The aim of the study was to determine the levels of selected cytokines and chemokines in the serum of multiple myeloma (MM) patients treated with bortezomib-based regimens. A total of 71 MM patients were examined: 41 with primary refractory disease (17) or early relapse (28), and 30 who were bortezomib sensitive with no progression for at least six months. Patients who demonstrated CR or PR after bortezomib-based therapies longer than six months after treatment discontinuation were designated bortezomib sensitive. Serum cytokine levels were assayed with Bio-Rad Bio-Plex Pro Human Cytokine 27-Plex Assay on the MAGPIX Multiplex Reader and the Bio-Plex® 200 System (Bio-Rad). Higher levels of MIP-1α and lower levels of MIP-1β and IL-9 were associated with better responses to bortezomib-based treatment, and higher levels of IL-1ra and IL-8 were associated with bone involvement. MCP-1 was elevated in patients with hemoglobin<10 g/dl compared to those without anemia. The levels of IL-8, MIP-1α, and TNF-α were significantly higher in patients with renal insufficiency. Only MIP-1α was elevated in patients with hypercalcemia compared to patients with normal calcium levels. In conclusion, distinct cytokines are involved in the pathogenesis of MM and may play a prominent role in the prediction of treatment response. However, a single measurement of serum cytokines should be interpreted with caution and further studies are needed.

No abstract

Bortezomib is the first-in-class proteasome inhibitor, commonly used in the treatment of multiple myeloma (MM). The mechanisms underlying acquired bortezomib resistance in MM are poorly understood. Several cell-free miRNAs have been found to be aberrantly regulated in MM patients. The aim of this pilot study was to identify a blood-based miRNA signature that predicts bortezomib-based therapy efficacy in MM patients. Thirty MM patients treated with bortezomib-based regimens were studied, including 19 with refractory disease and 11 who were bortezomib sensitive. Serum miRNA expression patterns were identified with miRCURY LNA miRNA miRNome PCR Panels I+II (Exiqon/Qiagen). Univariate analysis found a total of 21 miRNAs to be differentially expressed in patients with MM according to bortezomib sensitivity. Multivariate logistic regression was created and allowed us to discriminate refractory from sensitive patients with a very high AUC of 0.95 (95%CI: 0.84–1.00); sensitivity, specificity and accuracy were estimated as 0.95, 0.91, and 0.93. The model used expression of 3 miRNAs: miR-215-5p, miR-181a-5p and miR-376c-3p. This study is the first to demonstrate that serum expression of several miRNAs differs between patients who are bortezomib refractory and those who are sensitive which may prove useful in studies aimed at overcoming drug resistance in MM treatment.

Introduction: Autologous hematopoietic stem cell transplantation (AHSCT) is an acknowledged and effective treatment method for hematopoietic system diseases. MicroRNAs were reported to impact the bone marrow niche microenvironment and regulate proliferation and survival of the hematopoietic stem cells in such a manner may also influence bone marrow convalescence after AHSCT. The project aimed to identify changes in the signature of miRNAs freely circulating in the serum during AHSCT related to chemotherapy-induced injury and further bone marrow recovery using next-generation sequencing. Patients and methods: Serum samples from 10 patients undergoing ASCT were collected. Blood samples were taken from each patient at four time points: (T1) before conditioning with high dose chemotherapy, (T2) on the day of AHSCT (day 0), on day +7 (T3), and on +14 day after AHSCT (T4). The myeloablative conditioning regimen for patients with MM was melphalan 200 mg/m 2, while in lymphoma patients, BEAM was used. Total RNA was extracted from 200 μl serum using miRNeasy Serum/Plasma Advanced Kit (QIAGEN) following manufacture instructions. Libraries were prepared from 5 μl of total RNA using QIAseq® miRNA Library Kit. The libraries were pooled in equimolar concentrations and sequenced on a NextSeq 550 System using a single-end read length of 75 nucleotides at an average of 10 million reads per sample (Illumina). In bioinformatics analysis, after adapter cut-off, filtration and mapping, miRNAs were counted based on mapping to reference miRbase 22 (tools: fastp, bowtie, samtools, picard). MicroRNAs were filtered to have at least 10 counts-per-million (CPM) of classified sequences in at least two samples. MiRNAs expression levels between time points were compared using paired t-test with Bonferroni correction. Results: The study group consisted patients with multiple myeloma (N=4), Hodgkin lymphoma (HL, N=3), and non-HL (N=3) aged 48±13 years. There was a significant decrease in the hematological parameters during ASCT with a nadir at T3, including hemoglobin (T1 vs. T3, p&lt;0.0012), white blood cell count (p&lt;0.0001), neutrophil count (p=0.0003), and platelet count (p&lt;0.0001). Similarly, the decrease was observed in hsa-miR-223-3p (T1 vs. T3, p=0.048) and hsa-miR-18a-5p (T2 vs. T3, p=0.033) with a nadir at T3. On the other hand, an increase with a peak at T3 was observed in the expression of hsa-miR-320b (T1 vs. T3, p=0.007), hsa-miR-320c (T1 vs. T3, p=0.007), hsa-miR-320a-3p (T1 vs. T3, p=0.009), and hsa-miR-320d (T1 vs. T3, p=0.042). Interestingly, we have observed a gradual decrease across study timepoints in the expression of hsa-let-7f-5p, hsa-let-7i-5p, and hsa-miR-155-5p with a nadir at T4 (T1 vs. T4, p=0.004, p=0.01, and p=0.019, respectively). Similar changes were observed in the expression of hsa-miR-486-5p, but the statistically significant decline was only noted between T3 and T4 (p=0.024). Conversely, a gradual decrease was also seen in the expression of hsa-miR-96-5p, but there was a significant increment between T3 and T4 (p=0.036). Figure 1 presents the heatmaps for the miRNAs with significant expression changes and corresponding hematological parameters during AHSCT. Conclusion: Several significant changes in the miRNA expression profile were identified, both related to the chemotherapy-induced injury and subsequent bone marrow recovery. Figure 1 Figure 1. Disclosures Wierzbowska: Novartis: Consultancy; Abbvie: Consultancy; Jazz: Research Funding; Janssen: Consultancy; Astellas: Consultancy; Celgene/BMS: Consultancy.

Proteasome inhibitors, like bortezomib, play a key role in the treatment of multiple myeloma (MM); however, most patients eventually relapse and eventually show multiple drug resistance, and the molecular mechanisms of this resistance remain unclear. The aim of our study is to assess the expression of previously described genes that may influence the resistance to bortezomib treatment at the mRNA level (ABCB1, CXCR4, MAF, MARCKS, POMP, PSMB5, RPL5, TXN, and XBP1) and prognosis of MM patients. mRNA expression was determined in 73 MM patients treated with bortezomib-based regimens (30 bortzomib-sensitive and 43 bortezomib-refractory patients) and 11 healthy controls. RPL5 was significantly down-regulated in multiple myeloma patients as compared with healthy controls. Moreover, POMP was significantly up-regulated in MM patients refractory to bortezomib-based treatment. In multivariate analysis, high expression of PSMB5 and CXCR and autologous stem cell transplantation were independent predictors of progression-free survival, and high expression of POMP and RPL5 was associated with shorter overall survival.