Patients with hematologic malignancies still face a significant risk of severe coronavirus disease 2019 (COVID-19). The severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2)-neutralizing monoclonal antibody combination tixagevimab/cilgavimab (TIX/CGB) could be administered to immunocompromised patients for pre-exposure prophylaxis (PrEP) before the emergence of TIX/CGB-resistant COVID-19 Omicron variants. TIX/CGB application could be carried out regardless of the host’s immune response to previous active SARS-CoV-2 vaccinations or infections. Because the efficacy of COVID-19 PrEP remains unclear, especially in SARS-CoV-2-seropositive patients, German national guidelines recommended TIX/CGB PrEP only for SARS-CoV-2-seronegative patients in addition to an intensified active vaccination schedule. Having followed these guidelines, we now report the characteristics and outcomes of 54 recipients of TIX/CGB PrEP in SARS-CoV-2-seronegative patients with hematological disease from a German tertiary medical center and compare them to 125 seropositive patients who did not receive any PrEP. While the number of patients with B-cell lymphomas was significantly higher in the seronegative cohort (33 (61%) vs. 18 (14%) cases, p < 0.01), patients with myeloid diseases were significantly more frequent in the seropositive cohort (51 (41%) vs. 5 (9%) cases, p < 0.01). Strikingly, patients who had undergone allogeneic hematopoietic stem cell transplantation were significantly more likely (forty-nine (39%) vs. six (11%) cases, p < 0.01) to be SARS-CoV-2 seropositive. We observed that prophylactic application of TIX/CGB PrEP to a highly vulnerable group of SARS-CoV-2-seronegative patients resulted in a similar number of COVID-19 breakthrough infections compared to the untreated seropositive control group (16 (32%) vs. 39 (36%), p = 0.62) and comparable COVID-19-related outcomes like hospitalization and oxygen requirement throughout an extended follow-up period of 12 months. In conclusion, our results support the tailored approach of administering TIX/CGB PrEP only to SARS-CoV-2-seronegative patients during the COVID-19 pandemic and might provide a rationale for similar strategies during future outbreaks/diseases, especially in times of initial limited availability and/or financial constraints.

Proteasome inhibition is a highly effective treatment for multiple myeloma (MM). However, virtually all patients develop proteasome inhibitor resistance, which is associated with a poor prognosis. Hyperactive small ubiquitin-like modifier (SUMO) signaling is involved in both cancer pathogenesis and cancer progression. A state of increased SUMOylation has been associated with aggressive cancer biology. We found that relapsed/refractory MM is characterized by a SUMO-high state, and high expression of the SUMO E1-activating enzyme (SAE1/UBA2) is associated with poor overall survival. Consistently, continuous treatment of MM cell lines with carfilzomib (CFZ) enhanced SUMO pathway activity. Treatment of MM cell lines with the SUMO E1-activating enzyme inhibitor subasumstat (TAK-981) showed synergy with CFZ in both CFZ-sensitive and CFZ-resistant MM cell lines, irrespective of the TP53 state. Combination therapy was effective in primary MM cells and in 2 murine MM xenograft models. Mechanistically, combination treatment with subasumstat and CFZ enhanced genotoxic and proteotoxic stress, and induced apoptosis was associated with activity of the prolyl isomerase PIN1. In summary, our findings reveal activated SUMOylation as a therapeutic target in MM and point to combined SUMO/proteasome inhibition as a novel and potent strategy for the treatment of proteasome inhibitor-resistant MM.

Deubiquitylases (DUBs) are therapeutically amenable components of the ubiquitin machinery that stabilize substrate proteins. Their inhibition can destabilize oncoproteins that may otherwise be undruggable. Here, we screened for DUB vulnerabilities in multiple myeloma, an incurable malignancy with dependency on the ubiquitin proteasome system and identified OTUD6B as an oncogene that drives the G1/S‐transition. LIN28B, a suppressor of microRNA biogenesis, is specified as a bona fide cell cycle‐specific substrate of OTUD6B. Stabilization of LIN28B drives MYC expression at G1/S, which in turn allows for rapid S‐phase entry. Silencing OTUD6B or LIN28B inhibits multiple myeloma outgrowth in vivo and high OTUD6B expression evolves in patients that progress to symptomatic multiple myeloma and results in an adverse outcome of the disease. Thus, we link proteolytic ubiquitylation with post‐transcriptional regulation and nominate OTUD6B as a potential mediator of the MGUS‐multiple myeloma transition, a central regulator of MYC, and an actionable vulnerability in multiple myeloma and other tumors with an activated OTUD6B‐LIN28B axis.

Rationale Multiple myeloma (MM) cells synthesize large amounts of paraproteins, making radiolabeled amino acids promising candidates for PET imaging of MM patients. Methods We compare tumor uptake of the two amino acid analogs [ 18 F]-fluoroethyltyrosine and [ 18 F]-FACBC in a MM xenograft model and show the feasibility of PET imaging with [ 18 F]-FACBC in a MM patient. Results Preclinically [ 18 F]-FACBC showed superior performance, mainly due to the uptake via the ASC-system. In a subsequent proof-of-concept investigation [ 18 F]-FACBC PET was performed in a MM patient. It allowed identification of both lesions with and without CT correlate (SUVmean 8.0 or 7.9) based on higher uptake compared to normal bone marrow (SUVmean 5.7). Bone signal was elevated compared to non-MM patients, and, thus [ 18 F]-FACBC potentially allows the assessment of bone marrow infiltration. Conclusion The FDA/EMA approved PET agent [ 18 F]-FACBC is promising for imaging MM and should be further evaluated in prospective clinical studies.

Background: Background: Multiple myeloma (MM) is a genetically and clinically heterogeneous neoplastic plasma cell disorder. Treatment regimens for MM include proteasome inhibitors (PIs) which are routinely combined with dexamethasone and chemotherapeutics or immunomodulatory drugs (IMiDs). While such regimens have resulted in significant improvement of disease control and survival, the development of drug resistance remains a major clinical problem. Relapsed/refractory MM (r/r MM) is associated with particularly poor prognosis. New therapeutic strategies to further improve the management of MM and to overcome drug resistance are therefore urgently needed. SUMOylation is a post-translational protein modification pathway closely related to ubiquitylation. Hyperactive SUMOylation (SUMO) signalling is involved in both cancer pathogenesis and cancer progression. A state of increased SUMOylation has been associated with aggressive cancer biology. Aims: Aims: We sought to identify stress response mechanisms associated with adverse prognosis and PI resistance. The overall goal was to develop novel therapy combinations targeting molecular vulnerabilities associated with advanced and/or PI-resistant MM. Methods: Methods: RNA-sequencing, unbiased mass spectrometry-based proteomics and bioinformatics analyses were used to interrogate the biology and in particular stress response pathways in MM. Cell culture finding were informed with primary MM patient data from gene expression studies and and survival data from clinical trials. Novel drug combination treatments were tested in MM cell-based models in vitro, in MM xenograft models in vivo, and in primary MM patient samples. Here, we found that r/r MM is characterized by a state of activated SUMOylation, a ubiquitin-related post-translational protein modification pathway. High expression of the SUMO E1 activating enzyme (SAE1/UBA2) was associated with poor overall survival. Induced resistance to the second generation PI carfilzomib (CFZ) enhanced SUMO pathway activity. Accordingly, CFZ-pretreated patients showed enhanced SUMO pathway activity in the MM compartment. Treatment of MM cell lines with subasumstat (also TAK-981), a novel small-molecule inhibitor targeting the SUMO E1 activating enzyme, showed synergistic treatment efficacy with CFZ in both PI-sensitive and PI-resistant MM cell lines irrespective of the TP53 state. Combination therapy was effective in two murine MM xenograft models, where in vivo growth was significantly inhibited, and in patient-derived primary MM cells in vitro. Mechanistically, combinatorial treatment of subasumstat and CFZ enhanced genotoxic and proteotoxic stress and apoptosis. SUMO/proteasome a the of MM.

Biomarkers that predict response to lenalidomide maintenance therapy in patients with multiple myeloma (MM) have remained elusive. We have shown that immunomodulatory drugs (IMiDs) exert anti-MM activity via destabilization of MCT1 and CD147. In this study, cell samples of 654 patients with MM who received lenalidomide (n = 455), thalidomide (n = 98), or bortezomib (n = 101) maintenance were assessed by gene expression profiling and RNA sequencing, followed by correlation of MCT1 and CD147 expression with data for progression-free survival (PFS) and overall survival (OS). Patients with high expression levels of MCT1 showed significantly reduced PFS (31.9 months vs 48.2 months in MCT1high vs MCT1low; P = .03) and OS (75.9 months vs not reached [NR] in MCT1high vs MCT1low; P = .001) in cases with lenalidomide maintenance, whereas MCT1 expression had no significant impact on PFS or OS in cases with bortezomib maintenance. We validated the predictive role of MCT1 for IMiD-based maintenance in an independent cohort of patients who received thalidomide (OS, 83.6 months vs NR in MCT1high vs MCT1low; P = .03). Functional validation showed that MCT1 overexpression in human MM cell lines significantly reduced the efficacy of lenalidomide, whereas no change was observed with bortezomib treatment, either in vitro or in a MM xenograft model. Our findings have established MCT1 expression as a predictive marker for response to lenalidomide-based maintenance in patients with MM.

BACKGROUND:Multiple myeloma is the second most common hematologic malignancy, which to date remains incurable despite advances in treatment strategies including the use of novel substances such as proteasome inhibitors, immunomodulatory drugs, and monoclonal antibodies. SUMMARY:The bone marrow-based disease is preceded by the 2 sequential premalignant conditions: monoclonal gammo-pathy of undetermined significance and smoldering myeloma. Plasma cell leukemia and extramedullary disease occur, when malignant clones lose their dependency on the bone marrow. Key genetic features of these plasma cell dyscrasias include chromosomal aberrations such as translocations and hyperdiploidy, which occur during error-prone physiologic processes in B-cell development. Next-generation sequencing studies have identified mutations in major oncogenic pathways and tumor suppressors, which contribute to the pathogenesis of multiple myeloma and have revealed insights into the clonal evolution of the disease, particularly along different lines of therapy. More recently, the importance of epigenetic alterations and the role of the bone marrow microenvironment, including immune and osteogenic cells, have become evident. Key Messages: We herein review the current knowledge of the pathogenesis of multiple myeloma, which is crucial for the development of novel targeted therapeutic strategies. These can contribute to the endeavor to make multiple myeloma a curable disease.

High-dose chemotherapy followed by autologous stem cell transplantation is a major component in the treatment of patients with multiple myeloma. As a prerequisite, the successful collection of a sufficient number of viable peripheral blood hematopoietic CD34+ cells is critical. A common standard protocol for mobilization is currently not defined and critically discussed especially in German-speaking Europe. In times of the Covid-19 pandemic, safe and effective strategies have to be chosen to minimize hospitalization times and severe courses. In this single-center retrospective analysis, safety and efficacy of cyclophosphamide plus etoposide (CE) and growth-factor support (n = 33) was compared to cyclophosphamide mono treatment and growth-factor support (n = 49) in 82 patients with multiple myeloma at first diagnosis. CE was superior to cyclophosphamide mono with a significantly higher number of collected CD34+ cells (15.46 × 106 CD34+ cells/kg vs. 9.92 × 106 CD34+ cells/kg), significantly faster engraftment of granulocytes after stem cell transplantation (day 10.5 vs. day 11.6), shorter duration of the inpatient stay (17.47 days vs. 19.16 days) and significantly less transfusions (8.82 % vs. 30.61 % patients receiving transfusions). The safety profile was comparable in both groups and in line with published data. We conclude that CE is a safe and highly effective mobilization protocol in patients with multiple myeloma at first diagnosis and appears to be superior to the commonly used cyclophosphamide mono regimen.

The complex architecture of transmembrane proteins requires quality control (QC) of folding, membrane positioning, and trafficking as prerequisites for cellular homeostasis and intercellular communication. However, it has remained unclear whether transmembrane protein-specific QC hubs exist. Here we identify cereblon (CRBN), the target of immunomodulatory drugs (IMiDs), as a co-chaperone that specifically determines chaperone activity of HSP90 toward transmembrane proteins by means of counteracting AHA1. This function is abrogated by IMiDs, which disrupt the interaction of CRBN with HSP90. Among the multiple transmembrane protein clients of CRBN-AHA1-HSP90 revealed by cell surface proteomics, we identify the amino acid transporter LAT1/CD98hc as a determinant of IMiD activity in multiple myeloma (MM) and present an Anticalin-based CD98hc radiopharmaceutical for MM radio-theranostics. These data establish the CRBNAHA1-HSP90 axis in the biogenesis of transmembrane proteins, link IMiD activity to tumor metabolism, and nominate CD98hc and LAT1 as attractive diagnostic and therapeutic targets in MM.

Hypomethylating agents (HMAs) are widely used in patients with higher-risk myelodysplastic syndromes (MDS) not eligible for stem cell transplantation; however, the response rate is <50%. Reliable predictors of response are still missing, and it is a major challenge to develop new treatment strategies. One current approach is the combination of azacytidine (AZA) with checkpoint inhibitors; however, the potential benefit of targeting the immunomodulator indoleamine-2,3-dioxygenase (IDO-1) has not yet been evaluated. We observed moderate to strong IDO-1 expression in 37% of patients with high-risk MDS. IDO-1 positivity was predictive of treatment failure and shorter overall survival. Moreover, IDO-1 positivity correlated inversely with the number of infiltrating CD8+ T cells, and IDO-1+ patients failed to show an increase in CD8+ T cells under AZA treatment. In vitro experiments confirmed tryptophan catabolism and depletion of CD8+ T cells in IDO-1+ MDS, suggesting that IDO-1 expression induces an immunosuppressive microenvironment in MDS, thereby leading to treatment failure under AZA treatment. In conclusion, IDO-1 is expressed in more than one-third of patients with higher-risk MDS, and is predictive of treatment failure and shorter overall survival. Therefore, IDO-1 is emerging as a promising predictor and therapeutic target, especially for combination therapies with HMAs or checkpoint inhibitors.

Ziel/Aim Increased amino acid supply and dysregulated integrin signaling constitute two hallmarks of tumor biology and are pivotal for metastatic transformation of cells. In line with its involvement in both pathophysiological processes, overexpression of the cluster of differentiation 98 heavy chain (CD98hc) is clinically observed in various cancers, rendering it a promising tumor target. Hence, development of a Zr-89-ImmunoPET radiopharmaceutical that targets corresponding heterodimeric amino acid transporters holds promise for deeper insight into tumor physiology and as a potential tool for in vivo diagnostics.

Introduction: Cereblon (CRBN) is the target for immunomodulatory drugs (IMiDs) such as thalidomide and its derivatives lenalidomide and pomalidomide, which are key therapeutics for hematologic malignancies such as multiple myeloma (MM) and del(5q) myelodysplastic syndrome (MDS). We have previously described a ubiquitin-independent chaperone-like function of CRBN, which stabilizes the transmembrane proteins CD147 and MCT1. IMiDs interfere with this chaperone-like function of CRBN in a competitive manner to mediate both their anti-tumor and their teratotoxic effects (Eichner et al. Nature Medicine 2016). So far, the underlying mechanisms of transmembrane protein maturation, the global impact of CRBN on the cell surface proteome and the precise molecular mechanism of IMiDs, especially their clinically well-established synergy with proteasomal inhibitors remain unclear. Methods: Novel CRBN-clients were identified by cross-validation of the CRBN-interactome with a cell surface proteomic screen. Various molecular and cell biological methods including immunofluorescence, flow cytometry, immunoprecipitations, GST-pulldowns, amino acid transport and proliferation assays were used to decipher underlying mechanisms. In vitro assays and in vivo xenograft experiments were performed using MM cell lines and patient-derived CD138+ MM cells. 18FDG- and 18FET-PET was used for imaging of xenografted tumors. Results: Our unbiased screening approaches imply a global role of CRBN in transmembrane protein maturation. In particular, we identify the amino acid transporter LAT1 and its functional subunit CD98hc as novel CRBN client proteins, which are frequently overexpressed in MM to drive cell proliferation. CD98hc/LAT1 become destabilized and inactivated upon IMiD treatment, which attenuates MM cell proliferation, tumor formation and perturbs the uptake of essential amino acids, thereby further linking IMiD-activity to tumor-metabolism. CD98hc/LAT1 destabilization only occurs in IMiD-sensitive patients and cell lines, thus being a potential biomarker to predict IMiD-response. Moreover, inhibition of LAT1 is cytotoxic in both IMiD-sensitive and -resistant cells, which makes it an attractive therapeutic option for IMiD-resistant and -refractory patients. Mechanistically, we show CRBN to function as a new selective co-chaperone of HSP90, which facilitates transmembrane protein maturation in a ubiquitin-independent way, which is impaired by IMiD-treatment. Conclusion: We establish CRBN as a transmembrane protein-specific co-chaperone for HSP90 and identify modulation of the CRBN-CD98hc/LAT1 axis as crucial means by which IMiDs mediate their anti-tumor activity. Notably, we specify CD98hc/LAT1 as valuable biomarkers for IMiD-response and druggable targets for IMiD-resistant and -refractory MM patients and beyond. Furthermore, this ubiquitin-independent mechanism solves the paradox regarding the well documented synergistic anti-myeloma activity of IMiDs and proteasomal inhibitors. Disclosures Götze: AbbVie: Membership on an entity's Board of Directors or advisory committees. Bassermann:Celgene: Consultancy, Research Funding.

Immunomodulatory drugs (IMiDs), such as thalidomide and its derivatives lenalidomide and pomalidomide, are key treatment modalities for hematologic malignancies, particularly multiple myeloma (MM) and del(5q) myelodysplastic syndrome (MDS). Cereblon (CRBN), a substrate receptor of the CRL4 ubiquitin ligase complex, is the primary target by which IMiDs mediate anticancer and teratogenic effects. Here we identify a ubiquitin-independent physiological chaperone-like function of CRBN that promotes maturation of the basigin (BSG; also known as CD147) and solute carrier family 16 member 1 (SLC16A1; also known as MCT1) proteins. This process allows for the formation and activation of the CD147-MCT1 transmembrane complex, which promotes various biological functions, including angiogenesis, proliferation, invasion and lactate export. We found that IMiDs outcompete CRBN for binding to CD147 and MCT1, leading to destabilization of the CD147-MCT1 complex. Accordingly, IMiD-sensitive MM cells lose CD147 and MCT1 expression after being exposed to IMiDs, whereas IMiD-resistant cells retain their expression. Furthermore, del(5q) MDS cells have elevated CD147 expression, which is attenuated after IMiD treatment. Finally, we show that BSG (CD147) knockdown phenocopies the teratogenic effects of thalidomide exposure in zebrafish. These findings provide a common mechanistic framework to explain both the teratogenic and pleiotropic antitumor effects of IMiDs.