MKC-3946 is an IRE1α endoribonuclease inhibitor, which triggers modest cytotoxicity in MM cells. (A) Formal chemical structure of MKC-3946. (B) RPMI 8226 cells were treated with or without Tm (5 μg/mL) in combination with MKC-3946 (0-10μM) for 3 hours. Total RNA was extracted; XBP1 and β-actin mRNA were evaluated by RT-PCR. (C) RPMI 8226 cells were treated with Tm (5 μg/mL) in the presence or absence of MKC-3946 (10μM) for the indicated times. Total RNA was extracted, and XBP1 and β-actin mRNA were evaluated by RT-PCR. Whole-cell lysates were subjected to Western blotting using anti-IRE1α, phospho-IRE1α (p-IRE1α), BiP/GRP78, and actin Abs. (D) RPMI 8226 cells were cultured with or without MKC-3946 (10μM) for 8 hours. XBP1 target genes, such as SEC61A1, p58IPK, and ERdj4, were determined by real-time quantitative PCR. Data represent mean ± SD fold changes relative to β-actin mRNA in triplicate samples. *P < .001. (E) MM cell lines were cultured with MKC-3946 (0-12.5μM) for 48 hours. Cell viability was assessed by MTT assay of triplicate cultures, expressed as a percentage of untreated control. Data represent mean ± SD. (F) Primary MM cells isolated from patients (Pt) were cultured with or without MKC-3946 (10μM) for 6 hours. Total RNA was extracted and subjected to RT-PCR for analysis of XBP1 splicing.