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Introduction: Recent data suggests that PD-1 and its ligands PD-L1/PD-L2 mediate immune evasion in CLL. However, recent work (Ding, BLOOD 2017) demonstrates that pembrolizumab (pembro) alone is ...ineffective in patients (pts) with CLL (ORR 0%, median PFS 2.4 months). In 5 pts with relapsed/refractory (r/r) CLL, 3 responded to the combination of ibrutinib / nivolumab (Jain ASH, 2016). A key interaction exists between PI3K signaling and immune checkpoint surveillance by which inhibition of PI3K decreases PD-L1 tumor expression. Thus, we hypothesized synergistic activity with PD-1 + P13K blockade. We tested the safety and activity of umbralisib, a next generation, highly-specific PI3K-δ inhibitor, in combination with pembro and the glycoengineered anti-CD20 monoclonal antibody ublituximab in r/r CLL, representing the first reported combination of a PD-1 inhibitor with a PI3K-δ inhibitor.
Methods: This is a Ph I (3+3 design), multicenter study to assess the safety of pembro in combination with umbralisib and ublituximab in pts with r/r CLL. Treatment involved three stages: Eligible pts received umbralisib (800mg daily) and ublituximab (900mg 3 out of 4 weeks) for the first two cycles (Induction). Pembro (dose level 1=100mg, dose level 2=200mg) was then initiated every 3 wks in combination with umbralisib daily and ublituximab (900mg week 2 of cycle 4 and 6) for cycles 3-6 (Consolidation). Upon completion of cycle 6, pts continue umbralisib 800 mg daily until progressive disease (PD) or unacceptable toxicity (Maintenance). The primary endpoint is safety of pembro, umbralisib, and ublituximab; efficacy is a secondary endpoint. Response assessments are based on the iwCLL 2008 criteria and performed after umbralisib + ublituximab (2 mos), umbralisib, ublituximab, pembro (6 mos) and during maintenance with umbralisib at month 12. Peripheral blood and/or bone marrow biopsy were obtained for correlative analyses at screening, month 2 and month 6. For correlative analysis, mononuclear cells were enriched and cryopreserved. Cryopreserved cells were subjected to multicolor immunophenotyping to analyze: B cells (CD5, CD38, CD3, HLA-DR, CD19, PDL2, CD27, PDL1 and viability) and T/NK cells (CD8, CD56, CCCR7, CD3, CD4, IgG4, CD19, TIM-3, CD25, PD1 and viability).
Results: Ten pts were initially treated: 9 with CLL (3 on the 100 mg pembro cohort and 6 in the 200 mg pembro cohort) and 1 with Richter's Transformation (100 mg pembro cohort). Herein we report on the 9 CLL pts who were evaluable for safety and efficacy. Baseline demographics were as follows: male/female (5/4), median age 71 years (range 60-81), median prior therapies 1 (1-3), 78% refractory. 56% were treated with a BTK inhibitor (ibrutinib or acalabrutinib) prior to study enrollment, all of which were refractory to BTK therapy. 78% had at least 1 high risk genetic feature (del17p, del11q, TP53 mut, Notch1 mut or complex karyotype). All grade and grade ≥ 3 AEs during cycles 3-6 (umbralisib, ublituximab, and pembro combination) are listed in Table 1. Of note, only 1 DLT occurred (ALT/AST elevation - 200 mg pembro cohort) which trigged expansion to 6 pts, with no additional DLTs reported. The MTD was not reached and therefore the primary study endpoint was met. We did not observe an increase in expected grade ≥ 3 PI3Kδ-associated toxicities including pneumonitis, colitis, and transaminitis (1 event). ORR was 75% for the non-BTK refractory pts (3/4) and 60% in BTK refractory pts (3/5). At the time of this analysis, only 1 pt experienced PD and no deaths were observed (Figure 1). Eight of nine pts remain on study in follow up (range 4 - 21+ mos). Of note, patient 1 elected not to participate in the maintenance phase and has achieved an ongoing 21+ month PFS without therapy. Figure 2 includes the swimmers plot for all 9 CLL study pts. Four pts have had detailed correlative analyses. In one pt, there was a 20-fold increase in bone marrow CLL cells expressing PD-L1 /PD-L2 post treatment with pembro. The proportions of the major T cell subsets (including Tregs) and PD1 levels did not change appreciably during therapy.
Conclusion: These are the first data of a PI3K-δ inhibitor in combination with anti PD-1 therapy in CLL. The triplet combination of umbralisib + ublituximab + pembro was well-tolerated with durable responses in pts refractory to BTK inhibitor therapy. Enrollment is ongoing in both the CLL and Richter's Transformation cohorts. Correlative analyses for all subjects will be reported.
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Mato:Regeneron: Research Funding; Janssen: Consultancy; AstraZeneca: Consultancy; DTRM: Research Funding; AbbVie: Consultancy, Research Funding; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Portola: Research Funding; Acerta: Research Funding; Kite: Consultancy; Pharmacyclics: Research Funding; TG Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead Sciences, Inc.: Membership on an entity's Board of Directors or advisory committees. Dorsey:TG Therapeutics, Inc.: Consultancy. Schuster:Janssen: Consultancy, Honoraria; Gilead: Consultancy, Research Funding; Seattle Genetics: Consultancy; Bristol-Myers Squibb: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; Merck: Research Funding; Nordic Nanovector: Consultancy; Celgene: Consultancy, Research Funding. Svoboda:Celgene: Research Funding; Pharmacyclics: Research Funding; Merck: Research Funding; Seattle Genetics: Consultancy, Research Funding; Kite: Consultancy; BMS: Consultancy, Research Funding. Becker:GlycoMimetics, Inc.: Research Funding. Dwivedy Nasta:Incyte: Research Funding; Immunogen: Research Funding; Takeda: Research Funding. Landsburg:Takeda: Research Funding; Curis: Consultancy, Research Funding. Purdom:TG Therapeutics, Inc.: Employment, Equity Ownership. Paskalis:TG Therapeutics, Inc.: Employment, Equity Ownership. Sportelli:TG Therapeutics, Inc.: Employment, Equity Ownership. Miskin:TG Therapeutics, Inc.: Employment, Equity Ownership. Weiss:TG Therapeutics, Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Shadman:Merck: Research Funding; Celgene: Research Funding; Acerta Pharma: Research Funding; AbbVie: Other: advisory board; Genentech: Consultancy, Research Funding; Gilead: Research Funding; Emergent: Research Funding; PLEXXIKON: Research Funding; TG Therapeutics: Research Funding; Pharmacyclics: Other: advisory board, Research Funding.
Umbralisib is an oral, once-daily inhibitor of PI3Kδ and CK1ε that is highly selective for the delta isoform. The UNITY-NHL trial (NCT02793583) is a multicenter, open-label, Phase 2b study evaluating ...umbralisib in previously treated NHL patients. Herein, we present results from the UNITY-NHL study.
208 patients with R/R marginal zone, follicular, or small lymphocytic lymphoma (MZL, FL, SLL) unresponsive to prior treatments (≥1 MZL; ≥2 FL/SLL), including ≥1 anti-CD20–based therapy, were administered umbralisib 800mg orally once-daily until disease progression, unacceptable toxicity, or study withdrawal. The primary endpoint of the study was overall response rate (ORR) as assessed by an independent review committee, according to the revised IWG criteria.
At a median follow up of 27.8 months, MZL patients (n=69) had an ORR of 49.3% (95% CI 37.0% - 61.6%) with 15.9% achieving a complete response (CR), and a Disease Control Rate (DCR: CR+PR+SD) of 82.6%. Median time-to-response (TTR): 2.8 months (95% CI 2.7 - 2.9). No patient who achieved CR has progressed to date. At a median follow up of 27.5 months, FL patients (n=117) had an ORR of 45.3% (95% CI 36.1% - 54.8%) with 5.1% achieving a CR, and a DCR of 79.5%. Median TTR: 4.6 months (95% CI 3.0 - 5.6). At a median follow up of 29.3 months, SLL patients (n=22) had an ORR of 50.0% (95% CI 28.2 - 71.8) with 4.5% achieving a CR, and a DCR of 86.4%. Median TTR: 2.7 months (95% CI 2.4 - 2.8). At least one grade ≥3 treatment-emergent adverse event (TEAE) was reported in 53.4% of patients. TEAEs led to umbralisib discontinuation in 32 patients (15.4%). Grade ≥3 TEAEs reported in ≥10% of patients: neutropenia (11.5%) and diarrhea (10.1%). Increased ALT/AST (grade ≥3) occurred in 6.7%/7.2% of patients.
Umbralisib achieved meaningful clinical activity in a heavily pretreated iNHL population. The safety profile was manageable, with a relatively low incidence of immune-mediated toxicities and AE-related discontinuations. These results suggest umbralisib has a favorable benefit-risk profile in this patient population and have supported the Accelerated Approval of umbralisib in previously treated MZL and FL patients.
Time limited, response-adapted venetoclax, umbralisib and ublituximab is safe and yields high rates of undetectable MRD for relapsed CLL.
Many patients with chronic lymphocytic leukemia (CLL) will ...develop treatment resistance to Bruton’s tyrosine kinase (BTK) inhibitors. Phosphatidylinositol-3-kinase (PI3K) inhibitors, including umbralisib, have significant clinical activity in relapsed/refractory CLL, but prolonged exposure is associated with potential toxicities. Due to the synergistic anti-tumor effects of combined PI3K and BCL-2 inhibition, we sought to explore the feasibility of response-adapted, time-limited therapy to optimize disease control while mitigating the risks of prolonged treatment. We conducted a phase 1/2 clinical trial to determine the safety and efficacy of venetoclax in combination with umbralisib and the anti-CD20 monoclonal antibody, ublituximab, (U2-VeN) in patients with relapsed/refractory CLL (N=46) and Richter’s transformation (RT, N =5). After 12 cycles, treatment was stopped for patients with CLL who achieved undetectable minimal residual disease (uMRD). Adverse events of special interest included diarrhea in 50% of patients (11% grade 3/4), and AST and/or ALT elevation in 15 patients (33%), with 3 (7%) grade 3/4. There were no cases of tumor lysis syndrome (TLS) related to venetoclax, with outpatient initiation in 96% of patients. The intent-to-treat overall response rate for CLL was 98% with 100% of evaluable patients responding with 38% complete responses. The end of treatment rate of uMRD at 10-4 in bone marrow was 77% (30 out of 39), including a 71% uMRD rate among 14 patients refractory to prior BTK inhibitor. Time-limited venetoclax and U2 is safe and highly effective combination therapy for relapsed/refractory CLL patient including those who have been previously treated with covalent BTK inhibitors. This trial was registered on ClinicalTrials.gov under identifier: NCT03379051
Introduction: First generation PI3Kδ inhibitors such as idelalisib and duvelisib are active in patients (pts) with lymphoid malignancies but are often associated with significant immune-mediated ...adverse events, including transaminitis, diarrhea/colitis, and pneumonitis, as well as an increased risk of serious infections. These toxicities can be severe, and frequently lead to treatment discontinuation. Umbralisib (TGR-1202) is a next generation, once-daily, oral PI3Kδ inhibitor, that is active in pts with relapsed/refractory (R/R) lymphoid malignancies, with a 94% ORR in previously treated CLL (Burris et al, 2015). Umbralisib has a markedly different chemical structure and pharmacologic profile compared to idelalisib and duvelisib, and kinome-profiling further differentiates umbralisib from these agents. All three compounds are selective for the PI3Kδ isoform, however the PI3Kγ isoform, which promotes pro-inflammatory cytokines and increased recruitment and cytotoxicity of T cells, is not inhibited by umbralisib (Kaneda et al, Nature 2016). Umbralisib also has an additional effect on casein kinase-1 epsilon (CK-1ε), a protein which may have an inhibitory effect on regulatory T-cell function (Deng et al, 2016). Collectively, these pharmacologic features provide a potential rationale for the differentiated toxicity profile of umbralisib compared to other PI3Kδ inhibitors. Here, we present an integrated safety analysis for pts dosed with umbralisib either as monotherapy or in combination with other agents.
Methods: Safety data were pooled from 5 completed or ongoing Phase 1 or 2 studies containing umbralisib. All studies shared similar key eligibility criteria: pts had R/R lymphoid malignancies with an ECOG PS ≤ 2 without limit to number of prior therapies. Umbralisib was dosed daily until progression or unacceptable toxicity, while dosing of combination agents varied. Adverse event grading was done by CTCAE v4.03 criteria.
Results: A total of 336 patients were included in the analysis. Patients received the following regimens: umbralisib as monotherapy (135 pts) or umbralisib in combination with: the glycoengineered anti-CD20 mAb ublituximab (“U2”, 98 pts), ibrutinib (32 pts), ublituximab + ibrutinib (38 pts), or ublituximab + bendamustine (33 pts). Among the 336 patients, 32% had CLL/SLL, 35% DLBCL, 22% indolent NHL, and 12% other lymphoma. Median and mean duration of exposure to umbralisib was 5 and 9 months respectively, with the longest patient on daily umbralisib for 4+ years, and a cumulative duration of drug exposure across all 336 patients of over 240 years. Common all-grade adverse events regardless of causality are described in Table 1, and Grade 3/4 adverse events are described in Table 2. The most common all-grade non-hematologic toxicities were: diarrhea (44%), nausea (39%), and fatigue (35%). All-grade hematologic toxicities included neutropenia (22%), anemia (20%), and thrombocytopenia (18%). Grade 3/4 adverse events were infrequent, with the most common non-hematologic events including diarrhea (4%) and dyspnea (3%), and hematologic toxicities of neutropenia (17%), anemia (6%), and thrombocytopenia (5%). Key adverse events prevalent amongst prior generation PI3Kδ inhibitors were also infrequent: transaminitis (9% All; G3/4 <3%), colitis (<1.5% All; G3/4 <1%), and pneumonitis (<1.5% All; G3/4 <0.5%). Serious adverse events that occurred in >1% of patients were pneumonia (5%), febrile neutropenia (3%), sepsis (2%), and pyrexia (2%). Discontinuations due to adverse events were rare at under 10% for all studies.
Conclusions: In this integrated safety analysis of 336 patients, many of whom were on treatment for over a year, umbralisib exhibits a differentiated safety profile compared to prior generation PI3kδ inhibitors. Improved tolerability with very few discontinuations of umbralisib due to AEs has allowed patients to remain on continuous dosing to achieve and sustain promisingly high rates of response. The mechanism for decreased immune-mediated toxicity is still being elucidated through ongoing pre-clinical and correlative studies examining umbralisib's selectivity for PI3Kδ over PI3Kγ, complimentary CK1ε inhibition, and enhancement of regulatory T-cell function.
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Davids:TG Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Astra-Zeneca: Consultancy; Merck: Consultancy; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Infinity: Consultancy, Research Funding; Pharmacyclics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; InCyte: Membership on an entity's Board of Directors or advisory committees. Flinn:Beigene: Research Funding; Agios: Research Funding; Janssen: Research Funding; Constellation: Research Funding; Takeda: Research Funding; Forty Seven: Research Funding; Infinity: Research Funding; Portola: Research Funding; Janssen: Research Funding; Verastem: Research Funding; Genentech: Research Funding; Novartis: Research Funding; Pharmacyclics: Research Funding; Merck: Research Funding; KITE: Research Funding; Pfizer: Research Funding; TG Therapeutics: Research Funding; Pharmacyclics LLC: Research Funding; Seattle Genetics: Research Funding; Celgene: Research Funding; Curis: Research Funding; Trillium: Research Funding; AbbVie Company: Research Funding; Acerta: Research Funding; Calithera: Research Funding; Incyte: Research Funding; Gilead: Research Funding. Mato:Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; DTRM: Research Funding; Portola: Research Funding; Acerta: Research Funding; Gilead Sciences, Inc.: Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Research Funding; TG Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy; AstraZeneca: Consultancy; Kite: Consultancy; Pharmacyclics: Research Funding; Regeneron: Research Funding. O'Connor:Celgene: Honoraria, Research Funding; Trillium Therapeutics: Research Funding. Brander:Genentech: Consultancy; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Teva Pharmaceuticals, Genentech, AbbVie, Pharmacyclics: Consultancy; TG Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees. Lunning:Juno: Consultancy; AbbVie: Consultancy; Pharmacyclics: Consultancy; Celgene: Consultancy; Onyx: Consultancy; Gilead: Consultancy; TG Therapeutics: Consultancy; Genentech: Consultancy; Epizyme: Consultancy; Spectrum: Consultancy; BMS: Consultancy. Nastoupil:Celgene: Honoraria, Research Funding; Karus Therapeutics: Research Funding; Genentech: Honoraria, Research Funding; Gilead: Honoraria; Abbvie: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; TG Therapeutics: Honoraria, Research Funding. Flowers:Eastern Cooperative Oncology Group: Research Funding; National Cancer Institute: Research Funding; Millennium/Takeda: Research Funding; OptumRx: Consultancy; Pharmacyclics LLC, an AbbVie Company: Research Funding; Bayer: Consultancy; Prime Oncology: Research Funding; Gilead: Consultancy; Research to Practice: Research Funding; Educational Concepts: Research Funding; Abbvie: Consultancy, Research Funding; Spectrum: Consultancy; National Institutes Of Health: Research Funding; Janssen Pharmaceutical: Research Funding; Celgene: Consultancy, Research Funding; TG Therapeutics: Research Funding; Acerta: Research Funding; Clinical Care Options: Research Funding; V Foundation: Research Funding; Burroughs Welcome Fund: Research Funding; Seattle Genetics: Consultancy; Onyx: Research Funding; Genentech/Roche: Consultancy, Research Funding; Infinity: Research Funding. Brown:Celgene: Consultancy; AbbVie: Consultancy, Honoraria; AstraZeneca: Consultancy; Janssen: Consultancy; Janssen Oncology: Honoraria; Infinity Pharmaceuticals: Consultancy; Sun BioPharma: Consultancy, Research Funding; Pharmacyclics: Consultancy; Astellas Pharma: Consultancy; Roche/Genentech: Consultancy; Pfizer: Consultancy; Redx: Consultancy; Gilead: Consultancy, Research Funding. Ghosh:Pharmacyclics: Consultancy, Honoraria, Research Funding, Speakers Bureau; TG Therapeutics: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Honoraria, Research Funding; Gilead: Consultancy, Honoraria, Speakers Bureau; Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; AbbVie: Consultancy, Honoraria, Speakers Bureau; Jassen: Consultancy, Honoraria, Research Funding; Kite Pharma: Consultancy, Honoraria; Seattle Genetics: Consultancy, Honoraria, Research Funding. Lansigan:Spectrum Pharmaceuticals: Consultancy, Research Funding; Seattle Genetics: Consultancy. Cheson:AbbVie, Roche-Genentech, Pharmacyclics, Acerta: Consultancy; Acerta, Pharmacyclics, Epizyme, Gilead, Roche, AbbVi: Other: Institution receives research support . Barr:AbbVie: Consultancy, Research Funding; Novartis: Consultancy; Celgene: Consultancy; Gilead: Consultancy; Pharmacyclics LLC, an AbbVie Company: Consultancy, Research Funding; Seattle Genetics: Consultancy; Infinity: Consultancy. Pagel:Actinium Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees. Pinilla-Ibarz:ARIAD: Consultancy, Honoraria; BMS: Honoraria, Speakers Bureau; Pfizer: Honoraria, Speakers Bureau. Burke:Celgene: Consultancy; Bayer: Consultancy; Incyte: Consultancy; Gilead: Consultancy; Genentech: Consultancy. Siddiqi:Pharmacyclics, an AbbVie Company: Other: Steering committee for ibrutinib, Speakers Bureau; Ju
Background: Constitutively activated PI3K/AKT/mTOR pathway plays a critical role in the proliferation and survival of cancer cells, through the expression of numerous pro-survival and proliferative ...genes. Specific inhibitors of the various isoforms of PI3K have shown promising activity in the treatment of indolent B-cell lymphoma. However, they have not shown similar activity in aggressive lymphoma, potentially because the expression of many PI3K/AKT/mTOR dependent pro-survival genes may be activated by alternative signals, for example, NF-kappaB (NF-kB). While specific inhibitors of NF-kB are currently not available in the clinic, proteasome inhibitors are well known to inhibit NF-kB, among its pleiotropic mechanisms of action. We hypothesize that if cancer cell survival is dependent on a complex protein network involving PI3K and NF-kB, then complimentary inhibition of PI3K and the proteasome will become highly synergistic. Our results provide strong support for the hypothesis, by demonstrating that TGR-1202, a PI3Kdelta inhibitor, and carfilzomib, a proteasome inhibitor, are unique among the combinations of PI3K and proteasome inhibitors in potently suppressing the mTORC1 complex and NF-kB, and are highly effective in models of B- and T-cell lymphoma.
Methods: Cytotoxicity of PI3K and proteasome inhibitors was studied in a panel of B and T-cell lymphoma cell lines. Growth inhibition was determined using Cell Titer Glo that measures ATP produced by live and proliferating cells. Cell death was determined by flow cytometry, using the probes Annexin V and propidium iodide. Drug: drug synergy was determined by calculating the combination index (CI) and relative risk ratio (RRR). Values below 1 indicate synergy, with smaller RRR and CI values indicating higher levels of synergy. The mechanistic basis of the synergy is determined through in-depth interrogation of the PI3K/AKT/mTOR pathway and its downstream targets and interacting pathways.
Results: We used a high-throughput platform to screen the cytotoxicity of TGR-1202, idelalisib/CAL-101, bortezomib, and carfilzomib, in a panoply of B- and T-cell lymphoma cell lines. We found that TGR-1202 and idelalisib as single agent caused only minimal to mild cytotoxicity in lymphoma cells. Bortezomib and carfilzomib as single agent caused dose dependent inhibition of cell growth. Surprisingly, when TGR-1202 or Cal-101 were combined with bortezomib or carfilzomib in a 2x2 design, the combination of TGR-1202 and carfilzomib was consistently the most synergistic, while the other 3 combinations were either much less or not at all synergistic. For example, Table 1 clearly demonstrated that TGR-1202 and carfilzomib demonstrated much higher inhibitory effect and stronger synergy than Cal-101 and bortezomib in two T-ALL cell lines. We have systematically interrogated the effects of these drug combinations on the PI3K/AKT/mTOR pathway, and found that the combination of TGR-1202 and carfilzomib was able to uniquely and potently inhibit components of the mTORC1 complex, causing inhibition of mTORC1 dependent signals such as phosphorylated 4E-BP1, leading to markedly reduced expression of critical genes such as c-myc (Figure 1). Furthermore, the combination of TGR-1202 and carfilzomib uniquely and potently inhibited activation of NF-kB, resulting in markedly suppressed expression of Bcl-xL. The complementary inhibition of the PI3K/AKT/mTOR and NF-kB pathways resulted in robust growth inhibition and apoptosis (Figure 1), and warrants further clinical evaluation for aggressive B- and T-cell lymphoma.
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Sportelli:TG Therapeutics: Employment, Equity Ownership. Miskin:TG Therapeutics, Inc.: Employment, Equity Ownership. Viswanadha:Incozen: Employment.
The constitutively activated PI3K/AKT/mTOR pathway plays a key role in the proliferation and survival of cancer cells. Specific inhibitors of the delta isoform of PI3K, such GS-1101 (idelalisib), ...have shown promising activity in the treatment of B-cell lymphoma. In contrast, some data from models of T-cell lymphoma (TCL) have suggested that these diseases may require inhibition of both PI3Kdelta and PI3Kgamma for optimal cytotoxicity. Proteasome inhibitors, such as carfilzomib, potently inhibit the activation of NF-kappaB (NF-kB) by preventing degradation of the NF-kB inhibitor IkB. We hypothesize that inhibition of this pathway at both the proximal (PI3K) and distal (NF-kB) aspects, using both inhibitors of PI3K and proteasome, could lead to a synergistic cyototoxic effect in models of lymphoma.
Cytotoxicity of TGR-1202 and carfilzomib was studied in a panel of B and T-cell lymphoma cell lines. Growth inhibition was determined using Cell TiterGlo that measures ATP produced by live and proliferating cells. Cell death was determined by flow cytometry, using the probes Annexin V and propidium iodide. Drug: drug synergy was determined by calculating relative risk ratio (RRR). Values below 1 indicate synergy, with smaller RRR values corresponding to higher levels of synergy.
TGR-1202 as a single agent active against a panel of diverse B- and T-lymphomas. By the Cell TiterGlo assay, the concentration required to inhibit growth by 50% (IC50) following 48-hour exposure ranged from 10 uM to 15 uM in most of the cell lines. Maver and H9, representing mantle cell lymphoma (MCL) and TCL respectively, were more resistant to TGR-1202. Carfilzomib, on the other hand, is a potent inhibitor of B- and T cell lymphomas, with an IC50 in the range of 2-8 nM most lymphomas (Table 1). Remarkably, when TGR-1202 is combined with carfilzomib at minimally, or mildly inhibitory concentrations, there was a highly synergistic inhibition in both B and T-cell lymphomas. Following a 24 hour exposure, carfilzomib alone achieved only 20% growth inhibition of the MCL cell line Jeko-1, while TGR-1202 at concentrations ranging from 2.5 to 15 uM did not produce any growth inhibition (Figure 1). However, the combination of carfilzomib and TGR-1202 markedly inhibited growth of Jeko-1, with a synergy index, RRR, as low as 0.10. In contrast, the synergy of carfilzomib and TGR-1202 in the TCL cell line H9 following a 48-hour exposure, as shown in Figure 2, demonstrated remarkable synergy with a RRR as low as 0.02. To further confirm the synergy of these two drugs in lymphoma, apoptosis was determined in H9 cells treated either with each drug as a single agent or together in combination. Figure 4 illustrates that the combination of these two drugs at marginally active concentrations caused apoptosis in as many as 90% of the H9 cells. The synergy of TGR-1202 and carfilzomib stood out among the combinations tested, as being among one of the most synergistic combinations explored. Table 2 contains a summary of the synergy seen with TGR-1202 and carfilzomib, in contrast to the effects seen with other drug: drug combinations.
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Table 1IC50 of carfilzomib in B- and T-cell lymphoma cells.Cell lineLymphomaIC50 (nM)LY1DLBCL8.1LY10DLBCL1.8Rec-1MCLNRHBL2MCL2.4MaverMCL5.6HHCTCL3.9H9CTCL4.0
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In conclusion, the combination of the PI3K delta inhibitor TGR-1202 and the proteasome inhibitor carfilzomib remarkably and distinctively synergize to kill both B and T-cell lymphoma cells, and represents a promising therapeutic strategy in the treatment of these diseases.
C-: Negative control
carfilzomib
TGR-1202
Relative risk ratio. Values below 1 indicate synergy.
C-: Negative control
carfilzomib
TGR-1202
Relative risk ratio. Values below 1 indicate synergy.
IC50 was not reached at the highest tested concentrations of 32 nM.
Diffuse large B cell lymphoma
Mantle cell lymphoma
Cutaneous T cell lymphoma
SportelliTG Therapeutics, Inc. : Employment, Equity Ownership. Miskin:TG Therapeutics, Inc.: Employment, Equity Ownership. Vakkalanka:3Rhizen Pharmaceuticals: Employment, Equity Ownership. Viswanadha:Incozen Therapeutics Pvt. Ltd.: Employment, Equity Ownership.
The phosphatidylinositol 3-kinase (PI3K) pathway is consistently activated in relapsed/refractory Hodgkin lymphoma (HL), suggesting that TGR-1202, a novel inhibitor of the delta isoform of PI3K ...(PI3K-δ), in clinical development for patients with hematologic malignancies, might represent an attractive therapeutic option. The anti-CD30 monoclonal antibody Brentuximab Vedotin (BV) conjugated to the microtubule-disrupting agent monomethyl auristatin E (MMAE) has recently been reported to induce an overall response rate of 75% in relapsed/refractory HL, but is associated with limited response duration. Combination therapies aimed at enhancing the anti-tumor activity of BV and reducing its side effects may have significant clinical impact in the treatment of relapsed/refractory HL. The present study was aimed at investigating the activity and mechanism(s) of action of the PI3K-δ inhibitor TGR-1202, in combination with BV in non-clinical models of HL.
Three HL cell lines, including L-540, KM-H2 and L-428, were used to test the effects of TGR-1202 alone, BV alone, or the combination of TGR-1202 with BV. Cell cycle effects and cell survival were determined by flow cytometry and Western blotting (WB). Additionally, WB was used to assess modulating effects of TGR-1202 on the PI3K/AKT pathway as well as microtubule interacting proteins. Cyclin B1, p21, and α-tubulin were detected by indirect immunofluorescence microscopy. The activity of TGR-1202 and/or BV on microtubule distribution and polymerization were quantified using a three-dimensional volume rendering technique.
TGR-1202 and BV used as single agents were able to induce a time- and dose-dependent inhibition of cell proliferation and induction of cell death in all cell lines. Compared to the single agent effects, the combination of TGR-1202 (10 µM) and BV (10 ng/ml) synergistically inhibited the mean (±SEM) growth of L-540, KM-H2, and L-428 cell lines (TGR-1202: 40 ± 4%; BV: 30 ± 2%; TGR-1202/BV: 85 ± 1%). Inhibition of cell proliferation induced by the 2-drug combination was associated with a dramatic G2/M cell cycle arrest. Upon TGR-1202/BV treatment, the mean (±SEM) percentages of cells in G2/M phases were increased by 4-fold (72 ± 3%) as compared to TGR-1202 (18 ± 1%) or BV (18 ± 1%) alone. This finding was paralleled by a 3-fold reduction of cells in S phase (TGR-1202: 25 ± 1%; BV: 23 ± 1%; TGR-1202/BV: 9 ± 1%, mean ± SEM) and a marked Cyclin B1 and p21 overexpression. In comparison to each drug as a single agent, the TGR-1202/BV combination led to a synergistic cell death induction. In fact, upon TGR-1202/BV treatment, mean (±SEM) cell death values detected in L-540, KM-H2, and L-428 cell lines were increased by 3-fold over TGR-1202 or BV alone (TGR-1202: 27 ± 2%; BV: 27 ± 2%; TGR-1202/BV: 75 ± 2%). Analysis of caspase-3 and PARP cleavage and blocking experiments with the pan-caspase inhibitor Z-VAD-FMK revealed a caspase-dependent cell death mechanism. In addition, the antiproliferative and cytotoxic effects of TGR-1202 were associated with a marked time-dependent inhibition of PI3K/Akt pathway and dephosphorylation of GSK-3β, Aurora kinases, and stathmin, suggesting that modulation of molecules associated with microtubule polymerization are critically involved in TGR-1202/BV-triggered cell death. To asses potential effects on microtubule dynamics, HL cells were treated with TGR-1202, BV, or the combination for 24 hours, and the effect on microtubules was determined by α-tubulin staining. Compared with controls, TGR-1202 and BV treatment alone led to a modest loss of microtubules (TGR-1202: 11%; BV: 9%), while the combined TGR-1202/BV treatment resulted in a potent synergistic microtubule disruption (mean values of α-tubulin inhibition of 40%, P ≤.0001), supported by a diffuse stain and irregular microtubule fragments throughout the cytosol. Additionally, TGR-1202/BV was found to interfere with the mitotic spindle integrity which may suggest that the G2/M arrest and cytotoxicity of the combined TGR-1202/BV treatment primarily arises from the inhibition of tubulin polymerization.
Novel PI3K-δ inhibitor TGR-1202 enhances the anti-tumor activity of BV by increasing drug-induced apoptosis and tubulin disruption in all HL cell lines analyzed in the present study. Our data provides a strong rationale for evaluating TGR-1202 in combination with BV in patients with relapsed/refractory HL.
Sportelli:TG Therapeutics, Inc.: Employment, Equity Ownership.
PI3K-δ is highly expressed in cells of hematopoietic origin and is often upregulated in lymphoid malignancies. TGR-1202 is a novel, next generation PI3K-δ inhibitor shown to inhibit Akt ...phosphorylation and induce apoptosis in lymphoma and leukemia cell lines, displaying activity in numerous pre-clinical models with potentially superior pharmacokinetic (PK) properties to other PI3K-δ inhibitors in development, including an extended half-life observed in pre-clinical animal studies. In addition, the chemical structure of TGR-1202 was designed specifically to avoid heterocylic nitrogen moieties in the backbone of the molecule, known to interact with hepatic enzymes, in an effort to mitigate a class effect of hepatotoxicity. Herein we present preliminary results of a Phase I, first in human, open label study of the oral PI3K-δ inhibitor, TGR-1202.
The dose escalation portion of the study will determine the maximum tolerated dose (MTD) of TGR-1202 using a standard 3+3 dose escalation design. Patients (pts) with a confirmed diagnosis of B-cell non-Hodgkin lymphoma, chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), peripheral T-cell lymphoma, or other lymphoproliferative disorders previously treated with at least one prior therapy are eligible. Additional eligibility criteria include ECOG PS ≤ 2 and measurable/evaluable disease. Primary endpoints are safety and PK measurement; secondary endpoints include pharmacodynamic (PD) analysis and efficacy (overall response rate). TGR-1202 will be administered orally QD until unacceptable toxicity, disease progression, or withdrawal from treatment. Treatment cycles are 28 days. Efficacy evaluations are planned every 8 weeks. AEs are assessed using the CTCAE v4.0. For pts with CLL/SLL, hematologic toxicity is graded according to IWCLL guidelines.
Thirteen patients have been enrolled to date across 4 dose levels: QD 50mg, 100mg, 200mg, and 400mg. Pts are 83% male, ECOG 0/1: 54%/46%, with a median age of 69 yrs (range: 49-82). Patients had a median of 2 (range: 1-5) prior treatment regimens, and 38% were refractory to prior treatment. Lymphoma histologies include follicular lymphoma (5 pts), CLL/SLL (4 pts), and lymphoplasmacytic lymphoma, mantle cell lymphoma, DLBCL, and atypical hairy cell leukemia (1 pt each). Twelve of the thirteen patients are evaluable for DLT assessment (one patient in Cohort 1 was replaced within first week of treatment due to rapid disease progression). Of the 12 evaluable patients, 8 are currently on study; 4 patients discontinued treatment due to disease progression. To date, there have been no DLTs. One patient experienced G3 neutropenia and thrombocytopenia which resulted in a dose reduction. No other G3/4 related hematologic or non-hematologic toxicities have been observed. Notably, aside from unrelated G1 elevation of GGT in one patient, there has been no hepatotoxicity. Of the 7 patients who completed 2 cycles of treatment (8 wks) at 200mg daily dosing or less, 3 (43%) showed SD and 4 (57%) had PD. Five additional patients have not reached their first response assessment.
TGR-1202's preliminary mean pharmacokinetic parameters determined on Day 1 of Cycle 1 (table) are: median Tmax of 2 hrs (range 1-8hrs), harmonic mean t1/2 of 13 (± 8.06) hr, and CL/F of 48.97 (± 15.6) L/hr. CL values over the 4 dose levels are independent of dose (r = 0.07, p > 0.20). A linear relationship (Spearman's) exists between dose and both AUC (r = 0.95) and Cmax (r = 0.80). Steady-state levels are reached by Day 15. The ave. accumulation Cmin ratio between Cycle 1 t24hr & Cycle 2 t0hr is 8 (± 2.65).
Preliminary PK data from the first 4 cohorts is summarized below.Dose50 mg(n=4)100 mg(n=3)200 mg(n=3)400 mg(n=3)Cmax (ng/mL)107 (± 47)179 (± 41)157 (± 12.5)585 (± 243)AUC (ug x hr/mL)1.3 (± 0.61)2.1 (± 0.14)4.2 (± 1.01)6.2 (± 3.56)
To date, TGR-1202 has been well tolerated in patients with relapsed/refractory hematologic malignancies. There have been no DLTs observed and toxicities have been minimal to date. Enrollment continues at higher dose cohorts. Updated safety, efficacy, PK, and PD data will be presented.
Lanasa:Rhizen Pharmaceuticals S A: Research Funding. Kuhn:TG Therapeutics, Inc.: Consultancy. Miskin:TG Therapeutics, Inc.: Employment, Equity Ownership. Sportelli:TG Therapeutics, Inc. : Employment, Equity Ownership. Vakkalanka:Rhizen Pharmaceuticals S A: Employment, Equity Ownership.
Abstract 5018
The bone marrow microenvironment contributes to the pathogenesis of Multiple Myeloma (MM) by promoting the oncogenic process including drug resistance. High expression levels of the ...PI3Kδ isoform in patient MM cells implicate this target as a novel and attractive intervention strategy aimed at attenuating the progression of the disease. Herein, we describe the biological and pharmacokinetic properties of TGR-1202, a novel and small molecule PI3Kδ inhibitor with scope to be developed as a clinical candidate for patients with relapsed and/or refractory MM.
Activity of TGR-1202 on individual PI3K isoforms was determined in enzyme, cell, and whole blood based assays. Potency of the compound was confirmed viaMM cell viability and apoptosis assays besides testing for inhibition of pAkt, a downstream kinase regulating cell survival and growth. Additionally, the compound was tested for potency in viability, apoptosis, and Akt phosphorylation assays using immortalized and MM patient derived primary cells. ADME and pharmacokinetic properties of the molecule were also determined.
TGR-1202 demonstrated significant potency against PI3Kδ (22. 2 nM) with several fold selectivity over the α (>10000), β (>50), and γ (>48) isoforms. Additionally, the compound inhibited B-cell proliferation (24. 3 nM) and FcεR1 induced CD63 expression in human whole blood basophils (68. 2 nM) indicating specificity towards the delta isoform. The addition of 5 μM TGR-1202 to dexamethasone (0. 55 μM) caused a dramatic shift in GI50 of MM-1S cells compared to that of either TGR-1202 (9. 7 μM) or dexamethasone (18. 3 μM) alone. Further viability testing demonstrated that the compound caused a dose-dependent inhibition (>50% @ 1–3 μM) in growth of immortalized (U266B1 and MM-1R) as well as patient-derived MM cells. Reduction in viability was accompanied by a reduction in pAKT (>80% @ 3 μM) along with induction of apoptosis in both cell lines and patient samples. Pharmacokinetic studies across species indicated good oral absorption (>40% bioavailability for mice, rat, and dog) with favorable plasma concentrations (3–10 μM @ 20 mg/kg for mice, rat, and dog) relevant for efficacy.
Data demonstrate the therapeutic potential of TGR-1202 in MM viainhibition of the PI3Kδ pathway. Data from immortalized as well as primary patient-derived cells justify further evaluation of this molecule in MM. Combination therapy of TGR-1202 and dexamethasone in a mouse MM-1S xenograft model is currently ongoing.
Vakkalanka:Rhizen Pharmaceuticals S A: Employment, Equity Ownership. Viswanadha:Incozen Therapeutics: Employment. Sportelli:TG Therapeutics, Inc.: Employment, Equity Ownership. Miskin:TG Therapeutics, Inc.: Employment, Equity Ownership.
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