1. |
- Alkan, Aylin, et al.
(author)
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Biomarkers and cell-based models to predict the outcome of neoadjuvant therapy for rectal cancer patients
- 2021
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In: Biomarker Research. - : Springer Science and Business Media LLC. - 2050-7771. ; 9
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Journal article (peer-reviewed)abstract
- Rectal cancer constitutes approximately one-third of all colorectal cancers and contributes to considerable mortality globally. In contrast to colon cancer, the standard treatment for localized rectal cancer often involves neoadjuvant chemoradiotherapy. Tumour response rates to treatment show substantial inter-patient heterogeneity, indicating a need for treatment stratification. Consequently researchers have attempted to establish new means for predicting tumour response in order to assist in treatment decisions. In this review we have summarized published findings regarding potential biomarkers to predict neoadjuvant treatment response for rectal cancer tumours. In addition, we describe cell-based models that can be utilized both for treatment prediction and for studying the complex mechanisms involved.
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2. |
- Almobarak, Bilal, et al.
(author)
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Exposure to nonanoic acid alters small intestinal neuroendocrine tumor phenotype
- 2023
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In: BMC Cancer. - : Springer Science and Business Media LLC. - 1471-2407. ; 23:1
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Journal article (peer-reviewed)abstract
- BackgroundSmall intestinal neuroendocrine tumors (SI-NET) are highly differentiated and genetically stable malignant tumors, yet they often present with advanced metastatic spread at the time of diagnosis. In contrast to many other types of malignant tumors, primary SI-NET are often asymptomatic and typically smaller in size compared to adjacent lymph node metastases. This study explores the hypothesis that stimulating the chemosensing olfactory receptor 51E1 (OR51E1) decreases SI-NET proliferation suggesting a mechanism that explains a difference in proliferative rate based on tumor location.MethodsClinical data was used to address difference in tumor size depending on location. A SI-NET tissue microarray was used to evaluate expression of OR51E1 and olfactory marker protein (OMP). Primary cultured tumor cells from 5 patients were utilized to determine the effect of OR51E1 agonist nonanoic acid on metabolic activity. The SI-NET cell line GOT1 was used to determine effects of nonanoic acid on the transcriptome as well as long-term effects of nonanoic acid exposure with regards to cell proliferation, serotonin secretion, alterations of the cell-cycle and morphology.ResultsTumor size differed significantly based on location. OR51E1 and OMP were generally expressed in SI-NET. Primary SI-NET cells responded to nonanoic acid with a dose dependent altered metabolic activity and this was replicated in the GOT1 cell line but not in the MCF10A control cell line. Nonanoic acid treatment in GOT1 cells upregulated transcripts related to neuroendocrine differentiation and hormone secretion. Long-term nonanoic acid treatment of GOT1 cells decreased proliferation, induced senescence, and altered cell morphology.ConclusionOur results raise the possibility that exposure of intraluminal metabolites could represent a mechanism determining aspects of the SI-NET tumor phenotype. However, we could not causally link the observed effects of nonanoic acid exposure to the OR51E1 receptor.
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3. |
- Andersson, Ellinor, et al.
(author)
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Expression profiling of small intestinal neuroendocrine tumors identifies subgroups with clinical relevance, prognostic markers and therapeutic targets.
- 2016
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In: Modern Pathology. - : Elsevier BV. - 0893-3952 .- 1530-0285. ; 29:6, s. 616-629
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Journal article (peer-reviewed)abstract
- We wanted to define the transcriptome of small intestinal neuroendocrine tumors in order to identify clinically relevant subgroups of tumors, prognostic markers and novel targets for treatment. Genome-wide expression profiling was conducted on tumor biopsies from 33 patients with well-differentiated neuroendocrine tumors of the distal ileum and metastatic disease at the time of diagnosis. Unsupervised hierarchical clustering analysis identified three groups of tumors. The largest group, comprising half of the tumors, was characterized by longer patient survival and higher expression of neuroendocrine markers, including SSTR2. Tumors with higher grade (G2/3) or gain of chromosome 14 were associated with shorter patient survival and increased expression of cell cycle-promoting genes. Pathway analysis predicted the prostaglandin E receptor 2 (PTGER2) as the most significantly activated regulator in tumors of higher grade, whereas Forkhead box M1 (FOXM1) was the most significantly activated regulator in tumors with gain of chromosome 14. Druggable genes identified from expression profiles included clinically proven SSTR2 and also novel targets, for example, receptor tyrosine kinases (RET, FGFR1/3, PDGFRB and FLT1), epigenetic regulators, molecular chaperones and signal transduction molecules. Evaluation of candidate drug targets on neuroendocrine tumors cells (GOT1) showed significant inhibition of tumor cell growth after treatment with tyrosine kinase inhibitors or inhibitors of HDAC, HSP90 and AKT. In conclusion, we have defined the transcriptome of small intestinal neuroendocrine tumors and identified novel subgroups with clinical relevance. We found specific gene expression patterns associated with tumor grade and chromosomal alterations. Our data also suggest novel prognostic biomarkers and therapies for these patients.
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4. |
- Elf, Anna-Karin, et al.
(author)
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NAMPT inhibitor GMX1778 enhances the efficacy of 177Lu-DOTATATE treatment of neuroendocrine tumors.
- 2017
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In: Journal of Nuclear Medicine. - : Society of Nuclear Medicine. - 0161-5505 .- 2159-662X. ; 58:2, s. 288-292
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Journal article (peer-reviewed)abstract
- Neuroendocrine tumors (NETs) can be treated by peptide receptor radionuclide therapy using radiolabeled somatostatin analogs. However, the efficacy of such treatment is low and needs to be optimized.To evaluate the potential radiosensitizing effects of NAMPT inhibition on (177)Lu-DOTATATE treatment in a NET model.Nude mice xenografted with the human NET cell line GOT1 were treated with semi-efficient doses of (177)Lu-DOTATATE (7,5 MBq, i.v.) and/or GMX1778 (100 mg/kg/week, p.o.).Median time to tumor progression (tumor volume larger than at day 0) was 3 days for controls, 7 days for single dose GMX1778, 28 days for single dose (177)Lu-DOTATATE and 35 days for 3 weekly doses of GMX1778. Combined treatment with (177)Lu-DOTATATE and GMX1778 x1 resulted in a median time to progression of 98 days. After (177)Lu-DOTATATE and 3 weekly doses of GMX1778 none of the tumors progressed within 120 days.The NAMPT inhibitor GMX1778 enhances the efficacy of (177)Lu-DOTATATE treatment and induces a prolonged antitumor response. Combinations of radiolabeled somatostatin analogs and radiosensitizing drugs should be further evaluated to optimize the efficacy of peptide receptor radionuclide therapy in NETs.
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5. |
- Hofving, Tobias, 1989, et al.
(author)
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177 Lu-octreotate therapy for neuroendocrine tumours is enhanced by Hsp90 inhibition
- 2019
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In: Endocrine-Related Cancer. - 1479-6821 .- 1351-0088. ; 26:4, s. 437-449
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Journal article (peer-reviewed)abstract
- Lu-177-octreotate is an FDA-approved radionuclide therapy for patients with gastroenteropancreatic neuroendocrine tumours (NETs) expressing somatostatin receptors. The Lu-177-octreotate therapy has shown promising results in clinical trials by prolonging progression-free survival, but complete responses are still uncommon. The aim of this study was to improve the Lu-177-octreotate therapy by means of combination therapy. To identify radiosensitising inhibitors, two cell lines, GOT1 and P-STS, derived from small intestinal neuroendocrine tumours (SINETs), were screened with 1224 inhibitors alone or in combination with external radiation. The screening revealed that inhibitors of Hsp90 can potentiate the tumour cell-killing effect of radiation in a synergistic fashion (GOT1; false discovery rate < 3.2 x 10(-11)). The potential for Hsp90 inhibitor ganetespib to enhance the anti-tumour effect of Lu-177-octreotate in an in vivo setting was studied in the somatostatin receptor-expressing GOT1 xenograft model. The combination led to a larger decrease in tumour volume relative to monotherapies and the tumour-reducing effect was shown to be synergistic. Using patient-derived tumour cells from eight metastatic SINETs, we could show that ganetespib enhanced the effect of Lu-177-octreotate therapy for all investigated patient tumours. Levels of Hsp90 protein expression were evaluated in 767 SINETs from 379 patients. We found that Hsp90 expression was upregulated in tumour cells relative to tumour stroma in the vast majority of SINETs. We conclude that Hsp90 inhibitors enhance the tumour-killing effect of Lu-177-octreotate therapy synergistically in SINET tumour models and suggest that this potentially promising combination should be further evaluated.
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7. |
- Hofving, Tobias, 1989, et al.
(author)
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SMAD4 haploinsufficiency in small intestinal neuroendocrine tumors
- 2021
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In: BMC Cancer. - : Springer Science and Business Media LLC. - 1471-2407. ; 21:1
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Journal article (peer-reviewed)abstract
- Background: Patients with small intestinal neuroendocrine tumors (SINETs) frequently present with lymph node and liver metastases at the time of diagnosis, but the molecular changes that lead to the progression of these tumors are largely unknown. Sequencing studies have only identified recurrent point mutations at low frequencies with CDKN1B being the most common harboring heterozygous mutations in less than 10% of all tumors. Although SINETs are genetically stable tumors with a low frequency of point mutations and indels, they often harbor recurrent hemizygous copy number alterations (CNAs) yet the functional implications of these CNA are unclear. Methods: Utilizing comparative genomic hybridization (CGH) arrays we analyzed the CNA profile of 131 SINETs from 117 patients. Two tumor suppressor genes and corresponding proteins i.e. SMAD4, and CDKN1B, were further characterized using a tissue microarray (TMA) with 846 SINETs. Immunohistochemistry (IHC) was used to quantify protein expression in TMA samples and this was correlated with chromosome number evaluated with fluorescent in-situ hybridization (FISH). Intestinal tissue from a Smad4+/− mouse model was used to detect entero-endocrine cell hyperplasia with IHC. Results: Analyzing the CGH arrays we found loss of chromosome 18q and SMAD4 in 71% of SINETs and that focal loss of chromosome 12 affecting the CDKN1B was present in 9.4% of SINETs. No homozygous loss of chromosome 18 was detected. Hemizygous loss of SMAD4, but not CDKN1B, significantly correlated with reduced protein levels but hemizygous loss of SMAD4 did not induce entero-endocrine cell hyperplasia in the Smad4+/− mouse model. In addition, patients with low SMAD4 protein expression in primary tumors more often presented with metastatic disease. Conclusions: Hemizygous loss of chromosome 18q and the SMAD4 gene is the most common genetic event in SINETs and our results suggests that this could influence SMAD4 protein expression and spread of metastases. Although SMAD4 haploinsufficiency alone did not induce tumor initiation, loss of chromosome 18 could represent an evolutionary advantage in SINETs explaining the high prevalence of this aberration. Functional consequences of reduced SMAD4 protein levels could hypothetically be a potential mechanism as to why loss of chromosome 18 appears to be clonally selected in SINETs.
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8. |
- Hofving, Tobias, 1989
(author)
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Small intestinal neuroendocrine tumours - Disease models, tumour development, and remedy
- 2019
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Doctoral thesis (other academic/artistic)abstract
- Small intestinal neuroendocrine tumours (SINETs) are malignant neoplasms which at the time of diagnosis often present with distant metastasis. The field of SINET research faces several challenges. There is a lack of preclinical models for studying SINETs, and it is unclear how well currently available models actually recapitulate the tumour disease. The genetic changes that underlie SINET tumour development are largely unknown and, lastly, curative therapy is rarely achieved. Novel therapies, such as the recently FDA-approved 177Lu-octreotate therapy and up-and-coming immunotherapies need to be further investigated to deliver better response rates for SINET patients. In our first two papers (papers I and II), we sought to evaluate frequently used and readily available gastroenteropancreatic neuroendocrine tumour (GEPNET) cell lines as models of neuroendocrine tumour disease. We investigated the characteristics of these cell lines in terms of their neuroendocrine phenotype, genomic background, and therapeutic sensitivity. While several cell lines exhibited an expected neuroendocrine differentiation and harboured genetic alterations characteristic of the GEPNET disease, three cell lines did not. In fact, it turned out that one of the most frequently used cell lines in the field – KRJ-I, together with the cell lines L-STS and H-STS, were incorrectly identified and instead lymphoblastoid cell lines (EBV-immortalised B-lymphocytes). This might have led to the incorrect use and potentially faulty conclusions in a number of GEPNET studies. Among authentic cell lines, we performed a large-scale inhibitor sensitivity screening and predicted that SINETs would be more sensitive to HDACi compared to pancreatic neuroendocrine tumours (PanNET) and PanNET more sensitive to MEKi compared to SINET. The prediction was supported by subsequent experiments with primary tumour cells. In our third paper (paper III), we evaluated a mechanism by which hemizygous loss of SMAD4 could lead to SINET initiation and/or progression by acting as a haploinsufficient tumour suppressor. We found that loss of SMAD4 was associated with a decrease in corresponding mRNA and protein, and that this correlated to patient survival. We also found that the amount of SMAD4 protein in the primary tumour could predict whether the patient presented with distant metastasis. In our last papers (papers IV and V), we investigated the potential for two novel treatment strategies for SINETs. In paper IV we identified an inhibitor, the heat shock protein 90 inhibitor ganetespib, that could synergistically enhance the 177Lu-octreotate therapy for SINETs. Ganetespib was initially found to sensitise SINETs to radiation in a large-scale inhibitor synergy screening, and its radiosensitising effect for radionuclide treatment of SINETs was validated both in mouse xenografts and in primary patient tumours. Lastly, in paper V we characterised the SINET immune microenvironment. Using immunohistochemistry and flow-cytometry we detailed the immune cell composition of the SINET immune microenvironment and could demonstrate the successful isolation and expansion of tumour-infiltrating lymphocytes. We saw that after infiltrating lymphocytes were expanded they could degranulate when challenged with autologous tumour cells. In conclusion, these studies have provided a thorough characterisation of authentic, and provided important information regarding misidentified, frequently used gastroenteropancreatic cell lines. It has also investigated the role of hemizygous SMAD4 loss in the development of SINETs and demonstrated the potential of two novel therapies for SINETs: 177Lu-octreotate combined with Hsp90i ganetespib and immunotherapy.
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9. |
- Hofving, Tobias, 1989, et al.
(author)
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The Microenvironment of Small Intestinal Neuroendocrine Tumours Contains Lymphocytes Capable of Recognition and Activation after Expansion
- 2021
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In: Cancers. - : MDPI AG. - 2072-6694. ; 13:17
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Journal article (peer-reviewed)abstract
- Simple Summary The body's immune system can recognize tumors because they often contain proteins that are either different from or more abundant than in normal cells. Here, we characterised the immune cells of a rare tumor type called small-intestinal neuroendocrine tumors (SINET). We find that so called tumour-infiltrating lymphocytes (TILs) can be grown in the laboratory and activated by challenging them with digested tumor. This study provides insights into the largely unknown SINET immune landscape and reveals the anti-tumour reactivity of TILs, which might merit adoptive T cell transfer as a feasible treatment option for patients with SINET. Traditionally, immune evasion and immunotherapy have been studied in cancers with a high mutational load such as melanoma or lung cancer. In contrast, small intestinal neuroendocrine tumours (SINETs) present a low frequency of somatic mutations and are described as genetically stable tumours, rendering immunotherapies largely unchartered waters for SINET patients. SINETs frequently metastasise to the regional lymph nodes and liver at the time of diagnosis, and no curative treatments are currently available for patients with disseminated disease. Here, we characterised the immune landscape of SINET and demonstrated that tumour-infiltrating lymphocytes (TILs) can be expanded and activated during autologous tumour challenge. The composition of lymphocyte subsets was determined by immunophenotyping of the SINET microenvironment in one hepatic and six lymph node metastases. TILs from these metastases were successfully grown out, enabling immunophenotyping and assessment of PD-1 expression. Expansion of the TILs and exposure to autologous tumour cells in vitro resulted in increased T lymphocyte degranulation. This study provides insights into the largely unknown SINET immune landscape and reveals the anti-tumour reactivity of TILs, which might merit adoptive T cell transfer as a feasible treatment option for patients with SINET.
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10. |
- Hofving, Tobias, 1989, et al.
(author)
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The neuroendocrine phenotype, genomic profile and therapeutic sensitivity of GEPNET cell lines
- 2018
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In: Endocrine-Related Cancer. - : Bioscientifica. - 1351-0088 .- 1479-6821. ; 25:3, s. 367-380
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Journal article (peer-reviewed)abstract
- Experimental models of neuroendocrine tumour disease are scarce, and no comprehensive characterisation of existing gastroenteropancreatic neuroendocrine tumour (GEPNET) cell lines has been reported. In this study, we aimed to define the molecular characteristics and therapeutic sensitivity of these cell lines. We therefore performed immunophenotyping, copy number profiling, whole-exome sequencing and a large-scale inhibitor screening of seven GEPNET cell lines. Four cell lines, GOT1, P-STS, BON-1 and QGP-1, displayed a neuroendocrine phenotype while three others, KRJ-I, L-STS and H-STS, did not. Instead, these three cell lines were identified as lymphoblastoid. Characterisation of remaining authentic GEPNET cell lines by copy number profiling showed that GOT1, among other chromosomal alterations, harboured losses on chromosome 18 encompassing the SMAD4 gene, while P-STS had a loss on 11q. BON-1 had a homozygous loss of CDKN2A and CDKN2B, and QGP-1 harboured amplifications of MDM2 and HMGA2. Whole-exome sequencing revealed both disease-characteristic mutations (e.g. ATRX mutation in QGP-1) and, for patient tumours, rare genetic events (e.g. TP53 mutation in P-STS, BON-1 and QGP-1). A large-scale inhibitor screening showed that cell lines from pancreatic NETs to a greater extent, when compared to small intestinal NETs, were sensitive to inhibitors of MEK. Similarly, neuroendocrine NET cells originating from the small intestine were considerably more sensitive to a group of HDAC inhibitors. Taken together, our results provide a comprehensive characterisation of GEPNET cell lines, demonstrate their relevance as neuroendocrine tumour models and explore their therapeutic sensitivity to a broad range of inhibitors.
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