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Proteasome inhibitors (PIs) can preferentially restore bone in bone-defective lesions of patients with multiple myeloma (MM) who respond favorably to these drugs. Most prior in vitro studies on PIs used continuous exposure to low PI concentrations, although pharmacokinetic analysis in patients has shown that serum concentrations of PIs change in a pulsatile manner. In the present study, we explored the effects of pulsatile treatment with PIs on bone metabolism to simulate in vivo PI pharmacokinetics. Pulsatile treatment with bortezomib, carfilzomib, or ixazomib induced MM cell death but only marginally affected the viability of osteoclasts (OCs) with F-actin ring formation. Pulsatile PI treatment suppressed osteoclastogenesis in OC precursors and bone resorption by mature OCs. OCs robustly enhanced osteoblastogenesis in cocultures with OCs and MC3T3-E1 pre-osteoblastic cells, indicating OC-mediated coupling to osteoblastogenesis. Importantly, pulsatile PI treatment did not impair robust OC-mediated osteoblastogenesis. These results suggest that PIs might sufficiently reduce MM cell-derived osteoblastogenesis inhibitors to permit OC-driven bone formation coupling while suppressing OC differentiation and activity in good responders to PIs. OC-mediated coupling to osteoblastogenesis appears to be a predominant mechanism for preferential occurrence of bone regeneration at sites of osteoclastic bone destruction in good responders.

Multiple myeloma (MM) preferentially expands and acquires drug resistance in the bone marrow (BM). We herein examined the role of histone deacetylase 1 (HDAC1) in the constitutive activation of the master transcription factor IRF4 and the prosurvival mediator PIM2 kinase in MM cells. The knockdown or inhibition of HDAC1 by the class I HDAC inhibitor MS-275 reduced the basal expression of IRF4 and PIM2 in MM cells. Mechanistically, the inhibition of HDAC1 decreased IRF4 transcription through histone hyperacetylation and inhibiting the recruitment of RNA polymerase II at the IRF4 locus, thereby reducing IRF4-targeting genes, including PIM2. In addition to the transcriptional regulation of PIM2 by the HDAC1-IRF4 axis, PIM2 was markedly upregulated by external stimuli from BM stromal cells and interleukin-6 (IL-6). Upregulated PIM2 contributed to the attenuation of the cytotoxic effects of MS-275. Class I HDAC and PIM kinase inhibitors cooperatively suppressed MM cell growth in the presence of IL-6 and in vivo. Therefore, the present results demonstrate the potential of the simultaneous targeting of the intrinsic HDAC1-IRF4 axis plus externally activated PIM2 as an efficient therapeutic option for MM fostered in the BM.

The aim of this study is to investigate the association between chemotherapy-induced fever with neutropenia less than 1500/μL (FwN) and oral bacteria and/or oral immunity in patients with hematological cancer. Thirty-two patients with hematological cancer were enrolled in the study. Secretory immunoglobulin A (sIgA) in saliva and the anaerobic bacteria in tongue coating of each subject were assessed before the first chemotherapy. Eleven subjects had an onset of FwN and 21 subjects did not during the observation periods. It was revealed by the Cox-proportional hazard model analysis that the levels of sIgA were low (HR 0.98, p

Background We are now in the new era of immunotherapies against multiple myeloma (MM) such as anti-CD38 and anti-SLAMF7 therapeutic monoclonal antibodies (mAbs); however, MM progression falls into attenuation and exhaustion of effector cells via immune checkpoint axes such as PD-1/PD-L1 and TIGIT/CD155, leading to hamper the clinical efficacy of these therapeutic mAbs. Methods To overcome this issue, we have been studying the utilization of Vδ2 γδT cells in peripheral blood mononuclear cells, which can be expanded and induced as Th1-like phenotype ex vivo using aminobisphosphonates in combination with IL-2, as effectors for MM treatment. The present study was aimed to develop the novel therapeutic strategies of γδT cells in combination with anti-MM mAbs. Results The next generation IMiDs, cereblon E3 ligase modulators (CELMoDs) of iberdomide and CC-92480 have been developed. CELMoDs was able to expand and activate γδT cells at much lower dose compared to IMiDs of lenalidomide and pomalidomide, in combination with zoledronic acid (ZA) ex vivo. Since the expanded Th1-like γδT cells highly expressed CD16, FcγRIIIa without expression of checkpoint molecules such as PD-1, TIGIT, CTLA-4, and LAG-3, we next examined whether anti-MM mAbs, elotuzumab (ELO), daratumumab (DARA), and isatuximab (ISA), induced ADCC in the presence of the γδT cells. DARA and ISA did not induce cytotoxicity against CD38-expressing MM cells in the presence of the γδT cells; however, addition of ELO further increased cytotoxicity of the γδT cells against SLAMF7-expressing MM.1S and OPM-2 cells but not RPMI 8226 cells with marginal SLAMF7 expression, suggesting induction of ELO's ADCC with the γδT cells. We next examined the cytotoxic effect of the combinatory treatment on osteoclasts (OCs) because MM cells resides in the bone marrow and the interaction between MM cells and OCs forms vicious cycle of MM cell growth and bone destruction. Of note, OCs, which can be induced from monocytes using M-CSF and RANK ligand, highly expressed SLAMF7 along with elevation of NFATc1 transcription factor. Although the expanded γδT cells can target OCs, addition of ELO augmented the cytotoxic effect of γδT cells on OCs, suggesting the emergence of ELO's ADCC with Th1-like γδT cells towards OCs. As γδT cells recognize not only neoantigens but also phosphoantigens (pAgs), we treated MM cells and OCs with ZA to enhance the expression of pAgs, and γδT cells then incubated with the ZA-treated cells. ZA-stimulation augmented the cytotoxicity of γδT cells towards MM cells and OCs even in the presence of ELO. Conclusions Taken together, our findings demonstrated the effective usage of ELO and Th1-like γδT cells with ZA against MM cells and ambient cells in MM bone marrow. Further study is warranted on the effective revitalization and expansion of Th1-like γδT cells using CELMoDs.

Background Although liver is often involved in systemic AL amyloidosis, there are few modalities to differentiate amyloidosis from common hepatic disorders with hepatomegaly and serum ALP elevation. Because liver biopsy is an invasive procedure that may cause complications such as bleeding, a non-invasive imaging modality is wanted to diagnose and assess hepatic AL amyloidosis. Ultrasound shear wave elastography (SWE) is a novel imaging modality to evaluate tissue elasticity, which is currently applied to the quantitative assessment of liver stiffness in patients with diffuse liver disease. Herein, we investigated the efficacy of ultrasound SWE for diagnosis of hepatic involvement and assessment of hepatic organ response in patients with AL amyloidosis. Methods Thirteen patients with systemic AL amyloidosis (6 males and 7 females) with a median age of 65 years old (51-84) were studied. Hepatic involvement of AL amyloidosis was observed in 5 out of 13 patients without liver complications to affect liver stiffness. The ultrasound SWE was carried out for the right lobe of the liver, using an M-probe placed on the intercostal space. The long diagonal liver span was 18.24 ± 2.76 vs 13.11 ± 1.15 cm (p=0.003); serum ALP (normal range: 106 - 322 U/L) was 820.0 ± 537.62 vs 233.25 ± 30.52 U/L (p=0.013); and the shear wave velocity in a region of interest in SWE images was 2.02 ± 0.22 vs 1.33 ± 0.10 m/sec (p=0.001) in patients with and without hepatic involvement of AL amyloidosis, respectively. Results The shear wave velocity corresponded well to the severity of hepatic amyloid involvement judged by the liver size and serum ALP levels. The shear wave velocity was decreased after attaining hepatic organ response in 2 patients, while unchanged in those without hepatic response. Conclusions These results suggest that non-invasive ultrasound SWE is instrumental in diagnosis of hepatic involvement and assessment of hepatic organ response in AL amyloidosis, and provides unique information on amyloid deposition in the liver.

Although a therapeutic outcome with extended survival has been improved in patients with multiple myeloma (MM) owing to implementation new anti-MM agents, MM eventually relapses and still remains incurable. MM-initiating cells or MM progenitors are generally accepted to contribute to disease relapse through their drug-resistant nature. In order to overcome this drug resistance, various forms of immunotherapies have been studied. One such approach is the application of γδT cells. Vγ9Vδ2γδT (Vδ2 γδT) cells are important effectors in the first-line defense and tumor surveillance, and can be activated and expanded from peripheral blood mononuclear cells (PBMCs) upon treatment with zoledronic acid (Zol) and IL-2. New generation IMiDs, namely lenalidomide (LEN) and pomalidomide (POM), not only exert direct anti-MM actions, but also activate immune cells surrounding MM cells. In the present study, we aimed to clarify the effects of new generation IMiDs, LEN and POM, on activation and expansion of Vδ2 γδT cells in combination with Zol and the cytotoxic effects of ex-vivo expanded Vδ2 γδT cells against MM progenitors. After PBMCs isolated from normal donors were incubated for one week with Zol at 1 microM and LEN at 1 microM in combination, Vδ2 γδT cells were robustly expanded to be a majority of CD3-positive T cells as observed with Zol at 1 microM and IL-2 at 100 U/ml in combination. The expanded Vδ2 γδT cells expressed intracellular IFN-γ but not Foxp3 along with increased surface expression of NKG2D and DNAM-1, molecules known to be associated with NK or cytotoxic T cell-mediated cytotoxic activity, indicating induction of Th1-like Vδ2 γδT cells. In the same way as LEN, POM at 0.01, 0.1 and 1 microM was able to expand Vδ2 γδT cells from PBMCs isolated from all 10 normal donors. The expansion efficiency was slightly better with POM over LEN in some donors. The expansion efficiency with Zol plus these IMiDs appeared to be weaker in PBMCs from a majority of patients with MM than those from normal donors, as previously reported with Zol plus IL-2. Intriguingly, these IMiDs further expanded upon treatment with Zol plus IL-2 Vδ2 γδT cells from PBMCs from some donors. However, these IMiDs alone only marginally expanded Vδ2 γδT cells in the absence of Zol, suggesting costimulation of antigen-primed Vδ2 γδT cells by these IMiDs. Thus, LEN or POM appears to be able to expand Th1-like Vδ2 γδT cells in the presence of Zolwithout exogenous addition of IL-2. Such expanded Th1-like Vδ2 γδT cells exerted potent cytotoxic activity against MM cells. Interestingly, the expanded Vδ2 γδT cells also markedly minimized the sizes of side populations in RPMI8226 and KMS-11 cells, and suppressed their clonogenic capacity as determined by in vitro colony formation and tumorigenic capacity in SCID mice, suggesting targeting a drug-resistant clonogenic MM cells. Panobinostat, a pan-HDAC inhibitor, upregulated surface expression of NKG2D ligands, ULBPs and MICA/B, on MM cells, while impairing MM cells. Pretreatment with panobinostat may further potentiate the cytotoxic activity of Vδ2 γδT cells against MM cells. Combination with novel anti-MM agents warrants further study in terms of further enhancement of anti-MM effects with the ex-vivo expanded Th1-like Vδ2 γδT cells. The precise underlying mechanisms for ex-vivo expansion of Th1-like Vδ2 γδT cells should be clarified to make the best use of these new generation IMiDs with this unique and interesting phenomenon. Disclosures No relevant conflicts of interest to declare.