ALBERT

Description: Abstract 286 It is well established that deregulation of the PI3K signaling pathway plays an important role in the etiology of human malignancies including those of hematologic origin. In 30–50% of solid tumors, oncogenic activation of the PI3K pathway is the result of gain-of-function mutations in the PI3K p110α isoform or due to the loss-of-function of the PTEN phosphatase that is responsible for PI3K downregulation. In B cell malignancies these mutations are rarely observed in spite of the fact that PI3K pathway activation is commonly observed and often essential for tumor cell growth and survival. In this case, PI3K pathway activation has been shown to result from constitutive B cell receptor (BCR) activation and/or from exposure to survival factors present in the microenvironment. The activation of the PI3K pathway by cell surface receptors is directly mediated by the Class I isoforms (α, β, δ, and γ), however, their relative contribution in B cell tumors is unknown. Interestingly, genetic and pharmacological approaches that specifically inactivate the p110δ isoform have demonstrated its important role in normal B cell signaling in response to multiple factors including antigen, CD40L, BAFF, SDF-1 and CXCR13 making it an attractive target for therapeutic intervention in B cell malignancies. CAL-101 is an oral p110δ specific inhibitor which is currently being evaluated in a phase I clinical trial for the treatment of patients with select hematologic malignancies. This compound is a novel potent p110δ inhibitor with an IC50 of 2.5 nM against purified p110δ and EC50 of 65 nM in p110δ-mediated basophil activation in whole blood. CAL-101 demonstrates 300-, 200-, and 40-fold selectivity over the other class I family members (α, β, and, γ respectively) and no activity against class II and III PI3K family members or other PI3K-related proteins including mTOR and DNA-PK. Furthermore, a kinome-wide screen failed to detect activity against any additional kinases. To investigate the potential role of p110δ in B cell hematologic tumors we screened a wide range of leukemia and lymphoma cell lines for constitutive PI3K pathway activation. The expression of p110δ was observed in 〉90% of these cell lines and in many cases was accompanied by constitutive Akt phosphorylation. In this context, CAL-101 was able to reduce p-Akt levels and block additional downstream effectors such as p-S6, and GSK-3β in cells that represent a range of tumor types including follicular lymphoma, acute lymphoblastic leukemia (ALL), diffuse large B-cell lymphoma, and mantle cell lymphoma (MCL). Furthermore, treatment with CAL-101 resulted in growth suppression and induction of apoptosis which was accompanied by PARP and caspase-3 cleavage. Growing evidence suggests that signals from the microenviroment are essential for the expansion, homing, and survival of malignant B cells, in addition to promoting resistance to conventional drug therapy. To investigate the potential role p110δ plays during B cell signaling via interactions with the microenvironment, we examined invoked stimulation of leukemia and lymphoma cell lines with CXCL12, CXCL13, BAFF, or BCR crosslinking in the presence or absence of CAL-101. Stimulation with either chemokines or growth factors resulted in the phosphorylation of Akt which was inhibited by CAL-101 in a dose dependent manner. Moreover, p110δ inhibition with CAL-101 inhibits the chemotaxis of ALL and MCL cell lines to CXCL12. These studies have now been extended to the analysis of primary patient B-ALL and MCL cells to further establish preclinical proof of concept for therapeutic application of CAL-101. In summary, CAL-101 is a potent and selective p110δ kinase inhibitor with broad anti-tumor activity against cancer cells of hematologic origin. Our results demonstrate that selective inhibition of p110δ with CAL-101 inhibits malignant B cell growth, survival, and migration. Furthermore, p110δ inhibition may enhance the effect of cytotoxic drugs or overcome cell mediated drug resistance by inhibiting the protective signals of the microenviroment, providing a rational for combination therapy. These data suggest that p110δ may play an important role in regulating signals between malignant B cells and their microenvironment thereby providing the preclinical rationale for clinical evaluation of CAL-101 alone or in combination with cytotoxics or biologics in patients with hematologic malignancies. Disclosures: Lannutti: Calistoga Pharmaceuticals: Employment. Meadows:Calistoga Pharmaceuticals: Employment. Kashishian:Calistoga Pharmaceuticals: Employment. Steiner:Calistoga Pharmaceuticals: Employment. Johnson:Calistoga Pharmaceuticals: Research Funding. Giese:Calistoga Pharmaceuticals: Employment.

Description: The class I phosphatidylinositol 3-kinases (PI3K) regulate a variety of cellular functions including motility, metabolism, proliferation, growth, and survival, depending on cellular milieu. Deregulation of the PI3K/Akt pathway is one of the most frequently observed defects in human malignancies including those of hematological origin and has been shown to play an important role in tumor progression. Therefore, selective targeting of PI3K signaling in hematological tumor cells could provide an effective treatment strategy while limiting potential undesirable effects of pan-inhibitors that broadly block PI3K signaling in all cells. Of the class IA PI3Ks (p110a, p110b, p110d), p110d’s expression is largely restricted to cells of hematopoietic origin and is essential for PI3K signaling in lymphocytes. Here, we report on the characterization of a novel p110d specific inhibitor, CAL-101. This compound is a potent PI3K inhibitor with an IC50 of 1–10 nM against the purified p110d subunit and 30–70 nM cellular potency against p110d-mediated basophil activation in whole blood. Importantly, CAL-101 plasma concentrations of 500–5000 nM that greatly exceed those needed for p110d inhibition in blood were safely maintained in a 7 day multidose normal human volunteer study. CAL-101 demonstrates 〉30-fold selectivity over other class I, II and III PI3K family members as well as selectivity over other PI3K-related proteins including mTOR and DNA-PK. Furthermore, a genome wide screen of 〉350 protein kinases did not detect any activity. To investigate the potential role of p110d in hematologic tumors we screened a wide range of leukemia and lymphoma cell lines for p110 isoform expression and constitutive PI3K pathway activation. The expression of p110d was observed in 〉90% of these cell lines that was in many cases accompanied by constitutive Akt phosphorylation. In this context, CAL-101 was able to reduce p-Akt levels and block additional downstream effectors such as p-p70S6K, p-GSKb, and p-Bad in cells that represent a range of tumor types including acute myeloid leukemia, acute lymphoblastic leukemia (ALL), and diffuse large B-cell lymphoma among others. Recent studies have demonstrated the importance of PTEN loss and enhanced PI3K signaling in primary T-ALL cells. We report high levels of p110d protein and activated Akt in 6 of 6 ALL cell lines evaluated. Inhibition of p110d with CAL-101 treatment of both T-ALL and B-ALL cell lines resulted in a reduction of Akt and GSK-3b phosphorylation and a decrease in cellular proliferation that was accompanied by cell death demonstrating an essential role of PI3K signaling independent of PTEN status. Treatment of T-ALL cell lines with CAL-101 induced processing of pro-caspase-3 and cleavage of PARP supporting a role for caspase mediated cell death. These studies have now been extended to the analysis of primary patient blast samples to further establish preclinical proof of concept for therapeutic application of CAL-101 for the treatment of ALL. In summary, CAL-101 is a highly potent and selective p110d kinase inhibiter with broad anti-tumor activity against cancer cells of hematologic origin. Clinical studies in normal human volunteers demonstrated good tolerability with high drug exposure and favorable steady-state pharmacokinetic properties. Taken together, these data support the on going Phase 1 clinical trial that includes a wide range of hematological malignancies.

Description: Abstract 48 BCR signaling is increasingly recognized as a central mechanism for disease progression in CLL. PI3K transmits various external, microenvironment-derived signals that influence survival, drug resistance, and cell migration of CLL cells. PI3K isoforms have distinct functions and tissue distributions. P110δ mutant mice display defective BCR signaling, along with impaired B cell development and differentiation. Because of this unique role of p110δ in BCR signaling, p110δ inhibitors have been developed for treatment of B cell malignancies and autoimmune disorders. CAL-101, a potent and selective p110δ showing an IC50 of 2.5 nM in vitro and 〉200-fold selectivity relative to other PI3K isoforms, displays pre-clinical and clinical activity in CLL. Prior in vitro CLL studies revealed that CAL-101 induces caspase-dependent apoptosis, and inhibits CD40L-, BAFF-, TNF-alpha- and fibronectin-derived survival signals. Because of the importance of p110δ in BCR signaling, we tested the effects of CAL-101 on BCR-derived CLL cell activation, and confirmed our findings in correlative studies related to an ongoing CAL-101 phase I clinical trial. We found that BCR cross-linking with anti-IgM significantly increased CLL cell viability to 121 ± 5 % of controls (mean ± SEM, n=15, *P〈 0.05).This pro-survival effect was abrogated by CAL-101, which reduced CLL cell viability to 85± 3 % of controls at 48 hours (mean ± SEM, n=15, *P〈 0.05). CLL cell viability in co-culture with NLC was also significantly reduced by CAL-101 to 64± 6% of untreated controls at 48 hours (mean ± SEM, n=10, *P〈 0.05). BCR cross-linking induces secretion of the chemokines CCL3 and CCL4 by CLL cells, which was quantified in CLL supernatants by ELISA. CAL-101 significantly reduced supernatant CCL3 concentrations from 4060 ± 1392 pg/mL to 2901 ± 1220 pg/mL, and CCL4 levels from 5721 ± 1789 pg/mL to 3223 ± 1311 pg/mL (mean ± SEM, n=6, *P

Description: Background Myelofibrosis (MF) is characterized by activation of the JAK-STAT pathway, with the JAK2 V617F mutation found in 50-60% of patients. Although JAK inhibitors, such as FDA-approved ruxolitinib, have been effective in reducing splenomegaly and mitigating symptoms, patients uniformly exhibit “disease persistence” which is equated with a lack of hematologic or molecular remissions, or with loss of clinical improvement over time. Prior studies using cell lines or primary patient samples have shown that the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway is dysregulated in MPNs and is a potential therapeutic target (Kamishimoto et al, Cell Signal 2011; Huang et al, ASH 2009 Abstract 1896; Vannucchi et al, ASH 2011 Abstract 3835; Khan et al, Leukemia 2013). In CLL and other B-cell malignancies, the PI3K pathway is constitutively upregulated and is dependent on PI3Kδ. Idelalisib is a δ-isoform-specific PI3K inhibitor that is efficacious in patients with CLL and indolent NHL. Herein, the specific aims of our study were: 1) to determine whether the PI3Kδ isoform is expressed in progenitor cells from MF patients, and 2) to evaluate the inhibitory effects of idelalisib on basal and thrombopoietin (TPO)-stimulated AKT/S6RP phosphorylation (p-AKT/p-S6RP) in cell lines and in primary samples from MF patients who were either on chronic ruxolitinib (RUX) therapy or were not exposed to ruxolitinib (RUX-naïve or off-therapy at the time of sample collection). Methods To evaluate isoform expression, CD34+ cells from the peripheral blood of MF patients were sorted by FACSAria and cell lysates were analyzed by Simple Western using Peggy (ProteinSimple) with recombinant protein as a positive control. For cell line studies, BaF3/MPL W515L and UT-7/TPO cells were stimulated with recombinant human TPO and incubated with idelalisib. Whole cell lysates were analyzed by Western blot to quantify the % of p-AKT and p-S6RP levels compared to idelalisib-untreated cells. For MF patient samples, PBMCs were isolated from the whole blood of MF patients who were either RUX-naïve or on chronic RUX therapy and treated for 2 hours with idelalisib. Antibodies specific to p-AKT Ser473 and pS6RP Ser235/236 were used to quantify the proportion of p-AKT and pS6RP in basal and TPO-stimulated CD34+/CD3-/CD14-/CD19-/CD66- gated cells. Results The PI3Kδ isoform was found to be the predominant isoform expressed in 3 of 3 RUX-naïve and 4 of 4 chronic RUX patients tested; PI3Kβ was expressed at lower levels and no PI3Kα or γ was detected (Figure 1). In BaF3/MPL cells, p-AKT levels decreased by 51%, 64% and 67%, with 0.1, 1.0, 2.0 µM idelalisib, respectively, when compared to idelalisib-untreated cells; p-S6RP levels decreased by 24%, 27%, and 41%, respectively. Similarly, for UT-7/TPO cells, p-AKT decreased by 11%, 44%, and 55%, and p-S6 decreased by 13%, 28% and 48%, respectively. In CD34+ cells from RUX-naïve patients (n=3), p-AKT and p-S6RP levels decreased with increasing concentrations of idelalisib (0.02, 0.2, 2 µM). All patients on chronic RUX treatment demonstrated decreased p-AKT (n=3) and p-S6RP (n=4 basal, n=3 TPO-induced; patient 4 was only tested for basal) levels with increasing concentrations of idelalisib in both basal (Figure 2A) and TPO-stimulated (Figure 2B) assays. All 4 chronic RUX and 2 of 3 RUX-naïve patients tested carried the JAK2 V617F mutation. Conclusions The PI3Kδ isoform was identified as the predominant isoform expressed in CD34+ cells from MF patients. In both cell lines and patient samples, idelalisib inhibits the PI3K/AKT pathway, with a dose-dependent decrease of p-AKT and p-S6RP. Inhibition was observed for both RUX-naïve and chronic RUX-treated patients. Studies are underway to evaluate the effects of idelalisib on progenitor colony formation and induction of cell cycle arrest and apoptosis. * Meadows and Nguyen are first co-authors Disclosures: Meadows: Gilead: Employment, Equity Ownership. Queva:Gilead: Employment, Equity Ownership. Lannutti:Gilead, Acetra, Effector: Consultancy, Employment, Equity Ownership, Membership on an entity’s Board of Directors or advisory committees. Kashishian:Gilead: Employment, Equity Ownership. Jun:Gilead: Employment, Equity Ownership. Coutre:Gilead: Research Funding. Dansey:Gilead: Employment, Equity Ownership. Gotlib:Gilead: Consultancy, Research Funding.

Description: GS-1101 (CAL-101) is an oral PI3Kδ-specific inhibitor that has shown preclinical and clinical activity in non-Hodgkin lymphoma and chronic lymphocytic leukemia. To investigate the potential role of PI3Kδ in Hodgkin lymphoma (HL), we screened 5 HL cell lines and primary samples from patients with HL for PI3Kδ isoform expression and constitutive PI3K pathway activation. Inhibition of PI3Kδ by GS-1101 resulted in the inhibition of Akt phosphorylation. Cocultures with stroma cells induced Akt activation in HL cells, and this effect was blocked by GS-1101. Conversely, production of the stroma-stimulating chemokine, CCL5, by HL cells was reduced by GS-1101. GS-1101 also induced dose-dependent apoptosis of HL cells at 48 hours. Reductions in cell viability and apoptosis were enhanced when combining GS-1101 with the mTOR inhibitor everolimus. Our findings suggest that excessive PI3Kδ activity is characteristic in HL and support clinical evaluation of GS-1101, alone and in combination, as targeted therapy for HL.

Description: Abstract 3730 Background: The PI3K/Akt/mTOR pathway plays a critical role in cellular proliferation and survival through transduction of signals from cell-surface receptors to proteins involved in cell cycling (eg, cyclin D1) and mRNA translation (eg, ribosomal protein S6 and translation initiation factor 4E-binding protein 1 [4E-BP1]). MCL is an aggressive B-cell non-Hodgkin lymphoma. Overexpression of cyclin D1 has prompted clinical evaluation of mTOR inhibitors (everolimus, temsirolimus) in MCL. While efficacy has been observed, the extent and duration of tumor control has been modest, encouraging assessment of additional methods of intervention. Among the several PI3K isoforms (p110α, β, δ, γ), we have previously shown a unique role for PI3Kδ in maintaining the survival of hematological cancers and have also shown that GS-1101, a highly selective oral PI3Kδ-isoform-specific inhibitor with no activity against mTOR, shows promising clinical activity in non-Hodgkin lymphoma (NHL) and chronic lymphocytic leukemia (CLL). These considerations prompted us to assess the specific role of PI3Kδ in MCL and to determine if dual PI3Kδ/mTOR inhibition might enhance antitumor effects. Results: In all evaluated MCL cell lines (Jeko, Mino, Granta, and NCEB), PI3Kδ levels were high while expression of PI3Kα, β, and γ were variable. PI3Kδ was functionally active, inducing phosphorylation of Akt (pAkt) in all tested cell lines and also in 4 of 4 primary MCL samples. Single-agent GS-1101 decreased pAkt in cell lines and primary samples. The pAkt decrease was associated with growth suppression and induction of apoptosis in all MCL cell lines. Consistent with these effects, immunoblotting showed that GS-1101 decreased cyclin D1 levels in MCL cell lines. Because signals from the tumor microenvironment are essential for homing, survival, and proliferation of malignant B cells, we investigated the potential role of PI3Kδ in mediating these signals by stimulating MCL lines with CXCL12 (SDF-1), CXCL13 (BCA-1), BAFF, or BCR crosslinking in the presence or absence of GS-1101. These stimuli all resulted in the phosphorylation of Akt, which was inhibited by GS-1101 in a dose-dependent manner. Furthermore, selective inhibition of PI3Kd was able to attenuate the upregulation of Akt phosphorylation and the secreation of CCL17 and CCL22 associated with coculturing MCL cells and stromal cells. Treatment with GS-1101 also decreased S6 phosphorylation, but levels of phospho-4E-BP1 remained high, suggesting that mTOR signaling was not completely inhibited. However, the combination of GS-1101 and everolimus suppressed phosphorylation of both S6 and 4E-BP1, inhibiting MCL viability and enhancing apoptosis relative to treatment with either GS-1101 or everolimus alone. Conclusions: Our findings indicate that excessive PI3Kδ activity is characteristic in MCL and GS-1101-mediated PI3Kδ inhibition reduces MCL growth and survival. Combination therapy to address the molecular complexity associated with the convergence of the PI3Kδ-Akt and mTOR pathways may provide a novel treatment approach for MCL. Disclosures: Meadows: Gilead Sciences: Employment. Kashishian:Gilead Sciences: Employment. Johnson:Gilead Sciences: Employment. Ulrich:Gilead Sciences: Employment. Miller:Gilead Sciences: Employment. Lannutti:Gilead Sciences: Employment.

Description: Abstract 3926 Phosphatidylinositide 3-kinases (PI3Ks) are a family of lipid kinases that are involved in signaling events which control a diverse number of cellular processes. The class I kinases contain 4 isoforms designated p110α, β, δ, γ, and are activated by cell surface receptors. Aberrant regulation of the PI3K signaling pathway is frequently observed in human malignancies including those of hematological origin. CAL-101 is an oral p110δ-specific inhibitor which has shown preclinical and clinical activity in non-Hodgkin lymphoma (NHL) and chronic lymphocytic leukemia (CLL). This compound is a potent p110δ inhibitor (EC50 of 65 nM in a whole-blood assay) with 〉200-fold selectivity over the other class I PI3K isoforms and no activity against Class II and III PI3K family members or other PI3K-related proteins, including mTOR and DNA-PK. Prior in vitro NHL studies revealed that CAL-101 induces caspase-dependent apoptosis, and inhibits CD40L-, BAFF-, CXCL12- and CXCL13-derived survival signals in cellular models (Lannutti BJ, et al., Blood 2010). To investigate the potential role of p110δ in Hodgkin lymphoma (HL) we screened a number of HL cell lines for p110δ isoform expression and constitutive PI3K pathway activation. We report high levels of p110δ protein and activated Akt in 5 of 5 HL cell lines evaluated (L428, L540, L591, L1236, KM-H2). Inhibition of p110δ with CAL-101 treatment of cell lines resulted in a reduction of Akt phosphorylation and a decrease in cellular viability. Because previous studies have established the importance of signals from the microenvironment for the survival and proliferation of malignant cells as well as for their resistance to standard therapies, we investigated the effect of p110δ inhibition by CAL-101 in HL cell line-stroma cocultures. In this setting, CAL-101 overcame tumor cell growth induced by coculture of HL cells with bone marrow stromal cells. In addition, CAL-101 induced dose-dependent apoptosis of HL cells at 48 hours. Furthermore, stromal cell coculture resulted in increased CCL5, CCL17, and CCL22 levels; productions of these chemokines by HL cells cultured in the presence of stromal cells were reduced by CAL-101 in a dose-dependent manner. These results indicate that specific inhibition of p110δ may disrupt signals between HL cells and their microenvironment, thereby providing the preclinical rationale for clinical evaluation of CAL-101 as a novel therapeutic approach in patients with Hodgkin lymphoma. Disclosures: Meadows: Calistoga Pharmaceuticals: Employment. Kashishian:Calistoga Pharmaceuticals: Employment. Johnson:Calistoga Pharmaceuticals: Employment. Lannutti:Calistoga Pharmaceutical Inc.: Employment.

Description: Abstract 3876 Background: Chronic lymphocytic leukemia (CLL) is the most common leukemia in the Western World and remains incurable with standard therapies. Despite significant advances, novel treatments are essential to improve outcomes. A number of therapeutic agents have recently been developed including some that have shown significant activity and tolerability in clinical trials. Among these drugs, are small molecule kinase inhibitors that inhibit B-cell receptor (BCR)-mediated signaling pathways and disrupt essential CLL cell-microenvironment interactions. Specifically, when primary CLL cells are treated with GS-1101, a PI3 kinase delta-specific isoform (PI3Kδ) inhibitor, Bruton's tyrosine kinase (Btk) inhibitors or spleen tyrosine kinase (Syk) inhibitors, inhibition of signaling pathways downstream of the BCR, cell killing, and disruption of chemokine-mediated CLL cell migration are observed. Although significant clinical activity has been observed in patients treated with single agents that target these critical pathways, very little is known about the effects of inhibiting multiple nodes in the BCR pathway. Simultaneous inhibition of multiple pathways downstream of the BCR has the potential to result in a synergistic response that may overcome the resistance observed with single compound use. These considerations prompted us to assess the effects of PI3Kd and Syk inhibition and to determine if dual inhibition might enhance antitumor effects in CLL. Methods and Results: CLL patient samples were assessed for growth inhibition, chemokine release, and pathway activation status using a tetrazolium-based MTS assay, ELISA, and flow cytometry, respectively. PBMCs were isolated from primary patient samples using a Ficoll-Paque gradient. GS-1101, a PI3Kδ inhibitor, and GS-9973, a novel Syk inhibitor, were plated either alone in concentrations known to span their IC50 (.6nM to 10uM), or in combination using equimolar concentrations of each drug (.6nM to 10uM) in 96-well plates. 5×105 primary cells were cultured in triplicate in HS-5 stromal cell conditioned media and growth inhibition was determined after 72 hours. To explore potential additive, synergistic, or antagonistic interactions between GS-1101 and GS-9973, dose response and interaction indices were calculated using R (Lee et al. 2007). Given the importance of microenvironmental signals in the ability of malignant cells to survive, proliferate, and resist standard therapies, we investigated the effect of combining PI3Kd and Syk inhibitors for use on CLL cells in the presence of stroma-conditioned media. Immunoblotting and flow cytometry were used to measure the inhibitory effects of each drug alone or in combination on downstream targets known to be associated with BCR activation, including AKT, ERK, and S6 in Ramos cells, a Burkitt's lymphoma cell line, and in primary CLL cells. Of the 14 primary CLL samples treated with the single agent GS-9973, the median cell viability IC50 for all samples was 3.7 μM and an IC50 of ≤ 2μM was obtained in 7 samples (50%). Interaction indices were calculated from combination studies using GS-1101 and GS-9973 in 7 samples. At least one significantly synergistic concentration of the combination of GS-1101 and GS-9973 was found in five of the seven samples. The two remaining CLL samples showed additive responses when treated with the combination. Patient samples cultured in the presence of conditioned media resulted in increased CCL2, CCL3, and CCL4 levels. Production of these chemokines by CLL cells was reduced by both GS-1101 and GS-9973, alone and in combination. Furthermore, treatment with individual inhibitors decreased S6 and ERK, phosphorylation, an effect further enhanced by the combination of PI3Kd and Syk inhibition. Conclusion: Our findings indicate that both PI3Kd and Syk inhibition reduces CLL survival. Dual targeting can also induce synergistic growth inhibition and further disrupt chemokine signaling. Given the complexity of BCR signaling pathways, simultaneous targeting of multiple kinases has the potential to significantly increase clinical activity. Since inhibition of BCR mediated kinases has demonstrated good patient tolerability, combination therapy targeting both PI3Kd and Syk may provide a novel treatment approach, especially in patients with poor risk disease that may not respond optimally to single agents. Disclosures: Clarke: Gilead Sciences Inc: Employment. Meadows:Gilead sciences: Employment. Loriaux:Gilead Sciences Inc: Research Funding. Maciejewski:Gilead Sciences Inc: Employment. Di Paolo:Gilead Sciences Inc: Employment. Lannutti:Gilead Sciences Inc: Employment. Druker:Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Cylene: Consultancy, Membership on an entity's Board of Directors or advisory committees. Spurgeon:Gilead: Research Funding.

Description: Phosphatidylinositol-3-kinase p110δ serves as a central integration point for signaling from cell surface receptors known to promote malignant B-cell proliferation and survival. This provides a rationale for the development of small molecule inhibitors that selectively target p110δ as a treatment approach for patients with B-cell malignancies. We thus identified 5-fluoro-3-phenyl-2-[(S)-1-(9H-purin-6-ylamino)-propyl]-3H-quinazolin-4-one (CAL-101), a highly selective and potent p110δ small molecule inhibitor (half-maximal effective concentration [EC50] = 8nM). Using tumor cell lines and primary patient samples representing multiple B-cell malignancies, we have demonstrated that constitutive phosphatidylinositol-3-kinase pathway activation is p110δ-dependent. CAL-101 blocked constitutive phosphatidylinositol-3-kinase signaling, resulting in decreased phosphorylation of Akt and other downstream effectors, an increase in poly(ADP-ribose) polymerase and caspase cleavage and an induction of apoptosis. These effects have been observed across a broad range of immature and mature B-cell malignancies, thereby providing a rationale for the ongoing clinical evaluation of CAL-101.