Safety and Clinical Efficacy Of Rapidly-Generated Virus-Specific T Cells With Activity Against Adv, EBV, CMV, HHV6 and BK Virus Administered After Allogeneic Hematopoietic Stem Cell Transplant

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 148-148 ◽  
Author(s):  
Anastasia Papadopoulou ◽  
Usha L Katari ◽  
Ulrike Gerdemann ◽  
Caridad Martinez ◽  
Kathryn Leung ◽  
...  
Keyword(s):  
T Cells ◽  
Viral Load ◽  
De Novo ◽  
Bk Virus ◽  
Morbidity And Mortality ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Bladder Pain ◽  
Allogeneic Hsct ◽  
Viral Antigens

Abstract Viral infections remain a major cause of morbidity and mortality after allogeneic HSCT. We and others have demonstrated that the adoptive transfer of virus-specific T cells (VSTs) specific for EBV, CMV and Adv antigens can treat infections that are impervious to conventional therapies, but broader implementation and extension to additional problematic viruses has been limited by competition between viral antigens and time-consuming/laborious manufacturing. We therefore developed a simplified 10-day system for generating a single preparation of VSTs with activity against 12 antigens from 5 viruses (EBV, CMV, Adv, BK, HHV6) that commonly cause post-transplant morbidity and mortality. We report our initial clinical results using these pentavalent pVSTs. With NHLBI-PACT support, we prepared 35 clinical-grade pVSTs from PBMCs (3x107 cells/sample) that we exposed to overlapping peptide libraries spanning immunogenic Adv (Hexon, Penton), CMV (pp65, IE1), EBV (LMP2, EBNA1, BZLF1), BK (Large T, VP1) and HHV-6 (U11, U14, U90) antigens. Exposure was followed by a 9-11 day expansion phase in a G-Rex device in the presence of IL4+7, producing a mean of 374x106 T cells (range 99-713x106). These lines were polyclonal, comprising both CD4+ (57±5%) and CD8+ (35±5%) cells and retained expression of the memory markers CD45RO+CD62L+ (58±8%). Their specificity was dependent on the prior viral exposure of the cell donor; 32/35 lines had activity against Adv (Hexon: 446±153; Penton:317±108 SFC/2x105), 20/35 against CMV (IE1: 337±141; pp65 1059±479), 26/35 against EBV (LMP2: 175±87; EBNA1: 116±44; BZLF1: 129±88), 18/35 against BK (Large T: 130±67; VP1: 231±104) and 21/35 against HHV-6 (U90: 66±50; U11: 36±18; U14: 82±21). None of the lines reacted against recipient PHA blasts (mean Cr51release of 1% at a 20:1 E:T ratio). We have administered pVSTs to 10 allogeneic HSCT recipients in a dose escalation study; 4 on DL1 (5x106/m2), 4 on DL2 (1x107/m2) and 2 on DL3 (2x107/m2). There were no immediate infusional toxicities, and no de novo acute GvHD, demonstrating the in vivo safety of these pVSTs even after a single exposure to viral antigens in vitro. Three patients received the cells as viral prophylaxis (days 38-43 post-HSCT) and all remain well and virus-infection free at up to 3 months post-treatment. The other 7 patients received the cells as treatment for one or more active infections between days 59-139 post-HSCT. Based on viral load measurements by day 42 post-infusion, the pVSTs were successful in controlling active CMV (1 complete (CR) and 1 partial response (PR)), EBV (2 CRs, including a case of frank PTLD); Adv (1 CR); HHV6 (1 CR); and BK (3 CR, 1 PR, 1NR) infections. Of note, 3 of our BK virus responders had tissue disease with severe hemorrhagic cystitis and all had marked improvement or disappearance of hematuria following infusion. One subsequently had an episode of transient but severe bladder pain in association with inflammation seen on cytoscopy coincident with a 6 log fall in urine BK viral load. Our only non-responder was a patient with BK infection whose line lacked activity for this virus, likely reflecting the serostatus of the donor. In addition, 3 patients subsequently reactivated other viruses than those for which they were initially treated, but all cleared these infections by week 12, without requiring additional cell infusions (CMV: 1CR; EBV: 1CR; BK: 1CR; HHV6: 1CR). Finally, 1 patient received pVSTs under a single patient protocol as an emergency treatment for widespread and bulky rituximab-resistant EBV-PTLD. Post pVST there was an immediate decline in her EBV viral load with complete and sustained resolution of PTLD, coincident with an increase in circulating EBV-specific T cells. However, the profound anti-tumor activity mediated by the rapidly-expanding EBV-directed T cells also produced a transient systemic inflammatory response syndrome, which was controlled with steroids and anti-TNFR antibody, with no long term adverse effects. Thus, infusion of pVSTs as prophylaxis or treatment has been safe and is associated with the appearance of virus-reactive T cells in peripheral blood that have been able to control infection with all 5 targeted viruses. We are currently exploring the extension of this platform to include additional clinically relevant viruses and are planning to assess the activity of these cells in the 3rd party setting for broader implementation. Disclosures: Off Label Use: Virus-specific CTLs manufactured under an investigator-initiated IND. Vera:Wilson Wolf Corporation: Consultancy, Research Funding.

Intravesicular Cidofovir in the Treatment of BK Virus-Associated Hemorrhagic Cystitis (BKV-HC) Following Allogeneic Hematopoietic Stem Cell Transplant (HSCT)

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 2085-2085
Author(s):  
Graham Tooker ◽  
Ashraf Z. Badros ◽  
Jennifer Nishioka ◽  
David Riedel
Keyword(s):  
Viral Load ◽  
Median Time ◽  
Research Funding ◽  
Lymphoblastic Leukemia ◽  
Bk Virus ◽  
Response To Treatment ◽  
Cell Transplant ◽  
Initial Report ◽  
Hematopoietic Stem ◽  
Common Diagnosis

Abstract Background: BKV HC is a well-known complication following allo-SCT. Supportive care with bladder irrigation and blood transfusions were the only available treatment. Since our initial report (Bridges B et al. Am J Hematol 2006;81:535), several studies confirmed that intravesicular cidofovir is a potential effective treatment for BKV HC. In this study, we report a large series of consecutive patients who developed BKV HC following allo SCT and received intravesicular cidofovir. Methods: We conducted a retrospective review of allo SCT patients who developed BKV HC and were prescribed intravesicular cidofovir from 2012 to 2017. Results: 33 patients were diagnosed with BKV HC. The median age was 50 years (range=23-73), and 18 (55%) were male. Acute myeloid leukemia (n=12, 35%) was the most common diagnosis followed by non-Hodgkin lymphoma (n=7, 21%) and B cell acute lymphoblastic leukemia (n=4, 12%). Conditioning regimens were myeloablative (n=19, 58%) or reduced-intensity (n=14, 42%); 15 (45%) patients received cyclophosphamide, and 22 (67%) received total body irradiation. The median time to onset of HC symptoms following SCT was 37 days (range: 8-178); 17 (52%) patients had acute graft vs. host disease. HC symptom severity ranged from grade 0-4 (median=2). The median BK urine viral load pre-treatment was 100,000,000 IU/ml. Patients received a median of 2 intravesicular treatments (range=1-7) at a dose of 5 mg/kg. Four patients (12%) were also treated concurrently with intravenous cidofovir. 19 (59%) patients demonstrated complete clinical resolution of symptoms, 9 (28%) demonstrated partial response to treatment, and 4 (13%) had no change in symptoms following treatment. These improvements in clinical status were independent of viral load, though most had reductions in the viral load. The median time to symptom resolution was 17 days (range=7-53; n=28). 82% of patients had no recurrent symptoms of HC. The main side effect of intra-vesicular instillation was increased discomfort and bladder spasms; severe in 3 patients (9%). No patient had impaired renal function directly attributable to intra-vesicular cidofovir. At 12 months after BKV HC diagnosis, 26 (79%) patients were alive. Conclusions: To our knowledge this is the largest study of intravesicular cidofovir for BKV HC reported to date; 77% of patients with BVK HC achieved clinical improvement of symptoms with minimal side effects. Clinical trials of intravesicular cidofovir could provide further evidence for adding intravesicular cidofovir as a standard tool for the treatment of BKV HC. Disclosures Badros: Celgene: Consultancy, Research Funding; Karyopharm: Research Funding; GSK: Research Funding.

Adenovirus and BK Virus Co-Infection in the Pediatric Recipient of Stem Cell Transplant with CMV Infection.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 5010-5010
Author(s):  
Suradej Hongeng ◽  
Siriorn Watcharananan ◽  
Usanarat Anurathapan ◽  
Wisutwadee Piyatuctsanawong ◽  
Samart Pakakasama ◽  
...  
Keyword(s):  
Stem Cell ◽  
Viral Load ◽  
Bk Virus ◽  
Diagnostic Methods ◽  
Hemopoietic Stem Cell ◽  
Cell Transplant ◽  
Cmv Infection ◽  
Allogeneic Hsct ◽  
Number Of Patients ◽  
Peak Viral Load

Abstract Background: Viral infections are a common cause of complication following hemopoietic stem cell transplantation (HSCT). Cytomegalovirus (CMV) is known to be a concerning pathogen and most transplant centers have implemented CMV monitoring as a standard of care. With improvement of virological diagnostic methods, the role of other viruses including adenovirus (ADV) and polyomaviruses (PMV) as pathogens causing major infection are also increasingly appreciated, however, evidence to support routine monitoring of these viruses following HSCT is lacking. In addition, there is a paucity of data on the incidence of ADV and/or PMV co-infection in children who develop CMV infection following HSCT. The aim of this study was to uncover the incidence of CMV and ADV or PMV co-infection in pediatric recipients of allogeneic- HSCT and to correlate the finding with degree of CMV viremia. Methods: We retrospectively performed Q-RT- PCR of ADV, BKV and JCV in 219 blood samples from 77 pediatric patients following allogeneic- HSCT, initially taken for quantitative real-time PCR of CMV (monthly post engraftment, as a routine CMV monitoring, or weekly if CMV syndrome was suspected). These patients were known to have varying degree of CMV viremia.We defined high graded, low to moderate graded and undetectable CMV viremia as CMV viral load > 10,000, 400-< 10,000 and < 400 copies/ml.The lower limit of viral load detection is 400 and 10 copies/ml for CMV and for ADV, BKV and JCV. Results: Twenty-eight patients (36%) had CMV viremia, of which were high graded (peak viral load 32,000->100,000 copies/ml) in 13 patients (17%), in at least one moment of follow up. Twelve patients (15%) had ADV viremia, whereas 40 (52%) had BKV viremia. ADV and BKV viremia occurred in 3 (23%) and 10 (77%) of patients with high-graded CMV viremia and they were mostly high graded (ADV and BKV peak viral load 11,400–367,000 and 10–2,120 copies/ml). In patients with non-high graded CMV viremia, most of them had lower graded ADV and BKV viremia (peak viral load 1,830–14,800 and 10–441 copies/ml), except for one patient who had ADV viral load 3,140,000 copies/ml and BKV viral load of 10 copies/ml. Five patients (6.5%) had CMV, ADV and BKV co-viremia, which were most frequent in patients with high-graded CMV viremia (15%).Two patients (2.5%) had very low JCV viremia (viral load 10 and 13 copies/ml). See Table for summarized results. Conclusion: In pediatric allogeneic HSCT, CMV, BKV and ADV infection is common. High-graded, BKV, ADV or BKV and ADV co-viremia occurred most frequently in patients with high-graded CMV viremia. Low-graded BKV and ADV viremia mostly occurred in patient without high-graded CMV viremia. JCV viremia was not common and occurred irrespective of CMV infection. From our study, monitoring of ADV and BKV should be considered in patients with high-graded CMV viremia following pediatric allogeneic HSCT. CMV viral load (copies/ml) >10,000 400- <10,000 < 400 Number of patients (%) Total patients Cytomegalovirus 13(17%) 15(19%) 49(64%) 77(100%) Adenovirus 3(23%) 1(6%) 8(16%) 12(15%) BK virus 10(77%) 3(23%) 27(55%) 40(52%) JC virus 0 0 2(4%) 2(2.5%) Adenovirus and BK virus 2(15%) 1(6%) 2(4%) 5(6.5%)

Paradoxical Immunological Activity of Flagellin: A Novel Method to Reduce GvHD In Allogeneic HSCT Recipients.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1465-1465
Author(s):  
Mohammad S Hossain ◽  
Andrew T Gewirtz ◽  
John Roback ◽  
Edmund Waller
Keyword(s):  
T Cells ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Cell Transplant ◽  
Term Survival ◽  
Hematopoietic Stem ◽  
Allogeneic Hsct ◽  
Post Transplant ◽  
Donor T Cells ◽  
Ifn Γ

Abstract Abstract 1465 Background: Graft-vs-host disease (GvHD) is a major complication in allogeneic Hematopoietic Stem Cell Transplant (HSCT) recipients. Using flagellin, a bacterial protein that agonistically binds with TLR5, we previously reported that two doses of flagellin (before irradiation and after allogeneic HSCT with 5×106 splenocytes) in H-2K à H-2b model significantly reduced GvHD and had 67% long-term survival by 132 days post-transplant whereas control recipients had severe acute GvHD and 100% early mortality within 15 days post transplant. Here, we report the mechanism by which flagellin reduces GvHD using the same H-2K à H-2b HSCT murine model. Methods: 3×106 splenocytes and 5 ×106 T cell depleted bone marrow (BM) cells harvested from the congeneic H-2K donors were transplanted into lethally irradiated (11Gy) C57BL/6 (H-2b) recipients. 50-μg HPLC purified LPS-free flagellin diluted in ice-cold PBS were administered i.p 3 hours before irradiation and 24 hours after transplant. Control recipients were treated with PBS. Recipients (6/group) were sacrificed on day 4 and 10 after post transplant. Serum was collected to determine cytokines by ELISA and splenocytes were analyzed by FACS to determine immune cells phenotypes. Results: Although both Fla and PBS-treated recipients showed identical weight-loss within day 10 post transplant, surprisingly significantly lower numbers of cells/spleen were determined in the spleens of Fla recipients compared to control recipients [Fla 0.9 ± 0.1 (x106) vs PBS 1.7 ± 0.4(x106), p=0.002] on day 4 post transplant but not on day 10 [Fla 186.1 ± 35.1 (x106) vs PBS 151.2 ± 40.5(x106), p=0.43] post-transplant. We investigated the paradoxical immune response of flagellin on donor T cells on day 4-post transplant. First, we determined the numbers of donor spleen-derived Thy1.2+ T cells per spleen. The numbers of donor spleen-derived T cells per spleen were significantly lower in Fla recipients compared to PBS-treated recipients [Fla 0.005 ± 0.002 (x103) vs PBS 0.04 ± 0.03(x103), p=0.02]. Accordingly, donor spleen-derived both CD4 and CD8 T cells per spleen of Fla recipients were also found significantly lower compared to PBS-treated recipients (CD4, p=0.04; CD8, p= 0.003). The CD62L, a naïve and also markers for allo-reactive T cells that cause GvHD were found significantly lower in both CD4 and CD8 T cells (CD4, p= 0.03; CD8, p=0.003) and the inducible co-stimulatory molecule 1 (ICOS-1), another prominent T cells activation marker were also found significantly lower (CD4, p= 0.04; CD8, p=0.007) in Fla recipients compared to PBS-treated recipients. These lower immune phenotypes of donor T cells in Fla recipients may reduce the initiation of GvHD at the early time points of transplant. However, flagellin-induced reduction of donor T cells activity were not suppressed considerably as the numbers of donor spleen-derived CD4 T cells expressing activation markers such as CD25 {Fla 0.5 ± 0.2 (x103) vs PBS 4.5 ± 0.5 (x103), p=0.07} and CD69 {Fla 0.5 ± 0.2 (x103) vs PBS 6.5 ± 5.0 (x103), p=0.08} per spleen were not found significantly different. On the other hand, the numbers of CD8 T cells those expressed CD25 {Fla 0.2 ± 0.04 (x103) vs PBS 1.3 ± 0.7 (x103), p=0.01} and CD69 {Fla 0.4 ± 0.1 (x103) vs PBS 2.4 ± 1.7 (x103), p=0.03} per spleen were found significantly lower in Fla recipients compared to PBS-treated recipients. Although over 90% of donor spleen CD4 T cells of both Fla and PBS-treated recipients expressed PD-1, 24% and 32% respectively, expressed IFN-γ after vitro PMA stimulation. Similar results were found in case of CD8 T cells. The over expression of PD-1 in HSCT recipients, thus did not make the donor T cells exhausted as they expanded over 100 times within day 10 post transplant with the expression of PD-1. Although similar level of serum IFN-γ was determined between the Fla and PBS-treated recipients on day 10 post-transplant, IFN-γ level was found below detection limit on day 4 post-transplant. The serum levels of IL-1β and IL-10 were undetectable on both days 4 and 10 post-transplant. Serum LPS were found identical in both Fla and PBS-treated groups determined on day 4 and 10 days post transplant. Conclusion: Flagellin protected allogeneic HSCT recipients from lethal GvHD by reducing donor CD62L+ and ICOS-1+ T cells expansion and activation within 4 days post-transplant without compromising their normal immune responses. Disclosures: No relevant conflicts of interest to declare.

Intravesicular Cidofovir in the Treatment of BK Virus–Associated Hemorrhagic Cystitis Following Hematopoietic Stem Cell Transplantation

2019 ◽  
Vol 54 (6) ◽  
pp. 547-553 ◽  
Author(s):  
Graham M. Tooker ◽  
Kristen A. Stafford ◽  
Jennifer Nishioka ◽  
Ashraf Z. Badros ◽  
David J. Riedel
Keyword(s):  
Stem Cell ◽  
Viral Load ◽  
Hematopoietic Stem Cell ◽  
Hematopoietic Stem Cell Transplant ◽  
Stem Cell Transplant ◽  
Hemorrhagic Cystitis ◽  
Bk Virus ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Bladder Irrigation

Background: BK virus hemorrhagic cystitis (BKV-HC) is a common complication following hematopoietic stem cell transplant (HSCT); optimal management remains uncertain. Supportive care (bladder irrigation and blood transfusions) and intravenous and intravesicular cidofovir have all been used with varying success. Objective: The purpose of this study was to determine the safety and effectiveness of intravesicular cidofovir for BKV-HC following HSCT. Methods: A retrospective analysis of all HSCT patients with BKV-HC prescribed intravesicular cidofovir from 2012 to 2017. Results: 33 patients were treated for BKV-HC. The median age was 50 years (range 23-73), and 18 (55%) were male. The median HC symptom severity was 2, with a median BK urine viral load pretreatment of 100,000,000 IU/mL. Patients received a median of 2 intravesicular treatments (range 1-7) at a dosage of 5 mg/kg per instillation. In all, 19 (59%) patients demonstrated complete clinical resolution of symptoms; 9 (28%) had a partial response; and 4 (13%) had no change in symptoms. Patients with a high pretreatment BK viral load (>100 million) and high HC grade (2-4) had a lower frequency of complete remission. The main side effect of intravesicular instillation was severe bladder spasms in 4 patients (12%). Conclusion and Relevance: This is the largest study of intravesicular cidofovir treatment of BKV HC reported to date; 88% of patients with BVK-HC achieved clinical improvement of symptoms with minimal side effects. Clinical trials of intravesicular cidofovir could provide further evidence for this treatment for BKV-HC.

Safety and Clinical Efficacy of Rapidly-Generated Trivirus-Directed T Cells After Allogeneic Hematopoietic Stem Cell Transplant

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 223-223
Author(s):  
Ulrike Gerdemann ◽  
Usha L Katari ◽  
Jacqueline Keirnan ◽  
John A. Craddock ◽  
Janet Salinas ◽  
...  
Keyword(s):  
T Cells ◽  
Viral Load ◽  
T Cell ◽  
Cell Line ◽  
Peripheral Blood ◽  
Cell Transplant ◽  
Clinical Grade ◽  
Hematopoietic Stem ◽  
Post Infusion

Abstract Abstract 223 We have previously demonstrated that small numbers of ex vivo-expanded, trivirus-specific T cells targeting Epstein Barr virus (EBV), cytomegalovirus (CMV), and Adenovirus (Adv) are safe, proliferate in vivo and protect human subjects against all 3 viruses following HSCT. However, broader implementation is limited by the need for infectious virus (EBV) to establish an EBV-transformed B lymphoblastoid cell line (EBV-LCL), for clinical grade adenoviral vector, and by prolonged (6wks for EBV-LCL and 6wks for T cells) and complex manufacture. Moreover, competition between viral antigens limits extension to additional viruses. We now evaluate whether it is possible to make clinically effective T cell lines using methods that exclude all viral components and utilize simplified manufacturing technology. With NHLBI-Production Assistance for Cellular Therapies (PACT) support, 29 clinical-grade rCTL lines have been generated. From an initial 15×106 PBMCs, we prepared a median of 214±88 × 106 T cells (range 100–420×106) over 9–11 days by using dendritic cells (DCs) nucleofected with DNA plasmids encoding immunogenic EBV (LMP2, EBNA1 and BZLF1), Adv (Hexon and Penton), and CMV (pp65 and IE1) antigens, and expanding them with IL4+7 in gas permeable (G-Rex) devices. The rCTL lines were polyclonal, comprising both CD4+ (33±3%) and CD8+ (60.5±3%) cells, that expressed the activation and memory markers CD45RO+ CD62L+ (64.3±26.6%) and CD45RO+ CD62L- (17.4±14%). Twenty lines generated from donors that were seropositive for all three viruses demonstrated activity against all 3 targets - CMV (IE1: 359±100; pp65: 637±177 SFC/2×105), EBV (LMP2: 217±60, EBNA1: 67±19 and BZLF1: 111±31) and Adv (Hexon: 265±74, Penton: 191±53) - while 9 lines generated from donors who were CMV seronegative demonstrated activity exclusively against EBV (LMP2: 197±70, EBNA1: 145±51 and BZLF1: 239±84) and Adv (Hexon: 271±96, Penton: 254±90). None of the lines reacted against recipient PHA blasts (median spontaneous Cr51 release of 0% at a 20:1 effector to target cell ratio). To date we have administered these lines to 10 recipients of allogeneic HSCT. Five patients received dose level (DL) 1 (5×106/m2), 2 received DL2 (1×107/m2) and 3 had DL3 (2×107/m2) of this phase I/II study. Three patients were infused as treatment for CMV, 2 for Adv, 2 for EBV, 1 for EBV+Adv, and 2 for CMV+Adv. Our major anticipated concern was that these once stimulated, unselected rCTLs might cause GvHD in vivo, but that was not the case. One patient developed a skin rash 2 weeks after rCTLs but no other toxicity related to the infused cells was observed. Eight of the 10 treated patients including one patient with a biopsy-proven EBV lymphoma and the 3 patients with double reactivations had complete responses to rCTL therapy with a return of viral load to normal and resolution of all other symptoms. Response was associated with an increase in the frequency of virus-specific T cells detected in the peripheral blood against the infecting virus. For CMV there was an increase from a median of 0.5 to 96 and 1 to 277 SFC/4×105 IE1 and pp65-specific T cells respectively 3–6wks post-infusion; for Adv an increase from a mean of 0.5 to 137 and 0.5 to 99 SFC/4×105 Hexon and Penton-specific cells 2wks post-infusion, respectively, and for EBV an increase from 2.75 to 227, 1.5 to 39, and 1 to 188.5 SFC/4×105 EBNA1, LMP2, and BZLF1-specific T cells 2–4wks post-infusion, respectively. Two patients failed to respond. The first had a 3 year history of persistent CMV colitis despite high circulating CMV-specific precursors (297 IE1-specific and 193 pp65-specific T cells/4×105 PBMCs). Post rCTL we saw no increase in T cell precursor levels and no clinical improvement. The second was treated for an elevated EBV viral load but also had high pre-existing EBV-specific T cell precursors (60, 23, and 240 SFC/4×105 EBNA1, LMP2, and BZLF1-specific T cells). Again, post-rCTL we did not detect an increase in EBV-specific precursors and no response. Thus, infusion of rCTLs has been safe and in 8/10 patients was associated with the appearance of virus-reactive T cells directed against the infecting virus in peripheral blood and subsequent virus clearance. rCTLs have the potential to increase the availability of cell products for HSCT recipients and we are currently extending this platform to additional viruses, thereby broadening the spectrum of pathogens that can be targeted by adoptive transfer of a single T cell line. Disclosures: Off Label Use: IND cell therapy product.

scholarly journals Predicting Immune Pathology after Hematopoietic Stem Cell Transplant with Transcriptomics: Naïve CD4 T Cell Expansion at Day 100 Predicts Patients with De Novo Chronic Gvhd

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 38-39
Author(s):  
Ben Watkins ◽  
James Kaminski ◽  
Muna Qayed ◽  
Kayla Betz ◽  
Yvonne Suessmuth ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Cd4 T Cells ◽  
Research Funding ◽  
De Novo ◽  
Cd4 T Cell ◽  
Unrelated Donor ◽  
Cell Transplant ◽  
Hematopoietic Stem

Background: Chronic graft-versus-host disease (CGVHD) is the leading cause of long-term morbidity and mortality following hematopoietic stem cell transplant (HCT) and occurs in over 50% of patients undergoing unrelated donor HCT. Despite its frequency, the mechanisms driving this disease remain incompletely understood, making its prevention and successful treatment challenging. To address this issue, we have undertaken a transcriptomic analysis of T cell reconstitution after unrelated donor HCT, to dissect mechanisms driving CGVHD. Methods: The patients studied were enrolled on a Phase 2, randomized, placebo-controlled trial of abatacept for GVHD prevention in patients receiving 8/8 unrelated-donor HCT for hematologic malignancies (NCT01743131). All immune analyses in the current study were performed on patients randomized to standard GVHD prophylaxis with calcineurin inhibition + methotrexate alone (placebo cohort, n =69), and thus provide insights into the drivers of CGVHD during standard unrelated donor HCT. On Day +100, CD4+ T cells were purified from the peripheral blood of these patients, and then analyzed by RNASeq. To determine the transcriptomic drivers of CGVHD without the confounder of significant prior acute GVHD (AGVHD) or exposure to steroids, we focused on profiling the CD4+ transcriptome of de novo CGVHD (CGVHD which develops in the absence of prior grade II-IV AGVHD, n = 7) and compared these patients to those who were 'operationally tolerant' and never developed either grade II-IV AGVHD or any CGVHD (n= 4). Gene expression from the resulting transcriptomes was quantified using kallisto. Differentially expressed (DE) genes were identified using DESeq2 (threshold for DE, adjusted (for multiple testing) p &lt;0.05). Gene Set Enrichment Analysis (GSEA) was also performed, with genes ranked by Log2FC/std_error (Log2FC), and gene signatures with an adjusted p &lt;0.05 considered significantly enriched. Results: DE analysis identified 101 genes that were significantly upregulated in CD4+ T cells from de novo CGVHD group and 54 genes that were significantly upregulated in the 'operationally tolerant' group (Figure 1A). GSEA identified that the mostly highly enriched signatures in patients with de novo CGVHD encompassed naïve CD4+ transcriptional programing (Figure 1B-C), in agreement with flow cytometric analysis, which also demonstrated expansion of CD4+ naïve T cells at Day +100 in patients developing de novo CGVHD compared to those demonstrating operational tolerance (Figure 1D). Importantly, the naïve CD4+ T cell signatures that were identified were distinct from those defining CD4+ stem cell memory T cells (which did not enrich in the de novo CGVHD cohort). In contrast, the gene signature of the operationally tolerant patients were enriched for regulatory gene sets (Figure 1C), consistent with a large body of evidence demonstrating that Treg expansion can be protective against CGVHD. Discussion: This study represents, to our knowledge, the first interrogation of the transcriptomic features of patients developing de novo CGVHD versus those operationally tolerant patients who develop neither significant AGVHD nor CGVHD after HCT. These patients may represent a particularly effective cohort in which to study immunologic drivers of CGVHD, given their freedom from prior treatment with corticosteroids, which can confound downstream transcriptomic analyses. Our data provide compelling evidence for a prominent naïve CD4+ T cell signature in patients who develop moderate-to-severe CGVHD despite their lack of antecedent AGVHD. These results are provocative, as they implicate a cell subset that is often considered more quiescent (naïve T cells) as associated with patients who develop immune pathology associated with CGVHD. These results suggest that naïve CD4+ T cells may represent a potent reservoir for alloreactivity, that, once activated, can cause significant disease. This would be in agreement with the implications of previously reported trials of naïve T cell depletion, which resulted in significant control of CGVHD. These results suggest that strategies to restrain naïve T cell pathogenic activation after Day +100 may improve CGVHD outcomes, and that the CD4+ T cell transcriptomic signature at this timepoint could be developed into a robust immunologic biomarker for the risk of developing CGVHD versus operational tolerance after HCT. Figure 1 Disclosures Watkins: Bristol Myers Squib: Honoraria. Qayed:Novartis: Consultancy; Mesoblast: Consultancy. Blazar:Tmunity: Other: Co-founder; KidsFirst Fund: Research Funding; BlueRock Therapeutics: Research Funding; Childrens' Cancer Research Fund: Research Funding; BlueRock Therapeuetic: Consultancy; Magenta Therapeutics: Consultancy; Fate Therapeutics Inc.: Research Funding. Horan:Bristol Myers Squib: Honoraria, Research Funding. Langston:Kadmon Corporation: Research Funding; Astellas Pharmaceuticals: Research Funding; Jazz Pharmaceuticals: Research Funding; Incyte: Research Funding; Bristol Myers Squib: Research Funding; Chimerix: Research Funding; Takeda: Research Funding. Kean:fortyseven: Consultancy; regeneron: Research Funding; hifibio: Consultancy; kymab: Consultancy; Bristol Meyers Squibb: Research Funding; gilead: Research Funding; novartis: Consultancy; bluebird bio: Research Funding; magenta: Research Funding.

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