Abstract: The role of membrane estrogen receptor-α (mERα) in rapid nongenomic responses to 17β-estradiol (E 2 ) was tested in sublines of GH3/B6 rat prolactinoma cells selected for high (GH3/B6/F10) and low (GH3/B6/D9) mERα expression. E 2 elicited rapid, concentration-dependent intracellular Ca 2+ concentration ([Ca 2+ ] i ) increases in the F10 subline. Lack of inhibition by thapsigargin depletion of intracellular Ca 2+ pools, together with abrogation of the response in Ca 2+ -free medium, suggested an extracellular source of Ca 2+ for this response. The participation of voltage-dependant channels in the E 2 -induced [Ca 2+ ] i increase was confirmed by the specific L-type Ca 2+ channel inhibitor nifedipine. For comparison, the D9 mERα-depleted subline was insensitive to steroid action via this signaling mechanism. [Ca 2+ ] i elevation was correlated with prolactin (PRL) release in the F10 cell line in as little as 3 min. E 2 caused a much higher PRL release than KCl treatment (which caused maximal Ca 2+ elevation), suggesting that secretion was also controlled by additional mechanisms. Participation of mERα in these effects was confirmed by the ability of E 2 -peroxidase (a cell-impermeable analog of E 2 ) to cause these responses, blockage of the responses with the ER antagonist ICI 182 780, and the inability of the E 2 stereoisomer 17α-E 2 to elicit a response. Thus rapid exocytosis of PRL is regulated in these cells by mERα signaling to specific Ca 2+ channels utilizing extracellular Ca 2+ sources and additional signaling mechanisms.

Abstract: Background: Current therapies for multiple myeloma (MM) delay disease progression and prolong survival but most patients (pts) eventually relapse or become refractory (RR). Daratumumab (D), an anti-CD38 antibody (Ab), plus bortezomib (V), a proteasome inhibitor (PI) and dexamethasone (d), is approved for the treatment of MM in pts who have received ≥1 prior line of therapy. Venetoclax (Ven), a potent and selective oral BCL-2 inhibitor, demonstrated anti-myeloma activity in pts with t(11;14) RRMM. This 3-part Phase 1/2 study is investigating the combination therapy VenDd +/- V in pts with RRMM. Treatment of pts with t(11;14) RRMM using VenDd (part 1) and pts with RRMM (irrespective of t(11;14) status; part 2) with VenDVd demonstrated a tolerable safety profile and an overall response rate (ORR) of 95.8% and 91.7%, respectively (Bahlis N et al. J Clin Oncol 2021). Part 3 further evaluates the safety and efficacy of VenDd vs DVd in pts with t(11;14) RRMM, with preliminary results presented here. Methods: Part 3 of this Phase 1/2, multicenter, dose-escalation and expansion study (NCT03314181) evaluates the safety and efficacy of VenDd vs DVd in pts with t(11;14) RRMM. The study was expanded to further interpret the pt safety profile in light of the increased incidence of infections in pts in the Ven arm of the BELLINI study (Kumar SK et al. Lancet Oncol 2020). Pts were randomized 4:2:5 to receive VenDd at 400 (Ven400Dd) or 800 mg (Ven800Dd), or DVd. Randomization was not stratified due to small sample size. Eligible pts must have received ≥1 prior line of therapy, including an immunomodulatory agent (IMiD), and be non-refractory to PIs or anti-CD38 Ab. This interim analysis was conducted to evaluate the safety profile of pts in part 3 only. No statistical comparisons were conducted for safety or efficacy. Treatments in Part 3 were as follows: VenDd cycles (C) were 28-day: daily, oral Ven (400 mg or 800 mg) + D (1800 mg SC [Cycle, C1, 2: Days 1, 8, 15, 22; C3-6: Days 1, 15; C7+: Day 1]) + d (40 mg total weekly); DVd C1 - 8 were 21-day, C9+ were 28-day: D (1800 mg SC [C1 - 3: Days 1, 8, 15; C4 - 8: Day 1; C9+: Day 1] ) + V (1.3mg/m 2 [C1 - 8: Days 1, 4, 8 and 11]) + d (20 mg [C1 - 3: Days 1, 2, 4, 5, 8, 9, 11, 12 ,15; C4 - 8: Days 1, 2, 4, 5, 8, 9, 11,12; C9+: Day 1] ). Results: As of 10 May 2021, 11, 7 and 16 pts were enrolled in the Ven400Dd, Ven800Dd and DVd arms, respectively. The median age (range) was Ven400Dd: 58.0 (42 - 75); Ven800Dd: 57 (53 - 82); and DVd: 68.5 (51 - 77). Median prior lines of therapy (range) were 1.0 (1 - 6) in Ven400Dd; 1.0 (1 - 3) in Ven800Dd; and 2.0 (1 - 3) in DVd. Pts with ISS I%/II%/III% disease were Ven400Dd: 54.5/9.1/0; Ven800Dd: 57.1/14.3/0; DVd: 25.0/25.0/31.3. All pts in the Ven400Dd and Ven800Dd arms had an ECOG performance status of ≤1. In the DVd arm, 87.5% and 12.5% of pts had a ECOG performance status of ≤1 and 2, respectively. Prior PI%/IMiD%/anti-CD38 Ab% exposure were Ven400Dd: 100/90.9/0; Ven800Dd: 100/100/0; DVd: 93.8/100/0. The most common adverse events (AEs) occurring in ≥5% of pts in ≥2 treatment groups included insomnia, fatigue, diarrhea, and nausea (Table). Grade 3/4 AEs (≥5% of pts in ≥2 treatment groups) were mainly hematologic toxicities (Table). There were no grade 3/4 infections occurring in ≥2 treatment groups. Serious AEs were observed in a total of 6 pts. In the Ven400Dd arm, 1 pt had a femur fracture and 1 pt had non-cardiac chest pain. In the Ven800Dd arm, 1 pt had febrile neutropenia and 1 pt had tonsil cancer. In the DVd arm, 1 pt had pyrexia and upper respiratory tract infection, and a second pt had hyperglycemia, autonomic neuropathy, distributive shock, disseminated cryptococcosis, and cytomegalovirus infection. No deaths were reported in part 3 of the study. The median treatment duration based on D exposure at the time of data cut was 6.5, 5.6, and 3.9 months for the Ven400Dd, Ven800Dd, and DVd arms, respectively. The preliminary ORR was 72.7%, 100%, and 62.5% for the Ven400Dd, Ven800Dd, and DVd arms. The preliminary rate of very good partial response or better (≥VGPR) was 72.7%, 100%, and 31.3% for the Ven400Dd, Ven800Dd, and DVd arms. Follow-up is still immature, and responses may deepen with time. Conclusion: The preliminary results from part 3 of this novel randomized, phase 2 study of t(11;14)-selected RRMM pts treated with VenDd vs DVd demonstrate a tolerable safety profile. Updated analyses, including response rates with longer follow-up and minimal residual disease status, will be included at presentation. Figure 1 Figure 1. Disclosures Kaufman: Roche/Genetech, Tecnopharma: Consultancy, Honoraria; Heidelberg Pharma: Research Funding; Sutro, Takeda: Research Funding; Fortis Therapeutics: Research Funding; BMS: Consultancy, Research Funding; Amgen: Research Funding; Novartis: Research Funding; Incyte, TG Therapeutics: Membership on an entity's Board of Directors or advisory committees; Tecnofarma SAS, AbbVie: Honoraria; Incyte, celgene: Consultancy; Janssen: Honoraria; Genentech, AbbVie, Janssen: Consultancy, Research Funding. Quach: Janssen/Cilag: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; GlaxoSmithKline: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; CSL: Consultancy, Membership on an entity's Board of Directors or advisory committees; Karyopharm: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Antengene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Baz: GlaxoSmithKline: Consultancy, Honoraria; Oncopeptides: Consultancy; Merck: Research Funding; BMS, sanofi, Karyopharm, Janssen, AbbVie: Consultancy, Research Funding. Harrison: Janssen Cilag: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Roche/Genentech: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Eusa: Consultancy, Honoraria, Speakers Bureau; Terumo BCT: Consultancy, Honoraria; Celgene/ Juno/ BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Sanofi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Haemalogix: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; GSK: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Abbvie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Plesner: Janssen, Celgene, Takeda, Oncopeptides, AbbVie: Consultancy, Research Funding; Genmab, Genentech, Roche: Research Funding. Moreau: Oncopeptides: Honoraria; Amgen: Honoraria; Celgene BMS: Honoraria; Sanofi: Honoraria; Janssen: Honoraria; Abbvie: Honoraria. Gibbs: AbbVie: Consultancy; Janssen, Celgene, Amgen, Takeda, BMS and Pfizer: Consultancy, Honoraria. Sehgal: AbbVie: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company. Kang: AbbVie: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company. Ross: AbbVie: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company. Lash-Fleming: AbbVie: Current Employment, Current equity holder in publicly-traded company. Luo: AbbVie: Current Employment, Current equity holder in publicly-traded company. Bahlis: Takeda: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; Karyopharm: Consultancy, Honoraria; Genentech: Consultancy; BMS/Celgene: Consultancy, Honoraria; GlaxoSmithKline: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria. OffLabel Disclosure: Venetoclax is a potent and selective oral BCL-2 inhibitor being investigated in the treatment of relapsed/refractory multiple myeloma.

Abstract: Background: Venetoclax (VEN) is a selective, potent, orally bioavailable BCL-2 inhibitor FDA-approved for patients with del(17p) chronic lymphocytic leukemia (CLL) and who have received ≥1 prior therapy. Based on preclinical evidence of synergy, VEN plus rituximab is being assessed in an ongoing Phase 1b study. Methods: Patients with relapsed/refractory (R/R) CLL received daily VEN with stepwise ramp-up over 3-4 weeks to reach daily doses of 200-600mg. After 1 week at the target dose, monthly rituximab was added for 6 doses. Responses and progression were assessed by iwCLL criteria with CT scan and bone marrow biopsy. Bone marrow assessments were done at screening, completion of combination therapy (month 7), and 2 months after clinical/radiologic criteria of iwCLL response were met. Minimal residual disease (MRD) was assessed in peripheral blood and marrow aspirates using ≥4 color flow cytometry (min sensitivity: 0.01%). Data cutoff was 04March2016, with analysis focusing on updated safety of cytopenias experienced on the course of treatment. Results: Forty-ninepatients enrolled (48 CLL/1 SLL). Patients had received a median of 2 prior therapies (range: 1-5) and disease in 25 (51%) was considered refractory to the most recent therapy. Median time on study was 28 ( 〈 1-42) months, with 31 patients active on study. Eighteen patients discontinued: 11 due to disease progression, 3 due to toxicity (peripheral neuropathy [1], MDS [1] , and death due to TLS [1]), 3 withdrew consent, and 1 was lost to follow up. Across all doses, the most common AEs of any grade were diarrhea (57%), neutropenia (55%), upper respiratory tract infection (55%), and nausea (51%). Peripheral blood cytopenias were the most common Grade 3/4 AEs (neutropenia [53%] , thrombocytopenia [16%], anemia [14%] , febrile neutropenia [12%], and leukopenia [12%] ). Twenty-seven (55%) patients had a history of neutropenia, of whom 6 were receiving G-CSF support prior to starting VEN. Overall, in the first month of therapy, 15 (31%) experienced an AE of neutropenia (any grade). Thereafter, the rate of new AEs of neutropenia decreased over time. While there was individual patient variability, mean ANC was stable over time. Overall, 26 (53%) patients had Grade 3/4 neutropenia. Neutropenia was generally well tolerated and managed by G-CSF support in 24 patients, in addition to ≥1 dose modification in 11 of the 24 patients. Of 8 (16%) patients who experienced grade 3 infections, 2 were while neutropenic. There were no grade 4 infections. Among the 11 (22%) patients who developed any-grade thrombocytopenia, none occurred within 2 weeks of a reported bleeding-related AE. One patient had thrombocytopenia overlapping with disease progression on therapy. Objective response rate for all patients was 86% (n=42), with 51% (n=25) who had complete response (CR/CRi; 12 achieved CR/CRi by month 7). At the completion of combination therapy (month 7), 39 patients had evaluable bone marrow assessments. Thirty (77%) had no histologic evidence of CLL in the bone marrow and 22 patients (56%) had attained bone marrow MRD-negativity. In longer follow up at any point during treatment for all 49 patients, 37 (75%) patients achieved complete marrow clearance and 28 (57%) achieved marrow MRD-negativity. Conclusions: Transient manageable neutropenia was the most common AE, with first onset usually seen within the first month of treatment and the onset of new neutropenia AEs decreased over time. No patients discontinued the study due to cytopenias. Patients were able to continue on study and high rates of response to treatment were observed. VEN given with rituximab achieved rapid and profound reductions in disease burden in peripheral blood and bone marrow. 77% of evaluable patients achieved morphologic clearance by month 7, and 57% were MRD-negative at any point on study. Figure 1 Figure 1. Disclosures Brander: TG Therapeutics: Research Funding; Gilead: Honoraria. Roberts:AbbVie: Research Funding; Servier: Research Funding; Janssen: Research Funding; Genentech: Research Funding; Genentech: Patents & Royalties: Employee of Walter and Eliza Hall Institute of Medical Research which receives milestone payments related to venetoclax. Ma:Pharmacyclics, LLC, an AbbVie Company: Consultancy, Honoraria, Research Funding, Speakers Bureau; Janssen: Consultancy, Honoraria, Speakers Bureau; Gilead: Consultancy, Honoraria, Research Funding, Speakers Bureau; Genentech: Consultancy, Honoraria, Speakers Bureau; Novartis: Research Funding; Xeme: Research Funding; AbbVie: Research Funding. Lash:AbbVie: Employment. Verdugo:AbbVie: Employment, Other: may own stock. Zhu:AbbVie Inc.: Employment, Other: may own stock. Kim:AbbVie: Employment. Seymour:Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Gilead: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Genentech: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie Inc.: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau.

Abstract: Mechanisms of pain‐related plasticity in the amygdala, a key player in emotionality, were studied at the cellular and molecular levels in a model of arthritic pain. The influence of the arthritis pain state induced in vivo on synaptic transmission and N ‐methyl‐ d ‐aspartate (NMDA) receptor function was examined in vitro using whole‐cell voltage‐clamp recordings of neurones in the latero‐capsular part of the central nucleus of the amygdala (CeA), which is now defined as the ‘nociceptive amygdala’. Synaptic transmission was evoked by electrical stimulation of afferents from the pontine parabrachial area (part of the spino‐parabrachio‐amygdaloid pain pathway) in brain slices from control rats and from arthritic rats. This study shows that pain‐related synaptic plasticity is accompanied by protein kinase A (PKA)‐mediated enhanced NMDA‐receptor function and increased phosphorylation of NMDA‐receptor 1 (NR1) subunits. Synaptic plasticity in the arthritis pain model, but not normal synaptic transmission in control neurones, was inhibited by a selective NMDA receptor antagonist. Accordingly, an NMDA receptor‐mediated synaptic component was recorded in neurones from arthritic animals, but not in control neurones, and was blocked by inhibition of PKA but not protein kinase C (PKC). Exogenous NMDA evoked a larger inward current in neurones from arthritic animals than in control neurones, indicating a postsynaptic effect. Paired‐pulse facilitation, a measure of presynaptic mechanisms, was not affected by an NMDA‐receptor antagonist. Increased levels of phosphorylated NR1 protein, but not of total NR1, were measured in the CeA of arthritic rats compared to controls. Our results suggest that pain‐related synaptic plasticity in the amygdala involves a critical switch of postsynaptic NMDA receptor function through PKA‐dependent NR1 phosphorylation.

Abstract: Cortical information storage requires combined changes in connectivity and synaptic strength between neurons, but the signaling mechanisms underlying this two-step wiring plasticity are unknown. Because acute 17β-estradiol (E2) modulates cortical memory, we examined its effects on spine morphogenesis, AMPA receptor trafficking, and GTPase signaling in cortical neurons. Acute E2 application resulted in a rapid, transient increase in spine density, accompanied by temporary formation of silent synapses through reduced surface GluR1. These rapid effects of E2 were dependent on a Rap/AF-6/ERK1/2 pathway. Intriguingly, NMDA receptor (NMDAR) activation after E2 treatment potentiated silent synapses and elevated spine density for as long as 24 h. Hence, we show that E2 transiently increases neuronal connectivity by inducing dynamic nascent spines that “sample” the surrounding neuropil and that subsequent NMDAR activity is sufficient to stabilize or “hold” E2-mediated effects. This work describes a form of two-step wiring plasticity relevant for cortical memory and identifies targets that may facilitate recovery from brain injuries.