Refined classification of breast cancer: Development of non-invasive biomarkers based on DNA methylation
Prosjekt
- Prosjektnummer
- 2016036
- Ansvarlig person
- Thomas Fleischer
- Institusjon
- Oslo universitetssykehus HF
- Prosjektkategori
- Doktorgradsstipend
- Helsekategori
- Cancer
- Forskningsaktivitet
- 2. Aetiology
Rapporter
We have shown that epigenetic markers such as DNA methylation and miRNA expression can be used as clinically applicable biomarkers, and through epigenetic editing we have shown that enhancer methylation can inhibit target gene expression illustrating the causal regulatory role of epigenetic alteration in cancer.Breast cancer is a heterogeneous disease and can be viewed as a collection of different diseases, and prognosis and response to treatments will vary greatly from patient to patient. Analyses of cellular and molecular characteristics allow subgrouping of patients, indicating which therapy the patient should receive and the biological mechanisms driving pathogenesis. The most important subdivision of breast cancers is the over expression of the protein estrogen receptor (ER). In tumors over-expressing ER, the hormone estrogen binds to the ER, which further activates genes that promote cell proliferation and tumor growth. In ER negative tumors cell proliferation is driven by other mechanisms such as alterations to the DNA, and the importance of epigenetic alterations is more uncertain.
DNA methylation is a modification made to the DNA molecule which can affect the expression of genes. We have developed a prognostic signature for patients with Luminal A disease based on DNA methylation of 40 genes and termed this signature SAM40 (Fleischer, Klajic et al., Oncotarget 2017). Dividing patients with Luminal A tumors into two groups with different prognosis may give two benefits: patients with poor prognosis can get more aggressive treatment, and patients with good prognosis may get less treatment. Providing less treatment for patients with good prognosis may avoid the side effects of treatment and improve quality of life.
MicroRNAs (miRNAs) are small non-coding RNAs that may regulate gene expression by regulating translation of mRNAs into proteins. miR-34a is found to be frequently down-regulated in cancer and is thought to have a tumor suppressive role in many cancers, and miR-34a replacement therapy has been suggested. When we over-expressed miR34a in a triple negative breast cancer cell line, we found that genes related to the Serine-glycine biosynthesis pathway and glycolysis had altered gene expression. In a breast cancer patient cohort, we saw that the genes in these pathways were negatively correlated to the expression of miR-34a, and that high levels of miR-34a was associated to better prognosis in patients with basal-like tumors (Rosvold et al., manuscript in preparation).
We are also working with DNA methylation biomarkers from blood samples. Biomarkers from liquid biopsies may allow early detection as well as treatment monitoring. Using bioinformatic approaches, we have identified potent candidates for CpGs methylated only in breast tumors, and we have been able to identify this methylation in blood samples from the corresponding patients. We are setting up the Enhanced-ice-COLD-PCR (Tost 2016; PMID:26589575) method for detection of DNA methylation in blood samples.
To investigate the functional importance of the alterations we observe with the statistical and bioinformatic approach (above), we use CRISPR epigenetic editing to alter specific parts of the epigenome. We have established a system in the lab that allows us to first alter the methylation at specific target loci, then measure alterations in transcription factor binding and target gene expression. The results show that increased enhancer methylation inhibits expression of target genes.
Nei
We have shown that epigenetic markers such as DNA methylation and miRNA expression can be used as clinically applicable biomarkers, and we are developing DNA methylation biomarkers from blood samples. Epigenetic edition will shed light on the causal regulatory role of epigenetic alteration in cancer.Breast cancer is a heterogeneous disease and can be viewed as a collection of different diseases, and prognosis and response to treatments will vary greatly from patient to patient. Analyses of cellular and molecular characteristics allow subgrouping of patients, indicating which therapy the patient should receive and the biological mechanisms driving pathogenesis. The most important subdivision of breast cancers is the over expression of the protein estrogen receptor (ER). In tumors over-expressing ER, the hormone estrogen binds to the ER, which further activates genes that promote cell proliferation and tumor growth. In ER negative tumors cell proliferation is driven by other mechanisms such as alterations to the DNA, and the importance of epigenetic alterations is more uncertain.
DNA methylation is a modification made to the DNA molecule which can affect the expression of genes. We have developed a prognostic signature for patients with Luminal A disease based on DNA methylation of 40 genes and termed this signature SAM40 (Fleischer, Klajic et al., Oncotarget 2017). Dividing patients with Luminal A tumors into two groups with different prognosis may give two benefits: patients with poor prognosis can get more aggressive treatment, and patients with good prognosis may get less treatment. Providing less treatment for patients with good prognosis may avoid the side effects of treatment and improve quality of life.
MicroRNAs (miRNAs) are small non-coding RNAs that may regulate gene expression by regulating translation of mRNAs into proteins. miR-34a is found to be frequently down-regulated in cancer and is thought to have a tumor suppressive role in many cancers, and miR-34a replacement therapy has been suggested. When we over-expressed miR34a in a triple negative breast cancer cell line, we found that genes related to the Serine-glycine biosynthesis pathway and glycolysis had altered gene expression. In a breast cancer patient cohort, we saw that the genes in these pathways were negatively correlated to the expression of miR-34a, and that high levels of miR-34a was associated to better prognosis in patients with basal-like tumors (Rosvold et al., manuscript in preparation).
We are also working with DNA methylation biomarkers from blood samples. Biomarkers from liquid biopsies may allow early detection as well as treatment monitoring. Using bioinformatic approaches, we have identified potent candidates for CpGs methylated only in breast tumors, and we have been able to identify this methylation in blood samples from the corresponding patients. We are setting up the Enhanced-ice-COLD-PCR (Tost 2016; PMID:26589575) method for detection of DNA methylation in blood samples.
To investigate the functional importance of the alterations we observe with the statistical and bioinformatic approach (above), we use CRISPR epigenetic editing to alter specific parts of the epigenome. We have estabilished a system in the lab that allows us to first alter the methylation at specific target loci, then measure alterations in transcription factor binding and target gene expression. The results (in preparation) will shed further light in the causal role of DNA methylation in regulation of gene expression and tumor development.
Nei
We show that epigenetic markers such as DNA methylation and miRNA expression can be used as clinically applicable biomarkers, and we are developing DNA methylation biomarkers from blood samples. We are now using CRISPR epigenetic editing to change epigenetic properties for improved therapy.Breast cancer is a heterogeneous disease and can be viewed as a collection of different diseases, and prognosis and response to treatments will vary greatly from patient to patient. Analyses of cellular and molecular characteristics allow subgrouping of patients, indicating which therapy the patient should receive and the biological mechanisms driving pathogenesis. The most important subdivision of breast cancers is the over expression of the protein estrogen receptor (ER). In tumors over-expressing ER, the hormone estrogen binds to the ER, which further activates genes that promote cell proliferation and tumor growth. In ER negative tumors cell proliferation is driven by other mechanisms such as alterations to the DNA, and the importance of epigenetic alterations is more uncertain.
DNA methylation is a modification made to the DNA molecule which can affect the expression of genes. We have developed a prognostic signature for patients with Luminal A disease based on DNA methylation of 40 genes and termed this signature SAM40 (Fleischer, Klajic et al., Oncotarget 2017). Dividing patients with Luminal A tumors into two groups with different prognosis may give two benefits: patients with poor prognosis can get more aggressive treatment, and patients with good prognosis may get less treatment. Providing less treatment for patients with good prognosis may avoid the side effects of treatment and improve quality of life.
MicroRNAs (miRNAs) are small non-coding RNAs that may regulate gene expression by regulating translation of mRNAs into proteins. miR-34a is found to be frequently down-regulated in cancer and is thought to have a tumor suppressive role in many cancers, and miR-34a replacement therapy has been suggested. When we over-expressed miR34a in a triple negative breast cancer cell line, we found that genes related to the Serine-glycine biosynthesis pathway had altered gene expression. In a breast cancer patient cohort, we saw that the genes in the pathway were negatively correlated to the expression of miR-34a, and that high levels of miR-34a was associated to better prognosis in patients with basal-like tumors [Rosvold et al., soon to be submitted]
We are also working with DNA methylation biomarkers from blood samples. Biomarkers from liquid biopsies may allow early detection as well as treatment monitoring. Using bioinformatic approaches, we have identified potent candidates for CpGs methylated only in breast tumors, and we have been able to identify this methylation in blood samples from the corresponding patients. We are setting up the Enhanced-ice-COLD-PCR (Tost 2016; PMID:26589575) method for detection of DNA methylation in blood samples. This is ongoing work, and more patients will be included.
To investigate the functional importance of epigenetic alterations in ER positive breast cancer, we use CRISPR epigenetic editing to alter the methylation level of distal regulatory element that we think is regulating a gene controlled by estrogen receptor. This approach allows us to measure the effects on proliferation, tumor growth after epigenetic alteration [Rosvold, Wang, Ankill et al., in preparation].
nei
We show that epigenetic markers such as DNA methylation and miRNA expression can be used as clinically applicable biomarkers, and we are developing DNA methylation biomarkers from blood samples. We are now using CRISPR epigenetic editing to change epigenetic properties for improved therapy.Breast cancer is a heterogeneous disease and can be viewed as a collection of different diseases, and prognosis and response to treatments will vary greatly from patient to patient. Analyses of cellular and molecular characteristics allow subgrouping of patients, indicating which therapy the patient should receive and the biological mechanisms driving pathogenesis. The most important subdivision of breast cancers is the over expression of the protein estrogen receptor (ER). In tumors over-expressing ER, the hormone estrogen binds to the ER, which further activates genes that promote cell proliferation and tumor growth. In ER negative tumors cell proliferation is driven by other mechanisms such as alterations to the DNA, and the importance of epigenetic alterations is more uncertain.
DNA methylation is a modification made to the DNA molecule which can affect the expression of genes. We have developed a prognostic signature for patients with Luminal A disease based on DNA methylation of 40 genes and termed this signature SAM40 (Fleischer, Klajic et al., Oncotarget 2017). Dividing patients with Luminal A tumors into two groups with different prognosis may give two benefits: patients with poor prognosis can get more aggressive treatment, and patients with good prognosis may get less treatment. Providing less treatment for patients with good prognosis may avoid the side effects of treatment and improve quality of life.
MicroRNAs (miRNAs) are small non-coding RNAs that may regulate gene expression by regulating translation of mRNAs into proteins. miR-34a is found to be frequently down-regulated in cancer and is thought to have a tumor suppressive role in many cancers, and miR-34a replacement therapy has been suggested. When we over-expressed miR34a in a triple negative breast cancer cell line, we found that genes related to the Serine-glycine biosynthesis pathway had altered gene expression. In a breast cancer patient cohort, we saw that the genes in the pathway were negatively correlated to the expression of miR-34a, and that high levels of miR-34a was associated to better prognosis in patients with basal-like tumors [Rosvold et al., soon to be submitted]
We are also working with DNA methylation biomarkers from blood samples. Biomarkers from liquid biopsies may allow early detection as well as treatment monitoring. Using bioinformatic approaches, we have identified potent candidates for CpGs methylated only in breast tumors, and we have been able to identify this methylation in blood samples from the corresponding patients. We are setting up the Enhanced-ice-COLD-PCR (Tost 2016; PMID:26589575) method for detection of DNA methylation in blood samples. This is ongoing work, and more patients will be included.
To investigate the functional importance of epigenetic alterations in ER positive breast cancer, we use CRISPR epigenetic editing to alter the methylation level of distal regulatory element that we think is regulating a gene controlled by estrogen receptor. This approach allows us to measure the effects on proliferation, tumor growth after epigenetic alteration [Rosvold, Wang, Ankill et al., in preparation].
Nei
Improved classification of breast cancer is crucial for improving both patient survival as well as quality of life for survivors. We show that epigenetic markers such as DNA methylation and miRNA expression can be used as clinically applicable biomarkers, and we are developing DNA methylation biomarkers from blood samples.Luminal A breast cancers are most often hormone receptor positive, Her2 negative, and have relatively low cell proliferation. Patients with this tumor type have relatively good prognosis, but many patients may still relapsed within ten years. Basal-like breast cancer is often both hormone receptor and Her2 negative, and has high proliferation, and the patients with this tumor type has relatively poor prognosis. Our goals are therefore to develop methods for subdividing patients with Luminal A or basal-like tumors into new subgroups with clinical utility, and to use DNA methylation in blood samples to assess treatment response.
DNA methylation is a modifications made to the DNA molecule which can affect the expression of genes. We have developed a prognostic signature for patients with Luminal A disease based on DNA methylation of 40 genes and termed this signature SAM40 (Fleischer, Klajic et al., Oncotarget 2017). Dividing patients with Luminal A tumors into two groups with different prognosis may give two benefits: patients with poor prognosis can get more aggressive treatment, and patients with good prognosis may get less treatment. Providing less treatment for patients with good prognosis may avoid the side effects of treatment and improve quality of life.
MicroRNAs (miRNAs) are small non-coding RNAs that may regulate gene expression by regulating translation of mRNAs into proteins. miR-34a is found to be frequently down-regulated in cancer and is thought to have a tumor suppressive role in many cancers, and miR-34a replacement therapy has been suggested. When we over-expressed miR34a in a triple negative breast cancer cell line, we found that genes related to the Serine-glycine biosynthesis pathway had altered gene expression. In a breast cancer patient cohort we saw that the genes in the pathway were negatively correlated to the expression of miR-34a, and that high levels of miR-34a was associated to better prognosis in patients with basal-like tumors. Associations between miR-34a expression and metabolic profiles will be investigated. This manuscript is under preparation.
We are also working with DNA methylation biomarkers from blood samples. Biomarkers from liquid biopsies may allow early detection as well as treatment monitoring. Using bioinformatic approaches, we have identified potent candidates for CpGs methylated only in breast tumors, and we have been able to identify this methylation in blood samples from the corresponding patients. We are setting up the Enhanced-ice-COLD-PCR (Tost 2016; PMID:26589575) method for detection of DNA methylation in blood samples. This is ongoing work, and more patients will be included. In addition, we will analyze blood samples from a breast cancer clinical trial where patients have been treated with chemotherapy +/- bevacizumab, to assess whether blood samples can be used to predict and monitor treatment response.
We have developed a statistical approach to identify epigenetically regulated pathways in breast cancer, and we have identified that loss of enhancer methylation is a key feature of ER positive breast cancer. To determine the functional importance of these epigenetic alterations, we will perform precision epigenetic engineering and alter the epigenetic landscape at specific loci, followed by assessment of gene expression and changes in the phenotype. These methods are set up in the lab.
NEI
Improved classification of breast cancer is crucial for improving both patient survival as well as quality of life for survivors. We show that epigenetic markers such as DNA methylation and miRNA expression can be used as clinically applicable biomarkers, and we are developing DNA methylation biomarkers from blood samples.Luminal A breast cancers are most often hormone receptor positive, Her2 negative, and have relatively low cell proliferation. Patients with this tumor type have relatively good prognosis, but many patients may still relapsed within ten years. Basal-like breast cancer is often both hormone receptor and Her2 negative, and has high proliferation, and the patients with this tumor type has relatively poor prognosis. Our goals are therefore to develop methods for subdividing patients with Luminal A or basal-like tumors into new subgroups with clinical utility, and to use DNA methylation in blood samples to assess treatment response.
DNA methylation is a modifications made to the DNA molecule which can affect the expression of genes. We have developed a prognostic signature for patients with Luminal A disease based on DNA methylation of 40 genes and termed this signature SAM40 (Fleischer, Klajic et al., Oncotarget 2017). Dividing patients with Luminal A tumors into two groups with different prognosis may give two benefits: patients with poor prognosis can get more aggressive treatment, and patients with good prognosis may get less treatment. Providing less treatment for patients with good prognosis may avoid the side effects of treatment and improve quality of life.
MicroRNAs (miRNAs) are small non-coding RNAs that may regulate gene expression by regulating translation of mRNAs into proteins. miR-34a is found to be frequently down-regulated in cancer and is thought to have a tumor suppressive role in many cancers, and miR-34a replacement therapy has been suggested. When we over-expressed miR34a in a triple negative breast cancer cell line, we found that genes related to the Serine-glycine biosynthesis pathway had altered gene expression. In a breast cancer patient cohort we saw that the genes in the pathway were negatively correlated to the expression of miR-34a, and that high levels of miR-34a was associated to better prognosis in patients with basal-like tumors. Associations between miR-34a expression and metabolic profiles will be investigated. This manuscript is under preparation.
We are currently generating methylation profiles using Illumina EPIC arrays (850,000 CpGs) for 96 patients with Luminal A disease in the Oslo1 cohort. These patients have around 20 years follow-up data, and most of the patients have not received adjuvant treatment (neither hormone treatment nor chemotherapy). This data will allow us to assess the clinical validity and utility of the SAM40 prognostic signature. In addition, we generate miRNA for these patients, and we will compare and combine the SAM40 signature with the miRNA signature reported in Aure et al., BCR 2017.
We are also working with DNA methylation biomarkers from blood samples. Biomarkers from liquid biopsies may allow early detection as well as treatment monitoring. Using bioinformatic approaches, we have identified potent candidates for CpGs methylated only in breast tumors, and we have been able to identify this methylation in blood samples from the corresponding patients. This is ongoing work, and more patients will be included. In addition, we will analyze blood samples from a breast cancer clinical trial where patients have been treated with chemotherapy +/- bevacizumab, to assess whether blood samples can be used to predict and monitor treatment response.
Improved classification of breast cancer is crucial for improving both patient survival as well as quality of life for survivors. Epigenetic markers such as DNA methylation and miRNA expression has great potential for clinical utility, and may also give valuable insight into the etiology of breast cancer.Luminal A breast cancers are most often hormone receptor positive, Her2 negative, and have relatively low cell proliferation. Patients with this tumor type have relatively good prognosis, but many patients may still relapsed within ten years. Basal-like breast cancer is often both hormone receptor and Her2 negative, and has high proliferation, and the patients with this tumor type has relatively poor prognosis. Our goals are therefore to develop methods for subdividing patients with Luminal A or basal-like tumors into new subgroups with clinical utility, and to understand the underlying biological differences between aggressive and less aggressive tumors within each subtype.
DNA methylation is a modifications made to the DNA molecule which can affect the expression of genes. We have developed a prognostic signature based on DNA methylation of 40 genes and termed this signature SAM40. We applied the SAM40 signature to four different patient groups, and we saw that we could divide the patients into two groups. One group showed low levels of methylation of the 40 genes, and these patients had good prognosis; the other group had high levels of methylation, and these patients had worse prognosis. Dividing patients with Luminal A tumors into two groups with different prognosis may give two benefits: patients with poor prognosis can get more aggressive treatment, and patients with good prognosis may get less treatment. Providing less treatment for patients with good prognosis is an advantage because these patients may avoid the side effects of treatment that can lead to permanently impaired quality of life. The work so far has been published in Oncotarget, 8 (1), 1074-1082; PMID: 27911866.
MicroRNAs (miRNAs) are small non-coding RNAs that may regulate gene expression by regulating translation of mRNAs into proteins. miR-34a is found to be frequently down-regulated in cancer and is thought to have a tumor suppressive role in many cancers, and miR-34a replacement therapy has been suggested. When we over expressed miR34a in a triple negative breast cancer cell line, we found that genes related to the Serine-glycine biosynthesis pathway had altered gene expression. In a breast cancer patient cohort we saw that the genes in the pathway were negatively correlated to the expression of miR-34a, and that high levels of miR-34a was associated to better prognosis in patients with basal-like tumors. This manuscript will be finalized soon after the candidate returns from maternity leave.
In future work, we will associate miR-34a expression with metabolic profiles in another breast cancer cohort, and we will further study the clinical impact of miR-34a expression in patients with basal-like tumors. The SAM40 classification will be validated in more patients with clinical follow up, and clinical applications will be explored.
Bedre klassifisering av brystkreftpasienter kan gi store fordeler. I dette prosjektet bruker vi biomarkører basert DNA-metylering til å hjelpe beslutningsprosessene ved behandling av brystkreft, noe som kan innebære å gi mildere behandling til pasienter med veldig god prognose og dermed øke deres livskvalitet.Luminal A er en brystkrefttype som er kjennetegnet ved at tumorene er hormonreseptor positive, Her2-reseptor negative, og har relativt lav celledeling. Pasienter med denne tumortypen har relativt god prognose, men samtidig vet vi at mange pasienter får tilbakefall innen ti år. Vårt mål er derfor å utvikle en metode for å dele inn pasienter med Luminal A i nye subgrupper, og at denne klassifiseringen kan bidra til å bestemme hvilken behandling som blir gitt til pasienter.
DNA-metylering er modifikasjoner som gjøres til DNA-molekylet, og som kan påvirke utrykket av gener. Vi har utviklet en signatur basert på DNA-metylering av 40 gener, og vi har kalt denne metoden SAM40. Vi har anvendt SAM40-signaturen i fire forskjellige pasientgrupper, og vi ser at vi kan dele inn pasientene i to grupper. Den ene gruppen har lavt metyleringsnivå av de 40 genene, og disse pasientene har god prognose; den andre gruppen har høyt metyleringsnivå av de 40 genene, og disse pasientene har dårligere prognose. Det å kunne dele pasienter med Luminal A tumorer inn i to grupper med forskjellig prognose kan gi to fordeler: pasienter med dårlig prognose kan få mer aggressiv behandling, og pasienter med god prognose kan få mindre, eller slippe, behandling. Det å gi mindre behandling til pasienter med god prognose er en fordel fordi disse pasientene da unngår bivirkninger av behandlingen som kan medføre varig nedsatt livskvalitet. Arbeidet så langt er publisert i Oncotarget, 8 (1), 1074-1082; PMID: 27911866.
I det videre arbeidet vil vi først forsikre oss om at vår måte å dele inn pasienter med Luminal A tumorer er den beste. For å undersøke dette vil vi sammenligne vår signatur med andre metoder som også finner forskjeller innad i gruppen med Luminal A tumorer og som er publisert av andre forskningsgrupper. Vi vil finne ut hvilken signatur som er best, og om signaturene kan kombineres.
Vitenskapelige artikler
Fleischer T, Klajic J, Aure MR, Louhimo R, Pladsen AV, Ottestad L, Touleimat N, Laakso M, Halvorsen AR, Grenaker Alnæs GI, Riis ML, Helland Å, Hautaniemi S, Lønning PE, Naume B, Børresen-Dale AL, Tost J, Kristensen VN
DNA methylation signature (SAM40) identifies subgroups of the Luminal A breast cancer samples with distinct survival.
Oncotarget 2017 Jan 03;8(1):1074-1082.
PMID: 27911866
Deltagere
- Brock Christensen Prosjektdeltaker
- Zdenko Herceg Prosjektdeltaker
- Olav Engebråten Prosjektdeltaker
- Bjørn Naume Prosjektdeltaker
- Ragnhild Eskeland Prosjektdeltaker
- Olga Troyanskaya Prosjektdeltaker
- Anthony Mathelier Prosjektdeltaker
- Antoni Hurtado Rodriguez Prosjektdeltaker
- Jörg Tost Prosjektdeltaker
- Arnoldo Frigessi Prosjektdeltaker
- Vessela N. Kristensen Medveileder
- Marie Elise Engkvist Postdoktorstipendiat (finansiert av denne bevilgning)
- Thomas Fleischer Hovedveileder
eRapport er utarbeidet av Sølvi Lerfald og Reidar Thorstensen, Regionalt kompetansesenter for klinisk forskning, Helse Vest RHF, og videreutvikles av de fire RHF-ene i fellesskap, med støtte fra Helse Vest IKT
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