Computational protocol: Molecular responses to therapeutic proteasome inhibitors in multiple myeloma patients are donor-, cell type- and drug-dependent

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Protocol publication

[…] Total RNA was extracted from PBMCs using the NucleoSpin® RNA mini kit (Macherey Nagel) and quantified with a BioSpec-nano spectrophotometer (Shimadzu Inc.). RNA (100 ng) was converted to cDNA with the Maxima First Strand cDNA Synthesis Kit for RT-qPCR (Thermo Scientific). Real-time PCR (in duplicates) was performed using Maxima SYBR Green/ROX qPCR Master Mix (Thermo Scientific) and the PikoReal 96 Real-Time PCR System. All oligonucleotides used in this work were designed using the primer-BLAST tool (http://www.ncbi.nlm.nih.gov/tools/primer-blast/). The RT-PCR gene-specific primers are reported in . [...] Data were collected from the medical files of the enrolled patients. Disease-related measures (depth of response, PFS, OS) were recorded according to IMWG (International Myeloma Working Group) criteria []. PFS and OS were measured from the day of treatment initiation to disease progression or death from any cause. Assays were performed at least in duplicates. Statistical significance was evaluated by using Student’s T test (2-tailed) or by using ANOVA single factor (Alpha = 0.05). Error bars denote standard deviation (S.D.); significance at P<0.05 or P<0.01 is indicated on the graphs by one or two asterisks, respectively. For S.D. calculation in Excel the STDEV function was used and the output is recorded on the graphs. S.D. points at T1B refer to variation of the independent measurements; the mean of which was set to 100%. Pearson’s (r) and Spearman’s Rank (rho) correlation coefficient tests were employed to examine relationships between various parameters and clinical data. Analyses were performed using either Microsoft Excel or SPSS 23 software (IBM SPSS statistics for Windows, Version 23.0, IBM Corp, Armonk, NY, USA). […]

Pipeline specifications

Software tools Primer-BLAST, SPSS
Applications Miscellaneous, qPCR
Organisms Homo sapiens
Diseases Multiple Myeloma, Neoplasms, Hematologic Neoplasms