ALBERT

Description: Background. The TT approach has significantly improved the outcome of multiple myeloma (MM) by combining new drugs with a regimen that comprises induction, tandem autologous stem cell transplantation (ASCT), consolidation and maintenance. However, a group of 15% of patients with high risk multiple myeloma (HRMM) have derived little benefit despite similar response rates to induction chemotherapy and ASCT when compared to low risk MM. The poor outcome of HRMM is explained by early relapse post ASCT resulting in a short progression free survival (PFS) with only 15-20% of patients surviving long-term. Daratumumab (Dara) is a human IgG1k anti-CD38 monoclonal antibody that has shown favorable results in early single-arm studies and more recently in phase III studies for relapsed/refractory and newly diagnosed MM. In TT7, we introduced Dara during all phases of therapy, including immune consolidation early post ASCT, to improve responses rate and PFS in HRMM. Methods. Patients had newly diagnosed HRMM as defined by high risk cytogenetic abnormalities, presence of extramedullary disease, 〉3 focal lesions on CT-PET, elevated LDH due to MM, or ISS II/III with cytogenetic abnormality. Dara (16mg/kgx1) was added to induction with KTD-PACE (carfilzomib, thalidomide, dexamethasone; and four-day continuous infusions of cisplatin, doxorubicin, cyclophosphamide, etoposide). Conditioning for tandem autologous stem cell transplantation (ASCT) was with fractionated melphalan (50mg/m2x4) (fMEL) based on prior observations that patients with adverse cytogenetics fare better with fMEL rather than single high dose MEL200mg/m2.In the inter tandem ASCT period immunological consolidation with Dara (16mg/kg) alone for 2 doses was followed by Dara (16mg/kg) on day 1 combined with K (36mg/m2) and D (20mg) weekly for 2 cycles. DaraKD was administered to avoid treatment free periods allowing for myeloma regrowth. The 2nd ASCT was followed by further immunological consolidation with Dara (16mg/k) for 2 doses, and maintenance therapy for 3 yrs with 3-months block of alternating Dara-KD (dara 16mg/kg day 1; K 36mg/m2 and dex 20mg weekly) and Dara-lenalidomide (R)D (dara 16mg/kg day 1; R 15mg day 1-21 q28 and D 20mg weekly). Results. TT7 enrolled 43 patients thus far. The median follow-up was 11 months (range: 1-22). The median age was 61 yrs (range 44-73). Sixteen patients were ≥65 yrs (37.2%). A mean of 29.4x106 CD34+ cells/kg (range: 4.6-86.4) were collected. 36 patients completed ASCT #1 (83.7%) and 18 (41.9%) ASCT #2, whilst 14 patients have proceeded to the maintenance phase. R-ISS II/III or metaphase cytogenetic abnormalities were present in 85.1 and 58.1% of patients, respectively. Elevated LDH or 〉3FL on CT-PET were noted in 30 and 41.8%. The 1-yr cumulative incidence estimates for reaching VGPR and PR were 87 and 83%, respectively. A CR or sCR was achieved in 68 and 46%. The 1-yr estimates of PFS and OS were 91.6 and 87.2%. 40 subjects are alive, whilst 5 progressed on study therapy and 3 subsequently died. 38 patients are progression free at the time of reporting. Dara was well-tolerated and no subjects discontinued therapy due to dara-related side effects. The CR and sCR rates compared favorably to the predecessor HRMM TT5 protocol where CR and sCR rates were 59 and 27%. Conclusion. The early results of TT7 point to increased response rates of HRMM to a dara-based TT regimen with especially higher rates of CR and sCR. Longer follow-up is required to determine if these early results translate into superior PFS and OS. Figure Disclosures van Rhee: Karyopharm Therapeutics: Consultancy; Kite Pharma: Consultancy; Adicet Bio: Consultancy; Takeda: Consultancy; Sanofi Genzyme: Consultancy; Castleman Disease Collaborative Network: Consultancy; EUSA: Consultancy. Walker:Celgene: Research Funding. Morgan:Amgen, Roche, Abbvie, Takeda, Celgene, Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Other: research grant, Research Funding. Davies:Amgen, Celgene, Janssen, Oncopeptides, Roche, Takeda: Membership on an entity's Board of Directors or advisory committees, Other: Consultant/Advisor; Janssen, Celgene: Other: Research Grant, Research Funding.

Description: BACKGROUND and OBJECTIVE: Tumor lysis syndrome (TLS), a potentially fatal oncologic complication, can have a significant clinical and economic impact to patients and the healthcare system. The aim of this analysis was to examine healthcare resource utilization in patients treated with rasburicase for the management of tumor lysis in the outpatient versus inpatient setting. METHODS: Adult patients were selected from the Integra Connect Database (IC) if they were treated with rasburicase between January 1, 2017 and March 31, 2019. The IC database comprises clinical and financial records for more than 750,000 community oncology patients, including 25,000 active Oncology Care Model participants representing approximately 20% of the unique beneficiaries in this Medicare model. Patients treated in the outpatient setting were divided into 3 groups: Primary Prophylaxis- treatment with rasburicase administered days 0-2 of chemotherapy (Group A), Early Reactive- rasburicase administered days 3-5 of chemotherapy (Group B), and Late Reactive- rasburicase administered after day 5 of chemotherapy (Group C). Inpatients were divided into 2 groups: Inpatient TLS Treatment- patients admitted and treated for TLS with rasburicase and who had not received rasburicase as part of their outpatient chemotherapy regimen (Group D) and Inpatient Chemotherapy- patients admitted for chemotherapy who were given rasburicase (Group E). Demographic, clinical characteristics including tumor types, lab values, and dose information were collected. Total cost of rasburicase was calculated as mean drug cost per patient. All variables were summarized descriptively as mean (SD), median (min and max) or counts (percentages). RESULTS: A total of 265 patients treated with rasburicase were included in the analysis. Of those, 189 patients received rasburicase in the outpatient setting vs 76 in the inpatient setting. None of the 189 patients in Groups A, B, and C who received outpatient rasburicase required admission due to TLS. Patient demographic and clinical characteristics, as well as rasburicase utilization, were similar between cohorts (Table 1 and Table 2). Our results show that while 54% of patients in Groups B, C, and D were initially treated with allopurinol, these patients were switched to rasburicase, indicating failure of allopurinol alone. The total cost of rasburicase trended lower in the outpatient vs inpatient setting ($9,287 vs $11,959). However, a more appropriate comparator to this cost is the published data for charges incurred for inpatient treatment of TLS, shown to be $151,9171 CONCLUSIONS: TLS continues to impact patients with diagnoses and chemotherapy regimens at intermediate and high risk for development of this syndrome. This study demonstrates that allopurinol is frequently inadequate and replacement with rasburicase is needed. We found rasburicase to be effective from both a clinical and a cost perspective in preventing TLS-related hospitalization. While rasburicase is most efficiently employed as primary prevention, close monitoring of patients allows effective reactive utilization evidenced by none of the outpatients in this study who received rasburicase were admitted for TLS. This study highlights the opportunity for greater utilization of rasburicase in the outpatient setting, as a means to lower the total cost of care. 1. Pathak et al. Blood. 2017; 130:3390 Disclosures McBride: teva: Consultancy; Sandoz: Consultancy; Sanofi Genzyme: Consultancy.

Description: Abstract 3280 CD4+CD25+Foxp3+ regulatory T cells (Treg) play an important role in the maintenance of self-tolerance and immune homeostasis and Treg deficiency contributes to the development of autoimmune diseases. CD4Treg, conventional CD4 T cells (Tcon) and CD8 T cells are derived from lymphocyte progenitor cells that differentiate into distinct functional subsets in the thymus before export to the peripheral circulation. As T cells differentiate and expand in the periphery, each T cell subset is differentially regulated and subjected to distinct homeostatic signals. For example, interleukin-2 (IL-2) is a critical regulator of Treg development, expansion and survival and lack of IL-2 results in selective Treg deficiency. In regulating Treg homeostasis, IL-2 has multiple and distinct effects on Treg differentiation, proliferation and susceptibility to apoptosis. To determine the mechanism whereby IL-2 affects susceptibility of Treg to apoptosis, we used a new flow cytometry-based assay (BH3 profiling) to measure the mitochondrial membrane depolarization in response to a panel of pro-apoptotic BH3 peptides (BIM, BID, BAD, NOXA, PUMA, BMF, HRK). This assay allowed us to compare “priming” which we define as susceptibility to BH3 peptide-induced mitochondrial membrane depolarization in different T cell subsets, including CD4 Treg, CD4 Tcon and CD8 T cells. We also examined cell surface expression of CD95 death receptor (Fas) and cytoplasmic expression of Bcl-2 and Ki67 as additional measures of susceptibility to apoptosis and proliferation in each subset. In resting blood obtained from healthy donors (n=10), CD4 Treg were more “primed” than either CD4 Tcon or CD8 T cells when exposed to several BH3 peptides (PUMA, BMF and the combination of BAD+NOXA). CD4 Treg were also found to have decreased expression of Bcl-2 and increased expression of CD95 and Ki67 compared to CD4 Tcon or CD8 T cells. Thus, Treg in healthy individuals have higher proliferative activity and are more susceptible to apoptosis than other major T cell subsets through both mitochondrial and death receptor pathways. To establish the functional effects of TCR stimulation and IL-2, CD4 Treg, CD4 Tcon and CD8 T cells were purified by cell sorting and cultured for 5–6 days with or without TCR stimulation (1μg/ml anti-CD3 + 1μg/ml anti-CD28) and IL-2 (100 IU/ml). Results were compared to cells cultured in media alone. Results are summarized in the table below. CD4 Tcon and CD8 T cells responded in a similar fashion to either TCR stimulation alone or TCR plus IL-2. This response included increased BH3 priming, reduced expression of Bcl-2, increased expression of CD95 and increased proliferation (Ki-67). IL-2 alone had no effect on CD4 Tcon or CD8 T cells. In contrast, TCR stimulation alone had no effect on CD4 Treg but IL-2 alone reduced BH3 priming and increased expression of Bcl-2. Combined TCR stimulation plus IL-2 in Treg increased BH3 priming, reduced expression of Bcl-2, increased expression of CD95 and increased proliferation. Thus, TCR stimulation reversed the anti-apoptotic effects of IL-2 alone and markedly increased susceptibility of Treg to apoptosis. When compared with CD4 Tcon and CD8 T cells, these studies demonstrate distinct effects of TCR stimulation and IL-2 on both mitochondrial and death receptor pathways of apoptosis in CD4 Treg and define mechanisms whereby TCR stimulation and IL-2 interact to regulate Treg homeostasis. Table 1. Effects of in vitro TCR stimulation and IL-2 on apoptotic pathways of T cell subsets TCR Stimulation IL-2 TCR + IL2 BH3 priming Bcl-2 CD95 Ki67 BH3 priming Bcl-2 CD95 Ki67 BH3 priming Bcl-2 CD95 Ki67 CD4 Treg – – – – ↓ ↑ – – ↑ ↓ ↑ ↑ CD4 Tcon ↑ ↓ ↑ ↑ – – – – ↑ ↓ ↑ ↑ CD8 ↑ ↓ ↑ ↑ – – – – ↑ ↓ ↑ ↑ Disclosures: No relevant conflicts of interest to declare.

Description: CD4+CD25+Foxp3+ regulatory T cells (Treg) play an important role in the maintenance of self-tolerance and immune homeostasis. Interleukin-2 (IL-2) is critical for Treg development, expansion, activity and survival. Lack of either IL-2 or IL-2Rα (CD25) results in Treg deficiency and Treg impairment is associated with loss of tolerance, autoimmunity, and chronic GVHD. We previously demonstrated that daily administration of low-dose IL-2 resulted in selective expansion of Treg in vivo and clinical improvement of chronic GVHD (Koreth et al. NEJM 2011). We also reported that low-dose IL-2 enhanced Treg resistance to FAS-mediated apoptosis through the extrinsic pathway (Matsuoka et al. Sci Transl Med 2013). However susceptibility of Treg to apoptosis through the intrinsic pathway has not been examined. To examine the mechanisms whereby IL-2 affects susceptibility of Treg to apoptosis through the intrinsic pathway, we used a functional flow cytometry-based assay (BH3 profiling) to measure mitochondrial membrane depolarization in response to a panel of pro-apoptotic BH3 peptides (BIM, BID, BAD, NOXA, PUMA, BMF, HRK). Mitochondria that are more sensitive to the BH3 peptides are more 'primed' for apoptosis, and more sensitive to pro-apoptotic signaling. This assay allowed us to compare “priming” which we define as susceptibility to BH3 peptide-induced mitochondrial membrane depolarization in Treg and Tcon. We also examined cytoplasmic expression of Bcl-2, BclxL, Mcl1, Bim, XIAP, Ki67 and Staurosporine (STS) induced apoptosis as additional measures of susceptibility to apoptosis and proliferation in each subset. In resting blood obtained from healthy donors (n=25), Treg were more “primed” than Tcon when exposed to several BH3 peptides (Bim, Bad, PUMA and BMF). This BH3 peptide pattern suggests that the anti-apoptotic protein, Bcl2 plays a primary role in the priming of these cells. Analysis of pro- and anti-apoptotic proteins revealed that Treg expressed higher levels Bim and BclxL and lower levels of Bcl2 and XIAP than Tcon, but there was no significant difference in Mcl1. Treg expressed higher levels of Ki67 and were more susceptible to both STS- and FAS-induced apoptosis than Tcon. Thus, Treg in healthy individuals have higher proliferative activity and are more susceptible to apoptosis than Tcon through both mitochondrial and death receptor pathways. To examine the functional effects of IL-2 on T cell homeostasis, Treg and Tcon were purified by cell sorting and cultured with different concentrations of IL-2 (10, 100, 1000 IU/ml) with and without TCR stimulation (1ug/ml anti-CD3 + 1ug/ml anti-CD28). BH3 priming was measured after 3 days and results were compared to cells cultured in media alone. Low-concentration IL-2 (10 U/ml) lowered BH3 priming and increased Bcl2 expression in Treg. Similar effects were observed in Tcon, but higher concentrations of IL-2 (〉100 U/ml) were required to increase Bcl2 expression and decrease BH3 priming in Tcon. TCR stimulation alone induced higher BH3 priming in Tcon but had little effect in Treg. The combination of TCR stimulation plus IL-2 increased BH3 priming in both Tcon and Treg. TCR stimulation increased proliferation only in Tcon, but the TCR + IL-2 combination increased proliferation in both Tcon and Treg. Thus, IL-2 alone reduced BH3 priming in unstimulated cells but the TCR + IL-2 combination induced higher proliferation and higher BH3 priming. STS-induced apoptosis assays revealed that low-dose IL-2 reduced susceptibility to apoptosis only in Treg. IL-2 did not change the expression of BclxL, Mcl1, and Bim but XIAP expression was increased in both Treg and Tcon. To confirm the effects of low-dose IL-2 on Treg priming were mediated by Bcl2, we examined the effect of ABT-199, a selective Bcl2 inhibitor on BH3 priming in Treg and Tcon. As shown in Figure 1, ABT-199 enhanced priming and spontaneous apoptosis of both Treg and Tcon. IL-2 had no effect on ABT-199-induced priming or apoptosis in Tcon. In contrast, IL-2 reversed the effects of ABT-199 on Treg priming and apoptosis providing further evidence that the inhibition of intrinsic pathway apoptosis mediated by IL-2 in Treg is dependent on Bcl2. Disclosures: Letai: AbbVie: Consultancy; Dana-Farber Cancer Institute: Patents & Royalties.

Description: Abstract 225 CD4+CD25+Foxp3+ regulatory T cells (Treg) are known to play a central role in the maintenance of self-tolerance and immune homeostasis. After allogeneic stem cell transplantation, impaired recovery of Treg is associated with the development of cGVHD. Interleukin-2 (IL-2) is a critical regulator of Treg development, expansion and survival and lack of IL-2 results in Treg deficiency. In patients with cGVHD, we previously demonstrated that Treg proliferate at high levels but this subset is also highly susceptible to apoptosis leading to inadequate Treg numbers (Matsuoka et al. JCI 2010). We also reported that low-dose IL-2 administration resulted in selective expansion of Treg in vivo and clinical improvement of cGVHD (Koreth et al. NEJM 2011). To identify mechanisms responsible for increased Treg susceptibility to apoptosis in cGVHD we used a new flow cytometry-based assay to measure mitochondrial membrane depolarization in response to a panel of pro-apoptotic BH3 peptides (BIM, BID, BAD, NOXA, PUMA, BMF, HRK). This assessment allowed us to compare BH3 peptide-induced mitochondrial membrane depolarization (“priming”) in different T cell subsets, including CD4 Treg, conventional CD4 T cells (CD4 Tcon), and CD8 T cells. Expression of Bcl-2, CD95 and Ki67 were also studied in each T cell subset. We studied peripheral blood samples from 36 patients with hematologic malignancies (median age 59 yr) who are 〉 2 years post HSCT (27 patients with cGVHD and 9 patients without cGVHD) and 15 patients who received daily subcutaneous IL-2 for 8 weeks for treatment of steroid-refractory cGvHD. Severity of cGVHD was classified according to NIH criteria. In patients without cGVHD, BH3 priming was similar in all 3 T cell subsets (CD4 Treg, CD4 Tcon and CD8). In patients with cGVHD, CD4 Treg were more primed than CD4 Tcon when challenged with BIM, BAD, PUMA, BMF and the combination of BAD + NOXA peptides (p