Reversible Proteasome Inhibitor Research Articles

A Short Course of Standard Velcade/Dexamethasone Followed by Unlimited Weekly Maintenance Therapy Is an Effective Treatment in Relapsed/Refractory Multiple Myeloma

Background: Bortezomib (B), known as Velcade, is a reversible proteasome inhibitor approved for relapsed/refractory multiple myeloma (RRMM) patients (pts). The standard of care protocol includes eight cycles of intravenous push (IVP) injections of B and oral dexamethasone (D), which increases the toxicity. Here, we describe the results of an open-label, phase II clinical trial employing only four cycles of B/D. Methods: RRMM pts treated with at least one previous therapy qualified for the trial. Pts were treated with B 1.3 mg/m2 IVP or subcutaneous (SC) on day 1, 4, 8, and 11, followed by a 10-day rest, Q21 days for four cycles; followed by maintenance therapy with once weekly B 1.6 mg/m2 IVP or SC on day 1, 8, 15, and 22, followed by 13 days’ rest, repeated Q36 day. Pts received D 20 mg on the days of and days after B. Pts with a complete response (CR) were removed. Those with a partial response (PR) or stable disease (SD) were placed on maintenance therapy until progressive disease (PD), unacceptable toxicity, or pts’ decision to stop. Results: A total of 24 pts were enrolled. CR was observed in six pts (25%), PR in eight pts (33%), and SD in nine pts (37.5%). Moreover, 14 of the 24 pts (58.3%) had PR or better. Four pts had PD during induction. The grade 3 toxicities included fatigue (58%), sensory neuropathy (54%), and thrombocytopenia (50%); the grade 4 toxicities were thrombocytopenia (12.5%), fatigue (12.5%), and sensory neuropathy (12.5%). Conclusions: A short course of B/D, plus maintenance with B, is well tolerated in RRMM pts. Long-term maintenance with B/D could become an alternative to new agents.

Acute Pancreatitis Caused by Bortezomib: Report of one Case and Literature Review

Drug-induced pancreatitis has been rarely reported. Bortezomib is a selective and reversible proteasome inhibitor used for the treatment of patients with multiple myeloma (MM). We describe the case of a patient with multiple myeloma who developed acute pancreatitis after treatment with bortezomib, commonly used in the treatment of this disease. The patient was admitted to the hospital with symptoms of AP. Common etiological factors of AP were all excluded. The patient was later diagnosed as having bortezomib-induced pancreatitis. We reviewed the available medical literature on this topic and found eleven other similar cases.

In vivo activity of the second-generation proteasome inhibitor ixazomib against pediatric T-cell acute lymphoblastic leukemia xenografts

The overall survival rate of patients with T-cell acute lymphoblastic leukemia (T-ALL) is now 90%, although patients with relapsed T-ALL face poor prognosis. The ubiquitin-proteasome system maintains normal protein homeostasis, and aberrations in this pathway are associated with T-ALL. Here we demonstrate the in vitro and in vivo activity of ixazomib, a second-generation orally available, reversible, and selective proteasome inhibitor against pediatric T-ALL cell lines and patient-derived xenografts (PDXs) grown orthotopically in immunodeficient NOD.Cg-PrkdcscidIL2rgtm1Wjl/SzJAusb (NSG) mice. Ixazomib was highly potent in vitro, with half-maximal inhibitory concentration (IC50) values in the low nanomolar range. As a monotherapy, ixazomib significantly extended mouse event-free survival of five out of eight T-ALL PDXs in vivo.

Neurofilament light chain levels indicate acute axonal damage under bortezomib treatment

IntroductionBortezomib (BTZ) is a selective and reversible proteasome inhibitor and first line treatment for multiple myeloma (MM). One of the side effects is BTZ-induced peripheral neuropathy (BIPN). Until now there is no biomarker which can predict this side effect and its severity. Neurofilament light chain (NfL) is a neuron specific cytoskeletal protein, of which higher levels can be detected in peripheral blood in case of axon damage. In this study, we aimed to evaluate the relationship between NfL serum levels and characteristics of BIPN.MethodsWe performed a first interim analysis of a monocentric, non-randomized, observational clinical trial including 70 patients (DRKS00025422) diagnosed with MM in the inclusion period of June 2021 until March 2022. Two groups of patients—one with ongoing BTZ treatment at the time of recruiting, and one with BTZ treatment in the past—were compared to controls. NfL in serum was analyzed via the ELLA™ device.ResultsBoth patients with previous and ongoing BTZ treatment had higher serum NfL levels than controls, and patients with ongoing BTZ treatment had higher NfL levels than patients with BTZ treatment in the past. Serum NfL levels correlated with electrophysiological measures of axonal damage in the group with ongoing BTZ treatment.ConclusionElevated NfL levels indicate acute axonal damage under BTZ in MM patients.

The role of proteasome inhibitor MG132 in cisplatin resistant ovarian cancer

Platinum based combined chemotherapy have been proved to be the most effective drugs for the ovarian cancer treatment, but it is difficult to treat cisplatin resistant ovarian cancer. Carbobenzoxy-L-leucy-L-Leucyl-L-Leucinal (MG132) is a reversible tripeptide aldehyde proteasome inhibitor, the purpose of this study was to observe the effect of MG132 on cisplatin resistant ovarian cancer SKOV3 cell and OVCAR-3 cell the expression of autophagy and apoptosis related factors. The cells were divided into four groups: control, MG132, cisplatin, MG132 and cisplatin combination groups. Cell growth was detected by cell counting kit-8 (CCK-8) assay. The apoptotic rates of cells and the cell cycle were detected by a flow cytometer (FCM). The Beclin1, Light chain 3 (LC3) and Caspase3 was detected by western blotting and reverse transcription-polymerase chain reaction (RT-PCR). Detection of apoptotic bodies by 4,6-Diamidino-2-phenylindole dihydrochloride (DAPI) staining. CCK-8 assay demonstrated that cell survival rate in the combination groups was lower than monotherapy group. FCM showed that apoptotic rates in the combination groups was higher than monotherapy group (p < 0.05). Western blotting and RT-PCR detected that Beclin1, LC3 and Caspase3 in the combination group were higher than monotherapy group (p < 0.05). DAPI staining showed the production of apoptotic bodies in the combination group and MG132 group. In conclusion, MG132 can inhibit the growth of cisplatin resistant ovarian cancer SKOV3 and OVCAR-3 cells, its inhibitory effect is related to apoptosis and autophagy, and it is expected to be a synergistic antitumor effect with cisplatin.

Bortezomib advanced mechanisms of action in multiple myeloma, solid and liquid tumors along with its novel therapeutic applications.

Bortezomib (BTZ) is a first-in-class reversible and selective proteasome inhibitor. It inhibits the ubiquitin proteasome pathway that leads to the degradation of many intracellular proteins. Initially, BTZ was FDA approved for the treatment of refractory or relapsed multiple myeloma (MM) in 2003. Later, its usage was approved for patients with previously untreated MM. In 2006, BTZ was approved for the treatment of relapsed or refractory Mantle Cell Lymphoma (MCL) and, in 2014, for previously untreated MCL. BTZ has been extensively studied either alone or in combination with other drugs for the treatment of different liquid tumors especially in MM. However, limited data evaluated the efficacy and safety of using BTZ in patients with solid tumors. In this review, we will discuss the advanced and novel mechanisms of action of BTZ documented in MM, solid tumors and liquid tumors. Moreover, we will shed the light on the newly discovered pharmacological effects of BTZ in other prevalent diseases.

Subcutaneous injection of a bortezomib-loaded thermosensitive hydrogel for the treatment of multiple myeloma

Multiple myeloma (MM) is incurable as one of the most common hematologic malignancies. Bortezomib (BTZ) is a first-in-class reversible proteasome inhibitor in the clinic for the treatment of MM. BTZ was initially approved for the administrator by only the intravenous injection route in 2003. Then, subcutaneous administration was approved in the US and EU in 2012 in addition to the intravenous injection route. The rates and severity of peripheral neuropathy were significantly lower than those of intravenous injection administration. Injectable thermosensitive hydrogels are a potential drug delivery system for controlled release. But it is difficult to achieve the aim of local therapy of MM. A subcutaneous delivery system for a long time may be a potential therapy method due to the subcutaneous administration of BTZ. Here, we designed a BTZ-loaded injectable hydrogel system to sustain drug release for a period of days. It could lower the side effects, enhance the therapeutic effect, and reduce the injection time.

Phase II Trial of Ixazomib and Dexamethasone Versus Ixazomib, Dexamethasone and Lenalidomide, Randomized with NFKB2 Rearrangement. (Proteasome Inhibitor NFKB2 Rearrangement Driven Trial, PINR)

Introduction: Proteasome inhibitors (PI) and immunomodulatory drugs have become the backbone of therapy for multiple myeloma (MM). The oral boron-containing selective and reversible proteasome inhibitor ixazomib has shown to induce deep and durable responses (Kumar RK et la, Blood 2016. 128(20):2415-2422). Triplets containing ixazomib, has shown to be more efficacious than doublet regimens in the relapse setting (Moreau P, et al. N Engl J Med 2016. 374:1621-1634).However, to date, there is not companion diagnostics capable of predicting PI response. We have recently discovered that MM patients resistant to PI lack of the ankyrin (ANK) and death domain (DD) present in the 3'-end of NFKB2. Loss of NFKB2 3'end frequently resulted from a structural rearrangements. We found that NFKB2-ANK and -DD are crucial at initiating bortezomib's apoptotic signal by facilitating caspase-8 activation (unpublished data). Based on this findings, we designed this study to examine the efficacy of NFKB2 break apart FISH to predict the response to ixazomib and dexamethasone (Id) vs. ixazomib, lenalidomide and dexamethasone (IRd) in early relapse MM patients. Methods: In this phase 2 biomarker-driven open-label trial, relapsed patients with <4 lines of therapy were randomized to ixazomib 4 mg weekly 3 of 4 weeks and 40 mg weekly dexamethasone vs. Id plus 25 mg of lenalidomide daily days 21/28, based on the status of NFKB2 rearrangement in plasma cells. Patients were screened for NFKB2 rearrangement detected by NFKB2 break apart FISH. Patients without NFKB2 rearrangement received Id and patients with NFKB2 rearrangement were subsequently randomized in a 1:1 fashion to receive Id or IRd. The primary endpoint is to compare the response rate (MR or better) of Id or IRd at 4 cycles according to the rearrangement status of NFKB2 Results: At the moment of the interim analysis, 69 out of 71 patients have achieved 4 cycles of treatment. All treatment groups (NFKB2 FISH [-] Id, n=27, NFKB2 FISH [+] Id, n=21 and IRd, n=21) received a median of 2 prior lines of therapy. A trend to higher ORR was observed in NFKB2 FISH negative treated with Id compared with NFKB2 FISH positive (59% CI:43.6%-74.4% vs. 42% CI:17.5%-67.3%, P=0.2, target p-value goal 0.15), including significantly higher rates of ≥ very good partial response, ≥ partial response, ≥ minimal response (15%, 25%, 15% vs. 4.7%, 23.8%, 14.2%, respectively). ORR for IRd arm was higher the patients with an NFKB2 positive FISH treated with Id (65%:CI:48.5%-81.5% vs. 42% CI:17.5%-67.3%, P=0.15). Progression free survival (PFS) at 12 and 24 months was higher in patients treated with Id with negative NFKB2 FISH than those with positive FISH ( 40.7 % CI:20.3-60% and 35% CI:15.6-55% vs 21% CI 7-41%, p-value 0.012 and 35% CI:15.6-55% vs 10.7% CI: 1.8- 28.7%, p-value:0.002, respectively). Treatment with IRd in patients with positive NFKB2 FISH demonstrated higher PFS at 12 months compared to those treated with Id (46% CI: 23-67% vs 21% CI 7-41%, p-value of 0.004). However, this difference disappeared after 18 months. Interestingly, >G2 treatment related adverse events were higher in Arm A (66%) and Arm C (55%) vs Arm B (28%). The most common (≥10%) ≥ grade 3 include URI/pneumonia 15% and diarrhea 22% in Id NFKB2 FISH negative vs. URI 10% (10%) in the Id NFKB2 FISH positive arm. Treatment discontinuations only occurred in 3 NFKB2 FISH positive Id treated patient (13%). Conclusion: Interim analysis demonstrates a trend higher efficacy and significantly better PFS in ixazomib with dexamethasone in MM patients with negative NFKB2 break-apart FISH compared to those with a positive test. Efficacy and toxicity of the triplet regimen are somewhat lower than what is seen in the Tourmaline 1 trial. This study is registered clinicaltrials.gov# NCT02765854

Prolonged proteasome inhibition antagonizes TGFβ1-dependent signalling by promoting the lysosomal-targeting of TGFβ receptors

Impairing autophagy disrupts transforming growth factor beta 1 (TGFβ1) signalling and epithelial-mesenchymal transition (EMT) in non-small cell lung cancer (NSCLC). Since autophagy and proteasome-mediated degradation are interdependent, we investigated how prolonged downregulation of proteasomal catalytic activity affected TGFβ1-dependent signalling and EMT. Proteasome-dependent degradation was inhibited in A549 and H1299 NSCLC cells using MG132 and lactacystin, which are reversible and irreversible proteasome inhibitors, respectively. We observed that inhibiting proteasomal activity for 24 h decreased TGFβ-dependent nuclear accumulation of Smad2/3. Time course studies were then carried out to characterize the time frame of this observation. Short-term (< 8 h) proteasome inhibition resulted in increased receptor regulated Smad (R-Smad) phosphorylation and steady-state TGFβ receptor type II (TGFβRII) levels. However, prolonged (8–24 h) proteasome inhibition decreased TGFβ1-dependent R-Smad phosphorylation and steady-state TGFβRI and TGFβRII levels. Furthermore, proteasome inhibition blunted TGFβ-dependent E- to N-Cadherin shift, stress fiber formation, and increased cellular apoptosis via the TAK-1-TRAF6-p38 MAPK pathway. Interestingly, proteasome inhibition also increased autophagic flux, steady-state microtubule-associated protein light chain 3B-II and active uncoordinated 51-like autophagy activating kinase 1 levels, and co-localization of lysosomes with autophagy cargo proteins and autophagy-related proteins. Finally, we observed that proteasome inhibition increased TGFβRII endocytosis and trafficking to lysosomes and we conclude that prolonged proteasome inhibition disrupts TGFβ signalling outcomes through altered TGFβ receptor trafficking.

Abstract 416: A novel non-covalent and rapidly reversible proteasome inhibitor for multiple myeloma and various solid cancers

Abstract The existing proteasome inhibitors, such as bortezomib and ixazomib, are effective in multiple myeloma, but have little activity against solid tumors. These are covalent boronic acid-based compounds and are associated with undesired side effects, mainly hematologic toxicity and peripheral neuropathy. A variety of improved covalent proteasome inhibitors have been developed, but lack of oral availability and low distribution in tumors make them ineffective in solid tumors. In addition, they still have a narrow therapeutic window due to unexpected adverse effects such as cardiac and pulmonary toxicity. We report a non-covalent and rapidly reversible proteasome inhibitor as a potential anti-cancer agent against solid tumors as well as multiple myeloma. These novel inhibitors, called QL compounds, are non-covalent chymotrypsin-like selective proteasome inhibitors that can be taken orally. They effectively inhibited tumor growth in multiple myeloma xenograft model without hematologic toxicity through improved PK properties, especially partition of compound between plasma and RBC. Moreover, this improved PK properties of the QL compounds allowed for sufficient distribution outside the blood compartment and induced tumor growth inhibition in some types of solid cancer xenograft models that were sensitive to proteasome inhibition. These results indicate that non-covalent and rapidly reversible proteasome inhibitors are an ideal strategy for multiple myeloma and potential agents for solid tumors. Citation Format: Won-Gyun Ahn, Yeejin Jeon, Yeong-In Yang, Jaeseung Kim, Seung-Joo Lee, Uwe Koch, Gunther Zischinsky, Axel Choidas, Ayesha Pasha, Bert Klebl, Robert Huber, Michael Hamacher, Kiyean Nam. A novel non-covalent and rapidly reversible proteasome inhibitor for multiple myeloma and various solid cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 416.

Immune-mediated neuropathy related to bortezomib in a patient with multiple myeloma

Treatment options in multiple myeloma (MM) based on novel agents are often limited by dose-related neurotoxicity. Bortezomib, a highly active reversible proteasome inhibitor, frequently causes peripheral neuropathy (PN). Bortezomib-induced PN (BIPN) is characterized by a length-dependent, sensory, axonal polyneuropathy (PNP) with predominant small fiber-affection. Following dose reduction or drug discontinuation, BIPN resolves within 3-4 months in the majority of patients. The pathophysiological mechanisms of BIPN are unclear. Rare cases of a severe demyelinating or mixed BIPN with prominent motor involvement have been attributed to autoimmune or inflammatory reactions. A case report, including nerve pathology, is presented of a 59-year-old man with stage III IgG-κ MM who was treated with bortezomib on the occurrence of progressive disease. After the fourth cycle, he developed a painful distal symmetric sensory PNP followed by gait instability and muscle weakness increasing over 3 months despite early cessation of bortezomib. Neurological examination revealed a distal flaccid tetraparesis mainly of the lower limbs with sensory loss and severe ataxia, electrophysiological features of a mixed axonal-demyelinating PNP, and pathomorphological evidence of neuritis. Steroid treatment was initiated, and partial recovery of the neurological symptoms within 6 months was observed. While a neurotoxic effect may explain the initial distal sensory disturbances, the worsening of neurological dysfunction after bortezomib withdrawal and the clinical pattern with steroid-responsive muscle weakness predominantly of the legs are consistent with an immune-mediated mechanism. This is in line with the sural nerve biopsy findings. Toxic BIPN followed by an immune-mediated BIPN in the same patient has not been reported before.

Abstract P186: Preclinical evaluation of the proteasome inhibitor ixazomib (MLN2238) on nasopharyngeal cacinoma (NPC)

Abstract Background: Nasopharyngeal carcinoma (NPC) is a common cancer in the South Asia and is closely associated with EBV infection. The standard treatment for newly diagnosed NPC is radiotherapy alone or in combination with adjuvant chemotherapy and there is currently no targeted therapy available for NPC. About 30% of the NPC patient will progress within 5 years after treatment, so study on new therapeutic agents is in need. Proteasome inhibitor is emerged as an effective antitumor therapeutic agent in both haematological and solid tumour. Studies have reported that proteasome inhibitor can inhibit nuclear factor-kB (NF-kB) signalling pathway which was found constitutively activated in NPC. Here, we examined the antiproliferation effect and molecular mechanisms of the second-generation, selective and reversible proteasome inhibitor ixazomib (MLN2238) on NPC in-vitro and in-vivo. Ixazomib is the first oral proteasome inhibitor approved by the FDA for multiple myeloma treatment and has shown less side effect and more selective to proteasome than the first-generation proteasome inhibitor, bortezomib. Method: Three human EBV positive NPC cell lines (C666-1, NPC43 and C17) and two PDXs (Xeno113 and Xeno76) PDXs were used. Results: In our study, we showed that Ixazomib can inhibit cell viability and proliferation in EBV-associated NPC cells C666-1, NPC43 and C17 in a time- and dose-dependent manner. The IC50 of 72h treatment of ixazomib in NPC cells is 11-40nM which is more sensitive than other solid tumour, e.g. hepatocellular carcinoma (260-428nM) and breast cancer (43-1007nM). Induction of apoptosis by ixazomib was confirmed by flow cytometry analysis of Annexin V staining and western blotting of caspase-3/9 activation and PARP cleavage. S and G2/M cell cycle arrest was observed after Ixazomib treatment, which is demonstrated by flow cytometry analysis of PI staining. Evaluation of the expression level of EBV lytic genes by RT-qPCR and western blot showed that Ixazomib associates with EBV lytic cycle activation in NPC. Consistent with the previous studies, inhibition of canonical NF-kB signalling pathway was seen after 12h of ixazomib treatment in NPC cells. In animal study, 4 or 8mg/kg of ixazomib was administered orally 2 times per week for 3 weeks on NPC cells/PDX transplanted BALD/c-nu/nu and NOD/SCID mice and all showed statistical significant inhibition on NPC growth. Combination treatment of ixazomib and cisplatin showed additive effect in both in-vitro and in-vivo studies. Conclusions: Ixazomib is a potential therapeutic agent for NPC treatment and the preclinical data supports further evaluation and clinical trial of ixazomib for NPC. Citation Format: Ka Yee Li, Victor Ho Fun Lee, George Sai Wah Tsao. Preclinical evaluation of the proteasome inhibitor ixazomib (MLN2238) on nasopharyngeal cacinoma (NPC) [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P186.

The Combination of Bortezomib with Cyclophosphamide, Epirubicin, Etoposide and Prednisone (BCHEP) Regimen As First-Line Treatment for Untreated Patients with Peripheral T Cell Lymphoma: A Prospective, Single Arm, Phase II Study

Background: Peripheral T-cell lymphoma (PTCL) is a clinically and biologically heterogeneous disease with poor prognosis. The response rate of standard CHOP regimen (cyclophosphamide, doxorubicin, vincristine, and prednisolone) is only 50-60%, with a poor long-term survival rate of 10-30%. The addition of etoposide to CHOP increases response rate, but not progression free survival (PFS) or overall survival (OS). Recent study reported that nuclear factor kappa B (NF-κB) pathway plays a critical role in PTCL. Bortezomib, a potent and reversible proteasome inhibitor, can induce tumor cell apoptosis by inhibiting activation of NF-κB pathway and has been recommended as single agent option in relapse/refractory PTCL. We aimed to study the efficacy and safety of bortezomib in combination with cyclophosphamide, doxorubicin, etoposide, and prednisone (BCHEP) in newly diagnosed PTCL patients for the first time.METHODS: A prospective, single arm, phase 2 study was conducted (NCT04061772). This is an interim analysis. Patients with newly diagnosis of PTCL were treated with up to 6 cycles of BCHEP regimen every 3 weeks. Bortezomib was subcutaneously administered on Days 1 and 8 at a dose of 1.3 mg/m 2 in combination with CHEP， consisting of 100 mg/m 2 etoposide on Days 1 to 3, 750mg/m 2 cyclophosphamide on Day 1, 75mg/m 2 epirubicin on Day 1 and 100mg prednisone on Days 1 to 5. The primary endpoint of the study was ORR including complete response (CR) and partial response (PR). The secondary endpoints included progression free survival (PFS), overall survival (OS) and adverse events (AEs).RESULTS: Between February 2019 and January 2021, a total of twenty-six patients were enrolled. Median age was 57 years (range 37-69) and six (23.1%) were female. Pathological subtypes included ALK-positive anaplastic cell lymphoma (ALCL, n=2), ALK-negative ALCL (n=4), PTCL, not otherwise specified (PTCL-NOS, n=9) and angioimmunoblastic T-cell lymphoma (AITL, n=11). Nineteen patients had stage III/IV disease and eleven had B symptoms, including weight loss in three cases and fever in eight. Ten patients had elevated serum lactate dehydrogenase (LDH), eleven had IPI score higher than 2. All patients had completed BCHEP treatment for at least two cycles and received imaging evaluation. Ten patients received prophylaxis of intrathecal chemotherapy with methotrexate at least once. Three patients received consolidated radiotherapy for metabolic residuals after chemotherapy, while one received autologous hematopoietic stem cell transplantation as consolidation treatment. This study had reached the primary end point at this interim analysis. The ORR was 92.3% (24/26) with a CR rate of 57.7% (15/26). Two patients had progression of disease within two cycles of chemotherapy. After a median follow-up of 16.3 months, twelve patients had disease progression, and six died. Median PFS was 10.9 months and 1-year PFS rate was 65.4%. Median OS was 14.6 months and 1-year OS rate was 88.5%. No patient presented with Grade 5 AE. The most frequent all-grade hematological toxicity was leucopenia (42.3%,11/26), anemia (50%,13/26) and thrombocytopenia (23.1%, 6/26). Other common toxicity included intestinal infection or pneumonia (19.2%, 5/26), Grade 1 peripheral neuropathy (15.4%, 4/26) and nausea (7.7%, 2/26). Dose reduction was performed in eight patients.CONCLUSIONS: Interim results showed that bortezomib in combination with CHEP is associated with high response rate and manageable toxicity in patients with previously untreated PTCL. The BCHEP regimen may serve as a novel first-line treatment option for patients with PTCL. The study is going on to enroll patients and updating results, including the prognostic value of serum inflammatory factors. Larger trials will be necessary to further verify the efficacy of this regimen in treatment naïve PTCL patients and to overcome relapse after remission. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Phase II Trial of Ixazomib and Dexamethasone Versus Ixazomib, Dexamethasone and Lenalidomide, Randomized with NFKB2 Rearrangement. (Proteasome Inhibitor NFKB2 Rearrangement Driven Trial, PINR)

Phase II Trial of Ixazomib and Dexamethasone Versus Ixazomib, Dexamethasone and Lenalidomide, Randomized with NFKB2 Rearrangement. (Proteasome Inhibitor NFKB2 Rearrangement Driven Trial, PINR)

Open-Label Study to Determine the Feasibility of Ixazomib As Maintenance after Allogeneic Stem Cell Transplant for Multiple Myeloma

Introduction: Allogeneic stem cell transplantation is an appropriate treatment option in patients (pts) with high-risk multiple myeloma (MM), but carries a mortality risk as a consequence of graft-versus-host disease (GVHD) and relapsed disease. Proteasome inhibitors have been shown to stimulate allogeneic immune cells to eliminate myeloma cells and at the same time reduce the incidence and severity of GVHD. The use of proteasome inhibitors after stem cell transplant can therefore prolong remissions after transplant and reduce the incidence of GVHD. Ixazomib is an orally bioavailable, reversible inhibitor of the 20S proteasome that has also shown impressive anti-myeloma activity in both the relapsed/refractory and the upfront setting. This is a Phase II, open-label, multicenter, non-randomized study to determine the feasibility of Ixazomib as maintenance after allogeneic stem cell transplant for MM. Here we present results from pts in the three treatment cohorts.Methods: High risk pts (including chromosome 17p, partial deletion (del(17p), t(4;14), t(14;16), t(14;20), plasma cell leukemia, or progression-free survival of less than 2 years after autologous stem cell transplant) who received an allogenic stem cell transplant and had no evidence of >Grade 2 GVHD or any GVHD requiring methylprednisolone were eligible to enroll between Days 45 and 120 after transplant. Pts received a weekly oral ixazomib dose on Days 1, 8, and 15 of each 28-day cycle for 6 cycles. Pts were treated at 2.3 mg (Cohort 1), 3 mg (Cohort 2), and 4 mg (Cohort 3).Results: To date, 7 pts were enrolled (3 on Cohort 1, 3 on Cohort 2, and 1 on Cohort 3); median age 49 years; 5 pts (71%) female; 4 pts (57%); International Staging System Grade ≥ 2. The median number of prior systemic regimens was 4 (range 3-8). 100% of pts had received autologous stem cell transplant. Median number of prior autologous transplants was 2 (range 1-3). Conditioning regimen prior to allogeneic stem cell transplant included fludarabine + melphalan (n = 6) and fludarabine + total body irradiation (n = 1). All pts received peripheral blood stem cells, donors included matched related donors (n=2) and unrelated donors (n=5). The median number of weeks of treatment were Cohort 1, 24 wks; Cohort 2, 16 wks; Cohort 3, 6 wks. To date, there have been no dose-limiting toxicities, 1 dose interruption on Cohort 1 due to thrombocytopenia, and 1 dose reduction on Cohort 2 due to rash. 5 pts (71%) have discontinued treatment, 3 pts (43%) due to disease progression and 2 pts (29%) have completed treatment. 2 pts (29%) were still on active treatment at the time of data cut-off. 3 pts (43%) have died due to disease progression. The median time on study overall was 9 months (range 2-28 months). Of the 6 pts (86%) eligible for response analysis, 2 pts (33%) had a stringent complete response, 1 pt (17%) had a complete response, 2 pts (33%) had a very good partial response, and 1 pt (17%) had progressive disease as a best overall response. The most common ≥ Grade 3 adverse events (AEs) were thrombocytopenia (3 pts, 43%) and hypertension (2 pts, 29%). 3 pts (43%) had ≥ Grade 3 treatment-related AEs of thrombocytopenia (3 pts, 43%) and rash (1 pts, 14%). 1 pt (14%) had treatment-related serious adverse events (vomiting and fever). Acute GVHD was reported for 5 pts (maximum Grade 1), chronic GVHD occurred in 4 pts (limited: 2 pts, extensive: 2 pts).Conclusions: There have been no dose-limiting toxicities on these 3 cohorts. The maximum-tolerated dose has not been reached for this study and enrollment is continuing. Initial response data is promising. DisclosuresMatous:Celgene: Speakers Bureau; Multiple Myeloma Advisory Committee: Membership on an entity's Board of Directors or advisory committees. Berdeja:Vivolux: Research Funding; Constellation: Research Funding; Takeda: Research Funding; Curis: Research Funding; Teva: Research Funding; Novartis: Research Funding; Janssen: Research Funding; Celgene: Research Funding; BMS: Research Funding; Bluebird: Research Funding; Amgen: Research Funding; Abbvie: Research Funding.

Proteasome inhibitor MG132 induces apoptosis in human osteosarcoma U2OS cells.

MG132 is a potent, reversible, and cell-permeable 20S proteasome inhibitor and it is derived from a Chinese medicinal plant. The purpose of this study is to investigate the anticancer effects of MG132 against human osteosarcoma U2OS cells. We first performed MTT and colony formation assays to investigate the anti-proliferative effects of MG132. The results demonstrated that MG132 suppressed the proliferation of U2OS cells. Furthermore, we found that treatment with MG132 increased apoptosis and induced DNA damage in U2OS cells. Additionally, zymography, wound healing, and invasion assays showed that MG132 suppressed the enzymatic activity of matrix metalloproteinases, cell migration, and invasion, respectively of U2OS cells. Furthermore, western blotting assay was performed to investigate the apoptotic signaling pathways in MG132-treated U2OS cells. Our results showed that MG132 downregulated the expression of antiapoptotic proteins, including CDK2, CDK4, Bcl-xL, and Bcl-2, whereas it upregulated the expression of proapoptotic proteins, including p21, p27, p53, p-p53 (ser15, ser20, and ser46), cleaved forms of caspase-3, caspase-7, caspase-9, and PARP, and FOXO3 in U2OS cells. These results demonstrated that MG132 activated apoptotic signaling pathways in U2OS cells. Interestingly, MG132 downregulated the phosphorylation of Akt and Erk. Taken together, our results suggest that MG132 has anticancer effects in U2OS cells. Therefore, MG132 may be a potential therapeutic agent for the treatment of osteosarcoma.

Separation, identification and characterization of stress degradation products of bortezomib using HPLC and LC-MS.

Bortezomib (BT) is a reversible inhibitor of proteasome which is used in the treatment of hematological cancers. To study the degradation behavior, BT was subjected to acidic, basic, neutral, photolytic, oxidative and thermal degradation conditions as per ICH guideline Q1A (R2). A gradient HPLC method has been developed for separating all the degradation products formed under various degradation conditions on Waters XBridge C18 column (150 mm × 4.6 mm × 3.5 µm) using the mobile phase composed of ammonium formate and acetonitrile. A total of six degradation products were formed in various stress conditions and these were separated identified, and characterized using high performance liquid chromatography in combination with electrospray ionization tandem mass spectrometric studies.

Ixazomib inhibits myeloma cell proliferation by targeting UBE2K

PurposeIxazomib is a selective, effective, and reversible inhibitor of 20S proteasome and is approved for the treatment of multiple myeloma. Ubiquitin-conjugating enzyme E2 (UBE2K) is involved in the synthesis of K48-linked ubiquitin chains and is the target of certain drugs used for the treatment of tumors. The purpose of this study was to investigate the relationship between ixazomib and UBE2K in myeloma cells. MethodsWe used CCK-8 and Annexin V-FITC/propidium iodide kit to detect the effects of ixazomib on survival and apoptosis of RPMI-8226 and U-266 myeloma cell lines. Quantitative polymerase chain reaction and western blot were used to detect the change in gene and protein expression levels of myeloma cells treated with ixazomib. Furthermore, the regulatory effects of ixazomib on UBE2K and its downstream targets were investigated following the overexpression of UBE2K. ResultsIn myeloma cells, ixazomib decreased cell survival and increased apoptosis in a dose-dependent manner. Ixazomib significantly increased the expression of HIST1H2BD, MNAT1, NEK3, and TARS2, while decreasing the expression of HSPA1B and UBE2K. In addition, ixazomib inhibited the proliferation of myeloma cells, blocked cell cycle, induced cell apoptosis, and increased the production of reactive oxygen species by inhibiting UBE2K expression. Lastly, ixazomib regulates mitosis- and apoptosis-related genes by lowering UBE2K expression. ConclusionIn summary, ixazomib leads to impaired proliferation of myeloma cells by targeting UBE2K.

Repurposing bortezomib for choroidal neovascularization treatment via antagonizing VEGF-A and PDGF-D mediated signaling

Neovascular age-related macular degeneration (neoAMD) is the leading cause of blindness in AMD and manifests as choroidal neovascularization (CNV). Anti-vascular endothelial growth factor (VEGF) therapies are the mainstay treatments but with limited efficacy and cause detrimental effects on the retina after long-term application. These disadvantages warrant alternative strategy. Herein, we examined the effect on CNV by intravitreal injection of bortezomib, a reversible proteasome inhibitor, and further dissected the mechanism. Krypton red Laser was used to create CNV model in mice. The angiogenesis volume was assessed in choroidal flat-mount with isolectin GS-IB4 labeling and the leakage was examined with fluorescein fundus angiography. Injection of Borsub inhibited angiogenesis in the CNV model which was dose-dependent; the injection significantly inhibited leakage as well. Furthermore, Borsub injection reduced the contents of VEGF-A, macrophage chemotactic factor 1 (MCP-1), and platelet-derived growth factor (PDGF)-D but not PDGF-B, examined by enzyme-linked immunosorbent assay, in choroid/retinal pigment epithelium (RPE) tissue. These injections also reduced phospho-VEGFR-2 and phospho-PDGFRβ in choroid/RPE tissue examined by immunoblotting. Moreover, Borsub inhibited the recruitment of mural cells or macrophages to laser-injured spots. Injection of Borsub indicated negative effect on scotopic and photopic responses recorded by electroretinogram. Altogether, intravitreal injection of Borsub significantly reduced CNV by antagonizing VEGF-A/Flk-1 and PDGF-D/PDGFRβ pathways without impacting electroretinography parameters. Thus, Borsub may offer an invaluable therapy for the prevention and treatment of neoAMD.

Stress degradation study of bortezomib: effect of co-solvent, isolation and characterization of degradation products by UHPLC-Q-TOF-MS/MS and NMR and evaluation of the toxicity of the degradation products

Bortezomib (BTZ) is a first-in-class, potent reversible inhibitor of proteasome used in the treatment of multiple myeloma, the second most common hematological cancer.