Paradoxical expression of INK4c in proliferative multiple myeloma tumors: bi-allelic deletion vs increased expression

BACKGROUND: A high proliferative capacity of tumor cells usually is associated with shortened patient survival. Disruption of the RB pathway, which is critically involved in regulating the G1 to S cell cycle transition, is a frequent target of oncogenic events that are thought to contribute to increased proliferation during tumor progression. Previously, we determined that p18INK4c, an essential gene for normal plasma cell differentiation, was bi-allelically deleted in five of sixteen multiple myeloma (MM) cell lines. The present study was undertaken to investigate a possible role of p18INK4c in increased proliferation of myeloma tumors as they progress. RESULTS: Thirteen of 40 (33%) human myeloma cell lines do not express normal p18INK4c, with bi-allelic deletion of p18 in twelve, and expression of a mutated p18 fragment in one. Bi-allelic deletion of p18, which appears to be a late progression event, has a prevalence of about 2% in 261 multiple myeloma (MM) tumors, but the prevalence is 6 to10% in the 50 tumors with a high expression-based proliferation index. Paradoxically, 24 of 40 (60%) MM cell lines, and 30 of 50 (60%) MM tumors with a high proliferation index express an increased level of p18 RNA compared to normal bone marrow plasma cells, whereas this occurs in only five of the 151 (3%) MM tumors with a low proliferation index. Tumor progression is often accompanied by increased p18 expression and an increased proliferation index. Retroviral-mediated expression of exogenous p18 results in marked growth inhibition in three MM cell lines that express little or no endogenous p18, but has no effect in another MM cell line that already expresses a high level of p18. CONCLUSION: Paradoxically, although loss of p18 appears to contribute to increased proliferation of nearly 10% of MM tumors, most MM cell lines and proliferative MM tumors have increased expression of p18. Apart from a small fraction of cell lines and tumors that have inactivated the RB1 protein, it is not yet clear how other MM cell lines and tumors have become insensitive to the anti-proliferative effects of increased p18 expression

Gain/Amplification of Chromosome Arm 1q21 in Multiple Myeloma

Multiple myeloma (MM), a plasma cell neoplasm, is an incurable hematological malignancy characterized by complex genetic and prognostic heterogeneity. Gain or amplification of chromosome arm 1q21 (1q21+) is the most frequent adverse chromosomal aberration in MM, occurring in 40% of patients at diagnosis. It occurs in a subclone of the tumor as a secondary genomic event and is more amplified as the tumor progresses and a risk factor for the progression from smoldering multiple myeloma to MM. It can be divided into either 1q21 gain (3 copies) or 1q21 amplification (≥4 copies), and it has been suggested that the prognosis is worse in cases of amplification than gain. Trisomy of chromosome 1, jumping whole-arm translocations of chromosome1q, and tandem duplications lead to 1q21+ suggesting that its occurrence is not consistent at the genomic level. Many studies have reported that genes associated with the malignant phenotype of MM are situated on the 1q21 amplicon, including CKS1B, PSMD4, MCL1, ANP32E, and others. In this paper, we review the current knowledge regarding the clinical features, prognostic implications, and the speculated pathology of 1q21+ in MM, which can provide clues for an effective treatment approach to MM patients with 1q21+

Synergistic Effects of Venetoclax and Daratumumab on Antibody-Dependent Cell-Mediated Natural Killer Cytotoxicity in Multiple Myeloma

The prognosis of multiple myeloma (MM) has drastically improved owing to the development of new drugs, such as proteasome inhibitors and immunomodulatory drugs. Nevertheless, MM is an extremely challenging disease, and many patients are still refractory to the existing therapies, thus requiring new treatment alternatives. Venetoclax is a selective, orally bioavailable inhibitor of BCL-2 that shows efficacy in MM not only as a single agent but also in combination therapy, especially for MM patients with translocation t(11;14). However, many patients are refractory to this drug. Here, we treated the MM cell lines KMS12PE and KMS27 with a combination treatment of venetoclax targeting BCL-2 and daratumumab targeting CD38 to evaluate the synergistic cytotoxicity of these drugs in vitro. MM cell lines were co-cultured with natural killer (NK) cells at an effector:target ratio of 0.3:1 in the presence of serial concentrations of daratumumab and venetoclax, and the resulting apoptotic MM cells were detected by flow cytometry using annexin V. These results indicated that the antibody-dependent cell-mediated NK cytotoxicity was enhanced in KMS12PE and KMS27 cells harboring t(11;14) with a high BCL-2 expression, suggesting that the combination treatment of venetoclax and daratumumab should be especially effective in patients with these characteristics

Immunomodulatory and Metabolic Changes after Gnetin-C Supplementation in Humans

Gnetin-C is a naturally occurring stilbene derived from the seeds of Gnetum gnemon L., an edible plant native to Southeast Asia that is called melinjo. Although the biological properties and safety of G. gnemon extract, which contains nearly 3% Gnetin-C, have been confirmed in various human studies, whether or not pure Gnetin-C is safe for humans is unclear at present. We conducted a randomized, double-blind, placebo-controlled trial. Healthy subjects were randomly divided into two groups. The interventional group (n = 6) was given Gnetin-C, and the control group (n = 6) was provided a placebo, for 14 days. Lipid profiles, biomarkers of oxidative stress and circulating blood cells were assessed before and after the intervention. All subjects completed the study, with no side effects reported across the study duration. Gnetin-C supplementation demonstrated a statistically significant increase in the absolute number of circulating natural killer (NK) cells expressing the activating receptors NKG2D and NKp46. NK cells derived from subjects who received Gnetin-C for two weeks showed higher cytotoxicity against K562 target cells than those before receiving Gnetin-C. In addition, Gnetin-C also resulted in a significant decrease in the absolute neutrophil count in the blood compared with the placebo. Furthermore, Gnetin-C significantly reduced the levels of uric acid, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, total adiponectin, and high-molecular-weight adiponectin. These data indicate that Gnetin-C has biological effects of enhancing the NK activity on circulating human immune cells. The immunomodulatory effects are consistent with a putative improvement in cancer immunosurveillance via the upregulation of the NKG2D receptor. The study was registered with UMIN-CTR, number 000030364, on 12 December 2017

Establishment of Mucoepidermoid Carcinoma Cell Lines from Surgical and Recurrence Biopsy Specimens

Patients with advanced/recurrent mucoepidermoid carcinoma (MEC) have a poor prognosis. This study aimed to establish and characterize human mucoepidermoid carcinoma cell lines from the initial surgical specimen and biopsy specimen upon recurrence from the same patient to provide a resource for MEC research. MEC specimens from the initial surgical procedure and biopsy upon recurrence were used to establish cell lines. The established cell lines were cytogenetically characterized using multi-color fluorescence in situ hybridization and detection, and the sequence of the CRTC1-MAML2 chimeric gene was determined. Furthermore, the susceptibility of head and neck mucoepidermoid carcinoma to standard treatment drugs such as cisplatin, 5-fluorouracil, and cetuximab was investigated. We successfully established unique MEC cell lines, AMU-MEC1, from an initial surgical specimen and AMU-MEC1-R1 and AMU-MEC1-R2 from the recurrent biopsy specimen in the same patient. These cell lines exhibited epithelial morphology and developed in vitro-like cobblestones. They shared eight chromosomal abnormalities, including der(19)ins(19;11)(p13;?), which resulted in a chimeric CRTC1-MAML2 gene, indicating the same origin of the cell lines. The susceptibility of all cell lines to cisplatin and 5-fluorouracil was low. Interestingly, EGFR dependency for cell growth decreased in AMU-MEC-R1 and AMU-MEC-R2 but was retained in AMU-MEC1. These cytogenetic and biochemical findings suggest that the established cell lines can be used to investigate the disease progression mechanisms and develop novel therapeutics for MEC