Appendix1_IMed_FU_Questionnaire_revised – Supplemental material for Self-reported Improvement in Side Effects and Quality of Life With Integrative Medicine in Breast Cancer Patients

<p>Supplemental material, Appendix1_IMed_FU_Questionnaire_revised for Self-reported Improvement in Side Effects and Quality of Life With Integrative Medicine in Breast Cancer Patients by Carolin C. Hack, Janina Hackl, Nina B. M. Hüttner, Hanna Langemann, Judith Schwitulla, Svenja Dietzel-Drentwett, Peter A. Fasching, Matthias W. Beckmann and Anna-Katharin Theuser in Integrative Cancer Therapies</p

Data_Sheet_3.XLSX

<p>Neutropenia secondary to chemotherapy in breast cancer patients can be life-threatening and there are no biomarkers available to predict the risk of drug-induced neutropenia in those patients. We previously performed a genome-wide association study (GWAS) for neutropenia events in women with breast cancer who were treated with 5-fluorouracil, epirubicin and cyclophosphamide and recruited to the SUCCESS-A trial. A genome-wide significant single-nucleotide polymorphism (SNP) signal in the tumor necrosis factor superfamily member 13B (TNFSF13B) gene, encoding the cytokine B-cell activating factor (BAFF), was identified in that GWAS. Taking advantage of these existing GWAS data, in the present study we utilized a pathway-based analysis approach by leveraging knowledge of the pharmacokinetics and pharmacodynamics of drugs and breast cancer pathophysiology to identify additional SNPs/genes associated with the underlying etiology of chemotherapy-induced neutropenia. We identified three SNPs in the hyaluronan mediated motility receptor (HMMR) gene that were significantly associated with neutropenia (p < 1.0E-04). Those three SNPs were trans-expression quantitative trait loci for the expression of TNFSF13B (p < 1.0E-04). The minor allele of these HMMR SNPs was associated with a decreased TNFSF13B mRNA level. Additional functional studies performed with lymphoblastoid cell lines (LCLs) demonstrated that LCLs possessing the minor allele for the HMMR SNPs were more sensitive to drug treatment. Knock-down of TNFSF13B in LCLs and HL-60 promyelocytic cells and treatment of those cells with BAFF modulated the cell sensitivity to chemotherapy treatment. These results demonstrate that HMMR SNP-dependent cytotoxicity of these chemotherapeutic agents might be related to TNFSF13B expression level. In summary, utilizing a pathway-based approach for the analysis of GWAS data, we identified additional SNPs in the HMMR gene that were associated with neutropenia and also were correlated with TNFSF13B expression.</p

Data_Sheet_1.XLSX

<p>Neutropenia secondary to chemotherapy in breast cancer patients can be life-threatening and there are no biomarkers available to predict the risk of drug-induced neutropenia in those patients. We previously performed a genome-wide association study (GWAS) for neutropenia events in women with breast cancer who were treated with 5-fluorouracil, epirubicin and cyclophosphamide and recruited to the SUCCESS-A trial. A genome-wide significant single-nucleotide polymorphism (SNP) signal in the tumor necrosis factor superfamily member 13B (TNFSF13B) gene, encoding the cytokine B-cell activating factor (BAFF), was identified in that GWAS. Taking advantage of these existing GWAS data, in the present study we utilized a pathway-based analysis approach by leveraging knowledge of the pharmacokinetics and pharmacodynamics of drugs and breast cancer pathophysiology to identify additional SNPs/genes associated with the underlying etiology of chemotherapy-induced neutropenia. We identified three SNPs in the hyaluronan mediated motility receptor (HMMR) gene that were significantly associated with neutropenia (p < 1.0E-04). Those three SNPs were trans-expression quantitative trait loci for the expression of TNFSF13B (p < 1.0E-04). The minor allele of these HMMR SNPs was associated with a decreased TNFSF13B mRNA level. Additional functional studies performed with lymphoblastoid cell lines (LCLs) demonstrated that LCLs possessing the minor allele for the HMMR SNPs were more sensitive to drug treatment. Knock-down of TNFSF13B in LCLs and HL-60 promyelocytic cells and treatment of those cells with BAFF modulated the cell sensitivity to chemotherapy treatment. These results demonstrate that HMMR SNP-dependent cytotoxicity of these chemotherapeutic agents might be related to TNFSF13B expression level. In summary, utilizing a pathway-based approach for the analysis of GWAS data, we identified additional SNPs in the HMMR gene that were associated with neutropenia and also were correlated with TNFSF13B expression.</p

Data_Sheet_6.XLSX

<p>Neutropenia secondary to chemotherapy in breast cancer patients can be life-threatening and there are no biomarkers available to predict the risk of drug-induced neutropenia in those patients. We previously performed a genome-wide association study (GWAS) for neutropenia events in women with breast cancer who were treated with 5-fluorouracil, epirubicin and cyclophosphamide and recruited to the SUCCESS-A trial. A genome-wide significant single-nucleotide polymorphism (SNP) signal in the tumor necrosis factor superfamily member 13B (TNFSF13B) gene, encoding the cytokine B-cell activating factor (BAFF), was identified in that GWAS. Taking advantage of these existing GWAS data, in the present study we utilized a pathway-based analysis approach by leveraging knowledge of the pharmacokinetics and pharmacodynamics of drugs and breast cancer pathophysiology to identify additional SNPs/genes associated with the underlying etiology of chemotherapy-induced neutropenia. We identified three SNPs in the hyaluronan mediated motility receptor (HMMR) gene that were significantly associated with neutropenia (p < 1.0E-04). Those three SNPs were trans-expression quantitative trait loci for the expression of TNFSF13B (p < 1.0E-04). The minor allele of these HMMR SNPs was associated with a decreased TNFSF13B mRNA level. Additional functional studies performed with lymphoblastoid cell lines (LCLs) demonstrated that LCLs possessing the minor allele for the HMMR SNPs were more sensitive to drug treatment. Knock-down of TNFSF13B in LCLs and HL-60 promyelocytic cells and treatment of those cells with BAFF modulated the cell sensitivity to chemotherapy treatment. These results demonstrate that HMMR SNP-dependent cytotoxicity of these chemotherapeutic agents might be related to TNFSF13B expression level. In summary, utilizing a pathway-based approach for the analysis of GWAS data, we identified additional SNPs in the HMMR gene that were associated with neutropenia and also were correlated with TNFSF13B expression.</p

Data_Sheet_5.XLSX

<p>Neutropenia secondary to chemotherapy in breast cancer patients can be life-threatening and there are no biomarkers available to predict the risk of drug-induced neutropenia in those patients. We previously performed a genome-wide association study (GWAS) for neutropenia events in women with breast cancer who were treated with 5-fluorouracil, epirubicin and cyclophosphamide and recruited to the SUCCESS-A trial. A genome-wide significant single-nucleotide polymorphism (SNP) signal in the tumor necrosis factor superfamily member 13B (TNFSF13B) gene, encoding the cytokine B-cell activating factor (BAFF), was identified in that GWAS. Taking advantage of these existing GWAS data, in the present study we utilized a pathway-based analysis approach by leveraging knowledge of the pharmacokinetics and pharmacodynamics of drugs and breast cancer pathophysiology to identify additional SNPs/genes associated with the underlying etiology of chemotherapy-induced neutropenia. We identified three SNPs in the hyaluronan mediated motility receptor (HMMR) gene that were significantly associated with neutropenia (p < 1.0E-04). Those three SNPs were trans-expression quantitative trait loci for the expression of TNFSF13B (p < 1.0E-04). The minor allele of these HMMR SNPs was associated with a decreased TNFSF13B mRNA level. Additional functional studies performed with lymphoblastoid cell lines (LCLs) demonstrated that LCLs possessing the minor allele for the HMMR SNPs were more sensitive to drug treatment. Knock-down of TNFSF13B in LCLs and HL-60 promyelocytic cells and treatment of those cells with BAFF modulated the cell sensitivity to chemotherapy treatment. These results demonstrate that HMMR SNP-dependent cytotoxicity of these chemotherapeutic agents might be related to TNFSF13B expression level. In summary, utilizing a pathway-based approach for the analysis of GWAS data, we identified additional SNPs in the HMMR gene that were associated with neutropenia and also were correlated with TNFSF13B expression.</p

Presentation_1.pdf

<p>Neutropenia secondary to chemotherapy in breast cancer patients can be life-threatening and there are no biomarkers available to predict the risk of drug-induced neutropenia in those patients. We previously performed a genome-wide association study (GWAS) for neutropenia events in women with breast cancer who were treated with 5-fluorouracil, epirubicin and cyclophosphamide and recruited to the SUCCESS-A trial. A genome-wide significant single-nucleotide polymorphism (SNP) signal in the tumor necrosis factor superfamily member 13B (TNFSF13B) gene, encoding the cytokine B-cell activating factor (BAFF), was identified in that GWAS. Taking advantage of these existing GWAS data, in the present study we utilized a pathway-based analysis approach by leveraging knowledge of the pharmacokinetics and pharmacodynamics of drugs and breast cancer pathophysiology to identify additional SNPs/genes associated with the underlying etiology of chemotherapy-induced neutropenia. We identified three SNPs in the hyaluronan mediated motility receptor (HMMR) gene that were significantly associated with neutropenia (p < 1.0E-04). Those three SNPs were trans-expression quantitative trait loci for the expression of TNFSF13B (p < 1.0E-04). The minor allele of these HMMR SNPs was associated with a decreased TNFSF13B mRNA level. Additional functional studies performed with lymphoblastoid cell lines (LCLs) demonstrated that LCLs possessing the minor allele for the HMMR SNPs were more sensitive to drug treatment. Knock-down of TNFSF13B in LCLs and HL-60 promyelocytic cells and treatment of those cells with BAFF modulated the cell sensitivity to chemotherapy treatment. These results demonstrate that HMMR SNP-dependent cytotoxicity of these chemotherapeutic agents might be related to TNFSF13B expression level. In summary, utilizing a pathway-based approach for the analysis of GWAS data, we identified additional SNPs in the HMMR gene that were associated with neutropenia and also were correlated with TNFSF13B expression.</p

Univariate analysis of clinico-pathological parameters influencing tumour-specific survival in the group of grade^a 1 and 2 breast tumours.

<p>^aAccording to Bloom and Richardson, as modified by Elston and Ellis [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#pone.0122106.ref032" target="_blank">32</a>].</p><p>^bLog-rank test at the two-sided significance level of 0.05.</p><p>^cMedian immunoreactive score (IRS) according to Remmele and Stegner [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#pone.0122106.ref030" target="_blank">30</a>] was used as cut-off: AGR3 low (IRS 0–4), AGR3 high (IRS 6–12).</p><p>^dAccording to TNM classification by Sobin and Wittekind [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#pone.0122106.ref058" target="_blank">58</a>].</p><p>^eImmunoreactive score (IRS) according to Remmele and Stegner [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#pone.0122106.ref030" target="_blank">30</a>].</p><p>^fOverexpression of the <i>ERBB2</i> gene (Her-2/neu) was diagnosed analogously to the threshold of the DAKO-Score system based on IHC assay. Significant P-values are marked in bold face.</p><p>Univariate analysis of clinico-pathological parameters influencing tumour-specific survival in the group of grade<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#t001fn001" target="_blank">^a</a> 1 and 2 breast tumours.</p

Data_Sheet_4.XLSX

<p>Neutropenia secondary to chemotherapy in breast cancer patients can be life-threatening and there are no biomarkers available to predict the risk of drug-induced neutropenia in those patients. We previously performed a genome-wide association study (GWAS) for neutropenia events in women with breast cancer who were treated with 5-fluorouracil, epirubicin and cyclophosphamide and recruited to the SUCCESS-A trial. A genome-wide significant single-nucleotide polymorphism (SNP) signal in the tumor necrosis factor superfamily member 13B (TNFSF13B) gene, encoding the cytokine B-cell activating factor (BAFF), was identified in that GWAS. Taking advantage of these existing GWAS data, in the present study we utilized a pathway-based analysis approach by leveraging knowledge of the pharmacokinetics and pharmacodynamics of drugs and breast cancer pathophysiology to identify additional SNPs/genes associated with the underlying etiology of chemotherapy-induced neutropenia. We identified three SNPs in the hyaluronan mediated motility receptor (HMMR) gene that were significantly associated with neutropenia (p < 1.0E-04). Those three SNPs were trans-expression quantitative trait loci for the expression of TNFSF13B (p < 1.0E-04). The minor allele of these HMMR SNPs was associated with a decreased TNFSF13B mRNA level. Additional functional studies performed with lymphoblastoid cell lines (LCLs) demonstrated that LCLs possessing the minor allele for the HMMR SNPs were more sensitive to drug treatment. Knock-down of TNFSF13B in LCLs and HL-60 promyelocytic cells and treatment of those cells with BAFF modulated the cell sensitivity to chemotherapy treatment. These results demonstrate that HMMR SNP-dependent cytotoxicity of these chemotherapeutic agents might be related to TNFSF13B expression level. In summary, utilizing a pathway-based approach for the analysis of GWAS data, we identified additional SNPs in the HMMR gene that were associated with neutropenia and also were correlated with TNFSF13B expression.</p

Evaluation of the diagnostic performance of candidate serum biomarkers AGR3, AGR2 and their combination.

<p>Scatter plot demonstrating a highly significant elevation of (<b>A</b>) AGR3 and (<b>C</b>) AGR2 protein concentration in human serum samples from breast cancer patients (n = 40) in comparison with samples from healthy women (n = 40). ROC curve analysis for (<b>B</b>) AGR3, (<b>D</b>) AGR2 and (<b>E</b>) the combination of both proteins. ** <i>P</i> < 0.01, *** <i>P</i> < 0.001. ROC: receiver operating characteristic, AUC: area under the curve, 95% CI: 95% confidence interval.</p

Multivariate Cox regression analysis including all factors potentially influencing tumour-specific survival in grade^a 1 and 2 breast cancer samples.

<p>^aAccording to Bloom and Richardson, as modified by Elston and Ellis [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#pone.0122106.ref032" target="_blank">32</a>].</p><p>^bMedian immunoreactive score (IRS) according to Remmele and Stegner [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#pone.0122106.ref030" target="_blank">30</a>] was used as cut-off: AGR3 low (IRS 0–4), AGR3 high (IRS 6–12).</p><p>^cAccording to TNM classification by Sobin and Wittekind [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#pone.0122106.ref058" target="_blank">58</a>]. Significant P-values are marked in bold face. HR, hazard ratio; CI, confidence interval.</p><p>Multivariate Cox regression analysis including all factors potentially influencing tumour-specific survival in grade<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122106#t002fn001" target="_blank">^a</a> 1 and 2 breast cancer samples.</p