples
with Differential SLC7A7 Gene
Expression (All Glioma)
Number of samples in group:
Up-Regulated:
Intermediate:
Down-Regulated:
Log-rank p-value
Up-Regulated vs. Intermediate:
192
149
2
2.16x10-8
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 1 – SLC7A7 (y+LAT1)
Kaplan-Meier Survival Plot for
Samples with Differential SLC7A7
Gene Expression (Astrocytoma)
Number of samples in
group:
Up-Regulated:
51
Intermediate:
54
Log-rank p-value(for
significance of difference of
survival between group of
samples)
Up-Regulated vs.
Intermediate: 6.68x10-4
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Summary – Results 1
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SLC3A2
 Increased expression in GBM compared with normal brain
and Low grade glioma
 Up-regulation is associated with a significantly worse
prognosis when considering all glioma
 Particular prognostic significance in oligodendroglioma.
SLC7A5
 Expression not upregulated in GBM
 increased expression possibly associated with better
prognosis (Not significant)
SLC7A7
 Increased expression in GBM compared with normal brain
and Low grade glioma
 Particular prognostic significance in astrocytoma.
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Study Method 2
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Identified patients treated at the Queen Elizabeth Hospital,
Birmingham for low grade glioma who had an overall survival of
> 5 years (Long survival) and <5 years (short survival).
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RNA extraction from paraffin embedded tumour samples
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Performed whole exome sequencing (Oxford Gene Technology)
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Correlated gene expression of SLC3A2, SLC7A5 and SLC7A7 to
patient survival.
Identify if there was any association of LAT1 gene expression
with IDH1 mutation.
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 2 – SLC3A2 (Heavy chain)
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 2 – SLC7A5 (Light chain)
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 2 – SLC7A7
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Results 2 – IDH1 status
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Summary – Results 2
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SLC3A2
 Increased expression in these grade 2 glioma samples did
not appear to influence survival.
SLC7A5
 Increased expression in was associated with a significantly
worse prognosis. (p = 0.001)
SLC7A7
 Increased expression in was associated with a significantly
worse prognosis. (p = 0.016)
ICNCT-16, Helsinki, June 2014 / Desire Ngoga
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Glioma heterogeneity and the LAT-1: A first step to stratified BPA-based BNCT?
Discussion
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Though numbers of samples were small, we were able to show
the significant impact of the gene expression of the LAT1
transporter on the treatment and survival of glioma patients.
The fact that transporter up-regulation is associated with IDH1
wild type patients and a significantly worse prognosis offers the
potential to stratify these patients for BNCT
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Thank you for your attention
Further questions? [email protected]