scholarly journals In vivo effects of monoclonal antibodies to distinct epitopes of Qa-2 antigens.

1990 ◽  
Vol 171 (1) ◽  
pp. 211-219
Author(s):  
Y Sharabi ◽  
D H Sachs
Keyword(s):  
T Cell ◽  
Cell Depletion ◽  
Maximal Effect ◽  
T Cell Depletion ◽  
Natural Ligand ◽  
Significant Prolongation ◽  
Cell Immunity ◽  
Cell Clearance

The effects of in vivo treatment with anti-Qa-2 mAbs on in vivo and in vitro parameters of T cell immunity have been examined. Two anti-Qa-2 mAbs of the same isotype and with similar avidities but directed against distinct epitopes of the same Qa-2 molecules were studied. mAb 1-1-2 was found to induce rapid T cell depletion, with maximal effect observed within 2-3 d, while administration of mAb 1-9-9 caused little or no depletion in the first few days, and reached maximal effect only by day 8. Surprisingly, administration of both antibodies resulted in a depletion pattern similar to that of the nondepleting antibody 1-9-9. Consistent with these effects on T cell depletion, treatment with 1-1-2 caused significant prolongation of survival of allogeneic skin grafts placed 1 d after antibody administration, while treatment with 1-9-9 or with the combination of both antibodies caused no prolongation. In an attempt to determine the mechanism of this phenomenon, we examined Qa-2 expression on the cell surface by flow microfluorometry after treatment with each of the two mAbs. Our data indicate that mAb 1-9-9 mediates significantly greater modulation of Qa-2 expression from the surface of peripheral T cells within 1 d than does mAb 1-1-2. Apparently, therefore, modulation occurs more rapidly than cell clearance, and the efficiency of T cell depletion and consequent immune suppression is correlated inversely with the ability of each mAb to cause modulation. The ability of 1-9-9 to cause Qa-2 modulation suggests that it may react with a determinant on this molecule of physiological relevance to the natural ligand interactions of Qa-2 antigens.

Post-Transplant Immune Suppressive Therapy Is Not Necessary In Related and Unrelated Reduced Intensity Conditioning Stem Cell Transplantation Using Low Dose Alemtuzumab In Vivo T Cell Depletion Combined with Alemtuzumab-Mediated In Vitro T Cell Depletion of the Graft

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2328-2328
Author(s):  
P.A. Von Dem Borne ◽  
C.J.M. Halkes ◽  
C.W.J. Starrenburg ◽  
W.A.F. Marijt ◽  
J.H.F. Falkenburg
Keyword(s):  
T Cell ◽  
Low Dose ◽  
Chronic Gvhd ◽  
Cell Depletion ◽  
Suppressive Therapy ◽  
T Cell Depletion ◽  
Post Transplant ◽  
Immune Suppressive Therapy

Abstract Abstract 2328 Introduction T cell depletion with alemtuzumab administered in vivo to the patient reduces the risk of graft-versus-host disease (GVHD) and graft rejection following reduced intensity conditioning allogeneic stem cell transplantation (RIC SCT). However, high doses of alemtuzumab can result in delayed immune reconstitution, increased non relapse mortality (NRM) due to infections, and potential loss of graft versus tumor responses. Recently, the feasibility of T cell depletion with low dose in vivo alemtuzumab was demonstrated in HLA-identical related RIC SCT. Dose reduction of alemtuzumab to 30 mg combined with post-transplant immune suppressive therapy with cyclosporine tapered from 3 months after transplantation resulted in a low risk of GVHD, no increase in NRM and improved lymphocyte recovery (Chakraverty et al, Blood pre-published online June 29, 2010). Early immunotherapeutic intervention after SCT with donor lymphocytes may be hampered by the administration of post-transplant immune suppressive therapy. We investigated whether in RIC SCT using low dose in vivo alemtuzumab, post-transplant immune suppressive therapy can be replaced by alemtuzumab-mediated in vitro T cell depletion of the graft just prior to infusion (“Campath in the bag”). Patient and donor characteristics Between 2007 and 2009, 29 patients were transplanted with an unrelated donor, and 28 patients with a related donor using a RIC regimen consisting of fludarabine (50 mg/m2 p.o. day -10 to -5), busulphan (3.2 mg/kg i.v. day -6 and -5) and alemtuzumab (15 mg i.v. day -4 and -3), followed by infusion of the graft after in vitro incubation with 20 mg alemtuzumab. No additional immune suppressive therapy was used after SCT. Unrelated donors were matched for HLA-A, B, C, DR and DQ, three patient donor combinations had one HLA-DQ mismatch. Median patient age was 59 years (range 21–72). Remission status at the time of transplant was: 47% CR, 37% PR, 9% SD, 7% PD. Indications were diverse (19 AML, 16 myeloma, 6 CLL, 5 low grade NHL, 3 aggressive T-NHL, 2 aggressive B-NHL, 2 SAA, 1 CML, 1 myelofibrosis, 1 CMML, 1 ALL). The unrelated and related transplanted group were comparable regarding age, disease and remission status. The median Gratwohl transplantation risk score was 3 in the related group (range 1–5), and 5 in the unrelated group (range 3–6). Results All patients engrafted; platelet numbers of 50 × 109/L were reached after a median of 11 days (range 0–38 days), neutrophil numbers of 0.5 × 109/L were reached after a median of 18 days (range 0–161 days) post transplant. Two patients had secondary graft failure. In patients transplanted with a related donor, grade 1–2 and 3–4 acute GVHD was observed in 36% and 4% of evaluable patients, respectively, resolving in all patients without development of chronic GVHD. NRM was 0% at 3 months and 4% at 1 year. Overall survival was 100% at 3 months and 89% at 1 year. In patients transplanted with an unrelated donor more acute GVHD was observed (59% grade 1–2, 15% grade 3–4 of evaluable patients). 24% of evaluable patients developed chronic GVHD, which was limited in 75% and extensive in 25% of these patients. Chronic GVHD resolved in most patients, one patient has ongoing extensive chronic GVHD. NRM was 7% at 3 months and 24% at 1 year. Overall survival was 93% at 3 months and 55% at 1 year. Conclusions RIC SCT using low dose alemtuzumab in vivo T cell depletion combined with alemtuzumab-mediated in vitro T cell depletion of the graft without additional post-transplant immune suppressive therapy is feasible in patients transplanted with related and unrelated donors. Results are excellent in patients transplanted with related donors with low NRM and high overall survival. Although results are good in patients transplanted with unrelated donors considering the high Gratwohl score in this group, further improvement is being sought by increasing the efficiency of T cell depletion. This RIC SCT regimen without post-transplant immune suppressive therapy is an appropriate platform for early cellular immunotherapeutic interventions including donor lymphocyte infusion after SCT. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Haploidentical Allogeneic Stem Cell Transplantation in Sickle Cell Disease: A Systematic Review and Meta-Analysis

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 424-424
Author(s):  
Mesire Aydin ◽  
Elisabeth Dovern ◽  
Mariska M.G. Leeflang ◽  
Josu de la Fuente ◽  
Adetola A. Kassim ◽  
...  
Keyword(s):  
Overall Survival ◽  
T Cell ◽  
Research Funding ◽  
Graft Failure ◽  
Meta Analysis ◽  
Cell Depletion ◽  
T Cell Depletion ◽  
Related Mortality

Abstract Background Allogeneic hematopoietic stem cell transplantation (SCT) is the only established curative treatment option for patients with sickle cell disease (SCD). Transplantations with an HLA-identical matched sibling donor (MSD) have resulted in excellent disease-free survival of >90% and overall survival (OS) of >95%. However, lack of HLA-identical siblings is a limiting factor. The chance of finding a potential matched unrelated donor is low for patients with a non-Western ethnic background (< 20%). Haploidentical related donors are a promising pool of donors potentially extending SCT as a curative treatment to a larger group of SCD patients with no other meaningful treatment options. Myeloablative conditioning regimens (MAC) are not recommended for adult SCD patients, as cumulative SCD-related organ damage renders these patients susceptible to increased toxicity and higher risk of transplant-related mortality. Using reduced-intensity conditioning (RIC) or non-myeloablative conditioning (NMC) in both adult and pediatric patients has resulted in decreased transplant-related toxicity and mortality. However, while successful in hematologic malignancies, NMC has been associated with significant risk of graft failure in hemoglobinopathies. In the present study, we aimed to systematically review (1) the outcomes of haploidentical SCT (Haplo-SCT), (2) the effects of conditioning intensity and modes of T-cell depletion on Haplo-SCT outcomes and (3) comparative outcomes between matched sibling donor SCT (MSD-SCT) and Haplo-SCT in selected studies. Methods A comprehensive search was performed in MEDLINE/PubMed and Embase up to May 2021. Data was extracted by two reviewers independently and the Newcastle-Ottawa Quality Assessment Scale was used to assess the quality of the studies. Only studies reporting at least one of the outcomes: graft failure, OS, transplant-related mortality, and acute/chronic graft-versus-host disease (GvHD) were included. Fourteen studies met the inclusion criteria. To have an overview of the results of Haplo-SCT, we divided the included studies in four groups according to the conditioning intensity (MAC versus NMC/RIC) and the T-cell depletion method (in vivo (post-transplant cyclophosphamide (PTCy)) vs. in vitro). Results All included studies were observational cohort studies. A subgroup meta-analysis of the results of Haplo-SCT showed relatively low overall pooled proportions of graft failure (7%, 95% CI: 2 - 20), acute (4%, 95% CI: 2 - 12) and chronic (11%, 95% CI: 7 - 16) GvHD. Overall survival was high (91%, 95% CI: 85 - 94). Graft failure in MAC-in vitro, MAC-in vivo, NMC/RIC-in vitro and NMC/RIC-in vivo groups was 4% (95% CI: 1 - 14), 0% (95% CI: 0 - 100), 25% (95% CI: 10 - 51) and 11% (95% CI: 3 - 36) respectively (Figure 1). OS was 100% and 93% for MAC and NMC/RIC groups with PTCy (in vivo T-cell depletion) respectively. In patients with in vitro T-cell depletion, OS was 87% and 81% in MAC and NMC/RIC groups respectively (Figure 2). Based on a comparative meta-analysis of the three studies that included both haploidentical and MSD transplantation, graft failure was significantly higher in the haploidentical group than in the MSD group (odds ratio 5.3, 95% CI: 1.0 - 27.6). Overall survival, transplant-related mortality and acute/chronic GvHD were not significantly different between the groups. Conclusions This systematic review shows that modifications in the intensity of the conditioning regimen and improved T-cell depletion approaches in Haplo-SCT in SCD have led to reduced transplantation-related toxicity while keeping graft failure rates low. Both in vitro and in vivo (PTCy) T-cell depletions result in very low transplantation-related mortality, though in vitro T-cell depletion is associated with a higher incidence of viral reactivations and other infectious complications. Haploidentical stem cell transplantation is becoming a viable alternative curative option for SCD, extending the availability of allogeneic SCT as a treatment option to many more transplant eligible SCD patients. Novel immunosuppression (immunoablation) and improvement in supportive care have allowed the use of RIC or NMC regimens, resulting in low risk of transplantation-related complications and improvement in engraftment rates. Figure 1 Figure 1. Disclosures Biemond: Global Blood Therapeutics: Honoraria, Research Funding, Speakers Bureau; Novartis: Honoraria, Research Funding, Speakers Bureau; CSL Behring: Honoraria; Sanquin: Research Funding; Novo Nordisk: Honoraria; Celgene: Honoraria. Nur: Celgene: Speakers Bureau; Roche: Speakers Bureau; Novartis: Research Funding, Speakers Bureau.

Low Incidence of Post-Transplant EBV-Related Disease After Alemtuzumab-Mediated T Cell Depletion Is Explained by the Differential Susceptibility to Alemtuzumab of B Cells and Protective CD8 and CD4 T Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2330-2330
Author(s):  
Constantijn J.M. Halkes ◽  
Inge Jedema ◽  
Judith Olde Wolbers ◽  
Esther M van Egmond ◽  
Peter A. Von Dem Borne ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Cell Depletion ◽  
T Cell Depletion

Abstract Abstract 2330 In vivo T cell depletion with anti-thymocyte globulin (ATG) or alemtuzumab (anti-CD52) before reduced intensity allogeneic stem cell transplantation (alloSCT) in combination with in vitro T cell depletion with alemtuzumab reduces the risk of GVHD. Detectable levels of circulating antibodies are present up to several months after the alloSCT, leading to a delayed immune reconstitution which is associated with an increased incidence of opportunistic infections and early relapses. Prior to 2007, combined in vitro (Alemtuzumab 20 mg added “to the bag”) and in vivo T cell depletion with horse-derived ATG (h-ATG) resulted in good engraftment without GVHD in the absence of GVHD prophylaxis after reduced intensity alloSCT using conditioning with fludarabine and busulphan. Due to the unavailability of h-ATG, rabbit-derived ATG (r-ATG) 10–14 mg/kg was introduced in the conditioning regimen in 2007. Strikingly, in this cohort of patients, early EBV reactivation and EBV-associated post-transplantation lymphoproliferative disease (PTLD) was observed in 10 out of 18 patients at a median time of 6 weeks after alloSCT (range 5 to 11 weeks) in the absence of GVHD or immunosuppressive treatment. Analysis of T and B cell recovery early after transplantation revealed preferential depletion of T cells as compared to B cells, thereby allowing unrestricted proliferation of EBV infected B cells. Due to this unacceptable high incidence of EBV-related complications, in the conditioning regimen r-ATG was replaced by low dose alemtuzumab (15 mg i.v. day -4 and -3) in 2008. In this cohort of 60 patients, only 2 patients experienced transient EBV reactivation during the first 3 months after alloSCT and one patient developed an EBV-associated lymphoma 4 weeks after alloSCT. To investigate the mechanisms underlying the low incidence of EBV reactivation using alemtuzumab for T cell depletion, we studied the in vivo and in vitro effects of alemtuzumab on different lymphocyte subsets. First, lineage-specific reconstitution was studied in 20 patients from the alemtuzumab cohort with known CD52 negative diseases (11 AML and 9 multiple myeloma) to exclude the confounding effect of antibody absorption by malignant cells. Whereas at 3 weeks after alloSCT detectable numbers of circulating NK cells and T cells were observed (medians 71 (range 6–378), and 12 (range 1–1164)E6/L, respectively), no circulating B cells could be detected (median 0, range 0–1 E6/L). At 6 weeks after alloSCT, NK and T cell numbers further increased (medians 212 (52-813), and 130 (range 25–1509)E6/L, respectively), whereas B cell numbers still remained low in the majority of patients (median 15, range 0–813E6/L). In all patients, T cells were detectable before the appearance of circulating B cells. Furthermore, the expression of CD52 and the sensitivity to alemtuzumab-mediated complement-dependent cell lysis (CDC) of B cells, T cells and NK cells was measured in vitro. The highest CD52 expression was observed on B cells (mean fluorescence intensity (MFI) 120), resulting in 95% lysis after incubation with 10ug/mL alemtuzumab and rabbit complement. NK cells showed a significantly lower CD52 expression (MFI 41), which was also reflected by a lower susceptibility to alemtuzumab-mediated CDC (62% lysis). Interestingly, differential expression of CD52 was observed on CD4 and CD8 T cells (MFI 120 and 101, respectively). Cytotoxicity analysis revealed relative protection of CD8 compared to CD4 T cells against alemtuzumab-mediated CDC, resulting in 52% and 90% lysis, respectively. Based on these results, we investigated in detail the presence and phenotype of the CD4 and CD8 subsets and EBV-specific CD8 T cells using tetramer staining at 6 weeks after alloSCT. In accordance with the in-vitro expression and susceptibility data, circulating CD52+ CD8 T cells including EBV-specific T cells were detectable. Interestingly, the majority of circulating CD4 T cells (64-93%, n=4) lacked CD52 expression, explaining their capacity to persist in the presence of alemtuzumab. We conclude that in vivo and in vitro T cell depletion with alemtuzumab is associated with a relatively low risk of EBV-associated PTLD because of efficient B cell depletion and persistent EBV immunity allowed by the relative insusceptibility for alemtuzumab of CD8 T cells and the development of CD52 negative escape variants of CD4 T cells. Disclosures: No relevant conflicts of interest to declare.

In Vivo T Cell Depletion Using Rabbit Derived ATG Leads to An Increased EBV-PTLD Risk Due to An Induced Imbalance Between B and T Cell Recovery Which Is Not Seen After Horse Derived ATG or Alemtuzumab

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1979-1979 ◽  
Author(s):  
C.J.M. Halkes ◽  
J.H.F. Falkenburg ◽  
H.M. van Egmond ◽  
J. Olde Wolbers ◽  
C.W.J. Starrenburg ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Cd8 T Cells ◽  
Cell Depletion ◽  
Cell Recovery ◽  
Post Transplantation ◽  
T Cell Depletion

Abstract Abstract 1979 Control of replication of endogenous viruses like CMV and EBV is fully dependent on CMV or EBV specific T cells after allogeneic stem cell transplantation (alloSCT). In the absence of specific CD8 T cell control, proliferation of EBV infected B cells can lead to post transplantation lymphoproliferative disease (PTLD). In an initial cohort of patients treated with horse derived anti thymocyte globulin (h-ATG), no early PTLD was observed. However, due to unavailability in Europe, h-ATG had to be replaced by rabbit derived ATG (r-ATG), leading to an unacceptable high incidence of EBV-PTLD (26% during first 3 months after alloSCT). Replacement of r-ATG by alemtuzumab (ALT) significantly reduced the incidence of EBV-PTLD (3 months incidence of EBV-PTLD 2%). To determine the immunological basis of these findings we performed a detailed analysis of immune reconstitution in these three cohorts of transplanted patients. The first cohort (41 patients) received h-ATG (Lymphoglobulin) 10 mg/kg/day for 4 days. The second cohort (19 patients) received r-ATG (Thymoglobulin) 2.0 or 3.5 mg/kg/day for 4 days and the third cohort (60 patients) received ALT, 15 mg/day for 2 days. All grafts consisted of PBSC to which 20 mg of ALT was added for in vitro T cell depletion. All patients received a fludarabin and busulphan based conditioning regimen. No standard post transplantation immunosuppressive treatment was given. In the r-ATG cohort, early EBV-PTLD occurred after a median of 7 weeks (range 4–12 weeks) post alloSCT. Three r-ATG treated patients died while high levels of circulating EBV-DNA were present (> log 4.0 copies/mL). Incidence of CMV disease was not significantly different in the three cohorts (5%, 6% and 0%, respectively). In contrast to the other 2 cohorts, immune reconstitution in the r-ATG cohort was characterized by an imbalance between recovery of B cells and CD8 T cells. Already 3 weeks after alloSCT, the majority (67%) of r-ATG patients showed a more rapid reconstitution of B cells than CD8 T cells, leading to B cells outnumbering CD8 T cells. This was seen in only a small minority of patients after h-ATG and ALT (17% and 6%, respectively, p<0.01 versus r-ATG). Because rapid recovery of T cells in the alemtuzumab patients was frequently found in the presence of circulating ALT (mean concentration 0.43 μg/mL and 0.12 μg/mL after 3 and 6 weeks, respectively), the phenotype of circulating CD4 and CD8 T cells at 6 weeks after ALT was analyzed. The majority of circulating CD8 and CD4 T cells lacked CD52 expression (56% (range 0–99%) and 81% (range 0–93%), respectively). Using tetramer staining, cytotoxicity assays and analysis of cytokine production, we demonstrated the presence of functional CD52 negative as well as CD52 positive CMV and EBV specific CD8 T cells. Based on FLAER negativity, it was demonstrated that the CD52 negative T cells are GPI anchor deficient, representing a PNH-like clone escaping ALT induced cell lysis. Because almost half of the circulating CD8 T cells were CD52 positive, we examined expression of CD52 and the in-vitro sensitivity to ALT-mediated complement-dependent cell lysis (CDC) of B cells, CD4 and CD8 T cells of healthy donors. The highest CD52 expression was observed on B cells (mean fluorescence intensity (MFI) 120), resulting in 95% lysis after incubation with ALT and complement. Differential expression of CD52 was observed on CD4 and CD8 T cells, MFI 120 and 101 respectively, resulting in relative protection of CD52 positive CD8 compared to CD4 T cells against ALT-mediated CDC (52% and 90% lysis). We conclude that the high incidence of EBV-PTLD after in-vivo T cell depletion with r-ATG is caused by an induced imbalance between B and T cell recovery, which is not seen after h-ATG or ALT. In-vivo T cell depletion with ALT is associated with a relatively low risk of EBV disease because of efficient B cell depletion and persistent EBV immunity due to the relative insusceptibility for ALT of CD8 T cells and the development of functional CD52 negative escape variants of CD4 and CD8 T cells. Disclosures: Off Label Use: Alemtuzumab and Anti Thymocyte Globulin used for in vivo T cell depletion prior to allogeneic stem cell transplantation.

Improved Long Term Survival with Minimal GVHD after Myeloablative Unrelated Donor Stem Cell Transplantation Using In Vitro and In Vivo T Cell Depletion with CAMPATH-1H.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2761-2761
Author(s):  
Peter A. von dem Borne ◽  
Floor Beaumont ◽  
Ingrid Starrenburg ◽  
Machteld A. Oudshoorn ◽  
Geoff Hale ◽  
...  
Keyword(s):  
Stem Cell ◽  
T Cell ◽  
Acute Gvhd ◽  
Chronic Gvhd ◽  
Cell Depletion ◽  
Unrelated Donor ◽  
T Cell Depletion ◽  
Related Mortality

Abstract In allogeneic stem cell transplantation (SCT) T-cell depletion reduces transplant related mortality by diminishing GVHD. We have investigated a myeloablative regimen for matched unrelated donor SCT using both in vivo and in vitro CAMPATH-1H for effective T-cell depletion, utilising DLI at a later time point for graft versus tumor effect if necessary. Thirty patients (median age 33 years, range 18–48) were transplanted from January 1997 to June 2002. Diagnoses were: CML CP (n=9), CML AP (n=2), AML/MDS (n=9), ALL (n=8), NHL (n=1) and Fanconi anemia (n=1). Six patients had one HLA mismatch, the others were identical for HLA A, B, C, DR and DQ. Conditioning consisted of CAMPATH-1H 5mg/d on days −8 to −4, TBI 6 Gy on days −8 and −7 and cyclofosfamide 60 mg/kg on days −6 and −5. T-cell depletion was performed by in vitro incubation of the graft with 20 mg CAMPATH-1H for 30 minutes (Campath “in the bag”). Post-transplant GVHD prophylaxis consisted of cyclosporine A and methotrexate. The stem cell source was bone marrow in 19 patients (63%) and peripheral blood in 11 patients. One graft failure was observed, all other patients had sustained engraftment of donor cells. Acute GVHD was observed in 12 patients (40%), maximally grade I-II skin. No severe acute GVHD (grade III-IV) was experienced. Limited chronic GVHD developed in 2 patients, resolving after treatment. Only in one patient extensive chronic GVHD developed, which did not resolve. CMV reactivation occurred in 23% of patients, one patient developed CMV disease. No EBV disease was observed. Ten patients received donor lymphocyte infusion (DLI) at a median of 17.4 months after SCT (8 patients with relapsed CML, one patient with relapsed ALL, one patient with autoimmune hemolytic anemia). After DLI acute GVHD grade I-II developed in 4 patients, and GVHD grade III-IV in 3. Chronic GVHD developed in 5 patients, of which 2 extensive, resolving in all except one patient. With a median follow up of 37 (range 21–84) months 17 patients are alive (57%). One of the CML patients shows persistence of molecular disease not responding to increasing doses of DLI. All other patients are in CR with the CML patients in molecular remission. Five patients (17%) died because of relapsed disease (2 AML/MDS and 3 ALL). Treatment related mortality was 26% (1 rejection, 2 GVHD, 1 myocardial infarction, 4 infections). In conclusion, matched unrelated donor SCT following myeloablative conditioning using T-cell depletion with CAMPATH-1H in vivo as well as in vitro results in good engraftment, minimal grade I-II GVHD and an overall survival of 57%. Relapse rate was not increased with this strategy. This regimen appears to be successful for young adults with high-risk malignancies.

scholarly journals THER-05. GENETICALLY STABLE POLIOVIRUS VECTOR CARRYING H3.3K27M ANTIGEN FOR TREATMENT OF DIFFUSE MIDLINE GLIOMA BY INTRAMUSCULAR INJECTION

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii472-iii472
Author(s):  
Mubeen Mosaheb ◽  
Daniel Landi ◽  
Elena Dobrikova ◽  
Michael Brown ◽  
Yuanfan Yang ◽  
...  
Keyword(s):  
T Cell ◽  
Antigen Presentation ◽  
Intramuscular Injection ◽  
Cd8 T Cell ◽  
Cytokine Profiles ◽  
Cell Immunity ◽  
Novel Approach ◽  
Diffuse Midline Glioma

Abstract BACKGROUND H3 K27M-mutant diffuse midline glioma (DMG) is invariably lethal. Viruses naturally engage innate immunity, induce antigen presentation, and mediate CD8 T cell priming against foreign antigens. Polioviruses, in particular, are uniquely tropic for dendritic cells (DC) and potently activate DC, inducing Th1-dominant cytokine profiles, CD8 T cell immunity, and enhanced epitope presentation. Thus, poliovirus is ideally suited for vectored delivery of signature tumor neoantigens, e.g. the H3 K27M feature of DMG. However, poliovirus vector design is inherently limited by genetic instability and the underlying neuropathogenicity of poliovirus. METHODS We created a genetically stable, polio:rhinovirus chimera vector devoid of neuropathogenicity and modified for stable expression of the HLA-A2 restricted H3.3 K27M antigen (RIPO (H3.3)). RESULTS RIPO(H3.3) infects, activates, and induces H3.3K27M antigen presentation in DCs in vitro. Given intramuscularly in vivo, RIPO(H3.3) recruits and activates DCs with Th1-dominant cytokine profiles, efficiently primes H3.3K27M-specific CD8 T cells, induces antigen-specific CD8 T cell migration to the tumor site, delays tumor growth, and enhances survival in murine tumor models. CONCLUSION This novel approach leverages the unique ability of polioviruses to activate DCs while simultaneously introducing the H3.3 K27M antigen. In this way, DCs are activated optimally in situ, while being simultaneously infected to express/present tumor antigen. RIPO(H3.3), given by intramuscular injection, will be evaluated in a clinical trial for children with H3 K27M-mutant diffuse midline glioma.

scholarly journals 708 Novel ways to exploit IL-21 to augment adoptive T cell transfer therapy against tumors

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A737-A737
Author(s):  
Anna Cole ◽  
Guillermo Rangel RIvera ◽  
Aubrey Smith ◽  
Megan Wyatt ◽  
Brandon Ware ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Immune Cells ◽  
Striking Difference ◽  
T Cell Therapy ◽  
Cell Immunity ◽  
Emory University

BackgroundIL-21 enhances the anti-tumor capacity of adoptively transferred CD8+ T cells, while IL-2 and IL-15 impair T cell immunity by driving their expansion to a more differentiated status. Yet, these cytokines can act on many different immune cells. Given the potency of IL-21, we tested if this cytokine directly augments T cells or rather if it enhances other immune cells in the culture that indirectly improves T cell therapy.MethodsTo test this question, splenocytes from pmel-1 transgenic mice were used, as all CD8+ T cells express a transgenic TCR specific for tumor-antigen gp10025–33 overexpressed on melanoma. We then peptide activated naïve CD8+ T cells enriched or not from the spleen of pmel-1 mice and expanded them in the presence of IL-21 or IL-2 (10 ng/mL) for four days. Expanded pmel-1 from these various cultures were then restimulated with irradiated splenocytes pulsed with gp10025–33 and grown an additional seven days with IL-2 (10 ng/mL), irrespective of their initial cytokine condition. The in vitro memory phenotype, exhaustion profile, and cytokine secretion of these cultures were then assayed. Furthermore, mice bearing B16KVP melanoma tumors were infused with pmel-1 T cells expanded via these various approaches and compared for their relative capacity to engraft, persist, and regress tumor in vivo.ResultsInterestingly, we discovered that IL-21-treated T cells generated from bulk splenocytes are phenotypically and functionally distinct from IL-21-treated isolated T cells. Upon restimulation, IL-21-treated T cells from bulk splenocytes exhibited an exhausted phenotype that was like anergic IL-2-treated T cells. Moreover, few cells expressed CD62L but expressed heightened markers of suppression, including TIM3, PD-1, and EOMES. Moreover, they produced more effector molecules, including granzyme B and IFN-gamma. In vivo IL-21-treated T cells expanded from bulk splenocytes engrafted and persisted poorly, in turn mediating suboptimal regression of melanoma. Conversely, IL-21 dramatically bolstered the engraftment and antitumor activity of T cells only if they were first isolated from the spleen prior to their expansion and infusion into the animal.ConclusionsCollectively, our data shows that IL-21 may improve ACT therapy best when used directly on antitumor CD8+ T cells. Further studies will illuminate the mechanism behind this striking difference and determine whether other cell subsets reactive to IL-21 cause T cell dysfunction and/or reduced bioavailability. These findings are important for defining the best culture conditions in which to use IL-21 for ACT.AcknowledgementsWe would like to acknowledge Emory University, The Winship Cancer Institute, and the Pediatrics/Winship Flow Cytometry Core.Ethics ApprovalAll animal procedures were approved by the Institutional Animal Care and Use Committee of Emory University, protocol number 201900225.

scholarly journals Human bone marrow and peripheral blood T lymphocyte depletion: efficacy and effects of both T cells and monocytes on growth of hematopoietic progenitors

Blood ◽  
1985 ◽  
Vol 65 (3) ◽  
pp. 663-679
Author(s):  
L Levitt ◽  
TJ Kipps ◽  
EG Engleman ◽  
PL Greenberg
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocyte ◽  
Peripheral Blood ◽  
Cell Depletion ◽  
Target Cells ◽  
Facs Analysis ◽  
Hematopoietic Progenitors ◽  
T Cell Depletion

The efficacy of four separate methods of human bone marrow T lymphocyte depletion was assessed, and the effect of T cells and monocytes on in vitro growth of marrow (CFU-GEMM, BFU-E, and CFU-GM) and peripheral blood (BFU-E) hematopoietic progenitors was determined. Extent of T cell depletion was assessed by multiparameter fluorescent cell sorter (FACS) analysis and by functional studies. Cells staining positively by FACS analysis for one or more of three separate fluorescent pan-T cell monoclonal antibodies (MCAbs) comprised 8.4% to 9.5% of control marrow mononuclear cells (MNCs). T cells constituted 3.2% to 5.1% of marrow following single, sequential, or combination treatment with two different pan-T cell MCAbs (Leu 1 and TM1) plus complement, 1.5% to 2.2% of marrow following solid-phase immunoabsorption (“panning”), 0.2% of marrow after sheep cell rosetting, and only 0.05% of marrow after FACS selective cell sorting and gated separation. T cells made up 59% to 73% of control peripheral blood MNCs and 0.8% to 2.8% of peripheral MNCs following sheep cell rosetting plus treatment with Leu 1 MCAb and complement. Mitogen (PHA, Con A) and allogeneic MLC-induced blastogenic responses (stimulation indices, experimental/control or E/C) revealed a concordant decrement in marrow T cell function after MCAb plus complement (E/C of 3.9 to 9.0), after panning (E/C of 1.6 to 3.5) and after sheep cell rosetting (E/C of 0.7 to 1.3), compared with control marrow (E/C of 5.3 to 15.7). After T cell depletion, marrow BFU-E growth was 95% to 120% of control, CFU-GM growth was 90% to 108% of control, and CFU-GEMM growth was 89% to 111% of control. Marrow T cell and/or monocyte depletion did not alter erythropoietin-dependent BFU-E growth in the absence of Mo-conditioned medium (81% to 95% of control), and the addition of as many as 50 to 100 X 10(3) purified marrow monocytes or T cells to 10(5) autologous nonadherent T cell-depleted marrow target cells had a negligible (P greater than .1) effect on marrow BFU-E growth in vitro. Peripheral blood (PB) BFU-E/10(5) T- depleted target cells were 106% +/- 19% of expected; PB BFU-E growth was significantly diminished after monocyte depletion alone (7% +/- 6% of expected) or after monocyte plus T cell depletion (8% +/- 4% of expected).(ABSTRACT TRUNCATED AT 400 WORDS)

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