Ignite Creation Date:
2024-10-26 @ 3:42 PM
Last Modification Date:
2024-10-26 @ 3:42 PM
Study NCT ID:
NCT06642506
Status:
ACTIVE_NOT_RECRUITING
Last Update Posted:
None
First Post:
2024-10-11
Brief Title:
Oncometabolome and MALDI-MSI in Upper GI Carcinomas - Chemosensitivity in Esophageal Carcinoma
Sponsor:
None
Organization:
None
Study Overview
Official Title:
Oncometabolome and MALDI-MSI in Upper GI Carcinomas - Chemosensitivity in Esophageal Carcinoma
Status:
ACTIVE_NOT_RECRUITING
Status Verified Date:
2024-10
Last Known Status:
None
Delayed Posting:
No
If Stopped, Why?:
Not Stopped
Has Expanded Access:
No
If Expanded Access, NCT#:
N/A
Has Expanded Access, NCT# Status:
N/A
Acronym:
None
Brief Summary:
Locally advanced adenocarcinoma of the esophagus is a leading cause of death from malignant disease in Germany and has been characterized on a molecular level in recent years This retrospective observational study deals with patients after esophagectomy with different risk constellations of esophageal carcinoma An early and individualized therapy of this tumor in an approach of precision oncology significantly improves the prognosis The metabolomic profile plays a central role in tumor plasticity and oncological outcome At the same time these factors affect the efficacy of chemotherapy and need to be investigated in more detail at the molecular level A central element of this study is the investigation of phospholipid metabolism locally in tumor tissue in adjacent normal tissue in terms of the tumor microenvironment and systemically in blood plasma The focus lies on the validation of known oncometabolites that significantly influence tumor sensitivity to chemotherapy
By combining mass spectrometry imaging using matrix-assisted laser desorption ionization - mass spectrometry imaging MALDI-MSI with metabolomics using liquid chromatography tandem mass spectrometry LC-MSMS the metabolic profile of tumors can be analyzed in detail allowing conclusions to be drawn about chemo-insensitive and therapeutically challenging tumors Both mass spectrometric methods are used to understand the heterogeneous metabolism of the tumors and to describe possible constellations that are associated with increasing chemoresistance For precise investigation the cohort under investigation is divided into two patient collectives Patients with a regression grade 1 after four sessions of FLOT chemotherapy are compared with a regression grade 3 according to Becker in the postoperative pathological assessment This facilitates the development of personalized therapeutic approaches tailored to the individual oncological profiles of the tumors The study is complemented by conventional HE microscopic examinations of the tumor itself and the tumor microenvironment which allow to analyze the morphology and its correlation with metabolic alterations in the tissue We hypothesize that adenocarcinoma of the esophagus with regression grade 1 encompasses a fundamentally distinct metabolic profile than adenocarcinoma of the esophagus with regression grade 3 Consequently a stratification parameter within the local tumor metabolism and the tumor microenvironment exists which correlates with the systemic response to neoadjuvant chemotherapy in blood plasma
The primary aim of the study is to create a comprehensive metabolic profile that clearly identifies tumors with a regression grade 1 versus a regression grade 3 according to Becker This will be used to improve diagnostics and develop personalized treatment strategies that increase treatment efficiency and patients chances of survival
This is ultimately carried out with the intention of achieving an improved survival rate and a higher quality of life for patients with locally advanced esophageal cancer The comprehensive analysis of the tumor microenvironment and the morphological and metabolic profiles should provide new insights into the mechanisms of tumor progression and resistance which in turn will form the basis for future translational research and treatment approaches The findings from this study have the potential to change the way esophageal cancer is treated by contributing to the development of stratified therapeutic approaches tailored to the molecular subtype of esophageal cancer
By combining mass spectrometry imaging using matrix-assisted laser desorption ionization - mass spectrometry imaging MALDI-MSI with metabolomics using liquid chromatography tandem mass spectrometry LC-MSMS the metabolic profile of tumors can be analyzed in detail allowing conclusions to be drawn about chemo-insensitive and therapeutically challenging tumors Both mass spectrometric methods are used to understand the heterogeneous metabolism of the tumors and to describe possible constellations that are associated with increasing chemoresistance For precise investigation the cohort under investigation is divided into two patient collectives Patients with a regression grade 1 after four sessions of FLOT chemotherapy are compared with a regression grade 3 according to Becker in the postoperative pathological assessment This facilitates the development of personalized therapeutic approaches tailored to the individual oncological profiles of the tumors The study is complemented by conventional HE microscopic examinations of the tumor itself and the tumor microenvironment which allow to analyze the morphology and its correlation with metabolic alterations in the tissue We hypothesize that adenocarcinoma of the esophagus with regression grade 1 encompasses a fundamentally distinct metabolic profile than adenocarcinoma of the esophagus with regression grade 3 Consequently a stratification parameter within the local tumor metabolism and the tumor microenvironment exists which correlates with the systemic response to neoadjuvant chemotherapy in blood plasma
The primary aim of the study is to create a comprehensive metabolic profile that clearly identifies tumors with a regression grade 1 versus a regression grade 3 according to Becker This will be used to improve diagnostics and develop personalized treatment strategies that increase treatment efficiency and patients chances of survival
This is ultimately carried out with the intention of achieving an improved survival rate and a higher quality of life for patients with locally advanced esophageal cancer The comprehensive analysis of the tumor microenvironment and the morphological and metabolic profiles should provide new insights into the mechanisms of tumor progression and resistance which in turn will form the basis for future translational research and treatment approaches The findings from this study have the potential to change the way esophageal cancer is treated by contributing to the development of stratified therapeutic approaches tailored to the molecular subtype of esophageal cancer
Detailed Description:
Introduction Esophageal cancer is one of the most common types of cancer worldwide However the incidence of esophageal cancer varies significantly by region with higher rates observed in some countries such as Japan and South Korea 1 Tumors detected early have a better prognosis than advanced tumors The 5-year survival rates for patients with esophageal cancer vary depending on the stage from 90 for localized tumours to only 5 in advanced stages T2 N whereby patient always refers to both genders As the majority of patients are mainly diagnosed at a locally advanced stage multimodal treatment is currently recommended in most cases before curative resection 2 The success of the systemic therapy which in Germany is usually carried out as chemotherapy alone with the FLOT protocol or as radiochemotherapy with the CROSS protocol is assessed postoperatively in the final histological evaluation by means of the degree of regression According to Becker the resulting degrees of regression are graded into stages 1 to 3 with grade 1 reflecting almost complete regression after systemic therapy and grade 3 indicating a postoperative residual tumor with 50 vital tumor cells and thus a poor response to neoadjuvant therapy 3 4
The reason for favorable versus a poor response in esophageal adenocarcinoma cannot lie solely in the patients clinical characteristics and genetic subtype Rather the metabolic phenotype plays a decisive role However the metabolic phenotype of esophageal adenocarcinomas with different regression grade has not yet been sufficiently researched
Rationale The oncological outcome of the esophageal adenocarcinoma depends on various factors including in particular tumor size tumor metabolism and the growth pattern of the primary tumor and infiltrated lymph nodes 5 An advanced tumor size is associated with an increased risk of local and systemic metastases and a poorer prognosis At the same time tumor size is associated with altered metabolic activity particularly glycolysis 6 Tumor biology which can be mapped using metabolomics also plays an important role in the prognosis and treatment of these cancers The expression of biomarkers such as HER2 and PD-L1 can also be of decisive importance both in the choice of therapy and in predicting the response to a particular treatment 7 However locoregional growth response to chemotherapy and tumor size also play a significant role in patient survival 8
The tumor microenvironemt TME is frequently observed in esophageal carcinoma to induce cell proliferation and thus hardens the surrounding tissue and restricts blood flow to the tumor which in turn reduces the effectiveness of chemotherapy and radiotherapy 9 Another key feature of tumor cell metabolism is the ability to extract nutrients from a frequently nutrient-poor microenvironment and use these nutrients to meet the demands of growth and proliferation 10 In this way the TME gradually forms encompassing cancer cells the cytokine environment the extracellular matrix subgroups of fibroblasts and immune cells 11 Within this complex network of cancer cells and cancer-related metabolic products the pro-tumorigenic environment plays a central role in stimulating tumor angiogenesis and promoting its invasiveness and metastatic potential 12 Certain tumor-associated metabolites are linked to aggressive cancer phenotypes facilitated angiogenesis promoted mutagenesis and suppression of the immune system 13 However the migration of cancer-related metabolites from the tumor itself and its microenvironment into the systemic bloodstream can be detected and quantified using state-of-the-art clinical mass spectrometry 14 Therefore abnormal amino acid and phospholipid metabolism in various cancers plays a diverse and crucial role and the potential impact of metabolic control and regulation within the tumor microenvironment is gaining increasing importance 15
While metabolomics is already implemented in basic research early detection diagnosis and therapy for metabolic diseases such as type 2 diabetes mellitus there is currently a high demand for the application of targeted metabolomics in oncological diseases In various entities it has been observed that the metabolome within the TME harbors a significant alteration in amino acid and phospholipid metabolism 16 From a metabolic perspective current investigations focus particularly on an increase in membrane-bound polar phospholipids and a decrease in apolar phospholipids and phosphateidylserines to characterize for example the response to chemotherapy 17
In addition to this quantitative evaluation via targeted metabolomics of local metabolism in tumor tissue the TME and systemically in blood plasma methods of mass spectrometry imaging allows for a qualitative assessment of tumor samples Matrix-Assisted Laser DesorptionIonization MALDI is a method for ionizing samples where a matrix usually a small organic acid is mixed with the sample The matrix-protein complex is then irradiated with a laser leading to desorption and ionization of the molecules 18 In the case of esophageal carcinoma MALDI-MSI is used to study tumor heterogeneity and visualize specific areas with high abundance of oncometabolites 19 20 The application of MALDI-MSI in other oncologic entities such as the pancreatic adenocarcinoma PDAC has provided specific insights into lipid metabolic changes associated with tumor growth and progression For example it has been found that certain lipids such as phospholipids and sphingolipids play a significant role in tumor biology 21 These changes in lipid metabolism can promote cancer cell proliferation and survival 22 These findings open new possibilities for the development of targeted therapies aimed at the altered lipid metabolism For instance areas of PDAC with higher proliferative and metabolic activity have been identified using glycerophosphocholine 23 Additionally it was successfully observed that the glutamineglutamate metabolism in tumor samples is significantly downregulated indicating a fundamental reprogramming of cellular metabolism which can be adequately represented using MALDI-MSI 24 Consequently in blood and tissue samples from patients specifically diagnosed with esophageal carcinoma functional histological examinations are conducted in addition to metabolomic studies to depict oncometabolites in tumor tissue and the tumor microenvironment 24
The response to neoadjuvant therapy in the era of approaches such as NEOFLOTS is more of a standard than a rarity but to this day no pre-therapeutic analysis for patient identification serves as its foundation 25 In terms of surgery in the locally advanced setting which can lead significantly more patients to resection this appears to be of great relevance 26 Current studies particularly the correlation between the tumors pattern of spread and its metabolism show a clear association between these two characteristics and oncological outcome parameters 27 With sufficient knowledge of patient-specific tumor characteristics it is possible to identify patients who for example would not benefit from systemic therapy due to their tumor metabolome and should instead be directed to resection as quickly as possible 28
Modern medicine inevitably follows a multimodal therapeutic approach It is of great importance to individualize the final therapy decision as much as possible for each patient 29 This is only possible by considering all potential tumorigenic aspects The underlying study aims to bridge the gap between newer approaches such as metabolism and the pattern of tumor heterogeneity and spread 30 Up until now the metabolomic properties of these tumors have barely been incorporated into diagnosis and therapy Moreover the TME in the upper gastrointestinal tract remains largely unexplored However studies in lung carcinoma have already shown changes in chemotherapy response depending on the TME 31 A joint investigation of the metabolome cellular morphology in hematoxylin-eosin HE histological sections and molecular constellation using MALDI-MSI as well as an investigation of correlations between these two aspects has not yet been conducted in esophageal carcinoma 32 33
Hypothesis We hypothesize that I adenocarcinoma of the esophagus with regression grade 1 encompasses a fundamentally distinct metabolic profile than adenocarcinoma of the esophagus with regression grade 3 II Consequently a stratification parameter within the local tumor metabolism and the tumor microenvironment exists which correlates with the systemic response to neoadjuvant chemotherapy in blood plasma
Objectives and eligibility criteria Primary objective The primary objective is to evaluate phospholipids in three compartments as a new stratification parameter for patients with adenocarcinoma of the esophagus with different responses to chemotherapy after four cycles of neoadjuvant FLOT chemotherapy followed by surgery for locally advanced esophageal cancer
Primary endpoint The primary endpoint for beneficial response to neoadjuvant chemotherapy is defined by the regressions grad according to Becker The primary endpoint for phospholipids as a new parameter for response to chemotherapy is the increase of phosphatidylcholines in the local tumor tissue in the adjacent normal tissue and in blood samples
Secondary objectives To determine a metabolic profile which shows a significant correlation between patients with regression grade 1 after neoadjuvant chemotherapy compared to regression grade 3
To evaluate the expression and distribution of the identified oncometabolites in tumor tissue and adjacent tissue
Secondary endpoints The metabolic profile will be carried out by LC-MSMS following multivariate analysis The expression and distribution of oncometabolites will be evaluated by MALDI - MSI
Study design This is a retrospective observative open label study of the correlation between phospholipid metabolism and the regressions grade in study participants with locally advanced esophageal cancer after four cycles of neoadjuvant chemotherapy according to the FLOT protocol
Approximately 20 study participants with newly diagnosed adenocarcinoma of the esophagus with a tumor stage of uT2 uN cM0 or uT3 cNx cM0 will be first treated with FLOT following open or minimal-invasive esophagectomy Baseline medical imaging with CTMRI according to standard clinical practice is performed within 28 days before neoadjuvant treatment start First tumor assessment after neoadjuvant treatment with CT scanMRI is scheduled within two weeks before surgery A second tumor assessment is planned within two weeks before adjuvant treatment start Thereafter every 12 weeks - 2 weeks up to end of treatment visit In the period of follow up a CTMRI will be performed every 12 weeks - 2 weeks for a time of 24 months
Study population The study population consists of patients who underwent surgery for stomach or esophageal tumors between January 1 2018 and January 31 2024 at the Department of General Visceral and Transplantation Surgery One prerequisite is the availability of fresh-frozen tumor tissue in the archive of the Department of General Visceral and Transplantation Surgery as well as fresh-frozen tumor tissue from the institutions biobank sample material request has been approved and a complete clinical dataset with a five-year follow-up for each patient The final cohort consists of patients for whom both the frozen biobank specimen potentially with a serum sample and the paraffin-embedded tumor tissue are available
Inclusion criteria Signed Written Informed Consent Study participants must have signed and dated an IEC approved written informed consent form in accordance with regulatory and institutional guidelines
Study participants must be willing and able to comply with scheduled visits treatment schedule laboratory tests and other requirements of the study
Target Population
Histologically confirmed resectable adenocarcinoma of the esophagus uT2 uN cM0 or uT3 cNx cM0 with the following specifications
Medical and technical operability No preceding cytotoxic or targeted therapy No prior partial or complete tumor resection Male or female patients 18 years of age at time of study entry Eastern Cooperative Oncology Group ECOG Performance Status of 0-1 Life expectancy of at least 12 months Adequate normal organ function as defined below Screening laboratory values must meet the following criteria and should be obtained within 28 days prior to registration WBC 1500μL Neutrophils 1000μL Platelets 75 x103μL Hemoglobin 90 gdL Serum creatinine 15 x institutional ULN or calculated creatinine clearance 40 mLmin Cockcroft-Gault ASTALT 25 x institutional ULN Total Bilirubin 15 x institutional ULN Body weight 30kg Reproductive Status Women of childbearing potential must have a negative serum or urine pregnancy test within one until two weeks prior to the start of neoadjuvant treatment and after surgery before starting adjuvant treatment
Women 50 years of age would be considered post-menopausal if they have been amenorrheic for 12 months or more following cessation of all exogenous hormonal treatments had radiation-induced menopause with last menses 1 year ago had chemotherapy-induced menopause with last menses 1 year ago
Women must not be breastfeeding Men who are sexually active with WOCBP must use any contraceptive method with a failure rate of less than 1 per year
Exclusion criteria
Study participants with squamous cell carcinoma of the esophagus Prior treatment with chemotherapy targeted therapy or radiotherapy for treatment of advanced cancer disease less than 5 years
Enrollment is possible for patients with
Adequately treated non-melanoma skin cancer or lentigo maligna without evidence of disease Adequately treated carcinoma in situ without evidence of disease Any other serious or uncontrolled medical disorder active infections physical exam findings laboratory finding altered mental status or psychiatric condition that in the opinion of the investigator would limit a study participants ability to comply with the study requirements substantially increase risk to the study participant or impact the interpretability or study results
Active or prior documented autoimmune or inflammatory disorders The following are exceptions to this criterion
Patients with vitiligo or alopecia Patients with hypothyroidism stable on hormone replacement Any chronic skin condition that does not require systemic therapy Patients with celiac disease controlled by diet alone Inhaled or topical steroids and adrenal replacement steroid doses 10mg daily prednisone equivalent are permitted in the absence of active autoimmune disease
History of active primary immunodeficiency History of any allogenic organ transplantation with currently intake of immune suppressive treatment Patients with interstitial lung disease that is symptomatic or may interfere with the detection or management of suspected drug-related pulmonary toxicity FEV 1 75 Patients has known current symptomatic congestive heart failure unstable angina pectoris or cardiac arrhythmia Receipt of live attenuated vaccine within 30 days prior to the first dose of IP Note Patients if enrolled should not receive live vaccine whilst receiving IP and up to 30 days after the last dose of IP
Prisoners or study participants who are involuntarily incarcerated Pregnancy or breastfeeding females Female patients who are pregnant or breastfeeding or male or female patients of reproductive potential who are not willing to employ effective birth control from screening to 90 days after the last dose of durvalumab monotherapy or 180 days after the last dose of durvalumab tremelimumab combination therapy
Flow-Chart and Events Schedule A study design has been established for the conduct of the investigation The chromatographic mass spectrometric and immunohistochemical methods employed have already been validated and compliance with Good Clinical Practice GCP regarding data management is ensured through appropriate standard operating procedures SOPs Since this is a retrospective data collection the samples have already been stored in the in-house biobank at -80C at the time of the application Basic patient data are already stored in existing databases at our institution All patients underwent surgery in the Clinic and Polyclinic for General Visceral and Transplant Surgery at the LMU Munich Hospital and were informed through the register of the Study Documentation and Quality Center StuDoQ of the German Society for General and Visceral Surgery DGAV The sample collection has already been conducted by the in-house biobank Our patients have already consented to the collection and scientific utilization of samples during the initial sampling perioperatively Upon retrieval these samples are pre-processed in the laboratory for experimental surgery in preparation for metabolomic measurements The pre-processing includes the careful thawing of the samples over a period of 24 hours as well as the aliquoting of the samples in the surgical laboratory Subsequently the samples are transported to the location of the in-house MS core facility until measurement
Clinical characteristics At baseline a medical history have been obtained The baseline examinations include weight to be measured at the study site height ECOG Performance Status measured blood pressure BP heart rate HR temperature and oxygen saturation by pulse oximetry at rest within 28 days prior to first cycle of neoadjuvant FLOT chemotherapy Baseline signs and symptoms are those that are assessed within 14 days prior to first dose Concomitant medication were collected from within 14 days prior to first cycle
Baseline local laboratory assessments have been done within 14 days prior to first cycle and include complete blood count with differential liver function test ALT AST total bilirubin and AP serum urea level uric acid creatinine with creatinine clearance phosphate glucose serum albumin C-reactive protein amylase lipase TSH free T3 and free T4 tumor marker CEA and CA 19-9 Physical examinations on study weight ECOG performance status vital signs and oxygen saturation have been assessed prior to neoadjuvant chemotherapy Additional measures including non-study required laboratory tests should be performed as clinically indicated or to comply with local regulations Additional testing or assessments may be performed as clinically necessary or where required by institutional or local regulations
Screening or baseline assessments are to be performed within 28 days prior to treatment start External CT scansMRI performed before enrolment in the trial is feasible as baseline scan if performed no longer than 28 prior to first dose of study drug administration In addition to the neck chest abdomen and pelvis all known sites of disease should be assessed at baseline Subsequent assessments should include neck chest abdomen using the same imaging method and technique as used at baseline Disease assessment with contrast-enhanced computed tomography CT scans acquired on dedicated CT equipment is preferred for this study Conventional CT with IV contrast and MRI gadolinium should be performed with contiguous cuts of 10 mm or less slice thickness Spiral CT should be performed using a 3- or 5-mm contiguous reconstruction Should study participant have contraindication for CT MRI may be performed In cases application of CT IV contrast is contra-indicated tumor assessment should preferably be performed using a non-contrast CT scan of the chest and a contrast-enhanced MRI of the neck and abdomen
Tumor measurements should be made by the same investigator or radiologist for each assessment whenever possible Change in tumor measurements and tumor response will be assessed by the Investigator using the RECIST 11 criteria Bone scan PET scan and ultrasound is not adequate for assessment of RECIST 11 response in target lesions Complementary CT andor MRI or biopsy must be performed in such cases
Doubtful cutaneous subcutaneous or lymph node lesions that are suspected should be confirmed by biopsy Histological or cytological evidence of recurrence should be attempted in all cases except for brain metastases when safe and clinically feasible An example when obtaining a biopsy to confirm recurrence may not be safe and clinically feasible is brain metastases
Biomarker Assessment As the neoadjuvant setting offers the unique opportunity to achieve large tumor samples from patients after multimodal treatment the study includes a comprehensive analysis of immune responses to private and shared antigens These analyses rely on standardized collection of tissue and blood samples which allow the planned metabolomic massspectromeric and functional immunologic analyses
Unfixed tumor specimens will be taken to our Institute of Pathology immediately following resection Tumor tissue and normal tissue will be processed according to local diagnostic standards of procedure In case of sufficient material biopsies of tumor and normal tissue can be taken and stored in liquid nitrogen As determined by the pathologist additional fresh tumor tissue which is not needed for pathological staging will be transferred to the biobank These samples will be analyzed by immunohistochemistry and molecular analyses respectively
Blood 45mL EDTA plasma 20mL and serum 45mL samples have been obtained at screening and have been immediately transferred to the biobank
Targeted metabolomics The LC-MSMS method was validated based on the current guidelines for bioanalytical method validation from the European Medicines Agency 2011 and the US Food and Drug Administration 2018 For LC-MSMS measurements an Agilent 1290 HPLC coupled with a QTRAP 6500 MS system is used Data acquisition and quantification are performed using Analyst 163 The analytical column is an Acquity UPLC Premier HSS T3 18 μm 21x50mm and offline solid-phase extraction SPE is carried out using SepPak tC18 100 mg 96-well plates Methanol and water of MS grade are obtained from VWR International GmbH Formic acid ammonium acetate and acetic acid are sourced from Thermo Fisher Scientific Inc The extracted metabolites are eluted into a collection plate in two consecutive steps with 2 300 μl of methanol After complete solvent evaporation at 50C under a gentle nitrogen stream the samples are reconstituted in 150 μl of methanol and diluted with 150 μl of water A 10 μl aliquot of the extracted sample is injected into the HPLC system Samples with concentrations exceeding the upper limit of quantification are reanalyzed after incubation Each analysis run is preceded by a one-minute automatic re-equilibration For each analyte a quantifier and a qualifier transition are identified and optimized for maximum intensity Isobaric compounds are baseline-separated from the analyte peaks
The Biocrates MxP Quant 500 XL Kit Biocrates Life Sciences AG Innsbruck Austria is used for the measurements allowing the quantification of 106 small molecules in chromatography mode and 524 lipids in flow injection mode FIA-MSMS thereby covering the major metabolic pathways The kit includes a patented 96-well filter plate with pre-set internal standards calibration standards and quality controls as well as a test sample for system suitability testing The qualitatively and quantitatively obtained metabolites are exported as an Excel file and further processed in the surgical laboratory
Mass spectrometry imaging The tumor tissue and adjacent normal tissue samples are dissolved in a matrix and applied to a MALDI plate The samples are irradiated with a UV laser causing the molecules to become ionized The ionized molecules are then accelerated in an electric field and their flight time is measured to determine the mz ratio Imaging experiments are performed using a rapifleX MALDI Tissuetyper MALDI-TOFTOF mass spectrometer Bruker Daltonik GmbH equipped with a SmartBeam 3G laser Tissues are measured in positive reflector mode with a spatial resolution of 25 μm and a mass range of 600-3200 Da Each measurement is externally precalibrated using a commercial peptide calibration mixture on the same target as the tissues at multiple positions Tissues from all patients are measured on the rapifleX with a sampling rate of 125 GSs and a pulsed ion extraction of 160 ns After the MALDI-MSI measurements the HCCA matrix is removed by washing the slide in 70 ethanol followed by HE counterstaining High-resolution images of the stained sections were acquired using the Mirax Scan system Carl Zeiss MicroImaging and co-registered with the MALDI-IMS data for histological correlation
Data Protection The provisions of data protection legislation will be observed It is assured by the sponsor that all investigational materials and data will be irreversibly anonymized in accordance with data protection legislation before scientific processing Trial study participants will be informed that their anonymized data will be passed on in accordance with provisions for documentation and notification pursuant to 12 and 13 of the GCP Regulations to the recipients described there Study participants who do not agree that information may be passed on in this way will not be enrolled into the trial
References
1 Puhr HC et al Recent Advances in the Systemic Treatment of Localized Gastroesophageal Cancer Cancers Basel 2023 156
2 Wang Z et al Incidence and prognosis of gastroesophageal cancer in rural urban and metropolitan areas of the United States Cancer 2013 11922 p 4020-7
3 Otsuka R et al ypTNM staging is a potentially useful prognostic stratification tool in patients with advanced gastric cancer after preoperative chemotherapy Langenbecks Arch Surg 2023 4081 p 133
4 Becker K et al Histomorphology and grading of regression in gastric carcinoma treated with neoadjuvant chemotherapy Cancer 2003 987 p 1521-30
5 Schmidt DR et al Metabolomics in cancer research and emerging applications in clinical oncology CA Cancer J Clin 2021 714 p 333-358
6 Zhang J et al Esophageal cancer metabolite biomarkers detected by LC-MS and NMR methods PLoS One 2012 71 p e30181
7 Derks S et al Epithelial PD-L2 Expression Marks Barretts Esophagus and Esophageal Adenocarcinoma Cancer Immunol Res 2015 310 p 1123-1129
8 Peng X et al Molecular Characterization and Clinical Relevance of Metabolic Expression Subtypes in Human Cancers Cell Rep 2018 231 p 255-269 e4
9 Li W et al Molecular alterations of cancer cell and tumour microenvironment in metastatic gastric cancer Oncogene 2018 3736 p 4903-4920
10 King R et al Hypoxia and its impact on the tumour microenvironment of gastroesophageal cancers World J Gastrointest Oncol 2021 135 p 312-331
11 Dickerson LK et al Emerging interleukin targets in the tumour microenvironment implications for the treatment of gastrointestinal tumours Gut 2023 728 p 1592-1606
12 Meier A et al Hypothesis-free deep survival learning applied to the tumour microenvironment in gastric cancer J Pathol Clin Res 2020 64 p 273-282
13 Liu J et al The combined signatures of the tumour microenvironment and nucleotide metabolism-related genes provide a prognostic and therapeutic biomarker for gastric cancer Sci Rep 2023 131 p 6622
14 Pun C et al Prognostic Significance of Tumour Budding and Desmoplastic Reaction in Intestinal-Type Gastric Adenocarcinoma Int J Surg Pathol 2023 316 p 957-966
15 Pardoll DM Immunology beats cancer a blueprint for successful translation Nat Immunol 2012 1312 p 1129-32
16 Hatogai K et al Comprehensive immunohistochemical analysis of tumor microenvironment immune status in esophageal squamous cell carcinoma Oncotarget 2016 730 p 47252-47264
17 Wang Y et al Tissue-based metabolomics reveals metabolic signatures and major metabolic pathways of gastric cancer with help of transcriptomic data from TCGA Biosci Rep 2021 4110
18 Balluff B et al Direct molecular tissue analysis by MALDI imaging mass spectrometry in the field of gastrointestinal disease Gastroenterology 2012 1433 p 544-549 e2
19 Streitz JM Jr et al Analysis of protein expression patterns in Barretts esophagus using MALDI mass spectrometry in search of malignancy biomarkers Dis Esophagus 2005 183 p 170-6
20 Erich K et al Spatial Distribution of Endogenous Tissue Protease Activity in Gastric Carcinoma Mapped by MALDI Mass Spectrometry Imaging Mol Cell Proteomics 2019 181 p 151-161
21 Gaye MM et al Delineation of disease phenotypes associated with esophageal adenocarcinoma by MALDI-IMS-MS analysis of serum N-linked glycans Analyst 2017 1429 p 1525-1535
22 Kunzke T et al Native glycan fragments detected by MALDI-FT-ICR mass spectrometry imaging impact gastric cancer biology and patient outcome Oncotarget 2017 840 p 68012-68025
23 Jia K et al Novel circulating peptide biomarkers for esophageal squamous cell carcinoma revealed by a magnetic bead-based MALDI-TOFMS assay Oncotarget 2016 717 p 23569-80
24 Alexandrov LB et al Signatures of mutational processes in human cancer Nature 2013 5007463 p 415-21
25 Lorenzen S et al Feasibility of perioperative chemotherapy with infusional 5-FU leucovorin and oxaliplatin with FLOT or without FLO docetaxel in elderly patients with locally advanced esophagogastric cancer Br J Cancer 2013 1083 p 519-26
26 Gotze TO et al Preventive HIPEC in combination with perioperative FLOT versus FLOT alone for resectable diffuse type gastric and gastroesophageal junction type IIIII adenocarcinoma - the phase III PREVENT- FLOT9 trial of the AIO CAOGI ACO BMC Cancer 2021 211 p 1158
27 Lombardi PM et al Facing adenocarcinoma of distal esophagus and esophagogastric junction a CROSS versus FLOT propensity score-matched analysis of oncological outcomes in a high-volume institution Updates Surg 2023 754 p 921-930
28 Hingorani M et al Neoadjuvant Management of Adenocarcinoma of the Esophagus and Esophagogastric Junction Review of Randomized Evidence and Definition of Optimum Treatment Algorithm Oncology 2023 1019 p 553-564
29 Kudo T et al Nivolumab treatment for oesophageal squamous-cell carcinoma an open-label multicentre phase 2 trial Lancet Oncol 2017 185 p 631-639
30 Schwacke J et al Discrimination of normal and esophageal cancer plasma proteomes by MALDI-TOF mass spectrometry Dig Dis Sci 2015 606 p 1645-54
31 Berghmans E et al MALDI Mass Spectrometry Imaging Linked with Top-Down Proteomics as a Tool to Study the Non-Small-Cell Lung Cancer Tumor Microenvironment Methods Protoc 2019 22
32 Low DE et al Benchmarking Complications Associated with Esophagectomy Ann Surg 2019 2692 p 291-298
33 Schmidt HM et al Defining Benchmarks for Transthoracic Esophagectomy A Multicenter Analysis of Total Minimally Invasive Esophagectomy in Low Risk Patients Ann Surg 2017 2665 p 814-821
The reason for favorable versus a poor response in esophageal adenocarcinoma cannot lie solely in the patients clinical characteristics and genetic subtype Rather the metabolic phenotype plays a decisive role However the metabolic phenotype of esophageal adenocarcinomas with different regression grade has not yet been sufficiently researched
Rationale The oncological outcome of the esophageal adenocarcinoma depends on various factors including in particular tumor size tumor metabolism and the growth pattern of the primary tumor and infiltrated lymph nodes 5 An advanced tumor size is associated with an increased risk of local and systemic metastases and a poorer prognosis At the same time tumor size is associated with altered metabolic activity particularly glycolysis 6 Tumor biology which can be mapped using metabolomics also plays an important role in the prognosis and treatment of these cancers The expression of biomarkers such as HER2 and PD-L1 can also be of decisive importance both in the choice of therapy and in predicting the response to a particular treatment 7 However locoregional growth response to chemotherapy and tumor size also play a significant role in patient survival 8
The tumor microenvironemt TME is frequently observed in esophageal carcinoma to induce cell proliferation and thus hardens the surrounding tissue and restricts blood flow to the tumor which in turn reduces the effectiveness of chemotherapy and radiotherapy 9 Another key feature of tumor cell metabolism is the ability to extract nutrients from a frequently nutrient-poor microenvironment and use these nutrients to meet the demands of growth and proliferation 10 In this way the TME gradually forms encompassing cancer cells the cytokine environment the extracellular matrix subgroups of fibroblasts and immune cells 11 Within this complex network of cancer cells and cancer-related metabolic products the pro-tumorigenic environment plays a central role in stimulating tumor angiogenesis and promoting its invasiveness and metastatic potential 12 Certain tumor-associated metabolites are linked to aggressive cancer phenotypes facilitated angiogenesis promoted mutagenesis and suppression of the immune system 13 However the migration of cancer-related metabolites from the tumor itself and its microenvironment into the systemic bloodstream can be detected and quantified using state-of-the-art clinical mass spectrometry 14 Therefore abnormal amino acid and phospholipid metabolism in various cancers plays a diverse and crucial role and the potential impact of metabolic control and regulation within the tumor microenvironment is gaining increasing importance 15
While metabolomics is already implemented in basic research early detection diagnosis and therapy for metabolic diseases such as type 2 diabetes mellitus there is currently a high demand for the application of targeted metabolomics in oncological diseases In various entities it has been observed that the metabolome within the TME harbors a significant alteration in amino acid and phospholipid metabolism 16 From a metabolic perspective current investigations focus particularly on an increase in membrane-bound polar phospholipids and a decrease in apolar phospholipids and phosphateidylserines to characterize for example the response to chemotherapy 17
In addition to this quantitative evaluation via targeted metabolomics of local metabolism in tumor tissue the TME and systemically in blood plasma methods of mass spectrometry imaging allows for a qualitative assessment of tumor samples Matrix-Assisted Laser DesorptionIonization MALDI is a method for ionizing samples where a matrix usually a small organic acid is mixed with the sample The matrix-protein complex is then irradiated with a laser leading to desorption and ionization of the molecules 18 In the case of esophageal carcinoma MALDI-MSI is used to study tumor heterogeneity and visualize specific areas with high abundance of oncometabolites 19 20 The application of MALDI-MSI in other oncologic entities such as the pancreatic adenocarcinoma PDAC has provided specific insights into lipid metabolic changes associated with tumor growth and progression For example it has been found that certain lipids such as phospholipids and sphingolipids play a significant role in tumor biology 21 These changes in lipid metabolism can promote cancer cell proliferation and survival 22 These findings open new possibilities for the development of targeted therapies aimed at the altered lipid metabolism For instance areas of PDAC with higher proliferative and metabolic activity have been identified using glycerophosphocholine 23 Additionally it was successfully observed that the glutamineglutamate metabolism in tumor samples is significantly downregulated indicating a fundamental reprogramming of cellular metabolism which can be adequately represented using MALDI-MSI 24 Consequently in blood and tissue samples from patients specifically diagnosed with esophageal carcinoma functional histological examinations are conducted in addition to metabolomic studies to depict oncometabolites in tumor tissue and the tumor microenvironment 24
The response to neoadjuvant therapy in the era of approaches such as NEOFLOTS is more of a standard than a rarity but to this day no pre-therapeutic analysis for patient identification serves as its foundation 25 In terms of surgery in the locally advanced setting which can lead significantly more patients to resection this appears to be of great relevance 26 Current studies particularly the correlation between the tumors pattern of spread and its metabolism show a clear association between these two characteristics and oncological outcome parameters 27 With sufficient knowledge of patient-specific tumor characteristics it is possible to identify patients who for example would not benefit from systemic therapy due to their tumor metabolome and should instead be directed to resection as quickly as possible 28
Modern medicine inevitably follows a multimodal therapeutic approach It is of great importance to individualize the final therapy decision as much as possible for each patient 29 This is only possible by considering all potential tumorigenic aspects The underlying study aims to bridge the gap between newer approaches such as metabolism and the pattern of tumor heterogeneity and spread 30 Up until now the metabolomic properties of these tumors have barely been incorporated into diagnosis and therapy Moreover the TME in the upper gastrointestinal tract remains largely unexplored However studies in lung carcinoma have already shown changes in chemotherapy response depending on the TME 31 A joint investigation of the metabolome cellular morphology in hematoxylin-eosin HE histological sections and molecular constellation using MALDI-MSI as well as an investigation of correlations between these two aspects has not yet been conducted in esophageal carcinoma 32 33
Hypothesis We hypothesize that I adenocarcinoma of the esophagus with regression grade 1 encompasses a fundamentally distinct metabolic profile than adenocarcinoma of the esophagus with regression grade 3 II Consequently a stratification parameter within the local tumor metabolism and the tumor microenvironment exists which correlates with the systemic response to neoadjuvant chemotherapy in blood plasma
Objectives and eligibility criteria Primary objective The primary objective is to evaluate phospholipids in three compartments as a new stratification parameter for patients with adenocarcinoma of the esophagus with different responses to chemotherapy after four cycles of neoadjuvant FLOT chemotherapy followed by surgery for locally advanced esophageal cancer
Primary endpoint The primary endpoint for beneficial response to neoadjuvant chemotherapy is defined by the regressions grad according to Becker The primary endpoint for phospholipids as a new parameter for response to chemotherapy is the increase of phosphatidylcholines in the local tumor tissue in the adjacent normal tissue and in blood samples
Secondary objectives To determine a metabolic profile which shows a significant correlation between patients with regression grade 1 after neoadjuvant chemotherapy compared to regression grade 3
To evaluate the expression and distribution of the identified oncometabolites in tumor tissue and adjacent tissue
Secondary endpoints The metabolic profile will be carried out by LC-MSMS following multivariate analysis The expression and distribution of oncometabolites will be evaluated by MALDI - MSI
Study design This is a retrospective observative open label study of the correlation between phospholipid metabolism and the regressions grade in study participants with locally advanced esophageal cancer after four cycles of neoadjuvant chemotherapy according to the FLOT protocol
Approximately 20 study participants with newly diagnosed adenocarcinoma of the esophagus with a tumor stage of uT2 uN cM0 or uT3 cNx cM0 will be first treated with FLOT following open or minimal-invasive esophagectomy Baseline medical imaging with CTMRI according to standard clinical practice is performed within 28 days before neoadjuvant treatment start First tumor assessment after neoadjuvant treatment with CT scanMRI is scheduled within two weeks before surgery A second tumor assessment is planned within two weeks before adjuvant treatment start Thereafter every 12 weeks - 2 weeks up to end of treatment visit In the period of follow up a CTMRI will be performed every 12 weeks - 2 weeks for a time of 24 months
Study population The study population consists of patients who underwent surgery for stomach or esophageal tumors between January 1 2018 and January 31 2024 at the Department of General Visceral and Transplantation Surgery One prerequisite is the availability of fresh-frozen tumor tissue in the archive of the Department of General Visceral and Transplantation Surgery as well as fresh-frozen tumor tissue from the institutions biobank sample material request has been approved and a complete clinical dataset with a five-year follow-up for each patient The final cohort consists of patients for whom both the frozen biobank specimen potentially with a serum sample and the paraffin-embedded tumor tissue are available
Inclusion criteria Signed Written Informed Consent Study participants must have signed and dated an IEC approved written informed consent form in accordance with regulatory and institutional guidelines
Study participants must be willing and able to comply with scheduled visits treatment schedule laboratory tests and other requirements of the study
Target Population
Histologically confirmed resectable adenocarcinoma of the esophagus uT2 uN cM0 or uT3 cNx cM0 with the following specifications
Medical and technical operability No preceding cytotoxic or targeted therapy No prior partial or complete tumor resection Male or female patients 18 years of age at time of study entry Eastern Cooperative Oncology Group ECOG Performance Status of 0-1 Life expectancy of at least 12 months Adequate normal organ function as defined below Screening laboratory values must meet the following criteria and should be obtained within 28 days prior to registration WBC 1500μL Neutrophils 1000μL Platelets 75 x103μL Hemoglobin 90 gdL Serum creatinine 15 x institutional ULN or calculated creatinine clearance 40 mLmin Cockcroft-Gault ASTALT 25 x institutional ULN Total Bilirubin 15 x institutional ULN Body weight 30kg Reproductive Status Women of childbearing potential must have a negative serum or urine pregnancy test within one until two weeks prior to the start of neoadjuvant treatment and after surgery before starting adjuvant treatment
Women 50 years of age would be considered post-menopausal if they have been amenorrheic for 12 months or more following cessation of all exogenous hormonal treatments had radiation-induced menopause with last menses 1 year ago had chemotherapy-induced menopause with last menses 1 year ago
Women must not be breastfeeding Men who are sexually active with WOCBP must use any contraceptive method with a failure rate of less than 1 per year
Exclusion criteria
Study participants with squamous cell carcinoma of the esophagus Prior treatment with chemotherapy targeted therapy or radiotherapy for treatment of advanced cancer disease less than 5 years
Enrollment is possible for patients with
Adequately treated non-melanoma skin cancer or lentigo maligna without evidence of disease Adequately treated carcinoma in situ without evidence of disease Any other serious or uncontrolled medical disorder active infections physical exam findings laboratory finding altered mental status or psychiatric condition that in the opinion of the investigator would limit a study participants ability to comply with the study requirements substantially increase risk to the study participant or impact the interpretability or study results
Active or prior documented autoimmune or inflammatory disorders The following are exceptions to this criterion
Patients with vitiligo or alopecia Patients with hypothyroidism stable on hormone replacement Any chronic skin condition that does not require systemic therapy Patients with celiac disease controlled by diet alone Inhaled or topical steroids and adrenal replacement steroid doses 10mg daily prednisone equivalent are permitted in the absence of active autoimmune disease
History of active primary immunodeficiency History of any allogenic organ transplantation with currently intake of immune suppressive treatment Patients with interstitial lung disease that is symptomatic or may interfere with the detection or management of suspected drug-related pulmonary toxicity FEV 1 75 Patients has known current symptomatic congestive heart failure unstable angina pectoris or cardiac arrhythmia Receipt of live attenuated vaccine within 30 days prior to the first dose of IP Note Patients if enrolled should not receive live vaccine whilst receiving IP and up to 30 days after the last dose of IP
Prisoners or study participants who are involuntarily incarcerated Pregnancy or breastfeeding females Female patients who are pregnant or breastfeeding or male or female patients of reproductive potential who are not willing to employ effective birth control from screening to 90 days after the last dose of durvalumab monotherapy or 180 days after the last dose of durvalumab tremelimumab combination therapy
Flow-Chart and Events Schedule A study design has been established for the conduct of the investigation The chromatographic mass spectrometric and immunohistochemical methods employed have already been validated and compliance with Good Clinical Practice GCP regarding data management is ensured through appropriate standard operating procedures SOPs Since this is a retrospective data collection the samples have already been stored in the in-house biobank at -80C at the time of the application Basic patient data are already stored in existing databases at our institution All patients underwent surgery in the Clinic and Polyclinic for General Visceral and Transplant Surgery at the LMU Munich Hospital and were informed through the register of the Study Documentation and Quality Center StuDoQ of the German Society for General and Visceral Surgery DGAV The sample collection has already been conducted by the in-house biobank Our patients have already consented to the collection and scientific utilization of samples during the initial sampling perioperatively Upon retrieval these samples are pre-processed in the laboratory for experimental surgery in preparation for metabolomic measurements The pre-processing includes the careful thawing of the samples over a period of 24 hours as well as the aliquoting of the samples in the surgical laboratory Subsequently the samples are transported to the location of the in-house MS core facility until measurement
Clinical characteristics At baseline a medical history have been obtained The baseline examinations include weight to be measured at the study site height ECOG Performance Status measured blood pressure BP heart rate HR temperature and oxygen saturation by pulse oximetry at rest within 28 days prior to first cycle of neoadjuvant FLOT chemotherapy Baseline signs and symptoms are those that are assessed within 14 days prior to first dose Concomitant medication were collected from within 14 days prior to first cycle
Baseline local laboratory assessments have been done within 14 days prior to first cycle and include complete blood count with differential liver function test ALT AST total bilirubin and AP serum urea level uric acid creatinine with creatinine clearance phosphate glucose serum albumin C-reactive protein amylase lipase TSH free T3 and free T4 tumor marker CEA and CA 19-9 Physical examinations on study weight ECOG performance status vital signs and oxygen saturation have been assessed prior to neoadjuvant chemotherapy Additional measures including non-study required laboratory tests should be performed as clinically indicated or to comply with local regulations Additional testing or assessments may be performed as clinically necessary or where required by institutional or local regulations
Screening or baseline assessments are to be performed within 28 days prior to treatment start External CT scansMRI performed before enrolment in the trial is feasible as baseline scan if performed no longer than 28 prior to first dose of study drug administration In addition to the neck chest abdomen and pelvis all known sites of disease should be assessed at baseline Subsequent assessments should include neck chest abdomen using the same imaging method and technique as used at baseline Disease assessment with contrast-enhanced computed tomography CT scans acquired on dedicated CT equipment is preferred for this study Conventional CT with IV contrast and MRI gadolinium should be performed with contiguous cuts of 10 mm or less slice thickness Spiral CT should be performed using a 3- or 5-mm contiguous reconstruction Should study participant have contraindication for CT MRI may be performed In cases application of CT IV contrast is contra-indicated tumor assessment should preferably be performed using a non-contrast CT scan of the chest and a contrast-enhanced MRI of the neck and abdomen
Tumor measurements should be made by the same investigator or radiologist for each assessment whenever possible Change in tumor measurements and tumor response will be assessed by the Investigator using the RECIST 11 criteria Bone scan PET scan and ultrasound is not adequate for assessment of RECIST 11 response in target lesions Complementary CT andor MRI or biopsy must be performed in such cases
Doubtful cutaneous subcutaneous or lymph node lesions that are suspected should be confirmed by biopsy Histological or cytological evidence of recurrence should be attempted in all cases except for brain metastases when safe and clinically feasible An example when obtaining a biopsy to confirm recurrence may not be safe and clinically feasible is brain metastases
Biomarker Assessment As the neoadjuvant setting offers the unique opportunity to achieve large tumor samples from patients after multimodal treatment the study includes a comprehensive analysis of immune responses to private and shared antigens These analyses rely on standardized collection of tissue and blood samples which allow the planned metabolomic massspectromeric and functional immunologic analyses
Unfixed tumor specimens will be taken to our Institute of Pathology immediately following resection Tumor tissue and normal tissue will be processed according to local diagnostic standards of procedure In case of sufficient material biopsies of tumor and normal tissue can be taken and stored in liquid nitrogen As determined by the pathologist additional fresh tumor tissue which is not needed for pathological staging will be transferred to the biobank These samples will be analyzed by immunohistochemistry and molecular analyses respectively
Blood 45mL EDTA plasma 20mL and serum 45mL samples have been obtained at screening and have been immediately transferred to the biobank
Targeted metabolomics The LC-MSMS method was validated based on the current guidelines for bioanalytical method validation from the European Medicines Agency 2011 and the US Food and Drug Administration 2018 For LC-MSMS measurements an Agilent 1290 HPLC coupled with a QTRAP 6500 MS system is used Data acquisition and quantification are performed using Analyst 163 The analytical column is an Acquity UPLC Premier HSS T3 18 μm 21x50mm and offline solid-phase extraction SPE is carried out using SepPak tC18 100 mg 96-well plates Methanol and water of MS grade are obtained from VWR International GmbH Formic acid ammonium acetate and acetic acid are sourced from Thermo Fisher Scientific Inc The extracted metabolites are eluted into a collection plate in two consecutive steps with 2 300 μl of methanol After complete solvent evaporation at 50C under a gentle nitrogen stream the samples are reconstituted in 150 μl of methanol and diluted with 150 μl of water A 10 μl aliquot of the extracted sample is injected into the HPLC system Samples with concentrations exceeding the upper limit of quantification are reanalyzed after incubation Each analysis run is preceded by a one-minute automatic re-equilibration For each analyte a quantifier and a qualifier transition are identified and optimized for maximum intensity Isobaric compounds are baseline-separated from the analyte peaks
The Biocrates MxP Quant 500 XL Kit Biocrates Life Sciences AG Innsbruck Austria is used for the measurements allowing the quantification of 106 small molecules in chromatography mode and 524 lipids in flow injection mode FIA-MSMS thereby covering the major metabolic pathways The kit includes a patented 96-well filter plate with pre-set internal standards calibration standards and quality controls as well as a test sample for system suitability testing The qualitatively and quantitatively obtained metabolites are exported as an Excel file and further processed in the surgical laboratory
Mass spectrometry imaging The tumor tissue and adjacent normal tissue samples are dissolved in a matrix and applied to a MALDI plate The samples are irradiated with a UV laser causing the molecules to become ionized The ionized molecules are then accelerated in an electric field and their flight time is measured to determine the mz ratio Imaging experiments are performed using a rapifleX MALDI Tissuetyper MALDI-TOFTOF mass spectrometer Bruker Daltonik GmbH equipped with a SmartBeam 3G laser Tissues are measured in positive reflector mode with a spatial resolution of 25 μm and a mass range of 600-3200 Da Each measurement is externally precalibrated using a commercial peptide calibration mixture on the same target as the tissues at multiple positions Tissues from all patients are measured on the rapifleX with a sampling rate of 125 GSs and a pulsed ion extraction of 160 ns After the MALDI-MSI measurements the HCCA matrix is removed by washing the slide in 70 ethanol followed by HE counterstaining High-resolution images of the stained sections were acquired using the Mirax Scan system Carl Zeiss MicroImaging and co-registered with the MALDI-IMS data for histological correlation
Data Protection The provisions of data protection legislation will be observed It is assured by the sponsor that all investigational materials and data will be irreversibly anonymized in accordance with data protection legislation before scientific processing Trial study participants will be informed that their anonymized data will be passed on in accordance with provisions for documentation and notification pursuant to 12 and 13 of the GCP Regulations to the recipients described there Study participants who do not agree that information may be passed on in this way will not be enrolled into the trial
References
1 Puhr HC et al Recent Advances in the Systemic Treatment of Localized Gastroesophageal Cancer Cancers Basel 2023 156
2 Wang Z et al Incidence and prognosis of gastroesophageal cancer in rural urban and metropolitan areas of the United States Cancer 2013 11922 p 4020-7
3 Otsuka R et al ypTNM staging is a potentially useful prognostic stratification tool in patients with advanced gastric cancer after preoperative chemotherapy Langenbecks Arch Surg 2023 4081 p 133
4 Becker K et al Histomorphology and grading of regression in gastric carcinoma treated with neoadjuvant chemotherapy Cancer 2003 987 p 1521-30
5 Schmidt DR et al Metabolomics in cancer research and emerging applications in clinical oncology CA Cancer J Clin 2021 714 p 333-358
6 Zhang J et al Esophageal cancer metabolite biomarkers detected by LC-MS and NMR methods PLoS One 2012 71 p e30181
7 Derks S et al Epithelial PD-L2 Expression Marks Barretts Esophagus and Esophageal Adenocarcinoma Cancer Immunol Res 2015 310 p 1123-1129
8 Peng X et al Molecular Characterization and Clinical Relevance of Metabolic Expression Subtypes in Human Cancers Cell Rep 2018 231 p 255-269 e4
9 Li W et al Molecular alterations of cancer cell and tumour microenvironment in metastatic gastric cancer Oncogene 2018 3736 p 4903-4920
10 King R et al Hypoxia and its impact on the tumour microenvironment of gastroesophageal cancers World J Gastrointest Oncol 2021 135 p 312-331
11 Dickerson LK et al Emerging interleukin targets in the tumour microenvironment implications for the treatment of gastrointestinal tumours Gut 2023 728 p 1592-1606
12 Meier A et al Hypothesis-free deep survival learning applied to the tumour microenvironment in gastric cancer J Pathol Clin Res 2020 64 p 273-282
13 Liu J et al The combined signatures of the tumour microenvironment and nucleotide metabolism-related genes provide a prognostic and therapeutic biomarker for gastric cancer Sci Rep 2023 131 p 6622
14 Pun C et al Prognostic Significance of Tumour Budding and Desmoplastic Reaction in Intestinal-Type Gastric Adenocarcinoma Int J Surg Pathol 2023 316 p 957-966
15 Pardoll DM Immunology beats cancer a blueprint for successful translation Nat Immunol 2012 1312 p 1129-32
16 Hatogai K et al Comprehensive immunohistochemical analysis of tumor microenvironment immune status in esophageal squamous cell carcinoma Oncotarget 2016 730 p 47252-47264
17 Wang Y et al Tissue-based metabolomics reveals metabolic signatures and major metabolic pathways of gastric cancer with help of transcriptomic data from TCGA Biosci Rep 2021 4110
18 Balluff B et al Direct molecular tissue analysis by MALDI imaging mass spectrometry in the field of gastrointestinal disease Gastroenterology 2012 1433 p 544-549 e2
19 Streitz JM Jr et al Analysis of protein expression patterns in Barretts esophagus using MALDI mass spectrometry in search of malignancy biomarkers Dis Esophagus 2005 183 p 170-6
20 Erich K et al Spatial Distribution of Endogenous Tissue Protease Activity in Gastric Carcinoma Mapped by MALDI Mass Spectrometry Imaging Mol Cell Proteomics 2019 181 p 151-161
21 Gaye MM et al Delineation of disease phenotypes associated with esophageal adenocarcinoma by MALDI-IMS-MS analysis of serum N-linked glycans Analyst 2017 1429 p 1525-1535
22 Kunzke T et al Native glycan fragments detected by MALDI-FT-ICR mass spectrometry imaging impact gastric cancer biology and patient outcome Oncotarget 2017 840 p 68012-68025
23 Jia K et al Novel circulating peptide biomarkers for esophageal squamous cell carcinoma revealed by a magnetic bead-based MALDI-TOFMS assay Oncotarget 2016 717 p 23569-80
24 Alexandrov LB et al Signatures of mutational processes in human cancer Nature 2013 5007463 p 415-21
25 Lorenzen S et al Feasibility of perioperative chemotherapy with infusional 5-FU leucovorin and oxaliplatin with FLOT or without FLO docetaxel in elderly patients with locally advanced esophagogastric cancer Br J Cancer 2013 1083 p 519-26
26 Gotze TO et al Preventive HIPEC in combination with perioperative FLOT versus FLOT alone for resectable diffuse type gastric and gastroesophageal junction type IIIII adenocarcinoma - the phase III PREVENT- FLOT9 trial of the AIO CAOGI ACO BMC Cancer 2021 211 p 1158
27 Lombardi PM et al Facing adenocarcinoma of distal esophagus and esophagogastric junction a CROSS versus FLOT propensity score-matched analysis of oncological outcomes in a high-volume institution Updates Surg 2023 754 p 921-930
28 Hingorani M et al Neoadjuvant Management of Adenocarcinoma of the Esophagus and Esophagogastric Junction Review of Randomized Evidence and Definition of Optimum Treatment Algorithm Oncology 2023 1019 p 553-564
29 Kudo T et al Nivolumab treatment for oesophageal squamous-cell carcinoma an open-label multicentre phase 2 trial Lancet Oncol 2017 185 p 631-639
30 Schwacke J et al Discrimination of normal and esophageal cancer plasma proteomes by MALDI-TOF mass spectrometry Dig Dis Sci 2015 606 p 1645-54
31 Berghmans E et al MALDI Mass Spectrometry Imaging Linked with Top-Down Proteomics as a Tool to Study the Non-Small-Cell Lung Cancer Tumor Microenvironment Methods Protoc 2019 22
32 Low DE et al Benchmarking Complications Associated with Esophagectomy Ann Surg 2019 2692 p 291-298
33 Schmidt HM et al Defining Benchmarks for Transthoracic Esophagectomy A Multicenter Analysis of Total Minimally Invasive Esophagectomy in Low Risk Patients Ann Surg 2017 2665 p 814-821
Study Oversight
Has Oversight DMC:
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Is a FDA Regulated Drug?:
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Is an Unapproved Device?:
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Is a US Export?:
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Is an FDA AA801 Violation?:
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