anti-CD20 mAb Research Articles

Anti-CD20 monoclonal antibody therapy in postpartum women with neurological conditions.

Postpartum, patients with multiple sclerosis (MS) and neuromyelitis optica spectrum disorder (NMOSD) have increased risk for disease activity. Anti-CD20 IgG1 monoclonal antibodies (mAb) are increasingly used as disease-modifying therapies (DMTs). Patients may wish to both breastfeed and resume DMT postpartum. This study aimed to determine the transfer of anti-CD20 IgG1 mAbs, ocrelizumab, and rituximab (OCR/RTX), into mature breastmilk and describe maternal and infant outcomes. Fifty-seven cis-women receiving OCR/RTX after 59 pregnancies and their infants were enrolled and followed up to 12M postpartum or 90 days post-infusion. Breastmilk was collected pre-infusion and serially up to 90 days and assayed for mAb concentration. Medical records and patients' questionnaire responses were obtained to assess neurologic, breastfeeding, and infant development outcomes. The median average concentration of mAb in breastmilk was low (OCR: 0.08 μg/mL, range 0.05-0.4; RTX: 0.03 μg/mL, range 0.005-0.3). Concentration peaked 1-7 days post-infusion in most (77%) and was nearly undetectable after 90 days. Median average relative infant dose was <1% (OCR: 0.1%, range 0.07-0.7; RTX: 0.04%, range 0.005-0.3). Forty-three participants continued to breastfeed post-infusion. At 8-12 months, the proportion of infants' growth between the 3rd and 97th World Health Organization percentiles did not differ for breastfed (36/40) and non-breastfed (14/16, p > 0.05) infants; neither did the proportion with normal development (breastfed: 37/41, non-breastfed: 11/13; p > 0.05). After postpartum infusion, two mothers experienced a clinical relapse. These confirm minimal transfer of mAb into breastmilk. Anti-CD20 mAb therapy stabilizes MS activity before conception to the postpartum period, and postpartum treatments appears to be safe and well-tolerated for both mother and infant.

Prior cycles of anti-CD20 antibodies affect antibody responses after repeated SARS-CoV-2 mRNA vaccination.

BACKGROUNDWhile B cell depletion is associated with attenuated antibody responses to SARS-CoV-2 mRNA vaccination, responses vary among individuals. Thus, elucidating the factors that affect immune responses after repeated vaccination is an important clinical need.METHODSWe evaluated the quality and magnitude of the T cell, B cell, antibody, and cytokine responses to a third dose of BNT162b2 or mRNA-1273 mRNA vaccine in patients with B cell depletion.RESULTSIn contrast with control individuals (n = 10), most patients on anti-CD20 therapy (n = 48) did not demonstrate an increase in spike-specific B cells or antibodies after a third dose of vaccine. A third vaccine elicited significantly increased frequencies of spike-specific non-naive T cells. A small subset of B cell-depleted individuals effectively produced spike-specific antibodies, and logistic regression models identified time since last anti-CD20 treatment and lower cumulative exposure to anti-CD20 mAbs as predictors of those having a serologic response. B cell-depleted patients who mounted an antibody response to 3 vaccine doses had persistent humoral immunity 6 months later.CONCLUSIONThese results demonstrate that serial vaccination strategies can be effective for a subset of B cell-depleted patients.FUNDINGThe NIH (R25 NS079193, P01 AI073748, U24 AI11867, R01 AI22220, UM 1HG009390, P01 AI039671, P50 CA121974, R01 CA227473, U01CA260507, 75N93019C00065, K24 AG042489), NIH HIPC Consortium (U19 AI089992), the National Multiple Sclerosis Society (CA 1061-A-18, RG-1802-30153), the Nancy Taylor Foundation for Chronic Diseases, Erase MS, and the Claude D. Pepper Older Americans Independence Center at Yale (P30 AG21342).

Monoclonal antibodies binding to different epitopes of CD20 differentially sensitize DLBCL to different classes of chemotherapy.

Rituximab (R), an anti-CD20 monoclonal antibody (mAb) and the world's first approved antibody for oncology patients, was combined with the CHOP chemotherapy regimen and markedly improved the prognosis of all B- cell-derived lymphomas, the most common hematological malignancy worldwide. However, there is a 35% disease recurrence with no advancement in the first-line treatment since R was combined with the archetypal CHOP chemotherapy regimen nearly 30 years ago. There is evidence that R synergizes with chemotherapy, but the pharmacological interactions between R and CHOP or between newer anti-CD20 mAbs and CHOP remain largely unexplored. We used in vitro models to score pharmacological interactions between R and CHOP across various lymphoma cell lines. We compared these pharmacological interactions to ofatumumab, a second-generation anti-CD20 mAb, and CHOP. Lastly, we used RNA-sequencing to characterize the transcriptional profiles induced by these two antibodies and potential molecular pathways that mediate their different effects. We discovered vast heterogeneity in the pharmacological interactions between R and CHOP in a way not predicted by the current clinical classification. We then discovered that R and ofatumumab differentially synergize with the cytotoxic and cytostatic capabilities of CHOP in separate distinct subsets of B-cell lymphoma cell lines, thereby expanding favorable immunochemotherapy interactions across a greater range of cell lines beyond those induced by R-CHOP. Lastly, we discovered these two mAbs differentially modulate genes enriched in the JNK and p38 MAPK family, which regulates apoptosis and proliferation. Our findings were completely unexpected because these mAbs were long considered to be biological and clinical equivalents but, in practice, may perform better than the other in a patient-specific manner. This finding may have immediate clinical significance because both immunochemotherapy combinations are already FDA-approved with no difference in toxicity across phase I, II, and III clinical trials. Therefore, this finding could inform a new precision medicine strategy to provide additional therapeutic benefit to patients with B-cell lymphoma using immunochemotherapy combinations that already meet the clinical standard of care.

The Development of Ofatumumab, a Fully Human Anti-CD20 Monoclonal Antibody for Practical Use in Relapsing Multiple Sclerosis Treatment.

The importance of B cells in multiple sclerosis (MS) has been demonstrated through the advent of B-cell-depleting anti-CD20 antibody therapies. Ofatumumab is the first fully human anti-CD20 monoclonal antibody (mAb) developed and tested for subcutaneous (SC) self-administration at monthly doses of 20mg, and has been approved in the US, UK, EU, and other regions and countries worldwide for the treatment of relapsing MS. The development goal of ofatumumab was to obtain a highly efficacious anti-CD20 therapy, with a safety and tolerability profile that allows for self-administration by MS patients at home and a positive benefit-risk balance for use in the broad relapsing MS population. This development goal was enabled by the unique binding site, higher affinity to B cells, and higher potency of ofatumumab compared to previous anti-CD20 mAbs; these properties of ofatumumab facilitate rapid B-cell depletion and maintenance with a low dose at a low injection volume (20mg/0.4ml). The high potency in turn enables the selective targeting of B cells that reside in the lymphatic system via subcutaneous (SC) administration. Through a comprehensive dose-finding program in two phase 2 studies (one intravenous and one SC) and model simulations, it was found that safety and tolerability can be further improved, and the risk of systemic injection-related reactions (IRRs) minimized, by avoiding doses≥30mg, and by reaching initial and rapid B-cell depletion via stepwise weekly administration of ofatumumab at Weeks 0, 1, and 2 (instead of a single high dose). Once near-complete B-cell depletion is reached, it can be maintained by monthly doses of 20mg/0.4ml. Indeed, in phase 3 trials (ASCLEPIOS I/II), rapid and sustained near-complete B-cell depletion (largely independent of body weight, race and other factors) was observed with this dosing regimen, which resulted in superior efficacy of ofatumumab versus teriflunomide on relapse rates, disability worsening, neuronal injury (serum neurofilament light chain), and imaging outcomes. Likely due to its fully human nature, ofatumumab has a low immunogenic risk profile-only 2 of 914 patients receiving ofatumumab in ASCLEPIOS I/II developed anti-drug antibodies-and this may also underlie the infrequent IRRs (20% with ofatumumab vs. 15% with the placebo injection in the teriflunomide arm) that were mostly (99.8%) mild to moderate in severity. The overall rates of infections and serious infections in patients treated with ofatumumab were similar to those in patients treated with teriflunomide (51.6% vs. 52.7% and 2.5% vs. 1.8%, respectively). The benefit-risk profile of ofatumumab was favorable compared to teriflunomide in the broad RMS population, and also in the predefined subgroups of both recently diagnosed and/or treatment-naïve patients, as well as previously disease-modifying therapy-treated patients. Interim data from the ongoing extension study (ALITHIOS) have shown that long-term treatment with ofatumumab up to 4years is well-tolerated in RMS patients, with no new safety risks identified. In parallel to the phase 3 trials in which SC administration was carried out with a pre-filled syringe, an autoinjector pen for more convenient self-administration of the ofatumumab 20mg dose was developed and is available for use in clinical practice.

Health-economic benefits of anti-CD20 treatments in relapsing multiple sclerosis estimated using a treatment-sequence model.

In high-income countries, four anti-CD20 monoclonal antibodies (mAbs) are used or in the pipeline for relapsing MS: ocrelizumab, ofatumumab (both registered), ublituximab (awaiting registration) and rituximab (off-label). List prices differ significantly between registered and off-label drugs. Comparing differences in benefits between anti-CD20 mAbs from a health-economic and societal perspective. To reflect lifetime use of DMTs, we used a treatment-sequence model to compare ocrelizumab/ofatumumab and eight other drug classes in terms of health (lifetime relapses, time to Expanded Disability Status Scale [EDSS] 6, lifetime quality-adjusted life years) and cost-effectiveness (net health benefit). To become cost-effective compared to ocrelizumab, we modelled the list price of ublituximab and desired effect on EDSS progression of rituximab. Although drug sequences with ocrelizumab in first- and second-line were more cost-effective than ofatumumab, our probabilistic analysis suggests this outcome was very uncertain. To be more cost-effective than ocrelizumab, ublituximab needs to be about 25% cheaper whilst rituximab needs to equal the effect on disability progression seen with first-line treatments. Our model showed no clear difference in cost-effectiveness between ocrelizumab and ofatumumab. Hence, prescribing the least costly anti-CD20 mAb can democratise MS care without a loss in health benefits.

Important functional role of the protein osteopontin in the progression of malignant pleural mesothelioma.

Malignant Pleural Mesothelioma (MPM) is an aggressive cancer of the mesothelial lining associated with exposure to airborne non-degradable asbestos fibers. Its poor response to currently available treatments prompted us to explore the biological mechanisms involved in its progression. MPM is characterized by chronic non-resolving inflammation; in this study we investigated which inflammatory mediators are mostly expressed in biological tumor samples from MPM patients, with a focus on inflammatory cytokines, chemokines and matrix components. Expression and quantification of Osteopontin (OPN) was detected in tumor and plasma samples of MPM patients by mRNA, immunohistochemistry and ELISA. The functional role of OPN was investigated in mouse MPM cell lines in vivo using an orthotopic syngeneic mouse model. In patients with MPM, the protein OPN was significantly more expressed in tumors than in normal pleural tissues and predominantly produced by mesothelioma cells; plasma levels were elevated in patients and associated with poor prognosis. However, modulation of OPN levels was not significantly different in a series of 18 MPM patients receiving immunotherapy with durvalumab alone or with pembrolizumab in combination with chemotherapy, some of whom achieved a partial clinical response. Two established murine mesothelioma cell lines: AB1 and AB22 of sarcomatoid and epithelioid histology, respectively, spontaneously produced high levels of OPN. Silencing of the OPN gene (Spp1) dramatically inhibited tumor growth in vivo in an orthotopic model, indicating that OPN has an important promoting role in the proliferation of MPM cells. Treatment of mice with anti-CD44 mAb, blocking a major OPN receptor, significantly reduced tumor growth in vivo. These results demonstrate that OPN is an endogenous growth factor for mesothelial cells and inhibition of its signaling may be helpful to restrain tumor progression in vivo. These findings have translational potential to improve the therapeutic response of human MPM.

Efficacy and safety of add-on anti-CD20 monoclonal antibody to Bruton tyrosine kinase inhibitor treatment for chronic lymphocytic leukemia: a meta-analysis

The efficacy of Bruton tyrosine kinase inhibitors (BTKi) remains suboptimal in chronic lymphocytic leukemia (CLL) treatment. A systematic review and meta-analysis were conducted to compare the outcomes of combining anti-CD20 monoclonal antibodies (mAb) with BTKi therapy versus BTKi monotherapy for patients with CLL. We searched for relevant studies in the Pubmed, Medline, Embase, and Cochrane databases until December 2022. We estimated the effective results using a hazard ratio (HR) for survival outcomes and relative risk (RR) for response outcomes and safety. Four randomized controlled trials (including 1056 patients) were found until November 2022 and fulfilled the inclusion criteria. Progression-free survival was significantly improved with the addition of anti-CD20 mAb to BTKi over BTKi (HR 0.70, 95% confidence interval (CI) 0.51–0.97), whereas pooled analysis of overall survival did not favor combination therapy compared to BTKi monotherapy (HR 0.72, 95% CI 0.50–1.04). Combination therapy was related to a statistically better complete response (RR, 2.03; 95% CI 1.01 to 4.06) and an undetectable minimal residual disease rate (RR, 6.43; 95% CI 3.54 to 11.67). The risk of grade ≥ 3 adverse events was comparable between the two groups (RR, 1.08; (95% CI 0.80 to 1.45). Overall, adding anti-CD20 mAb to BTKi revealed superior efficacy than BTKi alone in untreated or previously treated CLL patients without affecting the safety of single-agent BTKi. Conducting further randomized studies to confirm our results and determine the optimal therapy for managing patients with CLL is essential.

Leveraging CD16 fusion receptors to remodel the immune response for enhancing anti-tumor immunotherapy in iPSC-derived NK cells

BackgroundThe cytotoxicity of NK cells is largely dependent on IgG Fc receptor CD16a, which mediates antibody-dependent cell-mediated cytotoxicity (ADCC). The high-affinity and non-cleavable CD16 (hnCD16) is developed and demonstrated a multi-tumor killing potential. However, the hnCD16 receptor activates a single CD16 signal and provides limited tumor suppression. How to exploit the properties of hnCD16 and incorporate NK cell-specific activation domains is a promising development direction to further improve the anti-tumor activity of NK cells.MethodsTo expand the applications of hnCD16-mediated ADCC for NK cell-based immunotherapy in cancer, we designed the hnCD16 Fusion Receptor (FR) constructs with the ectodomain of hnCD16 fused with NK cell-specific activating domains in the cytoplasm. FR constructs were transduced into CD16-negative NK cell line and human iPSC-derived NK (iNK) cells and effective FR constructs were screened. The up-regulation of immune activation- and cytokine-releasing-related pathways in FR-transduced NK cells was screened and validated by RNA sequencing and multiplex cytokines release assay, respectively. The tumor-killing efficiency was tested in vitro and in vivo via co-culture with tumor cell lines and xenograft mice-bearing human B-cell lymphoma, respectively.ResultsWe screened the most effective combination to kill B cell lymphoma, which was fused with the ectodomain of hnCD16a, NK-specific co-stimulators (2B4 and DAP10) and CD3ζ in cytoplasmic domains. The screened construct showed excellent cytotoxicity effects and sharp multiple cytokines releasing both in the NK cell line and iNK cells. The transcriptomic analysis and validation assays of hnCD16- and hnCD16FR-transduced NK cells showed that hnCD16FR transduction remodeled immune-related transcriptome in NK cells, where significant upregulation of genes related to cytotoxicity, high cytokines releasing, induced tumor cell apoptosis, and ADCC in comparison with hnCD16 transduction were highlighted. In vivo xenograft studies demonstrated that a single low-dose regimen of engineered hnCD16FR iPSC-derived NK cells co-administered with anti-CD20 mAb treatment mediated potent activity and significantly improved survival.ConclusionWe developed a novel hnCD16FR construct that exhibits more potent cytotoxicity than reported hnCD16, which is a promising approach to treat malignancies with improved ADCC properties. We also offer a rationale for NK activation domains that remodel immune response to enhance CD16 signaling in NK cells.

Overexpression of SIRT6 alleviates apoptosis and enhances cell viability and monoclonal antibody expression in CHO-K1 cells.

Chinese hamster ovary (CHO) cells are the most predominantly utilized host for the production of monoclonal antibodies (mAbs) and other complex glycoproteins. A major challenge in the process of CHO cell culture is the occurrence of cell death following different stressful conditions, which hinders the production yield. Engineering genes involved in pathways related to cell death is a remarkable strategy to delay apoptosis, improve cell viability and enhance productivity. SIRT6 is a stress-responsive protein that regulates DNA repair, maintains genome integrity, and is critical for longevity and cell survival in organisms. In this study, SIRT6 was stably overexpressed in CHO-K1 cells and the impact of its expression on apoptosis related gene expression profile, viability, apoptosis, and mAb productivity was investigated. While a significant increase was observed in Bcl-2 mRNA level, caspase-3 and Bax mRNA levels were decreased in the SIRT6 engineered cells compared to the parental CHO-K1 cells. Moreover, improved cell viability and decreased rate of apoptotic progression was observed in a SIRT6-derived clone in comparision to the CHO-K1 cells during 5 days of batch culture. anti-CD52 IgG1 mAb titers were improved up to 1.7- and 2.8-fold in SIRT6-derived clone during transient and stable expression, respectively. This study indicates the positive effects of SIRT6 overexpression on cell viability and anti-CD52 IgG1 mAb expression in CHO-K1 cells. Further studies are needed to examine the potential of SIRT6-engineered host cells for the production of recombinant biotherapeutics in industrial settings.

Pharmacogenomic Analyses Implicate B Cell Developmental Status and MKL1 as Determinants of Sensitivity toward Anti-CD20 Monoclonal Antibody Therapy.

Monoclonal antibody (mAb) therapy directed against CD20 is an important tool in the treatment of B cell disorders. However, variable patient response and acquired resistance remain important clinical challenges. To identify genetic factors that may influence sensitivity to treatment, the cytotoxic activity of three CD20 mAbs: rituximab; ofatumumab; and obinutuzumab, were screened in high-throughput assays using 680 ethnically diverse lymphoblastoid cell lines (LCLs) followed by a pharmacogenomic assessment. GWAS analysis identified several novel gene candidates. The most significant SNP, rs58600101, in the gene MKL1 displayed ethnic stratification, with the variant being significantly more prevalent in the African cohort and resulting in reduced transcript levels as measured by qPCR. Functional validation of MKL1 by shRNA-mediated knockdown of MKL1 resulted in a more resistant phenotype. Gene expression analysis identified the developmentally associated TGFB1I1 as the most significant gene associated with sensitivity. qPCR among a panel of sensitive and resistant LCLs revealed immunoglobulin class-switching as well as differences in the expression of B cell activation markers. Flow cytometry showed heterogeneity within some cell lines relative to surface Ig isotype with a shift to more IgG+ cells among the resistant lines. Pretreatment with prednisolone could partly reverse the resistant phenotype. Results suggest that the efficacy of anti-CD20 mAb therapy may be influenced by B cell developmental status as well as polymorphism in the MKL1 gene. A clinical benefit may be achieved by pretreatment with corticosteroids such as prednisolone followed by mAb therapy.

Incidence of second primary malignancies (SPMs) in patients with multiple myeloma (MM) receiving anti-CD38 monoclonal antibodies (mAbs): A systematic review and meta-analysis.

e24078 Background: The addition of anti-CD38 mAbs, daratumumab and isatuximab, to standard anti-myeloma regimens have drawn scrutiny towards long-term safety concerns, including the risk of SPMs. ICARIA-MM study reported SPMs in 3.9% patients (pts.) in the Isatuximab, pomalidomide and dexamethasone (Isa-Pd) arm compared to 0.7% in the Pd arm. We hypothesized that exposure to anti-CD38 mAbs could potentially increase risk of SPMs due to prolonged immunosuppression. An assessment of this association is warranted, given the improved life expectancy and growing prevalence of pts. with MM. Methods: We performed a meta-analysis on data obtained from RCTs published between January 2005 and April 2022 that compared anti-CD38 mAb-based regimens to other standard anti-myeloma regimens (control) in pts. with newly diagnosed or relapsed/refractory (r/r) MM. Total number of SPMs (including categories) were extracted for pts. in both groups. Malignant melanomas were included under solid SPM category. Binary outcome data were analyzed using fixed-effect Peto method. Treatment effect estimates were expressed as odds ratio (OR) and 95% confidence interval (CI). I2 &gt; 50% indicated substantial heterogeneity. Results: 10 RCTs involving 4980 pts. were included, with 2824 (57%) and 2156 (43%) pts. randomized to the anti-CD38 mAb and control arms, respectively (Table). Overall, 191 and 112 SPMs were reported in the anti-CD38 mAb and control arms, respectively. At a median follow-up of 35.3 months, anti-CD38 mAb exposure was associated with an increased risk of developing SPMs compared to control (6.8% vs. 5.2%; OR: 1.53 [95% CI: 1.20-1.95]; I2= 0%). Non-melanoma cutaneous cancers were the most common, and higher in the anti-CD38 mAb vs. control arms (92 versus 47; OR: 1.77 [95% CI: 1.25-2.51]; I2= 0%). The increase in solid cancers (including melanoma) (OR: 1.28 [95% CI: 0.85-1.95]) and hematologic SPMs (OR: 1.86; [95% CI: 0.81-4.27] was not significant. On sensitivity analysis, the total number of SPMs were higher in the anti-CD38 mAb arm vs. control arm, regardless of the presence of immunomodulatory drugs in combination regimen. Conclusions: This meta-analysis shows an increased risk of SPMs in pts. with MM exposed to daratumumab or isatuximab, particularly that of non-invasive cutaneous tumors. This association should be further investigated in real-world settings using large databases. [Table: see text]

Talquetamab (tal) + daratumumab (dara) in patients (pts) with relapsed/refractory multiple myeloma (RRMM): Updated TRIMM-2 results.

8003 Background: Tal is a T-cell redirecting bispecific antibody (BsAb) targeting G protein–coupled receptor family C group 5 member D. Dara is an anti-CD38 mAb with direct on-tumor and immunomodulatory actions. Combining immunomodulatory effects of tal + dara may lead to synergistic efficacy. Initial TRIMM-2 (NCT04108195) results showed that SC tal RP2Ds, 0.4 mg/kg QW or 0.8 mg/kg Q2W, + SC dara had promising efficacy and increased CD38+/CD8+ T cells and proinflammatory cytokines. We report updated results with additional pts and longer follow-up. Methods: Pts had MM, ≥3 prior lines of therapy (LOT; including a proteasome inhibitor [PI] and immunomodulatory drug [IMiD]) or were double refractory to a PI and IMiD, and had not received anti-CD38 therapy in ≤90 d. Pts received tal RP2Ds with step-up dosing + dara 1800 mg per approved schedule. AEs were graded per CTCAE v5.0; cytokine release syndrome (CRS) and immune effector cell–associated neurotoxicity syndrome (ICANS) were graded per ASTCT guidelines. Responses were assessed per IMWG criteria. Results: As of Dec 12, 2022 (N = 65), median follow-up was 11.5 mo (range 1.0–27.3). Median age was 63 y (range 37–81); 18% of pts had high-risk cytogenetics; 25% had extramedullary plasmacytomas. Median prior LOT was 5 (range 2–16): 63% penta-drug exposed; 58% triple-class refractory. Prior treatments included anti-CD38 (88% [77% refractory]), anti-BCMA (54% [38%]), BsAb (25% [25%]), and anti-BCMA CAR-T (17% [2%]) therapy. All pts had ≥1 AE (grade [Gr] 3/4 78%), most commonly CRS (78%; all Gr 1/2), dysgeusia (75%), dry mouth (55%), anemia (52%), fatigue (45%), and skin exfoliation (45%). CRS had a median time to onset of 1 d after the most recent dose with median duration of 2 d. 63% of pts had infections (Gr 3/4 22%; Gr 5 3% [n = 2 pneumonia, possibly related to tal + dara]). 38% of pts had neutropenia (Gr 3/4 26%). 85% had postbaseline IgG &lt; 500 mg/dL; of these, 32% received IVIg. ICANS occurred in 3 pts (5%; all Gr 1/2 and resolved in 1–2 d). ORR was 78% (66% ≥VGPR; 45% ≥CR) across RP2Ds (100% in anti-CD38 naïve pts), and responses deepened over time. In pts exposed/refractory to prior therapy, ORRs were 75%/76% for anti-CD38, 74%/64% for anti-BCMA, and 75%/75% for BsAb. Median time to first response was 1 mo (range 0.9–8.3); at 12 mo, 86% of responders (89% of pts with ≥CR) still had responses. At data cutoff, 84% of responders remain on therapy (83%/82% anti-CD38 exposed/refractory). mPFS was 19.4 mo; 12-mo PFS and OS rates were 76% and 93%, respectively. Conclusions: Steroid-sparing tal + dara showed deep and durable responses with promising mPFS in heavily pretreated pts with RRMM, including pts refractory to anti-CD38/BCMA and T-cell redirecting therapy, suggesting combined immunomodulatory actions can yield robust responses in pts with refractory disease. The safety profile was clinically manageable; no new signals were identified with longer follow-up.

17th International Conference on Malignant Lymphoma, Palazzo dei Congressi, Lugano, Switzerland, 13 - 17 June, 2023.

Introduction: CD19 is an established target of multiple B-cell lymphoma therapies. Blockade of CD47, a macrophage immune checkpoint inhibitory ligand, induces tumor-cell phagocytosis. TG-1801 is a CD19/CD47 bispecific antibody designed to selectively block CD47 on CD19+ cells, while retaining a functional IgG1 Fc domain. Herein we report updated results from the FIH study. Methods: The primary objectives of this phase I 3+3 study were to characterize the safety profile and determine the recommended phase 2 dose (RP2D) as monotherapy and in combination with ublituximab. Pharmacokinetics, pharmacodynamics, and preliminary antitumor activity were also assessed. Eligible patients (pts) had relapsed/refractory (R/R) B-cell lymphoma after ≥1 prior standard therapy. TG-1801 doses evaluated included: 20 mg, 60 mg, 180 mg, 360 mg, & 500 mg. In the monotherapy arm, treatment consisted of weekly fixed dosing 1h-infusions of TG-1801 in a 4-week cycle: day (D)1, D8, D15, and D22 of cycle (C) 1 and C2. Pts who had at least stable disease after 2 cycles continued with TG-1801 once every 4 weeks for 4 further infusions. Intra-pt dose escalations were permitted. A second arm explored the combination of TG-1801 with ublituximab; an anti-CD20 mAb. Pts received TG-1801 and ublituximab, IV every 4 weeks: D1, D8, and D15 of C1; D1 of C2-C6; and D1 of C9 and every 3 cycles thereafter, up to 24 cycles. 2 dose levels of TG-1801 (300 mg and 400 mg) with 900 mg of ublituximab were tested. Responses were assessed by Lugano 2014 criteria. Results: As of October 2022, 14 pts (MZL = 5, DLBCL including Richter’s transformation = 5, FL = 4) received monotherapy and 16 pts received combination therapy (DLBCL = 9, FL = 4, MZL = 2, MCL = 1). Pts had a median of 3 prior therapies (range, 1–8) with 16 pts (53%) refractory to their last therapy. One DLT of grade (G) 4 thrombocytopenia occurred at 500 mg monotherapy. Most frequent (>20%) treatment emergent adverse events (TEAEs) at monotherapy doses of 20 mg to 360 mg (n = 10) included fatigue, thrombocytopenia, and infusion-related reaction (3 pts each), with no G3/4 TEAEs occurring in >20% of pts. Combination therapy was also well tolerated (TG-1801 at 300 mg N = 3, 400 mg N = 13) without DLT. Most common TEAEs (≥20%) for combination therapy were anemia, headache, and fatigue (5 pts each), with no G3/4 TEAEs occurring in >20% of pts. 2 pts permanently discontinued therapy due to TEAEs (1 infusion reaction [500 mg monotherapy], 1 rash [360 mg monotherapy]). Among 13 evaluable monotherapy pts, there were 3 (ORR = 23%) partial responses (PR). In combination (n = 16), a 44% ORR was observed with 1 complete response (CR) in a pt with FL and 6 PR (5 pts with DLBCL and 1 pt with FL) for a 56% ORR in DLBCL and 50% ORR in FL. The research was funded by: TG Therapeutics Inc. Keywords: Combination Therapies, Immunotherapy, Targeting the Tumor Microenvironment Conflicts of interests pertinent to the abstract. E. A. Hawkes Consultant or advisory role: Advisory board and speaker fees (institutional or personal) from Roche, Merck Sharpe & Dohme, Astra Zeneca, Gilead, Antigene, Novartis, Regeneron, Janssen, Specialised Therapeutics, outside the submitted work Research funding: Roche, Bristol Myers Squibb, Merck KgA, Astra Zeneca and Merck K. L. Lewis Consultant or advisory role: Astrazeneca, Roche, Loxo/Lilly, IQVIA, MSD Honoraria: Astrazeneca, Janssen, Roche H. P. Miskin Employment or leadership position: TG Therapeutics Inc. Stock ownership: TG Therapeutics Inc. P. Sportelli Employment or leadership position: TG Therapeutics Inc. Stock ownership: TG Therapeutics Inc. K. S. Kolibaba Employment or leadership position: TG Therapeutics Inc. Stock ownership: TG Therapeutics Inc. E. Normant Employment or leadership position: TG Therapeutics Inc. Stock ownership: TG Therapeutics Inc. T. Turpuseema Employment or leadership position: TG Therapeutics Inc. Stock ownership: TG Therapeutics Inc. C. Y. Cheah Consultant or advisory role: Roche, Janssen, Gilead, AstraZenecca, Lilly, TG therapeutics, Beigene, Novartis, Menarini, Daizai, Abbvie, Genmab. BMS Honoraria: Roche, Janssen, Gilead, AstraZenecca, Lilly, TG therapeutics, Beigene, Novartis, Menarini, Daizai, Abbvie, Genmab. BMS Research funding: BMS, Roche, Abbvie; MSD, Lilly

Real-world evidence study of treatment patterns and outcomes following covalent BTKi discontinuation in a contemporary cohort of chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) patients.

e19510 Background: In the past decade, covalent Bruton Tyrosine Kinase inhibitors (BTKi) have transformed the treatment landscape for patients with CLL/SLL. Previous research described BTKi discontinuation in CLL/SLL cohorts where most patients received the BTKi after multiple prior lines of therapy. Information on outcomes of patients who received BTKi in first or second line (1L or 2L) and subsequently discontinued is limited. We examine a cohort of contemporary CLL/SLL patients who discontinued the BTKi, a majority of whom initially received BTKi in 1L or 2L. Methods: A retrospective chart review study of adult CLL/SLL patients who were treated with a covalent BTKi at Dana-Farber Cancer Institute and who were resistant/refractory or intolerant to BTKi was conducted. Abstraction took place 12/2021-12/2022. Data abstracted included demographics, clinical characteristics, treatment patterns, reasons for treatment discontinuation, and cause of death. Results: A total of 104 patients were included with median time from CLL/SLL diagnosis to first BTKi initiation (index date) of 7 years, median age of 67 years at index date, and median follow-up of 6 years. Prognostic marker testing revealed the following: del(13q): 58%, del(11q): 28%, del(17p): 26%, trisomy 12: 20%, complex karyotype: 16%; IGHV unmutated: 59%, TP53 mutation: 66%. The most common index BTKi was ibrutinib (92%: 59% as monotherapy and 12% in combination with anti-CD20 mAb). 27% of patients were treated with BTKi in 1L, 32% in 2L, and 41% in later lines of therapy. 102 (98%) patients discontinued index BTKi. The median duration of index BTKi treatment was 2 years. 53% of patients discontinued index BTKi due to adverse events, most commonly atrial fibrillation (26%) and bleeding events (13%); 40% discontinued due to disease progression; and 3% due to Richter’s transformation. Subsequent treatments were observed for 74% of patients who discontinued due to adverse events and all patients who discontinued due to progression. Following last BTKi treatment, 60% received subsequent therapies; among these, 86% received regimens with venetoclax from index BTKi initiation to data cutoff. 34% of patients died with the most frequent causes of death being disease progression (62%), infection (17%), and secondary malignancy (7%). Conclusions: In this retrospective chart review study of adult CLL/SLL patients, adverse events and disease progression were key drivers for BTKi discontinuation, as found in earlier studies in more heavily pre-treated patients. Following BTKi discontinuation, a high percentage of patients received venetoclax-based regimens, yet with a median follow-up of 6 years, 34% died. This suggests that novel treatment approaches are sorely needed for less heavily pre-treated CLL/SLL patients who discontinue covalent BTKi.

Incidence of second primary malignancies in patients with multiple myeloma receiving anti-CD38 monoclonal antibodies: A systematic review and meta-analysis

Anti-CD38 monoclonal antibodies (mAbs) are commonly used for treating newly diagnosed and relapsed/refractory (r/r) multiple myeloma (MM). However, concerns have been raised about the occurrence of second primary malignancies (SPMs) in patients receiving anti-CD38 mAbs. Assessing the safety data for rare adverse events like SPMs is challenging because individual clinical trials are typically focused on the primary endpoint. Therefore, we conducted a meta-analysis of randomized controlled trials (RCTs) published between January 2005 and April 2022, including patients with newly diagnosed or r/r MM. Our aim was to compare SPM rate with the use of anti-CD38 mAb-based regimens with other anti-myeloma regimens. After a median follow-up of 35.3 months (range: 8.2–56.2), we found that exposure to anti-CD38 mAbs was associated with an increased risk of developing SPMs compared to the control group (6.8% vs. 5.2%; Peto odds ratio [OR]: 1.53 [95% confidence interval (CI): 1.20–1.95]; I2= 0%, p-value for heterogeneity= 0.44). This increased risk was primarily driven by non-melanoma cutaneous cancers (92 vs. 47; Peto OR: 1.77 [95% CI: 1.25–2.51]; I2 = 0%, p-value for heterogeneity = 0.54). However, there was no significant difference in the incidence of solid tumors (including malignant melanoma) (OR: 1.28 [95% CI: 0.85–1.95]) or hematologic SPMs (OR: 1.86; [95% CI: 0.81–4.27]). In conclusion, the use of anti-CD38 mAb-based combination regimens is associated with a higher risk of non-invasive cutaneous SPMs, but not solid tumors or hematologic SPMs. The increased occurrence of non-invasive cutaneous SPMs may be due to enhanced monitoring resulting from longer treatment duration with anti-CD38 mAbs.

CD4 T cell deficiency narrows virus-neutralizing IgG coverage for Polyomavirus and selects for antibody-escape variants

Abstract CD4 T cell deficiencies predispose patients to progressive multifocal leukoencephalopathy (PML), a brain disease caused by JC polyomavirus (JCPyV). PML is linked to immunomodulatory therapies that preferentially lower CD4 T cells in the CSF (natalizumab); but how CD4 T cells control JCPyV is undefined. “Blind spots” in the ability to neutralize JCPyV variants with mutations in the VP1 capsid protein were described in PML patients. We recently reported that a VP1 mAb selects for VP1-mutant escape variants both in vitroand in vivo; some of these variant viruses are neurovirulent. We hypothesized that alterations in the Ab response due to CD4 T cell insufficiency underlie emergence of Ab-escape mouse polyomavirus (MuPyV) variants. T-cell independent (TI) VP1-specific IgG (via anti-CD4 mAb depletion, CD40L blockade, MHCII KO mice, IL21R KO mice) neutralizes MuPyV but has reduced titer and avidity than anti-VP1 IgGs in CD4 T cell-sufficient mice. Serial passaging of MuPyV in vitrowith sera from TI mice selected an Ab-escape VP1 mutant containing amino acid deletions at residue 145–146 in an external loop of VP1 (MuPyV.145–146). In contrast to sera from healthy MuPyV-infected mice, TI sera failed to neutralize MuPyV.145–146. Depletion of CD4 T cells during persistent MuPyV infection, as a model of acquired CD4 T cell insufficiency (i.e. onset of immunomodulatory mAbs) via anti-CD4 mAb depletion, CD40L blockade, and anti-CD20 mAb B cell depletion also reduced anti-VP1 IgG titer and avidity. Ongoing studies seek to determine if a similar loss in recognition of the 145–146 VP1 epitope occurs. Our data shows that a narrowed antiviral IgG response due to CD4 T cell deficiency is a critical antecedent for outgrowth of Ab-escape PyV VP1 mutant viruses. Supported by grants from the NIH (5R01NS088367, 5R01NS092662, and R35NS127217) (AEL) Judy S. Finkelstein Memorial Award (KNA) T32 CA06395 (KNA)

Quantifying in vitrocomplement-dependent cytotoxicity (CDC) using advanced flow cytometry

Abstract Complement dependent cytotoxicity (CDC) is a key Fc mediated function of monoclonal antibody (mAb) therapeutics. mAb binding to antigen triggers a complex molecular cascade with sequential recruitment of serum proteins, eventuating in target cell lysis. Here we demonstrate CDC quantification in a streamlined, in vitro, advanced flow cytometry assay. Anti-CD20 mAbs were incubated with target cell lines in 96- or 384-well plates. Human serum (15%) was then added to induce CDC. Cells were labeled with iQue® Cell Membrane Integrity (R/Red) Dye to enable assessment of cell death using the iQue® Advanced Flow Cytometry Platform. Induction of CDC by anti-CD20 mAbs correlated with target cell CD20 expression, with most cell death observed with high-CD20 expressing Ramos cells. Maximal Ramos cell death induced by Rituximab and Truxima® was 68% and 70%, respectively. Comparatively, cell death of mid-CD20 expressing Raji cells was lower at 34% and 24%. No CDC was induced with CD20 negative Jurkat cells. Another anti-CD20-IgG1 mAb was compared against two isotype mutants: IgG1fut (non-fucosylated) and IgG1NQ (non-glycosylated). Their activity was profiled towards three previously described effector functions: antibody-dependent cellular cytotoxicity (ADCC); antibody-dependent cellular phagocytosis (ADCP) and CDC. CDC induction was comparable between the three mAbs, however their ADCC and ADCP activity differed. The IgG1NQ did not exert ADCP or ADCC activity, whilst the IgG1fut showed more ADCC and less ADCP than the native. These data exemplify the use of advanced flow cytometry to quantify Fc function of mAbs towards a range of effector mechanisms, highlighting the potential to profile libraries of novel therapeutics in minimal time.

First dose infusion reaction during anti-CD20 monoclonal antibody therapy in chronic lymphocytic leukemia associates with high IP-10 levels amid cytokine release syndrome

Abstract Intravenous (IV) anti-CD20 monoclonal antibody (mAb) therapy for chronic lymphocytic leukemia (CLL) patients often produces a first dose infusion reaction (FDIR) within the first 2 h that requires careful monitoring to avoid serious complications and fatalities. To better understand FDIR, we studied blood samples collected from 37 treatment-naïve CLL patients in our clinical trial NCT03788291 undergoing IV treatment with 50 mg of anti-CD20 mAb (rituximab). We analyzed four time points: baseline (prior to infusion), 1 h later (during infusion), at the end of infusion (~2.5 h) and at 48 h. Infusion reactions (CTCAE (v5) grade ≥2 events) were managed by interrupting the infusion and with symptomatic treatment. 24 (65%) patients had a FDIR and all patients completed the infusion. Patient and CLL characteristics including measures of disease severity (Rai stage, IGHV mutation status, cytogenetic defects) did not correlate with FDIR. High levels of CLL cells and/or CD20 and their subsequent depletion resulting in decreased serum mAb and/or complement are thought to be risks for FDIR. However, measurements of these parameters did not correlate. Induction of cytokine release syndrome (CRS) may correlate with FDIR. However, CRS was observed in all samples after mAb infusion. The only significant associations with FDIR were higher levels of IP-10, IL-6, and IL-8, with IP-10 most significant. Anti-CD20 mAb activated cytotoxic effector cells, such as tissue-resident macrophages that ingest CLL cells via antibody-dependent cellular phagocytosis (ADCP), may be responsible for these events leading to FDIR. A mouse model of anti-CD20 mAb FDIR and in vitro ADCP cell systems will be useful to further understand the biology of FDIR. Supported by funding from Acerta/AstraZeneca, the Cadregari Foundation, a generous donation by Elizabeth Aaron, and the NIH NCI grant number R21CA267040.

Ublituximab, a Novel, Glycoengineered Anti-CD20 Monoclonal Antibody (mAb), Demonstrates Enhanced Antibody-Dependent Cellular Cytotoxicity (ADCC) Relative to Other Anti-CD20 mAbs (P7-3.011)

Ublituximab, a Novel, Glycoengineered Anti-CD20 Monoclonal Antibody (mAb), Demonstrates Enhanced Antibody-Dependent Cellular Cytotoxicity (ADCC) Relative to Other Anti-CD20 mAbs (P7-3.011)

Changing Treatment Trends for MS Patients Recently Initiated on Their First DMT (P12-3.012)

<h3>Objective:</h3> To evaluate if disease-modify therapy (DMT) choice has changed for treatment-naïve multiple sclerosis (MS) patients from 2020–2022. <h3>Background:</h3> Cross-sectional audit of MS patients was conducted annually. Data were reviewed by an independent market intelligence agency which specializes in tracking the neurology market. <h3>Design/Methods:</h3> US neurologists (2020: N=199; 2021: N=205; 2022: N=208) completed a survey and contributed chart reviews for a retrospective, cross-sectional audit on MS patients who started their first DMT within the prior 3 months (2020: N=1,006; 2021: N=1,018; 2022: N=1,095). MS patients were analysed as a whole, as well as by physician-reported MS subtypes for relapsing-remitting MS (RRMS; N=722, N=768, N=827), active secondary progressive MS (aSPMS; N=74, N=50, N=59), and primary progressive MS (PPMS; N=92, N=69, N=72). <h3>Results:</h3> Overall use of glatiramer acetate (GA) (32% in 2020, 16% in 2022) has declined; use of fumarate agents (13%, 21%) and anti-CD20 monoclonal antibody (mAb) DMTs (13%, 21%) expanded. Interferon (13% in 2020 and 2022) and S1P receptor modulator class (12%, 14%) use has remained stable. Use of both brand (18%, 11%) and generics (14%, 6%) of GA declined. For fumarates, use has shifted from branded dimethyl fumarate (DMF; 13%, 8%) to generic DMF (0%, 6%) and diroximel fumarate (0%, 6%). For S1Ps, use has shifted from fingolimod (10%, 6%) to ozanimod (0%, 4%) and ponesimod (0%, 1%). Changes in DMT use are similar for RRMS: GA use declined (31%, 16%); fumarates (17%, 23%) and anti-CD20 mAb DMT (9%, 19%) use expanded. GA use also declined for aSPMS (30%, 7%) and PPMS (30%, 8%); anti-CD20 mAb DMT use grew for aSPMS (19%, 41%). <h3>Conclusions:</h3> DMT prescribing trends are evolving for MS patients recently initiated on their first DMT. Use of GA agents has fallen across MS subtypes, while use of the high-efficacy anti-CD20 mAb DMT class has expanded. <b>Disclosure:</b> Dr. McFadden has nothing to disclose.