#METABOLOMICS WORKBENCH TatianaJoao_20220914_101058 DATATRACK_ID:3461 STUDY_ID:ST002295 ANALYSIS_ID:AN003749 PROJECT_ID:PR001307 VERSION 1 CREATED_ON September 14, 2022, 10:19 am #PROJECT PR:PROJECT_TITLE Biochemical Impact of Platinum and Palladium-based Anticancer Agents – PR:PROJECT_TITLE BioIMPACT PR:PROJECT_TYPE NMR-based metabolomics PR:PROJECT_SUMMARY Platinum (Pt(II)) drugs, e.g. cisplatin (cDDP), are some of the most used PR:PROJECT_SUMMARY chemotherapeutic agents, yet tumor acquired resistance and high toxicity are PR:PROJECT_SUMMARY still current drawbacks. Palladium (Pd(II))-complexes are alternatives due to PR:PROJECT_SUMMARY similar metal coordination and promising cytotoxic properties. Metabolomics can PR:PROJECT_SUMMARY measure the metabolic response of drug-exposed tissues, unveiling insight into PR:PROJECT_SUMMARY drug mechanisms and new markers of drug efficacy/toxicity. The present 1H NMR PR:PROJECT_SUMMARY metabolomics study aims to characterize the in vivo response of the impact of a PR:PROJECT_SUMMARY Pd(II)-complex with polyamine spermine (Pd2Spm), compared to cDDP, on the PR:PROJECT_SUMMARY metabolism of several organs from a cell-derived xenograft mouse model of PR:PROJECT_SUMMARY Triple-Negative Breast Cancer. PR:INSTITUTE University of Aveiro PR:DEPARTMENT Department of Chemistry and CICECO-Aveiro Institute of Materials PR:LABORATORY Metabolomics from Ana M. Gil PR:LAST_NAME Carneiro PR:FIRST_NAME Tatiana João PR:ADDRESS Campus Universitário de Santiago, Aveiro, Aveiro, 3810-193, Portugal PR:EMAIL tatiana.joao@ua.pt PR:PHONE +351 234 370 200 PR:FUNDING_SOURCE This research was developed within the scope of the CICECO—Aveiro Institute of PR:FUNDING_SOURCE Materials, with references UIDB/50011/2020, UIDP/50011/2020, and LA/P/0006/2020, PR:FUNDING_SOURCE financed by national funds through the Portuguese Foundation for Science and PR:FUNDING_SOURCE Technology (FCT/MEC) and when appropriate co-financed by European Regional PR:FUNDING_SOURCE Development Fund (FEDER) under the PT2020 Partnership Agreement. This work was PR:FUNDING_SOURCE also funded by the FCT through UIDB/50006/2020, UIDB/00070/2020, PR:FUNDING_SOURCE POCI-01-0145-FEDER-0016786, and Centro-01-0145-FEDER-029956 (co-financed by PR:FUNDING_SOURCE COMPETE 2020, Portugal 2020 and European Community through FEDER). We also PR:FUNDING_SOURCE acknowledge the Portuguese National NMR Network (PTNMR), supported by FCT funds PR:FUNDING_SOURCE as the NMR spectrometer used is part of PTNMR and partially supported by PR:FUNDING_SOURCE Infrastructure Project Nº 022161 (co-financed by FEDER through COMPETE 2020, PR:FUNDING_SOURCE POCI and PORL, and the FCT through PIDDAC). We also acknowledge European Social PR:FUNDING_SOURCE Fund of the European Union and national funds FCT/MCTES for the PhD grants PR:FUNDING_SOURCE PD/BD/135460/2017 and SFRH/BD/145920/2019. #STUDY ST:STUDY_TITLE Metabolic impact of anticancer drugs Pd2Spermine and Cisplatin on the lipophilic ST:STUDY_TITLE metabolome of liver from cell-derived xenograft mouse model of Triple-Negative ST:STUDY_TITLE Breast Cancer (part 2) ST:STUDY_TYPE NMR-based metabolomics ST:STUDY_SUMMARY Platinum (Pt(II)) drugs, e.g. cisplatin (cDDP), are some of the most used ST:STUDY_SUMMARY chemotherapeutic agents, yet tumor acquired resistance and high toxicity are ST:STUDY_SUMMARY still current drawbacks. Palladium (Pd(II))-complexes are alternatives due to ST:STUDY_SUMMARY similar metal coordination and promising cytotoxic properties. Metabolomics can ST:STUDY_SUMMARY measure the metabolic response of drug-exposed tissues, unveiling insight into ST:STUDY_SUMMARY drug mechanisms and new markers of drug efficacy/toxicity. The present 1H NMR ST:STUDY_SUMMARY metabolomics study aims to characterize the in vivo response of the impact of a ST:STUDY_SUMMARY Pd(II)-complex with polyamine spermine (Pd2Spm), compared to cDDP, on lipophilic ST:STUDY_SUMMARY metabolism of liver from cell-derived xenograft mouse model of Triple-Negative ST:STUDY_SUMMARY Breast Cancer. ST:INSTITUTE University of Aveiro ST:DEPARTMENT Department of Chemistry and CICECO-Aveiro Institute of Materials ST:LABORATORY Metabolomics from Ana M. Gil ST:LAST_NAME Carneiro ST:FIRST_NAME Tatiana João ST:ADDRESS Campus Universitário de Santiago, Aveiro, Aveiro, 3810-193, Portugal ST:EMAIL tatiana.joao@ua.pt ST:PHONE +351 234 370 200 ST:NUM_GROUPS 3 ST:TOTAL_SUBJECTS 22 ST:NUM_FEMALES 22 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN CBA nude (N:NIH(S)II-nu/nu) SU:AGE_OR_AGE_RANGE 6 to 7-weeks old SU:GENDER Female SU:ANIMAL_ANIMAL_SUPPLIER i3S Animal Facility (Porto, Portugal) SU:ANIMAL_HOUSING ICBAS-UP Rodent Animal House Facility (Porto, Portugal) SU:ANIMAL_LIGHT_CYCLE 12h light/dark cycles (7.00 AM lights on) SU:ANIMAL_FEED ad libitum SU:ANIMAL_WATER ad libitum SU:ANIMAL_INCLUSION_CRITERIA Healthy animails SU:SPECIES_GROUP CBA nude (N:NIH(S)II-nu/nu) #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - xeno_L_EL_C_11_1_2 Treatment_group:Control RAW_FILE_NAME=xeno_L_EL_C_11_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_C_17_1_2 Treatment_group:Control RAW_FILE_NAME=xeno_L_EL_C_17_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_C_20b_1_2 Treatment_group:Control RAW_FILE_NAME=xeno_L_EL_C_20b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_C_26b_1_2 Treatment_group:Control RAW_FILE_NAME=xeno_L_EL_C_26b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_C_27b_1_2 Treatment_group:Control RAW_FILE_NAME=xeno_L_EL_C_27b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_C_29b_1_2 Treatment_group:Control RAW_FILE_NAME=xeno_L_EL_C_29b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_A_7_1_2 Treatment_group:Cisplatin-treated RAW_FILE_NAME=xeno_L_EL_A_7_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_A_14_1_2 Treatment_group:Cisplatin-treated RAW_FILE_NAME=xeno_L_EL_A_14_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_A_16_1_2 Treatment_group:Cisplatin-treated RAW_FILE_NAME=xeno_L_EL_A_16_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_A_24b_1_2 Treatment_group:Cisplatin-treated RAW_FILE_NAME=xeno_L_EL_A_24b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_A_25_1_2 Treatment_group:Cisplatin-treated RAW_FILE_NAME=xeno_L_EL_A_25_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_A_28b_1_2 Treatment_group:Cisplatin-treated RAW_FILE_NAME=xeno_L_EL_A_28b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_A_32b_1_2 Treatment_group:Cisplatin-treated RAW_FILE_NAME=xeno_L_EL_A_32b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_A_33b_1_2 Treatment_group:Cisplatin-treated RAW_FILE_NAME=xeno_L_EL_A_33b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_B_8b_1_2 Treatment_group:Pd2Spm-treated RAW_FILE_NAME=xeno_L_EL_B_8b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_B_10b_1_2 Treatment_group:Pd2Spm-treated RAW_FILE_NAME=xeno_L_EL_B_10b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_B_13c_1_2 Treatment_group:Pd2Spm-treated RAW_FILE_NAME=xeno_L_EL_B_13c_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_B_18b_1_2 Treatment_group:Pd2Spm-treated RAW_FILE_NAME=xeno_L_EL_B_18b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_B_19b_1_2 Treatment_group:Pd2Spm-treated RAW_FILE_NAME=xeno_L_EL_B_19b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_B_22b_1_2 Treatment_group:Pd2Spm-treated RAW_FILE_NAME=xeno_L_EL_B_22b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_B_23b_1_2 Treatment_group:Pd2Spm-treated RAW_FILE_NAME=xeno_L_EL_B_23b_1_2 SUBJECT_SAMPLE_FACTORS - xeno_L_EL_B_30b_1_2 Treatment_group:Pd2Spm-treated RAW_FILE_NAME=xeno_L_EL_B_30b_1_2 #COLLECTION CO:COLLECTION_SUMMARY At day 39 post-implantation of MDA-MB-231 cells (25G needle, 5E6 cells in 150 CO:COLLECTION_SUMMARY microliters of PBS) in the left flank of mice, and, at day 9 post-treatment with CO:COLLECTION_SUMMARY metal-based drugs, the mice were sacrificed with isoflurane, organs were CO:COLLECTION_SUMMARY excised, snap-frozen and stored at -80 ºC. CO:SAMPLE_TYPE Liver CO:COLLECTION_DURATION Inferior to one minute CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY At day 25 post-implantation, 8 animals were randomly chosen into three groups to TR:TREATMENT_SUMMARY receive the treatment with either (i) vehicle (phosphate-buffered saline, PBS) - TR:TREATMENT_SUMMARY controls, (ii) cDDP (2 mg/kg/day), or (iii) Pd2Spm (5 mg/kg/day), via TR:TREATMENT_SUMMARY intraperitoneal injection, during five consecutive days. Two control animals TR:TREATMENT_SUMMARY were excluded from the study since they developed ulcerated tumors (at day 28 TR:TREATMENT_SUMMARY post-implantation) and needed to be prematurely euthanized. TR:TREATMENT_ROUTE Intraperitoneal injection TR:TREATMENT_DOSEVOLUME 500 microliters TR:TREATMENT_VEHICLE PBS (phosphate-buffered saline solution) #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The half of liver (comprising median and left lobes) from each mouse was SP:SAMPLEPREP_SUMMARY mechanically grounded in liquid nitrogen and samples were extracted using the SP:SAMPLEPREP_SUMMARY biphasic methanol/ chloroform/ water (2:2:1) method. Lipophilic phases were SP:SAMPLEPREP_SUMMARY recovered and dried. Previously to NMR acquisition, lipophilic extracts were SP:SAMPLEPREP_SUMMARY suspended in 650 µL of CDCl3, containing 0.03% tetramethylsilane (TMS). Samples SP:SAMPLEPREP_SUMMARY were then homogenized and transferred into 5mm NMR tubes. SP:PROCESSING_STORAGE_CONDITIONS -80℃ SP:EXTRACTION_METHOD Biphasic method (methanol/ chloroform/ water) SP:EXTRACT_STORAGE -80℃ #ANALYSIS AN:ANALYSIS_TYPE NMR AN:LABORATORY_NAME Metabolomics Ana M. Gil AN:SOFTWARE_VERSION Topspin 3.2 AN:ACQUISITION_DATE April 2021 #NMR NM:INSTRUMENT_NAME Avance III TM HD 500MHz NM:INSTRUMENT_TYPE FT-NMR NM:NMR_EXPERIMENT_TYPE 1D-1H NM:FIELD_FREQUENCY_LOCK Deuterated chloroform NM:SPECTROMETER_FREQUENCY 500MHz NM:NMR_PROBE TXI NM:NMR_SOLVENT CDCl3 NM:NMR_TUBE_SIZE 5mm NM:SHIMMING_METHOD Topshim NM:RECEIVER_GAIN 203 NM:TEMPERATURE 298K NM:NUMBER_OF_SCANS 512 NM:ACQUISITION_TIME 2.34s NM:RELAXATION_DELAY 2s NM:SPECTRAL_WIDTH 7002.801 NM:ZERO_FILLING 64k NM:BASELINE_CORRECTION_METHOD Manual NM:CHEMICAL_SHIFT_REF_STD TMS (tetramethylsilane) NM:NMR_RESULTS_FILE ST002295_AN003749_Results.txt UNITS:ppm #END