scholarly journals Successful Treatment of Persistent SARS-CoV-2 Infection in a B-Cell Depleted Patient with Activated Cytotoxic T and NK Cells: A Case Report

2021 ◽  
Vol 22 (20) ◽  
pp. 10934
Author(s):  
Jacek Jassem ◽  
Natalia Maria Marek-Trzonkowska ◽  
Tomasz Smiatacz ◽  
Łukasz Arcimowicz ◽  
Ines Papak ◽  
...  
Keyword(s):  
T Cells ◽  
Case Report ◽  
B Cell ◽  
Nk Cells ◽  
Cytotoxic T Cells ◽  
Viral Clearance ◽  
Lymphoma Patient ◽  
Cd20 Antibody ◽  
Anti Cd20 ◽  
B Cell Deficiency

We report a lymphoma patient with profound B-cell deficiency after chemotherapy combined with anti-CD20 antibody successfully treated with remdesivir and convalescent plasma for prolonged SARS-CoV-2 infection. Viral clearance was likely attributed to the robust expansion and activation of TCR Vβ2 CD8+ cytotoxic T cells and CD16 + CD56- NK cells. This is the first presentation of TCR-specific T cell oligoclonal response in COVID-19. Our study suggests that B-cell depleted patients may effectively respond to anti-SARS-CoV-2 treatment when NK and antigen-specific Tc cell response is induced.

scholarly journals Activation of natural killer cells by rituximab in granulomatosis with polyangiitis

2019 ◽  
Vol 21 (1) ◽  
Author(s):  
Doris Urlaub ◽  
Shuyang Zhao ◽  
Norbert Blank ◽  
Raoul Bergner ◽  
Maren Claus ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Nk Cells ◽  
Natural Killer ◽  
Mechanism Of Action ◽  
Granulomatosis With Polyangiitis ◽  
Color Flow ◽  
Cd20 Antibody ◽  
Anti Cd20

Abstract Objective In the last few years, anti-CD20 antibody rituximab profoundly changed the therapeutic landscape of granulomatosis with polyangiitis (GPA). Here, we investigated whether natural killer (NK) cells may play a role in rituximab’s mechanism of action in GPA. Methods B cell depletion, NK cell degranulation, and the expression of CD69 and CD16 on NK cells were measured in a series of in vitro experiments using peripheral blood mononuclear cells (PBMCs). In vivo activation of NK cells was investigated in patients receiving rituximab infusions. Cells were analyzed by seven-color flow cytometry. Results NK cells from GPA patients were activated by immobilized rituximab. Also soluble rituximab activated NK cells, provided that B cells were present. NK cells degranulated and expressed the activation marker CD69 while CD16 expression was decreased. This activation of NK cells by soluble rituximab was accompanied by a reduction of B cells. The next-generation anti-CD20 antibody obinutuzumab showed stronger effects compared to rituximab on both the reduction of B cells and the activation of NK cells. Finally, we found that rituximab led to the activation of NK cells in vivo, provided that B cells were not depleted due to prior rituximab infusions. Conclusion B cell-bound rituximab activates NK cells in GPA. While NK cells therefore participate in rituximab’s mechanism of action in humans, their potential may be more efficiently exploited, e.g., by Fc engineering of therapeutic antibodies.

Negative Impact Of KIR/HLA Interactions On NK Cell Activity Can Be Overcome By The Glycoengineered Antibody GA101 (Obinutuzumab)

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 376-376
Author(s):  
Grzegorz Terszowski ◽  
Christian Klein ◽  
Jakob Passweg ◽  
Martin Stern
Keyword(s):  
B Cell ◽  
Nk Cells ◽  
Target Cell ◽  
Cell Activation ◽  
Nk Cell ◽  
Negative Impact ◽  
Target Cells ◽  
Cd107a Expression ◽  
Cd20 Antibody ◽  
Anti Cd20

Abstract Antibody dependent cellular cytotoxicity (ADCC) is one of the mechanisms by which therapeutic antibodies mediate tumor cell killing. The anti-CD20 antibody rituximab is the current standard of care in the treatment of B-cell lymphomas. GA101, a novel anti-CD20 antibody, contains a glycoengineered Fc-portion allowing approximately 10-fold greater affinity to FcgR3A, the Fc-IgG receptor expressed on the majority of natural killer (NK) cells. NK cell function is also regulated by inhibitory killer-cell immunoglobulin-like receptors (KIR), which interact with HLA class I antigens (2DL1-HLA-C2; 2DL2/3-HLA-C1, 3DL1-HLA-Bw4). The KIR/HLA interaction during NK cell development leads to the acquisition of full effector function in the “licensing” process, but also provides one of the main mechanisms of NK cell tolerance. The present study analyzed how KIR/HLA interactions influence ADCC, and whether there are differences between conventional and glycoengineered antibodies. We analyzed the activation (in terms of the degranulation measured by the CD107a expression) and killing capacity of KIR-positive NK cells induced by rituximab, GA101, and the parental non-Fc modified (wild-type) GA101wt. Target cells included HLA-negative B-cell lymphoma lines or B-cell lines expressing one or more HLA molecules. We confirmed previous observations that the licensing status affects the potential for rituximab-induced ADCC (degranulation against HLA-deficient 721.221 in licensed cells 35 ± 4% versus 19 ± 3% of unlicensed cells, p<0.01); and that KIR/HLA interactions strongly and selectively inhibit the response to targets expressing cognate HLA ligands (e.g. CD107a expression in KIR3DL1+ NK cells 17 ± 3% against 721.221-Bw4 cells, compared to 32 ± 4% against 721.221, p<0.01). Next, we analyzed rituximab-induced NK cell activation in donors expressing one, two, or three KIR ligands after co-incubation with target B-cell lines expressing corresponding HLA molecules. These experiments showed that the inhibitory effect during target cell encounter dominates over the activating effect of NK cell licensing, which leads to unlicensed NK cells being the strongest effectors of ADCC with rituximab (Figure, Panel A). We next compared the effect of the KIR/HLA interaction on rituximab-, GA101wt- and GA101-induced ADCC. GA101 largely compensated the hyporesponsiveness of unlicensed cells and NK cell activation was independent of the presence of HLA KIR ligands on target cells (Figure, Panel A). Finally, we addressed the question of how levels of NK cell degranulation correspond to target cell elimination. Correlation between CD107a expression and target cell elimination was excellent for all antibodies (Figure Panel B). GA101 induced the highest level of activation and the most effective target elimination. In contrast to rituximab and GA101wt, no negative impact of KIR/HLA interaction on degranulation or target cell elimination could be detected for GA101. In summary, we show that KIR/HLA interactions are relevant for ADCC with rituximab, with the negative impact during target cell encounter dominating over the positive effect of licensing. In contrast, the novel glycoengineered GA101 antibody overrides the negative signals derived from the KIR/HLA interaction and activates all NK cell subsets. These data suggest that the Fc-modification to enhance ADCC can be an effective strategy to augment the efficacy of therapeutic monoclonal antibodies by recruiting NK cells irrespective of their inhibitory KIR expression. Disclosures: Terszowski: Roche: Research Funding. Klein:Roche Glycart AG: Employment. Stern:Roche: Research Funding.

631 Development of highly efficacious and safe targeted cancer immunotherapy via IL12-based TMEkine™ platform

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A667-A667
Author(s):  
Dahea Lee ◽  
Donggeon Kim ◽  
Soomin Ryu ◽  
Byoung Chul Lee
Keyword(s):  
T Cells ◽  
B Cell ◽  
Nk Cells ◽  
Cell Lymphoma ◽  
Specific Binding ◽  
B Cell Lymphoma ◽  
Human T Cells ◽  
Syngeneic Mouse Model ◽  
Anti Cd20

BackgroundWe developed tumor microenvironment-targeting immunocytokine or TMEkine™ utilizing strong anti-tumoral effect of interleukin 12 (IL-12). In this effort, we created a bi-specific 1+1 antibody fusion with conventional knob-in-hole technology where anti-CD20 was paired with IL-12 fc fusion arm. A couple of IL-12 muteins were used in our therapeutic molecules to reduce systemic toxicity. IL-12 has been known for a key orchestrator in immune response. The main actions of IL-12 include the induction of CD4+ Th0 cells toward Th1 type and enhancement of IFN-γ production, stimulation of cytotoxicity and growth of natural killer (NK) cells and CD8+ T cells. For these reasons, IL-12 has long been considered as a potential therapeutic molecule for treating cancers by enhancing immune activity toward tumor cells. However, systemic administration of IL-12 showed poor efficacy and severe adverse effects. With our therapeutic approach of tumor targeting and attenuated IL-12 mutein, we expect that our IL12-based TMEkine™ holds great promise for the future of cancer immunotherapy.In this study, we targeted CD-20 expressing cancers such as B-cell lymphoma with our anti-CD20/IL-12 mutein TMEkine. We evaluated the biological activity of our molecules with in vitro and in vivo efficacy and safety.MethodsThe target specific binding to CD20 and IL-12 receptor was analyzed by FACS and ELISA. Biological activities as signaling transduction and T cell activation were confirmed in vitro using HEKblue IL12 cell line, primary human T cells and NK cells. The anti-tumor efficacy of TMEkine (CD20-IL-12) was assessed in A20 lymphoma syngeneic mouse model. To demonstrate long term protection to A20, the cured five mice after TMEkine administration were re-challenged with A20 and 4T1 cells.ResultsFirst, we analyzed the specific binding of our TMEkine molecules to CD20 expressing B-cell lymphoma cell lines (such as Raji). We showed that TMEkine (CD20-IL-12) binds to Raji and Ramos, which express CD20, but not to Jurkat, which does not express CD20. We also showed that TMEkine molecules bind to IL-12 receptor in a dose-dependent manner. pSTAT4 alphaLISA assay revealed that TMEkine (CD20-IL-12) transduces STAT4 signaling. In our IL-12 mutein, key residues for heparin binding were mutated. The biological activity of our mutein molecule was attenuated due to this change in human PBMC. In addition, our TMEkine molecules significantly induced IFN-γ secretion from primary human T cells and NK cells. An A20 B-cell lymphoma syngeneic mouse model was utilized to investigate the anti-tumor activity of TMEkine (CD20-IL-12). TMEkine molecules were injected three times with Q3D intraperitoneally. Tumor growth was substantially reduced and no cytotoxicity was observed. To further investigate the underlying mechanism, we analyzed tumor infiltrating lymphocytes (TIL) and as expected, we observed the increase in the number of CD8+ T cells in TIL, compared to control group. Interestingly, our tumor re-challenge result demonstrates that TMEkine (CD20-IL-12) protected animals from tumor recurrence implying that immunologic memory response was generated upon our TMEkine (CD20-IL-12) treatment.ConclusionsAltogether, our data suggest that TMEkine (CD20-IL-12) as an efficacious tumor targeting cytokine opening up a new avenue for the treatment of B-cell lymphoma.

scholarly journals Optimizing Ex-Vivo Expanded NK Cell- Mediated Antibody-Dependent Cellular Cytotoxicity (ADCC) Combined with NKTR-255 in Chronic Lymphocytic Leukemia (CLL), Follicular Lymphoma (FL), and Burkitt Lymphoma (BL)

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 23-24
Author(s):  
Yaya Chu ◽  
Susiyan Jiang ◽  
Jian Jiang ◽  
Meijuan Tian ◽  
Dean Anthony Lee ◽  
...  
Keyword(s):  
T Cells ◽  
B Cell ◽  
Nk Cells ◽  
Board Of Directors ◽  
Research Funding ◽  
Type I ◽  
Type Ii ◽  
Advisory Committees ◽  
Anti Cd20

Background: The CD20 molecule is universally expressed by normal B cells in all stages of development, from the pre-B cell up to the mature plasma cell as well as by most B cell malignancies including CLL, FL and BL (Chu/Cairo, BJH, 2016). Rituximab, a monoclonal chimeric anti-CD20 antibody, has been widely used as a chemoimmunotherapeutic regimen in the frontline therapy for patients with CD20+ BL and diffuse large B-cell lymphoma. The addition of rituximab to the CHOP backbone or to standard FAB/LMB therapy has greatly improved outcomes without significantly increasing toxicity in patients with B-NHL (Goldman/Cairo, Leukemia, 2013, Coiffier et al, NEJM, 2002). However, patients who relapse have a poor clinical response to rituximab retreatment. Obinutuzumab is a humanized, type II anti-CD20 monoclonal antibody glycoengineered to enhance Fc receptor affinity. It has lower complement-dependent cytotoxicity than rituximab but greater ADCC, phagocytosis and direct B-cell killing effects (Chu/Cairo, BJH, 2018). Obinutuzumab has been successfully utilized in front-line therapy in FLL (Marcus, et al, NEJM, 2017) and CLL (Goede, et al, NEJM, 2014; Moreno, et al, Lancet, 2019). Our group has successfully expanded functional and active peripheral blood NK cells PBNKwith irradiated feeder cells to target B-NHL (Chu/Cairo, et al, Can Imm Res 2015). We previously demonstrated that obinutuzumab has significantly enhanced expanded PBNK mediated cytotoxicity against BL and pre-B-ALL cell lines compared to rituximab (Tiwari/Cairo et al, BJH, 2015). NKTR-255 is an IL-15 receptor agonist designed to activate the IL-15 pathway and expand natural killer (NK) cells and promote the survival and expansion of memory CD8+ T cells without inducing suppressive regulatory T cells (Kuo/Zalevsky, Cancer Res. 2017). NKTR-255 stimulates proliferation and survival of NK, CD8+ T cells, and enhances long-term immunological memory which may lead to sustained anti-tumor immune response. Objective: To investigate the effects of NKTR-255 on the ADCC of expanded NK cells with anti-CD20 type I and type II antibodies against CLL, FL and rituximab-resistant BL. Methods: NK cells were expanded with lethally irradiated K562-mbIL21-41BBL cells as previously described (Denman/Dean Lee, PLoS One, 2012). Expanded PBNK cells were isolated using Miltenyi NK cell isolation kit. NKTR-255 was generously provided by Nektar Therapeutics. In vitro cytotoxicity was examined using luminescence reporter-based assays. IFNg, granzyme B and perforin levels were examined by standard enzyme-linked immunosorbent assays as we previously described (Chu/Cairo, ASH, 2018). MEC-1 (CLL), PGA-1 (CLL), DOHH2 (FL) and Rituximab-resistant BL cells Raji-2R and Raji-4RH were used as target cells. Results: NKTR-255 significantly enhanced the in vitro cytotoxicity of expanded NK cells when combined with rituximab against MEC-1 (E:T=3:1, p&lt;0.001), PGA-1 (E:T=3:1, p&lt;0.001), and DOHH2 (E:T=3:1, p&lt;0.001) as compared to the control groups (Fig.1A). NKTR-255 also significantly enhanced granzyme and perforin release from expanded NK cells when combined with rituximab against MEC-1 (granzyme: p&lt;0.05; perforin: p&lt;0.001), PGA-1(granzyme: p&lt;0.05; perforin: p&lt;0.05), DOHH2 (granzyme: p&lt;0.05; perforin: p&lt;0.001) as compared to controls. NKTR-255 significantly enhanced the in vitro cytoxicity of expanded NK cells when combined with obinutuzumab agains rituximab-resistant BL cells like Raji-2R (E:T=3:1, p &lt;0.01), and Raji-4RH (E:T=3:1, p&lt;0.01) as compared to the control groups (Fig.1B). NKTR-255 also significantly enhanced IFN-g, granzyme and perforin release from expanded NK cells when combined with obinutuzumab against Raji-2R (E:T=3:1, IFN-g: p&lt;0.001, granzyme: p&lt;0.001 and perforin: p&lt;0.001) and Raji-4RH (E:T=3:1, IFN-g: p&lt;0.001, granzyme: p&lt;0.01 and perforin: p&lt;0.01) as compared to controls. Conclusion: We found that NKTR-255 significantly enhanced the ADCC of expanded NK cells with anti-CD20 type I and type II antibodies against CLL, FL and rituximab-resistant BL cells in vitro with enhanced IFN-g, granzyme B and perforin release. The in vivo effects of NKTR-255 with expanded NK cells and anti-CD20 type I and type II antibodies against CLL, FL and rituximab-resistant BL cells using humanized NSG models are under investigation. Disclosures Lee: Kiadis Pharma Netherlands B.V: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Madakamutil:Nektar Therapeutics: Current Employment. Marcondes:Nektar Therapeutics: Current Employment. Klein:Roche: Current Employment, Current equity holder in publicly-traded company, Patents & Royalties. Cairo:Nektar Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Research Funding; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Miltenyi: Research Funding; Technology Inc/Miltenyi Biotec: Research Funding.

scholarly journals Anti-CD27 Enhances Lymphoma Immunotherapy through Profound Myeloid Cell Recruitment

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3024-3024
Author(s):  
Anna H Turaj ◽  
Vikki L Field ◽  
Claude H.T. Chan ◽  
Christine A. Penfold ◽  
Jinny H. Kim ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Nk Cells ◽  
Research Funding ◽  
Therapeutic Potential ◽  
Cell Lymphoma ◽  
Myeloid Cells ◽  
B Cell Lymphoma ◽  
Anti Cd20

Abstract Direct-targeting monoclonal antibodies (mAb) such as anti-CD20 mAb are thought to elicit their anti-tumor function through antibody-dependent cellular phagocytosis (ADCP) mediated by myeloid cells (monocytes and macrophages), with little involvement of T cells. In contrast, immunomodulatory mAbs to TNFR superfamily members, CD27, OX40 and CD137, function by augmenting T-cell responses. We examined the therapeutic potential of combining anti-CD20 mAb with a panel of immunomodulatory mAbs (OX40, CD137, CD27, TIGIT, GITR, CTLA4, PD-1). In the syngeneic BCL1 B-cell lymphoma mouse model only an agonistic mAb to CD27, provided a synergistic effect when combined with anti-CD20. Anti-CD20 and anti-CD27 mAb individually provided modest therapeutic benefit (median survival 33 days and 62 days, respectively), but mice treated with the combination survived beyond 100 days. Similar synergistic survival benefit was observed in another B-cell lymphoma model, A31, and in BCL1-bearing human CD27 transgenic mice, when anti-CD20 was combined with varlilumab, an anti-human CD27 mAb currently under clinical investigation. We observed that in mice treated with anti-CD27, there was an early and substantial increase in intra-tumoral monocyte, neutrophil and macrophage infiltration. CD27 is expressed constitutively on T and NK cells but not myeloid cells or the tumor itself. To investigate whether CD27 agonism promotes intra-tumoral myeloid cell infiltration through T cells, we depleted T cells in the BCL1model. Surprisingly, CD4 or CD8 T-cell depletion had no effect on the survival of anti-CD20 and anti-CD27-treated mice, suggesting that the remaining CD27+ immune effector cells, NK cells, are required. To further probe the relative importance of these two sub-sets, we performed experiments in γ chain knockout mice, where activatory FcγR are not expressed. Here, anti-CD27 mediated T-cell activation can still occur via crosslinking from the inhibitory FcγRII, but effector function through NK cells, mediated through activatory FcγR, is abrogated. In this model, the therapeutic benefit of anti-CD27 was completely abolished, thereby supporting the requirement for NK cells. We hypothesize that anti-CD27 stimulates CD27+ NK cells to release chemokines that draw myeloid cells into the tumor, where they subsequently perform augmented anti-CD20 mediated ADCP. These data demonstrate the clear therapeutic potential of combining direct targeting and immunomodulatory mAb but that the therapeutic mechanism of action may differ to that expected; here involving a previously unheralded effect of anti-CD27 on myeloid infiltration. Based upon these data, we have implemented a phase II clinical trial examining rituximab and varlilumab in B-cell lymphoma, which will commence recruitment shortly. Disclosures Keler: Celldex Therapeutics: Employment, Equity Ownership. Johnson:Celldex Therapeutics: Research Funding. Al-Shamkhani:Celldex Therapeutics: Patents & Royalties: On therapeutic use of antibodies targeting anti-CD27 and another applied for anti-CD20/anti-CD27 use, Research Funding. Glennie:Celldex Therapeutics: Patents & Royalties: Patent on therapeutics use of antibodies targeting human CD27 and patent for anti-CD20+anti-CD27 applied. Cragg:Baxalta: Consultancy; Gilead Sciences: Research Funding; GSK: Research Funding; Roche: Consultancy, Research Funding; Bioinvent International: Consultancy, Research Funding. Lim:Celldex Therapeutics: Patents & Royalties: Patent for anti-CD20+anti-CD27 applied, Research Funding.

Targeting B-Cell Malignancies With Anti-CD20-Interleukin-21 Fusokine

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 377-377 ◽  
Author(s):  
Shruti Bhatt ◽  
Daxing Zhu ◽  
Xiaoyu Jiang ◽  
Seung-uon Shin ◽  
John M Timmerman ◽  
...  
Keyword(s):  
Cell Death ◽  
B Cell ◽  
Nk Cells ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Cd20 Antibody ◽  
Anti Cd20

Abstract The anti-CD20 antibody rituximab has revolutionized the treatment for B cell non-Hodgkin lymphomas (NHLs). However, rituximab has limited effectiveness as a single agent in some NHL subtypes and its clinical efficacy is compromised by acquired drug resistance. As a result, many patients still succumb to NHLs. Hence, strategies that enhance the activity of anti-CD20 antibody may improve patient outcome. Interleukin-21 (IL21), a member of the IL2 cytokine family, exerts diverse regulatory effects on natural killer (NK), T and B cells. IL21 has been reported to possess potent anti-tumor activity against a variety of cancers not expressing IL21 receptor (IL21R) through activation of the immune system and is in clinical trials for renal cell carcinoma and metastatic melanoma. We have recently reported that apart from immuno-stimulatory effects, IL21 exerts direct cytotoxicity on IL21R expressing diffuse large B cell lymphoma (DLBCL) and mantle cell lymphoma (MCL) cell lines and primary tumors both in vitro as well in vivo (Sarosiek et al Blood 2010; Bhatt et al AACR 2013). Herein we designed a fusion protein comprising IL21 linked to the N-terminus of anti-CD20 antibody (αCD20-IL21 fusokine) to improve efficacy of its individual components and prolong IL21 half-life. We have verified the expression of full length fusion protein and demonstrated that αCD20-IL21 fusokine retained binding ability to its individual components; CD20 and IL21R, as analyzed by immunofluorescence and flow-cytometry analyses. Similar to our previous study of IL21 in DLBCL, treatment of B cell lymphoma cell lines with fusokine lead to phosphorylation of STAT1 and STAT3, upregulation of cMYC and BAX and downregulation of BCL-2 and BCL-XL, implying the activation of IL21R dependent signaling to trigger cytotoxic effects. In vitro, direct cell death induced by αCD20-IL21 fusokine in DLBCL (RCK8, WSU and Farage) and MCL (Mino, HBL2 and SP53) cell lines was markedly increased compared to its individual components (IL21 and parent αCD20-IgG1 antibody). More importantly, fusokine treatment resulted in cell death of MCL cell lines (L128, G519 and UPN1) that were found to be resistant to IL21 alone treatment. Furthermore, treatment of freshly isolated primary NHL cells with the αCD20-IL21 fusokine also exhibited a 40-50% increase in direct cell death compared to its individual components. Previous studies reported that IL21 enhances antibody-dependent cellular cytotoxicity (ADCC) of therapeutic antibodies by activation of NK cells. ADCC assays using chromium release with purified human NK cells demonstrated that ADCC induced by the parent antibody was enhanced in the presence of IL21 while IL21 alone had minimal effect on the lysis of Raji, Daudi, and Jeko1 target cells. Notably, αCD20-IL21 fusokine demonstrated increased ADCC activity in comparison to parent antibody plus IL21 in Raji, Daudi and Jeko-1 cells (p<0.001, p<0.005 and p<0.001, respectively). Similar results were obtained in primary MCL tumor cells. Consistent with this finding, fusokine treatment resulted in enhanced activation of the NK cells as assessed by CD69 upregulation and CD16 downregulation using flow-cytometry. Complement dependent cytotoxicity (CDC) of the fusokine was similar to the parent antibody and rituximab in Raji cells. Studies analyzing in vivo effects of the fusokine are in progress and will be presented at the meeting. These data strongly suggest that together with direct apoptotic potential, an anti-CD20 IL21 fusokine retains the ability to trigger indirect cell killing mediated via activation of immune effector cells. These dual effects may give remarkable advantage to the fusokine over existing anti-CD20 antibodies for the treatment of NHL tumors. Collectively, our study demonstrates that anti-tumor effects of IL21 and anti-CD20 antibodies can be enhanced by conjugation of IL21 with anti-CD20 antibody that may serve as a novel anti-lymphoma therapy. Disclosures: Rosenblatt: Seattle Genetics, Inc.: Research Funding.

scholarly journals Imaging the mechanisms of anti-CD20 therapy in vivo uncovers spatiotemporal bottlenecks in antibody-dependent phagocytosis

2021 ◽  
Vol 7 (8) ◽  
pp. eabd6167
Author(s):  
Capucine L. Grandjean ◽  
Zacarias Garcia ◽  
Fabrice Lemaître ◽  
Béatrice Bréart ◽  
Philippe Bousso
Keyword(s):  
B Cell ◽  
Cell Lymphoma ◽  
Critical Path ◽  
B Cell Lymphoma ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Fluorescent Reporter ◽  
Cd20 Antibody ◽  
Anti Cd20 ◽  
Partial Depletion

Anti-CD20 antibody (mAb) represents an effective strategy for the treatment of B cell malignancies, possibly involving complement activity, antibody-dependent cellular cytotoxicity and phagocytosis (ADP). While ADP by Kupffer cells deplete circulating tumors, mechanisms targeting non-circulating tumors remain unclear. Using intravital imaging in a model of B cell lymphoma, we establish here the dominance and limitations of ADP in the bone marrow (BM). We found that tumor cells were stably residing in the BM with little evidence for recirculation. To elucidate the mechanism of depletion, we designed a dual fluorescent reporter to visualize phagocytosis and apoptosis. ADP by BM-associated macrophages was the primary mode of tumor elimination but was no longer active after one hour, resulting in partial depletion. Moreover, macrophages were present at low density in tumor-rich regions, targeting only neighboring tumors. Overcoming spatiotemporal bottlenecks in tumor-targeting Ab therapy thus represents a critical path towards the design of optimized therapies.

Anti-CD20 therapy corrects a CD8 regulatory T cell deficit in multiple sclerosis

2021 ◽  
pp. 135245852110033
Author(s):  
Quentin Howlett-Prieto ◽  
Xuan Feng ◽  
John F Kramer ◽  
Kevin J Kramer ◽  
Timothy W Houston ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
B Cell ◽  
Regulatory T Cell ◽  
Healthy Controls ◽  
Regulatory Cells ◽  
Long Term Treatment ◽  
Anti Cd20

Objective: To determine the effect of long-term anti-CD20 B-cell-depleting treatment on regulatory T cell immune subsets that are subnormal in untreated MS patients. Methods: 30 clinically stable MS patients, before and over 38 months of ocrelizumab treatment, were compared to 13 healthy controls, 29 therapy-naïve MS, 9 interferon-β-treated MS, 3 rituximab-treated MS, and 3 rituximab-treated patients with other autoimmune inflammatory diseases. CD8, CD28, CD4, and FOXP3 expression in peripheral blood mononuclear cells was quantitated with flow cytometry. Results: CD8+ CD28− regulatory cells rose from one-third of healthy control levels before ocrelizumab treatment (2.68% vs 7.98%), normalized by 12 months (13.5%), and rose to 2.4-fold above healthy controls after 18 months of ocrelizumab therapy (19.0%). CD4+ FOXP3+ regulatory cells were lower in MS than in healthy controls (7.98%) and showed slight long-term decreases with ocrelizumab. CD8+ CD28− and CD4+ FOXP3+ regulatory T cell percentages in IFN-β-treated MS patients were between those of untreated MS and healthy controls. Interpretation: Long-term treatment with ocrelizumab markedly enriches CD8+ CD28− regulatory T cells and corrects the low levels seen in MS before treatment, while slightly decreasing CD4+ FOXP3+ regulatory T cells. Homeostatic enrichment of regulatory CD8 T cells provides a mechanism, in addition to B cell depletion, for the benefits of anti-CD20 treatment in MS.

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