Multiple myeloma (MM), a malignant plasma cell disorder, is still an incurable disease. Thus, the identification of novel therapeutic targets is of utmost importance. Here, we evaluated the ...peripheral blood-based metabolic profile of patients with MM.
Peripheral blood plasma levels of 188 endogenous metabolites, including amino acids, biogenic amines, acylcarnitines, glycerophospholipids, sphingomyelins, and hexoses were determined in patients with plasma cell dyscrasias: monoclonal gammopathy of undetermined significance, a precursor stage of MM (MGUS, n = 15), newly diagnosed MM, (NDMM, n = 32), relapsed/refractory MM (RRMM, n = 19) and in 25 healthy controls by mass spectrometry.
Patients with NDMM, RRMM and MGUS have a substantially different metabolomic profile than healthy controls. The amount of eight plasma metabolites significantly differs between the NDMM and MGUS group: free carnitine, acetylcarnitine, glutamate, asymmetric dimethylarginine (ADMA) and four phosphatidylcholine (PC) species. In addition, the levels of octadecanoylcarnitine, ADMA and six PCs were significantly different between RRMM and MGUS patients. 13 different concentrations of metabolites were found between RRMM and NDMM patients (free carnitine, acetylcarnitine, creatinine, five LysoPCs and PCs). Pathway analyses revealed a distinct metabolic profile with significant alterations in amino acid, lipid, and energy metabolism in healthy volunteers compared to MGUS/MM patients.
We identified different metabolic profiles in MGUS und MM patients in comparison to healthy controls. Thus, different metabolic processes, potentially the immunoregulation by indoleamine 2,3 dioxygenase-1 (IDO), which is involved in cancer development and progression supporting inflammatory processes in the tumor microenvironment and glutaminolysis, can serve as novel promising therapeutic targets in MM.
Antiangiogenic effects of the proteasome inhibitor bortezomib were analyzed on tumor xenografts in vivo. Bortezomib strongly inhibited angiogenesis and vascularization in the chicken chorioallantoic ...membrane. Bortezomib's inhibitory effects on chorioallantoic membrane vascularization were abrogated in the presence of distinct tumor xenografts, thanks to a soluble factor secreted by tumor cells. Through size-exclusion and ion-exchange chromatography as well as mass spectroscopy, we identified GRP-78, a chaperone protein of the unfolded protein response, as being responsible for bortezomib resistance. Indeed, a variety of bortezomib-resistant solid tumor cell lines (PC-3, HRT-18), but not myeloma cell lines (U266, OPM-2), were able to secrete high amounts of GRP-78. Recombinant GRP-78 conferred bortezomib resistance to endothelial cells and OPM-2 myeloma cells. Knockdown of GRP78 gene expression in tumor cells and immunodepletion of GRP-78 protein from tumor cell supernatants restored bortezomib sensitivity. GRP-78 did not bind or complex bortezomib but induced prosurvival signals by phosphorylation of extracellular signal–related kinase and inhibited p53-mediated expression of proapoptotic Bok and Noxa proteins in endothelial cells. From our data, we conclude that distinct solid tumor cells are able to secrete GRP-78 into the tumor microenvironment, thus demonstrating a hitherto unknown mechanism of resistance to bortezomib.
Mechanisms mediating resistance against the proteasome inhibition by bortezomib (BTZ) in multiple myeloma (MM) cells are still unclear. We analyzed the activation of the unfolded protein response ...(UPR), induction of prosurvival, and apoptotic pathways after proteasome inhibition in BTZ-sensitive and -resistant cells. Thereafter, these findings from tissue culture were proofed on MM cells of BTZ-sensitive and BTZ-refractory patients.
Proteasomal and ABC transporter activities were measured in sensitive and resistant cell lines by the use of the respective substrates. TP53 gene loss and mutations were determined by cytogenetics and targeted NGS. UPR pathways, proteasome subunit levels and protein secretion were studied by Western Blot analysis, and apoptosis was determined by flow cytometry. MM cell lines were stably transfected with inducible GRP78 expression to study unfolded protein expression. Transient knock-down of GRP78 was done by RNA interference. Splicing of XBP1 and expression of GRP78 was studied by real-time PCR in CD138-enriched MM primary cells of BTZ-refractory and -sensitive patients.
BTZ-sensitive cells displayed lower basal proteasomal activities. Similar activities of all three major ABC transporter proteins were detected in BTZ-sensitive and resistant cells. Sensitive cells showed deficiencies in triggering canonical prosurvival UPR provoked by endoplasmic reticulum (ER) stress induction. BTZ treatment did not increase unfolded protein levels or induced GRP78-mediated UPR. BTZ-resistant cells and BTZ-refractory patients exhibited lower sXBP1 levels. Apoptosis of BTZ-sensitive cells was correlating with induction of p53 and NOXA. Tumor cytogenetics and NGS analysis revealed more frequent
deletions and mutations in BTZ-refractory MM patients.
We identified low sXBP1 levels and
abnormalities as factors correlating with bortezomib resistance in MM. Therefore, determination of sXBP1 levels and
status prior to BTZ treatment in MM may be beneficial to predict BTZ resistance.
Purpose: The forkhead box transcription factor FoxP3 is specifically expressed in T cells with regulatory properties (Treg). Recently,
high numbers of Treg were described to be associated with poor ...survival in different malignancies. The aim of the presented
study was determine the prognostic effect of FoxP3 mRNA expression (reflecting the tissue content of Treg) in ovarian carcinoma
and its relation with cytokines, such as IFN-γ.
Experimental Design: Total RNA was isolated from 99 ovarian carcinoma and from 14 healthy ovarian biopsies. Real-time PCR for FoxP3 was done and
correlated with IFN-γ-, CD3-, IRF-1-, SOCS-1-, HER-2-, and iNOS expression as well as patients' outcome. The mRNA data was
corroborated by FoxP3 immunohistochemistry.
Results: Quantitation of FoxP3 expression identified a patient subgroup (>81th percentile), which is characterized by a significantly
worse prognosis in terms of overall survival (27.8 versus 77.3 months, P = 0.0034) and progression-free survival (18 versus 57.5 months; P = 0.0041). FoxP3 expression correlated with IFN-γ, IRF-1, and CD3 expression. High FoxP3 expression represents an independent
prognostic factor for overall survival ( P = 0.004) and progression-free survival ( P = 0.004).
Conclusions: High expression levels of FoxP3 might represent a surrogate marker for an immunosuppressive milieu contributing to tumor
immune escape. Strategies selectively depleting Treg might improve the antitumor activity of endogenously arising tumor-reactive
T cells and immunotherapies using vaccines or antibodies.
Purpose: T cells constitutively expressing both CD4 and CD25are essential for maintenance of self-tolerance and therefore have been
referred to as regulatory T cells (Treg). Experimental tumor models ...in mice revealed that Tregs are potent inhibitors of an
antitumor immune response. The current study was designed to determine whether cancer patients exhibit an expanded Treg pool.
Experimental Design: The frequency of Tregs in the peripheral blood of 42 patients suffering from epithelial malignancies and from 34 healthy
controls was determined by flow cytometry. The immunoregulatory properties of CD4 + CD25 + and CD4 + CD25 − T cells were characterized by proliferation and suppression assays. Cocultures with natural killer (NK) cells were performed
to determine the impact of Tregs on NK-mediated cytotoxicity.
Results: Patients with epithelial malignancies show an increase of CD4 + CD25 + T cells in the peripheral blood with characteristics of Tregs, i.e. , they are CD45RA − , CTLA-4 + , and transforming growth factor β + . Notably, CD4 + T cells from cancer patients are characterized by an impaired proliferative capacity, which is restored to the extend of
CD25-depleted CD4 + T cells from control persons by prior removal of CD25 + T cells. In contrast to CD4 + CD25 − T cells, isolated CD4 + CD25 + T cells from cancer patients were anergic towards T cell receptor stimulation. In addition, CD4 + CD25 + T cells suppressed the proliferation of CD4 + CD25 − T cells. When cultured together with CD56 + NK-cells, CD4 + CD25 + T cells from cancer patients effectively inhibited NK-cell-mediated cytotoxicity.
Conclusions: Thus, we provide evidence of an increased pool of CD4 + CD25 + regulatory T cells in the peripheral blood of cancer patients with potent immunosuppressive features. These findings should
be considered for the design of immunomodulatory therapies such as dendritic cell vaccination.
Multiple myeloma (MM) is still incurable due to resistance against various therapies. Thus, the identification of biomarkers predicting progression is urgently needed. Here, we evaluated four ...biomarkers in bone marrow and peripheral blood of MM patients for their prognostic significance.
Bone marrow- and peripheral blood plasma levels of FLT3-L, soluble TIE2, endostatin, and osteoactivin were determined in patients with monoclonal gammopathy of undetermined significance (MGUS, n = 14/n = 4), patients with newly diagnosed MM (NDMM, n = 42/n = 31) and patients with relapsed/refractory MM (RRMM, n = 27/n = 16) by sandwich ELISA.
Median FLT3-L expression increased from MGUS (58.77 pg/ml in bone marrow; 80.40 pg/ml in peripheral blood) to NDMM (63.15 pg/ml in bone marrow; 85.05 pg/ml in peripheral blood) and was maximal in RRMM (122 pg/ml in bone marrow; 160.47 pg/ml in peripheral blood; NDMM vs. RRMM p<0.001). A cut-off value of FLT3-L >92 pg/ml in bone marrow and >121 pg/ml in peripheral blood was associated with relapse or refractoriness in MM patients. FLT3-L was found to be a high predictive marker for discrimination between NDMM and RRMM as well in bone marrow as in peripheral blood (AUC 0.75 in bone marrow; vs 0.84 in peripheral blood).
High levels of FLT3-L in bone marrow and peripheral blood of MM patients identify patients with progressive disease and are associated with relapse or refractoriness in MM patients. FLT3-L could be useful as a marker to identify RRMM patients and should be evaluated as target for future therapies.
In our single-center study, 357 myeloma and lymphoma patients between 2009 and 2019 were mobilized with granulocyte colony-stimulating factor (G-CSF 7.5 µg/kg bid for four days) plus a fixed dose of ...24 mg Plerixafor when indicated (Plerixafor Group,
= 187) or G-CSF alone (G-CSF Group,
= 170). The target CD34 cell yields were ≥2.0 × 10
CD34+ cells/kg in lymphoma and ≥4.0 × 10
CD34+ cells/kg in myeloma patients to enable putative second transplants in the latter. There were no significant differences in engraftment kinetics or transfusion requirements between the Plerixafor Group and the control group in the myeloma cohort, with lymphoma patients not requiring Plerixafor showing significantly faster neutrophil recovery, a trend to faster platelet recovery, and a significantly lower need for platelet transfusions, probably due to the significantly lower number of CD34-positive cells re-transfused. While in myeloma patients the outcome (overall survival, progression-free survival) following autologous stem cell transplantation (ASCT) was similar between the Plerixafor Group and the control group, hard to mobilize lymphoma patients had significantly poorer progression-free survival (47% vs. 74% at 36 months after ASCT,
= 0.003) with a trend also to poorer overall survival (71% vs. 84%). In conclusion, while there seem to be no differences in stemness capacity and long-term engraftment efficiency between the Plerixafor and the G-CSF Group in lymphoma as well as myeloma patients, poor mobilizing lymphoma patients per se constitute a high-risk population with a poorer outcome after ASCT. Whether disease characteristics and/or a more intense or stem cell-toxic pre-mobilization chemo-/radiotherapy burden in this cohort are responsible for this observation remains to be shown in future studies.
Primary CNS lymphoma (PCNSL) is a highly aggressive malignant disease with a high recurrence rate and a poor prognosis. We present the case of a 71-year-old woman diagnosed with PCNSL in June 2010. ...After 3 relapses and intensive treatment with multiple chemotherapy regimens and whole-brain radiotherapy, she received off-label treatment with the Bruton tyrosine kinase inhibitor ibrutinib, responded well, achieved a complete remission, and is progression-free for now >3 years.
Invasive fungal infections (IFI) remain a leading cause of morbidity and mortality in immunocompromised patients. This retrospective single-center study analyzed incidence, treatment and outcome of ...invasive fungal infections in 1,095 patients with hematological malignancies receiving either cytoreductive chemotherapy or autologous or allogeneic hematopoietic stem cell transplantation at our institution between 1995 and 2004. IFI occurred in 167/1,095 (15%) patients with a significant increase over time (12.7% between 1995 and 2000 vs. 18.1% in the later IFI cohort,
P
= 0.0134). Fifty-four (32%) patients had proven, 70 (42%) patients had probable, and 43 (26%) patients suffered from possible IFI according to EORTC/MSG criteria. In 108/124 (87%) cases with proven or probable IFI, moulds were the causative pathogens. Both,
Aspergillus fumigatus
(
n
= 46) and
Aspergillus
terreus
(
n
= 41) were predominant. Yeast infections (
Candida
spp.) were documented in 16/124 (10%) cases with proven or probable IFI. Median overall survival of the entire IFI cohort was 7 (3–17) months. Overall survival was significantly better in patients with probable or possible IFI (37 and 38%, respectively) compared with patients with proven IFI (28%,
P
= 0.019). In 35% of patients, IFI was the principal cause of death with a significant decrease over time (44% in time cohort 1995–2000 vs. 28% in the later IFI cohort,
P
= 0.018) accompanied by an increased use of novel antifungals. By multivariate analysis, only proven IFI was significantly predictive for death (HR 1.7,
P
= 0.018). A significant decrease in fungus-related deaths was observed despite a significant increase of IFI over time, probably due to improved diagnostic and therapeutic approaches.
Summary
The meeting focused in particular on new strategies such as chimeric antigen receptor (CAR)-T cells and bispecific antibodies. Updates of clinical trials regarding induction treatment in ...transplantable and non-transplantable status were presented. Furthermore, minimal residual disease negativity (MRD) or, in other words, a status characterized by no measurable disease, using standardized multicolor-flow cytometry or next-generation sequencing techniques becomes increasingly important as an endpoint in clinical trials. A subjectively assessed overview of the current contributions to the treatment of multiple myeloma is given here.
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