Immunomodulatory Drugs Lenalidomide Research Articles

Real-world treatment patterns and outcomes in relapsed/refractory multiple myeloma (1-3 prior lines): Flatiron database

AbstractIn the context of multiple myeloma (MM), early use of the immunomodulatory drug lenalidomide has led to an increased population of patients with lenalidomide-refractory MM in early-line settings, but their outcomes are not well characterized. Herein, we report treatment patterns, survival outcomes, prognostic variables, and attrition rates for patients with proteasome inhibitor–exposed, lenalidomide-refractory MM, treated with 1 to 3 prior lines of therapy (LOT). From 12 767 patients with MM in the Flatiron Health database between January 2016 and April 2022, 1455 met the inclusion criteria. The most common subsequent treatments were triplet combinations (41.6% of patients); daratumumab/pomalidomide/dexamethasone was the most common treatment regimen (13.2%). Median real-world progression-free survival (RW-PFS) and overall survival (OS) were 6.5 months and 44.4 months, respectively. RW-PFS was similar in patients with 1, 2, or 3 prior LOT. International Staging System stage III, Eastern Cooperative Oncology Group performance status of 1, hemoglobin <12 g/dL, high-risk cytogenetics, and refractoriness to anti-CD38 antibody at baseline were associated with worse RW-PFS and OS. Outcomes remained similar for patients who received National Comprehensive Cancer Network–preferred treatments and those who received treatments after 2020. In 561 patients with 1 prior LOT at inclusion, the cumulative attrition rate from LOT 2 to 5 was 85%, which included 25% patients who died and 60% with no further treatment. Patients with lenalidomide-refractory MM who have received 1 to 3 prior LOT have poor outcomes and progress rapidly through available therapies, highlighting the need for more effective treatments early in the disease course, before patients are lost to attrition.

Mantle Cell Lymphoma- A Rare Occurrence: Case Report

Mantle Cell Lymphoma (MCL) is a relatively uncommon yet distinct type of malignant lymphoma whose clinical and pathological characterization has been limited by the small numbers of cases published to date. It comprises 3-7% of all Hodgkin’s lymphoma cases, with a median diagnosis age of 68 years. Based on immunophenotypic and molecular data, MCL is a neoplasm composed of cells resembling those residing in follicular mantle zones. Although MCL usually presents as advanced disease some patients may present with more limited (stages I-I) disease. We present the case of a 41-year-old male who exhibited generalized lymphadenopathy and hepatosplenomegaly. Peripheral smear showed absolute lymphocytosis and the bone marrow biopsy and flow cytometry identified the condition as mantle cell lymphoma. The patient’s management required a multidisciplinary approach, incorporating haematological, molecular, and genetic evaluations. Treatment included hyper-CVAD chemotherapy. Proteasome inhibitors (Bortezomib), mTOR inhibitors (Temsirolimus), and immunomodulatory drugs (Lenalidomide) have recently been added to the treatment options in MCL.This case underscores the significance of comprehensive evaluation and customized therapeutic strategies and re looking an old diagnosis in managing complex haematological disorders.

Immunomodulatory imide drugs inhibit human detrusor smooth muscle contraction and growth of human detrusor smooth muscle cells, and exhibit vaso-regulatory functions

BackgroundThe immunomodulatory imide drugs (IMiDs) thalidomide, lenalidomide and pomalidomide may exhibit therapeutic efficacy in the prostate. In lower urinary tract symptoms (LUTS), voiding and storage disorders may arise from benign prostate hyperplasia, or overactive bladder. While current therapeutic options target smooth muscle contraction or cell proliferation, side effects are mostly cardiovascular. Therefore, we investigated effects of IMiDs on human detrusor and porcine artery smooth muscle contraction, and growth-related functions in detrusor smooth muscle cells (HBdSMC). MethodsCell viability was assessed by CCK8, and apoptosis and cell death by flow cytometry in cultured HBdSMC. Contractions of human detrusor tissues and porcine interlobar and coronary arteries were induced by contractile agonists, or electric field stimulation (EFS) in the presence or absence of an IMID using an organ bath. Proliferation was assessed by EdU assay and colony formation, cytoskeletal organization by phalloidin staining, ResultsDepending on tissue type, IMiDs inhibited cholinergic contractions with varying degree, up to 50 %, while non-cholinergic contractions were inhibited up to 80 % and 60 % for U46619 and endothelin-1, respectively, and EFS-induced contractions up to 75 %. IMiDs reduced viable HBdSM cells in a time-dependent manner. Correspondingly, proliferation was reduced, without showing pro-apoptotic effects. In parallel, IMiDs induced cytoskeletal disorganization. ConclusionsIMiDs exhibit regulatory functions in various smooth muscle-rich tissues, and of cell proliferation in the lower urinary tract. This points to a novel drug class effect for IMiDs, in which the molecular mechanisms of action of IMiDs merit further consideration for the application in LUTS.

Multicolumn Nanoflow Liquid Chromatography with Accelerated Offline Gradient Generation for Robust and Sensitive Single-Cell Proteome Profiling.

Peptide separations that combine high sensitivity, robustness, peak capacity, and throughput are essential for extending bottom-up proteomics to smaller samples including single cells. To this end, we have developed a multicolumn nanoLC system with offline gradient generation. One binary pump generates gradients in an accelerated fashion to support multiple analytical columns, and a single trap column interfaces with all analytical columns to reduce required maintenance and simplify troubleshooting. A high degree of parallelization is possible, as one sample undergoes separation while the next sample plus its corresponding mobile phase gradient are transferred into the storage loop and a third sample is loaded into a sample loop. Selective offline elution from the trap column into the sample loop prevents salts and hydrophobic species from entering the analytical column, thus greatly enhancing column lifetime and system robustness. With this design, samples can be analyzed as fast as every 20 min at a flow rate of just 40 nL/min with close to 100% MS utilization time and continuously for as long as several months without column replacement. We utilized the system to analyze the proteomes of single cells from a multiple myeloma cell line upon treatment with the immunomodulatory imide drug lenalidomide.

Differential Analysis of Cereblon Neosubstrates in Rabbit Embryos Using Targeted Proteomics

Targeted protein degradation is the selective removal of a protein of interest through hijacking intracellular protein cleanup machinery. This rapidly growing field currently relies heavily on the use of the E3 ligase cereblon (CRBN) to target proteins for degradation, including the immunomodulatory drugs (IMiDs) thalidomide, lenalidomide, and pomalidomide which work through a molecular glue mechanism of action with CRBN. While CRBN recruitment can result in degradation of a specific protein of interest (e.g., efficacy), degradation of other proteins (called CRBN neosubstrates) also occurs. Degradation of one or more of these CRBN neosubstrates is believed to play an important role in thalidomide-related developmental toxicity observed in rabbits and primates. We identified a set of 25 proteins of interest associated with CRBN-related protein homeostasis and/or embryo/fetal development. We developed a targeted assay for these proteins combining peptide immunoaffinity enrichment and high-resolution mass spectrometry and successfully applied this assay to rabbit embryo samples from pregnant rabbits dosed with three IMiDs. We confirmed previously reported in vivo decreases in neosubstrates like SALL4, as well as provided evidence of neosubstrate changes for proteins only examined in vitro previously. While there were many proteins that were similarly decreased by all three IMiDs, no compound had the exact same neosubstrate degradation profile as another. We compared our data to previous literature reports of IMiD-induced degradation and known developmental biology associations. Based on our observations, we recommend monitoring at least a major subset of these neosubstrates in a developmental test system to improve CRBN-binding compound-specific risk assessment. A strength of our assay is that it is configurable, and the target list can be readily adapted to focus on only a subset of proteins of interest or expanded to incorporate new findings as additional information about CRBN biology is discovered.

Maintenance therapy for chronic lymphocytic leukaemia.

Maintenance therapy for chronic lymphocytic leukaemia.

Tafasitamab in Combination with a CD20xCD3 Bispecific T-Cell Engager Significantly Prolongs Survival in Preclinical Lymphoma Models

R-CHOP remains the standard of care for newly diagnosed DLBCL patients. However, 30-40% of patients are refractory or relapse after initial response to the immunochemotherapy, indicating a high-unmet medical need for these patients. Tafasitamab is a CD19-targeting, Fc-modified humanized monoclonal antibody mediating antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP) and direct cytotoxicity. Tafasitamab was granted accelerated approval by the FDA in 2020 and conditional marketing authorization by the EMA in 2021 for the treatment of transplant-ineligible adult patients with relapsed or refractory (R/R) DLBCL in combination with the immunomodulatory drug lenalidomide. CD20xCD3 T-cell engagers are currently under clinical investigation or recently received FDA approval in 2L+ DLBCL (Thieblemont 2023; Dickinson 2022). CD19- and CD20-targeting antibodies might complement each other, as I) CD19 could compensate for CD20-low/ -negative settings and vice versa and II) monoclonal antibodies and T-cell engagers combine powerful and distinct immune pathways holding great potential to increase responses in patients who may need additional therapeutic options. The aim of this study was to investigate the combination of tafasitamab and MOR212, a proprietary proof-of-concept CD20xCD3 T-cell engaging antibody, in preclinical models of lymphoma. First, MOR212 was characterized in vitro and in vivo to ensure comparability with other CD20xCD3 bispecific antibodies (comp1-3) currently under clinical investigation. In in vitro cytotoxicity assays, MOR212 demonstrated similar (comp1-2), or improved (comp3) cytotoxicity (MOR212 vs comp1 and 2 p&amp;gt;0.05 vs comp3 p=0.03; exemplary data for EC50 in SU-DHL-4 cells; n=6), cytokine secretion (MOR212 vs comp1-3 p&amp;gt;0.05; exemplary data for EC50 TNFα; n=4) and T-cell activation (MOR212 vs comp1 p=0.05; vs comp2 p&amp;gt;0.05 vs comp3 p=0.003; exemplary data for % CD69 on CD8 T-cells; n=6) in cell lines expressing different levels of CD20. In a lymphoma in vivo model co-engrafted with human T-cells, MOR212 prolonged progression-free survival of mice to a similar extent as two comparator molecules that were tested head-to-head (MOR212 vs comp1 and 3 p&amp;gt;0.05;) at a dose of 0.1 mg/kg. Next, we investigated the combination of tafasitamab and MOR212 in vitro. Due to the differential killing kinetics of a monoclonal antibody and a T-cell engager, we made use of a real time live-cell imaging assay. Lymphoma cells were co-cultured with healthy donor peripheral blood mononuclear cells (PBMCs) and tafasitamab and MOR212 were added in different concentration combinations. Tumor cell killing was monitored every 3 hours for 72 hours and revealed improved target cell killing by the combination of both compounds (signal/background caspase 3/7 staining at 48h: MOR212 24.2 vs tafasitamab 28.2 vs MOR212+tafasitamab 111.5; n=1). To confirm these findings in vivo, we used human CD34+ engrafted NCG mice with a huIL15 boost allowing for the development of human T-cells as well as NK-cells, as essential effector cells for the combination of both compounds. Raji lymphoma cells were engrafted and treatment with the single or combination therapy was started once tumors reached a pulpable size. Combination of tafasitamab and MOR212 significantly improved progression-free survival in animals compared to monotherapy treatment (tafasitamab+MOR212 vs tafasitamab only p=0.0002 and MOR212 only p=0.0098). In addition, the combined treatment regimen appeared to be safe in mice as no side effects on both body weight loss and clinical score were recorded throughout the entire observation period. Our findings prove that the combination of tafasitamab and a CD20xCD3 T-cell engaging antibody significantly increases tumor cell eradication both in vitro and in vivo and lay the ground to potentially translate the results into improved outcome for patients with aggressive lymphoma in future clinical trials.

Salvage Autologous Transplant and Lenalidomide Maintenance Versus Continuous Lenalidomide/Dexamethasone for Relapsed Multiple Myeloma: Long Term Follow up Results of the Randomized GMMG Phase III Multicenter Trial Relapse

Introduction The randomized, controlled, multicenter, phase III GMMG ReLApsE trial compared lenalidomide/dexamethasone (Rd) re-induction, salvage high dose chemotherapy, autologous stem cell transplantation (HDCT/ASCT) and lenalidomide (R) maintenance with standard continuous Rd. We have previously reported the absence of a significant survival benefit in the primary analysis. However, landmark analyses from salvage HDCT performed due to the fact that ~30% of patients in the HDCT arm did not receive salvage HDCT/ASCT suggested a survival benefit in patients undergoing salvage HDCT/ASCT. Here we report long term follow up results from the GMMG ReLApsE trial. Methods Between 2010 and 2016, 282 patients were randomized of whom 277 constituted the intention-to-treat (ITT) population (arm B/A n=139/138). Arm B received 3 cycles of Rd (lenalidomide 25 mg, day 1-21; dexamethasone 40 mg, day 1, 8, 15, 22; 4 week cycles) re-induction, HDCT (melphalan 200 mg/m 2), ASCT and R maintenance (10 mg daily) until progression (PD). Arm A was treated with Rd until PD. Key inclusion criteria were 1-3 prior treatment lines, age ≤ 75 years, time to PD ≥ 12 months in case of front-line HDCT/ASCT and WHO PS ≤ 2. R-refractory disease was not permitted and only 30/277 patients (11%) were previously exposed to R. The primary endpoint was progression free survival (PFS). Protocol-specified long term follow up collection included overall survival (OS), further lines of treatment and further salvage HDCT/ASCT after trial participation. ISRCTN16345835, Eudra CT-No: 2009-013856-61. Results Within the extended follow up period of 99 months, 238 PFS events (86%) and 174 deaths (63%) occurred. Median PFS in the ITT population was 20.5 months in arm B and 19.3 months in arm A without a statistically significant difference (HR 0.98; 95% CI 0.76-1.27; p=0.9). Median OS was 67.1 months in arm B vs. 62.7 months in arm A (HR 0.89; 95% CI 0.66-1.20; p=0.44). The absence of a significant PFS and OS benefit was consistent among key disease subgroups according to age, R-ISS, renal function, frontline single vs. tandem HDCT/ASCT, prior lines of treatment and prior maintenance treatment post HDCT/ASCT. Time to progression after frontline HDCT/ASCT was significantly associated with PFS in the total ITT population (p=0.001) as well as in both trial arms independently, but was not associated with a survival benefit from arm B vs. A (interaction test PFS p=1.0, OS p=0.7). Multivariate analyses revealed significant associations of survival with age (PFS: HR 0.8, p=0.045), high risk cytogenetic aberrations (PFS: HR 2.1, p&amp;lt;0.001; OS: HR 2.5, p&amp;lt;0.001), ISS stage III vs. I (OS: HR 2.0, p=0.02), elevated LDH (PFS: HR 1.5, p=0.04; OS: HR 2.1, p&amp;lt;0.001), &amp;gt;1 prior lines of treatment (PFS: HR 2.3, p=0.003), and no prior HDCT/ASCT (PFS: HR 1.8, p=0.04; OS: HR 2.2, p=0.009), but not with treatment arm B vs. A (PFS: HR 0.9, p=0.4; OS: HR 0.9, p=0.37). In landmark analyses from HDCT and the contemporaneous Rd cycle 5 in arm A (median interval from randomization to HDCT/Rd cycle 5: 117/122 days; n=103[B]/114[A]) no significant differences in PFS (23.0 vs. 20.3 months; HR 0.91; 95% CI 0.68-1.22; p=0.52) or OS (76.3 vs. 66.0 months; HR 0.8; 95% CI 0.56-1.13; p=0.2) in arm B vs. A were observed. Further lines of treatment after trial participation were reported in 88/139 (63%) and 96/138 (70%) patients in arm B and A (p=0.31). Significantly more salvage HDCT/ASCT were administered after trial participation in arm A vs. B (48/138 [35%] vs. 19/139 [14%], p&amp;lt;0.001). Immunomodulatory drug (lenalidomide, pomalidomide) based regimens were used comparably after both trial arms. The rate of second primary malignancies was comparable in arm B and A (26/139 [19%] vs. 19/138 [14%], p=0.3). Conclusions The GMMG ReLApsE trial is the only randomized, controlled trial comparing salvage HDCT/ASCT with continuous novel agent treatment. With extended follow up, no significant PFS or OS difference is observed in the overall trial population and the previously reported survival benefit in patients who actually received salvage HDCT/ASCT is not confirmed. Time to progression after frontline HDCT/ASCT which is frequently used to stratify for salvage HDCT/ASCT in clinical practice is a general prognostic parameter for PFS, but does not predict benefit from salvage HDCT/ASCT vs. continuous Rd treatment. The GMMG ReLApsE trial does not support salvage HDCT/ASCT for relapsed or refractory multiple myeloma.

Treatment Patterns and Healthcare Resource Utilization Among Patients with Triple Class Exposed Multiple Myeloma: A Population-Based Cohort Study

Introduction: Outcomes of patients with relapsed or refractory multiple myeloma (MM) have improved substantially with access to novel therapeutic agents. However, patients with triple class exposed (TCE) MM continue to have poor outcomes. Though newer bispecific antibody and chimeric antigen receptor T-cell immunotherapies have improved outcomes of TCE patients, these therapies are inaccessible outside of clinical trials in many publicly funded healthcare systems, including Canada. This study aimed to describe treatment patterns, outcomes, unplanned health care utilization and quality-of-life impairments of TCE MM patients in Ontario, Canada. These data will provide a crucial benchmark to compare to as novel immunotherapies become available. Methods: This retrospective observational study utilized data from the Institute for Clinical Evaluative Sciences (IC/ES) administrative database, which contains all health records of patients treated within Ontario's publicly funded healthcare system. Drug exposure was determined using the Cancer Activity Level Reporting (ALR) and Ontario Drug Benefit Claims Registry (ODB) databases, housed within IC/ES, which contain data on intravenous and oral MM treatment exposure for standard of care and clinical trial regimens. Patients were defined as TCE if they had prior or current treatment with a regimen containing an immunomodulatory drug (lenalidomide or pomalidomide), a proteasome inhibitor (bortezomib, carfilzomib, or ixazomib), and an anti-CD38 monoclonal antibody (isatuximab or daratumumab). The TCE index regimen was identified as the treatment on which a patient was first identified as having met the TCE definition. Inpatient hospitalization visits did not include planned chemotherapy infusion visits. Quality of life was described using the Edmonton Symptom Assessment System Score with scores ³ 7 corresponding to severe symptoms. Overall survival (OS) was determined using the Kaplan-Meier method. Results: Of the 16,777 patients with a diagnosis of MM between 2007-2021 in Ontario, 1358 patients were classified as TCE during their treatment course. The TCE index regimen was a daratumumab-based regimen in most patients [daratumumab/velcade/dex (n=566, 42%), daratumumab/lenalidomide/dex (n=619, 46%)]. Of the TCE patients, 247 (18%) were still receiving treatment with the TCE index regimen at last follow up, 263 (19%) died without receiving any additional treatment, and 489 (36%) received a subsequent MM treatment after the TCE index regimen. Baseline characteristics of patients (n=489) treated with a subsequent MM regimen after post TCE are summarized in Table 1. The three most commonly administered therapies post TCE were carfilzomib-dexamethasone (n=111, 23%), pomalidomide-dexamethasone (n=77, 16%) and oral cyclophosphamide (n=49, 10%). The next-line treatment regimen was administered in an academic versus community treatment center in 235 (48%) versus 254 (52%) of patients, respectively. The next-line therapy was a clinical trial in only 37 (8%) patients. Regarding healthcare utilization, among TCE patients receiving subsequent therapy, 149 (30%) were evaluated in the emergency department (ED), 126 (26%) were admitted to hospital, and 48 (10%) were referred to palliative care while on treatment with the subsequent therapy. Among patients with available information, the most common severe patient-reported symptoms during the subsequent line of treatment included: tiredness (27%), pain (23%) and overall poor well-being (21%). Additionally, 35% of patients had a ECOG PS of ≥2 on next-line treatment. The median OS patients treated after the TCE index regimen was 1.05 year (95% CI 0.89-1.35) as shown in Figure 1. Of the patients that died during follow up, the majority died in hospital (n=147, 44%) and were referred to palliative care (217, 65%). Conclusion: In this real-world study, we showed that the overall outcomes of TCE MM patients remains poor in Canada, highlighting the urgent need for novel therapies. Furthermore, we demonstrated that TCE patients receiving subsequent therapy had significant unplanned health care utilization and poor quality of life. Future studies will need to assess whether the outcomes, healthcare utilization, and patient-reported quality of life improve as newer immunotherapies become accessible as standard of care treatments within our publicly funded healthcare system.

Outcomes of Second Line Therapy for Light Chain Amyloidosis after Initial Therapy with a Bortezomib Based Regimen

Introduction: There are a lack of randomized trials to guide therapy for relapsed AL amyloidosis with treatment regimens generally extrapolated from experience in multiple myeloma. Methods: We conducted a retrospective review of bortezomib exposed patients that received second line therapy for AL amyloidosis at three Amyloidosis centres in Australia (Brisbane, Perth and Sydney). Of 190 patients treated with bortezomib based therapy upfront, 98 (52%) had transitioned to second line therapy during the study period and comprised the study cohort. Results: Baseline characteristics for the study cohort were typical for a cohort with AL amyloidosis. Median age was 66 years, with 38% of patients being male. Cardiac and renal involvement was present in 70% of patients respectively. Seven patients (7%) had frontline therapy that included daratumumab. Median duration of frontline therapy was 5 months (range 2-18 months) and median time from diagnosis to second line therapy was 9 months (range 2-65 months). We categorized treatment regimens according to commonly used combinations and drug classes to further analyse outcomes. Six categories of therapy regimens were identified; autologous stem cell transplant (ASCT) (n=17), thalidomide based (n=12), lenalidomide based (n=13), proteasome inhibitor (PI) based (n=9), Anti-CD38 monoclonal antibody (mAB) based (n=38) and melphalan based (n=4). Five patients were excluded from this analysis as they received an atypical therapy/regimen not commonly used for AL amyloidosis. Overall response rate (ORR) for the whole cohort was 84%. The ORR was lowest for thalidomide and melphalan based therapy (67% and 50% respectively) and highest for patients receiving an ASCT or anti-CD38 mAB therapy (94% and 92% respectively), Figure 1a. Median event free survival (EFS) for the cohort from second line therapy was 17 months. Median duration of follow up from second line therapy for the cohort was 26 months . Median EFS was shortest for patients treated with thalidomide and melphalan based regimens (6 and 12 months respectively) and longest for those who received an ASCT (28 months) or anti- CD38 mAB therapy (not reached), Figure 1b. When comparing immunomodulatory drugs lenalidomide treated patients had a longer EFS (median 6 months for thalidomide vs 16 months for lenalidomide, p=0.09). The median overall survival (OS) for the cohort was 81 months. OS was longest amongst patients receiving an ASCT or anti-CD38 mAB therapy (median of 91 months and not reached, respectively). Survival was similar amongst patients treated with a PI or Immunomodulatory drug (median 41 months for thalidomide, 44 months for lenalidomide and 36 months for PI based therapy). Conclusion: In bortezomib exposed AL amyloidosis patients, second line therapy with anti CD38 mAB's induced deep responses and prolonged survival. Amongst immunomodulatory agents, the second generation agents appear to be more effective. In our cohort the small group of patients receiving repeat therapy with a PI also appeared to respond to treatment. Although an intensive treatment with significant risks, our data show that in select individuals an ASCT can induce deep and long lasting remissions. Our data highlights the variability in second line therapies for AL amyloidosis given the paucity of evidence to guide treatment. The recent addition of daratumumab as a standard of care for upfront therapy further limits options in the relapsed setting and highlights the need for trials using next generation novel anti plasma cell agents in this patient population.

The Spop/BRD4/c-Myc Axis Modulates IMiDs Sensitivity in Multiple Myeloma

The development of novel agents including immunomodulatory drugs (IMiDs) lenalidomide (Len) and pomalidomide (Pom) has led to improved patient outcomes in multiple myeloma (MM); however, MM patients inevitably experience relapse and drug resistance. The expression level of CRBN, CRBN-binding proteins, and CRL4 CRBN ubiquitin ligase is considered to be associated with IMiDs resistance. Clinical studies have shown that MM patients can exhibit IMiDs resistance even without CRBN abnormalities, suggesting the molecular mechanisms of intrinsic resistance to IMiDs are not fully understood. To delineate the molecular mechanisms underlying IMiDs resistance, we performed genome-wide knockout (KO) screening in IMiDs-sensitive MM cells. For in vitro work, we used the MM cell lines MM1S, and H929. We used sgRNA to KO SPOP and BRD4 in MM cells. After performing a CRISPR KO screen in MM cells, we discovered that in addition to CRBN and its associated genes, sgRNAs targeting SPOP (speckle-type POZ protein, a speckled zinc finger protein) were highly enriched after treatment with Poma. Importantly, SPOP KO MM cell lines acquired significant resistance to Pom and Len treatment. To examine whether SPOP KO-induced IMiDs resistance was CRBN-pathway dependent, we assessed CRBN and its downstream interacting protein levels. SPOP KO showed no effect on CRBN expression; moreover, IMiDs can still trigger IKZF1 and IKZF3 degradation, associated with the downregulation of IRF4, suggesting that SPOP mediates sensitivity to IMiDs in a mechanism independent of CRBN-IKZF1/3 axis.Compared to plasma cells from normal donors, we found that the expression levels of SPOP were significantly decreased in MGUS patients, smoldering myeloma patients, newly diagnosed MM patients, and refractory relapsed MM patients (p-values: 2.4e-2, 2.0e-4, 9.9e-3, 3.17e-6, respectively). SPOP is the adaptor protein of the Cullin3-RING ubiquitin ligase complex. Therefore, the function of SPOP is determined by the characteristics of the substrates it recruits. We performed immunoprecipitation with overexpressed SPOP in MM cells, followed by mass spectrometry analysis, and identified BRD4 as a candidate protein binding to SPOP in MM, which was confirmed by Co-IP. KO of SPOP led to an upregulation of BRD4 protein levels in MM cells. As a downstream regulator of BRD4, the oncogene c-Myc showed a significant increase in protein expression in IMiDs-resistant MM patients. Furthermore, the KO of SPOP notably enhanced c-Myc expression in MM cells, while the KO of BRD4 in SPOP KO MM cells decreaed c-Myc expression and restored MM cell sensitivity to IMiD treatment. These findings suggest that SPOP modulates c-Myc expression through BRD4 in a CRBN-IKZF1/3 independent manner, thereby mediating MM cell sensitivity to IMiDs. BRD4 plays a significant role in the development and progression of various tumors. The BRD4 inhibitor JQ1 has been shown to exhibit anti-tumor effects in multiple types of cancer. We found that combination treatment with JQ-1 and IMiDs reversed IMiDs resistance of SPOP KO MM cell lines and primary MM cells from relapsed patients. Our data show that SPOP is a CRBN-independent modulator of IMiDs sensitivity by regulating the BRD4/c-Myc axis and provides the preclinical rationale for combining IMiDs with BRD4 inhibitors to overcome IMiDs resistance and improve patient outcomes.

Venetoclax Resistance Results in Broad Resistance to Majority of Anti-MM Agents Due to the Suppression of Apoptosis but Can be Overcome By BCMA-Targeted Immunotherapy

Venetoclax represents the first example of personalized medicine in multiple myeloma (MM), as it has meaningful clinical activity in patients harboring the t(11;14) translocation. This subgroup of patients generally displays upregulated BCL-2 expression and/or a higher BCL-2/BCL-X L ratio, reflecting an imbalance between cell survival (promoted by BCL-2) and cell death (promoted by BCL-X L). The Bellini study demonstrated significantly superior response rate and progression-free survival (PFS) with combination of Venetoclax and Bortezomib in relapsed myeloma, however, the overall survival in the arm with venetoclax was inferior. Part of the decrease in overall survival (OS) was considered to be related with increased incidence of infections, however, OS is also significantly impacted by response to subsequent therapies. Here, we investigated whether acquired venetoclax resistance could lead to global resistance to multiple subsequent anti-MM therapies and in turn can explain the decreased OS. We exposed venetoclax-sensitive myeloma cells (KMS27 and KMS12PE) to high-dose venetoclax treatment and generated monoclonal drug-tolerant expanded persister (DTEP) clones from single cell with a three- to ten-fold increase in IC50 compared to the parental cells (4 single cell clones for each cell line). Interestingly, the venetoclax resistance evolved without genomic alterations, as the parental and resistant cells did not show marked differences in mutational frequencies or copy number variations, including the absence of genomic alterations in the BCL-2 gene, such as the Gly101Val mutation, 1q21 gain or 8p loss. Consistent with previous literature in other disease models, these resistant clones also exhibited reduced mitochondrial priming and had increased protein expression of anti-apoptotic regulators, including MCL-1, BCL-X L, and BCL-W, thus overcoming the venetoclax-induced inhibition of the anti-apoptotic regulator BCL-2, resulting in the sequestration of the pro-apoptotic ligand BIM. We next evaluated if the altered mitochondrial priming observed in the resistant cells was also responsible for a broader resistance to anti-cancer agents. We observed that venetoclax-resistant clones were resistant to most standard-of-care anti-MM agents including alkylating agents (Melphalan, cyclophosphamide, bendamustine), proteasome inhibitors (bortezomib and carfilzomib) or immunomodulatory drugs (lenalidomide and pomalidomide) than parental cells, suggesting a broad resistance to anti-cancer agents possibly due to reduced apoptotic signaling. This indicates that switching to these agents after the acquisition of venetoclax resistance may not be efficacious. Since these data suggested a functional substitution between BCL-2 family members, we next evaluated if MCL1 or BCL-XL represent a co-dependency in resistant cells that can be therapeutically targeted to overcome resistance. Simultaneous inhibition of MCL1 (via S63845) or BCL-XL (via A155463) and BCL2 (via venetoclax) increased BIM release and enhanced cell death in the resistant clones compared to single agents, with combination index (CI) values &amp;lt; 0.3 in all doses tested. We further investigated the efficacy of immunotherapeutic approaches to overcome venetoclax resistance as these immunotherapies do not rely exclusively on traditional apoptotic signaling to mediate cell death. We observed that both antibody-dependent cellular cytotoxicity (ADCC) induced by Daratumumab and BCMA targeting CAR-T cells induced comparable cell death in venetoclax-resistant clones and parental cells. Moreover, myeloma cells from a patient with t(11;14) MM progressing on venetoclax, showed significant cytotoxicity to anti-BCMA CAR T cells in vitro and achieved a deep response subsequently when treated with Cilta-cel. In conclusion, we report that resistance to venetoclax confers broad resistance to standard-of-care myeloma drugs and that immunotherapies, especially CAR-T cells and CD38 monoclonal antibody, may remain effective, thus conferring potential benefits in managing acquired resistance to venetoclax.

Nanocomposite hydrogel to deliver the immunomodulator lenalidomide and anti-inflammatory hesperidin locally to joints affected by rheumatoid arthritis

Rheumatoid arthritis is an autoimmune disease characterized by inflammatory cell infiltration, neovascularization in the synovial tissue, and progressive joint destruction. The immunomodulatory drug lenalidomide and anti-inflammatory anti-oxidant hesperidin show therapeutic potential against the disease as two complementary drugs, but they also show poor oral bioavailability and solubility as well as a short time in circulation. To overcome these disadvantages, the two drugs were loaded into polymeric micelles in their native forms and also, they were conjugated to polyethylene glycol to create acid-activated prodrugs. Lastly, the micelles alone or with the prodrugs were formulated into an injectable hydrogel. Of all formulations, the hydrogel containing micelles and prodrugs led to the greatest drug uptake by Raw264.7 macrophages in culture and inhibition of angiogenesis in a chick chorioallantoic membrane assay. In rats with collagen-induced arthritis, the hydrogel released the two drugs in a sustained manner, preferentially in joints with acidic pH. They dampened inflammatory responses, mitigating bone injury. This biocompatible, pH-responsive formulation based on two complementary drugs may provide a new strategy for treating rheumatoid arthritis.

C-FOS is an integral component of the IKZF1 transactivator complex and mediates lenalidomide resistance in multiple myeloma.

The immunomodulatory drug lenalidomide, which is now widely used for the treatment of multiple myeloma (MM), exerts pharmacological action through the ubiquitin-dependent degradation of IKZF1 and subsequent down-regulation of interferon regulatory factor 4 (IRF4), a critical factor for the survival of MM cells. IKZF1 acts principally as a tumour suppressor via transcriptional repression of oncogenes in normal lymphoid lineages. In contrast, IKZF1 activates IRF4 and other oncogenes in MM cells, suggesting the involvement of unknown co-factors in switching the IKZF1 complex from a transcriptional repressor to an activator. The transactivating components of the IKZF1 complex might promote lenalidomide resistance by residing on regulatory regions of the IRF4 gene to maintain its transcription after IKZF1 degradation. To identify unknown components of the IKZF1 complex, we analyzed the genome-wide binding of IKZF1 in MM cells using chromatin immunoprecipitation-sequencing (ChIP-seq) and screened for the co-occupancy of IKZF1 with other DNA-binding factors on the myeloma genome using the ChIP-Atlas platform. We found that c-FOS, a member of the activator protein-1 (AP-1) family, is an integral component of the IKZF1 complex and is primarily responsible for the activator function of the complex in MM cells. The genome-wide screening revealed the co-occupancy of c-FOS with IKZF1 on the regulatory regions of IKZF1-target genes, including IRF4 and SLAMF7, in MM cells but not normal bone marrow progenitors, pre-B cells or mature T-lymphocytes. c-FOS and IKZF1 bound to the same consensus sequence as the IKZF1 complex through direct protein-protein interactions. The complex also includes c-JUN and IKZF3 but not IRF4. Treatment of MM cells with short-hairpin RNA against FOS or a selective AP-1 inhibitor significantly enhanced the anti-MM activity of lenalidomide in vitro and in two murine MM models. Furthermore, an AP-1 inhibitor mitigated the lenalidomide resistance of MM cells. C-FOS determines lenalidomide sensitivity and mediates drug resistance in MM cells as a co-factor of IKZF1 and thus, could be a novel therapeutic target for further improvement of the prognosis of MM patients.

CD33 BiTE® molecule-mediated immune synapse formation and subsequent T-cell activation is determined by the expression profile of activating and inhibitory checkpoint molecules on AML cells

Bispecific T-cell engager (BiTE®) molecules recruit T cells to cancer cells through CD3ε binding, independently of T-cell receptor (TCR) specificity. Whereas physiological T-cell activation is dependent on signal 1 (TCR engagement) and signal 2 (co-stimulation), BiTE molecule-mediated T-cell activation occurs without additional co-stimulation. As co-stimulatory and inhibitory molecules modulate the strength and nature of T-cell responses, we studied the impact of the expression profile of those molecules on target cells for BiTE molecule-mediated T-cell activation in the context of acute myeloid leukemia (AML). Accordingly, we created a novel in vitro model system using murine Ba/F3 cells transduced with human CD33 ± CD86 ± PD-L1. T-cell fitness was assessed by T-cell function assays in co-cultures and immune synapse formation by applying a CD33 BiTE molecule (AMG 330). Using our cell-based model platform, we found that the expression of positive co-stimulatory molecules on target cells markedly enhanced BiTE molecule-mediated T-cell activation. The initiation and stability of the immune synapse between T cells and target cells were significantly increased through the expression of CD86 on target cells. By contrast, the co-inhibitory molecule PD-L1 impaired the stability of BiTE molecule-induced immune synapses and subsequent T-cell responses. We validated our findings in primary T-cell-AML co-cultures, demonstrating a PD-L1-mediated reduction in redirected T-cell activation. The addition of the immunomodulatory drug (IMiD) lenalidomide to co-cultures led to stabilization of immune synapses and improved subsequent T-cell responses. We conclude that target cells modulate CD33 BiTE molecule-dependent T-cell activation and hence, combinatorial strategies might contribute to enhanced efficacy.

Abstract 1936: Demographic and socioeconomic determinants to the access to multi-agent therapy in multiple myeloma: A National Cancer Database (NCDB) analysis of years 2004-2017

Abstract Background: Multi-agent therapy consisting of Immunomodulatory Drug (Lenalidomide), Proteasome Inhibitor (Bortezomib), and Dexamethasone revolutionized patient outcomes in Multiple Myeloma (MM). In this retrospective analysis, the NCDB database was used to evaluate the determinants that affected access to multi-agent therapy among MM patients who were treated in commission on cancer-accredited facilities across the USA. Methods: Using the NCDB, we identified N= 171,261 patients diagnosed with MM from 2004 to 2017. Multivariate logistic regression analysis was conducted to identify the predictive factors for receiving multi-agent therapy, using significance level of p&amp;lt0.05. Multivariable cox regression was used to determine factors affecting survival in patients receiving multi-agent therapy. Kaplan-Meier (KM) survival curves for single and multi-agents were produced. SAS version 9.4 was used to analyze the data. Results: Five potential disparity factors were identified by multivariate logistic regression analysis: race and ethnicity, facility type, insurance status, median household income and level of education. Non-Hispanic Black patients had lower odds of receiving multi-agent therapy compared to Non-Hispanic White patients (OR=0.93, p&amp;lt0.0001). Hispanic Black patients had higher odds of receiving multi-agent therapy compared to Non-Hispanic Black patients (Hispanic Black vs Non-Hispanic Black: OR=1.47, p=0.01). Patients who weren't treated in academic centers were less likely to be treated with multi-agent therapy (OR=0.75, p&amp;lt0.0001). Those with Medicaid or Medicare were less likely to receive multi-agent regimen than those with private insurance (OR=0.88, p&amp;lt0.0001, OR=0.86, p&amp;lt0.0001, respectively). Patients with lower median household incomes had lower access to multi-agent therapy (OR=0.86, p&amp;lt0.0001). Those with lower level of education were less likely to be treated with multi-agent regimens than those who were (OR=0.93, p=0.0133). KM survival curves showed significant survival benefit for patients who received multi-agent therapy. Our data indicates that when Black and Hispanic patients receive multi-agent MM therapy, they tend to have better outcomes compared to White and Non-Hispanic patients (HR=1 and 0.92 for White and Black patients respectively; HR=1 and 0.8 for Non-Hispanic and Hispanic patients respectively, p&amp;lt0.0001). Conclusion: In this large analysis of MM patients, we identified that Non-Hispanic Black patients and those with a lower socioeconomic status were less likely to receive the standard of care multi-agent therapy in this large cohort of real world data set from across the USA. These results further corroborate the need to provide equitable access to care, which will translate to better clinical outcomes. Citation Format: Ludovic Saba, Chakra Pani Chaulagain, Hong Liang, Barbara Dominguez, Chieh-Lin Fu. Demographic and socioeconomic determinants to the access to multi-agent therapy in multiple myeloma: A National Cancer Database (NCDB) analysis of years 2004-2017 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1936.

Management of Relapsed and Refractory Multiple Myeloma: Recent advances

AbstractMultiple myeloma (MM) accounts for ∼10% of total hematologic malignancies worldwide. In India, the incidence of MM has increased two-fold with marked heterogeneity. Significant improvements in terms of clinical outcomes have been observed in the management of MM in recent years. However, most patients develop a disease relapse with the first or subsequent treatments. A combination of immunomodulatory drugs (thalidomide and lenalidomide) and proteasome inhibitors (PIs; bortezomib) has been the mainstay for the therapeutic management of relapsed/refractory multiple myeloma (RRMM). This review highlights the management of RRMM with newer agents such as belantamab, carfilzomib, daratumumab, elotuzumab, ixazomib, mafadotin, selinexor, panobinostat, and venetoclax, with more focus on PIs. As a single agent and in combination with other drugs including dexamethasone and carfilzomib has been studied extensively and approved by the United States, European Union, and India. Clinical trials of these newer agents, either alone or in combination, for the treatment of RRMM in Western countries indicate survival, improved outcomes, and overall well-being. However, evidence in Indian patients is evolving from ongoing studies on carfilzomib and daratumumab, which will ascertain their efficacy and safety. Currently, several guidelines recommend carfilzomib-based, daratumumab-based, and panobinostat-based regimens in RRMM patients. Currently, with more accessible generic versions of these drugs, more Indian patients may attain survival benefits and improved quality of life.

“A.B.C.” of Immunotherapy in Hematological Malignancies…Promise and Perils

AbstractThe treatment landscape of hematological malignancies has been evolving at an extremely fast pace. Hematological malignancies are diverse and distinct from solid tumors. These constitute challenges, which are also unique opportunities for immunotherapy. The five categories of immunotherapies that have found success in the management of hematological malignancies are allogeneic hematopoietic stem cell transplant, monoclonal antibodies and innovative designs, immune checkpoint inhibitors, chimeric antigen receptor (CAR) T cells, and B cell targeting small immunomodulatory molecules. Allogeneic stem cell transplant rightly called our bluntest weapon is the oldest form of successful immunotherapy. Alternate donor transplants and improvement in supportive care have improved the scope of this immunotherapy option. Among monoclonal antibodies, rituximab forms the prototype on which over a dozen other antibodies have been developed. The bispecific T-cell engager (BiTE) blinatumomab engages cytotoxic CD3 T cells with CD19 acute lymphoblastic leukemia (ALL) cells, which is an effective treatment method for relapsed refractory ALL. Immune checkpoint inhibitors have established their role in hematological malignancies with high PD-L1 expression, including relapsed refractory Hodgkin's lymphoma and primary mediastinal B cell lymphoma (BCL). Small immunomodulatory drugs targeting the B cell receptor downstream signaling through BTK inhibitors, SYK inhibitors, PI3K inhibitors (idelalisib), and BCL-2 inhibitors (venetoclax), and immunomodulatory imide drugs (lenalidomide) have also emerged as exciting therapeutic avenues in immunotherapy. CAR T cells are one of the most exciting and promising forms of adoptive immunotherapy. CAR T cells are rightly called living drugs or serial killers to keep patients alive. CAR T cells are genetically engineered, autologous T cells that combine the cytotoxicity of T cells with the antigen-binding specificity of CARs. CARs are antigen-specific but major histocompatibility complex/human leukocyte antigen-independent. There are five approved CAR T cell products for the management of relapsed refractory leukemias, lymphoma, and multiple myeloma. The past and present of immunotherapy have been really exciting and the future looks incredibly promising. The challenges include widening the availability and affordability beyond specialized centers, identification of potentially predictive biomarkers of response, and experience in the management of complications of these novel agents. The combinational approach of multiple immunotherapies might be the way forward to complement the treatment strategies to harness the immune system and to improve survival with good quality of life.

Phase II Trial of Elotuzumab with Pomalidomide and Dexamethasone for Relapsed/Refractory Multiple Myeloma (RRMM) in the Post-Daratumumab Progression Setting

Introduction: Elotuzumab (Elo) is an immunostimulatory (via natural killer [NK] cell activation) monoclonal antibody (mAb) targeting signaling lymphocytic activation molecule F7 (SLAMF7) that has shown activity in combination with the immunomodulatory drugs (IMiDs) lenalidomide (Len) or pomalidomide (Pom) and dexamethasone (Dex), in patients (pts) with RRMM in the ELOQUENT 2 and 3 trials, respectively. However, in ELOQUENT 2, pts were not Len refractory or previously treated with daratumumab (Dara) and in ELOQUENT 3, <5% of patients were previously treated with Dara. The efficacy of Elo-based regimens in Dara-refractory pts needs prospective evaluation to understand optimal sequencing of regimens in RRMM. We report data from a phase 2 trial evaluating the efficacy of Elo-Pom-Dex (EPd) in pts with Dara-refractory RRMM. Methods: This phase 2 trial (NCT03713294) enrolled pts with RRMM who had disease progression and were refractory to Dara. Key eligibility criteria included: measurable disease as per IMWG criteria, at least one prior line of therapy, adequate marrow/organ function (neutrophil count ≥1.0 x 109, platelets ≥50x 109, creatinine clearance ≥30 mL/min), ECOG performance status ≤2, and disease not refractory to Pom. EPd was dosed as per the ELOQUENT 3 study. The trial utilized a two-stage design with an interim analysis to test the null hypothesis that the true overall response rate (ORR) in this patient population is most 20% vs. the alternative hypothesis that it is at least 40%, with 90% power and 9% type I error. Treatment was given until disease progression, intolerance, or patient refusal. Results: 37 pts were enrolled. Median follow-up time was 23.4 months (95% CI: 14.6-37.2). The median age was 70 years (range: 42-84), 12 (32.4%) pts had ISS III disease and 16 (43.2%) pts had high risk cytogenetics. Pts had received a median of 4 (range: 1-9) prior lines of therapy. Prior anti-MM agents included bortezomib (35; 94.6%), Len (34; 91.9%), carfilzomib (9; 24.3%), ixazomib (9; 24.3%), stem cell transplantation (18; 48.6%) and CAR-T cell therapy (1; 2.7%). The majority of pts (33; 89%) were previously exposed to both a proteasome inhibitor and Len. 25 (67.6%) pts got Dara-based therapy as the immediate line of therapy prior to EPd (after a median of 4 prior lines [range 1-7]) whereas 12 (32%) pts got Dara-based therapy ≥1 line prior to EPd (after a median of 6 prior lines [range 2-9]). ORR in all pts was 35.1% (13 pts), with no CRs, 1 (2.7%) VGPR, and 12 (32.4%) PRs. Four pts (10.8%) achieved an MR and 15 (40.5%) achieved SD. The study met its primary endpoint. The ORR in pts exposed to Len was 11/34 (32.4%) compared to 2/3 (66.7%) in patients not exposed to Len. The ORR in pts with Dara as the immediate prior line of therapy was 44% (11/25) compared to 16.7% (2/12) in pts who had received Dara-based therapy ≥1 line prior to EPd (p=0.15). The median PFS for the entire cohort was 3.7 mth (95% CI: 2.9-11.1) (Figure 1A). The median time to best response was 2.9 mth. The median time to next treatment was 8.5 mth (95% CI: 6.8-20.4). Median PFS of pts with Dara as the immediate prior line of therapy was 4.7 mth (95% CI: 3.2-19.6) compared to 2.8 mth (95% CI: 2.0-NA) in patients who had received Dara-based therapy ≥1 line prior to EPd (p=0.03; Figure 1B). The most common grade 3/4 adverse events were neutropenia (57%), anemia (22%), thrombocytopenia (16%), leucopenia (16%), and fatigue (8%). Conclusion: This was a positive study with EPd showing clinically meaningful benefit in pts with Dara-refractory RRMM. The reported PFS of 3.7 mths is less than the 10.3 mths reported in ELOQUENT 3 trial which suggests that Elo-based regimens are less effective in Dara-refractory pts. Interestingly, pts had a significantly superior PFS if they received EPd immediately following progression on Dara, however they were less heavily pretreated comparted to pts who got EPd ≥1 line after Dara failure. Nonetheless, EPd is an effective regimen in the post-Dara setting. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Lenalidomide and risk of second primary malignancy—is disease context key?

The development of the immunomodulatory drug lenalidomide, along with proteasome inhibitors, has revolutionised the treatment of multiple myeloma since the early 2000s. Lenalidomide is now used in triplet or quadruplet combination therapeutic approaches for patients with newly diagnosed and relapsed myeloma. Lenalidomide has also been shown to prolong progression-free survival and overall survival as maintenance therapy after autologous haematopoietic stem-cell transplantation (HSCT). The approved indications for lenalidomide now include other haematological malignancies such as myelodysplastic syndrome with an isolated del(5q), mantle cell lymphoma, and follicular lymphoma.