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Background: Eflapegrastim (Rolontis®, Efla) represents the first novel, long-acting granulocyte-colony stimulating factor (G-CSF) to be introduced in more than 15 years. Efla consists of a recombinant human G-CSF analog conjugated to a human IgG4 Fc fragment via a polyethylene glycol linker. Preclinical, clinical, and pharmacodynamic/pharmacokinetic data have shown increased potency for Efla versus pegfilgrastim (Peg). Both independent, randomized Phase 3 studies comparing Efla and Peg for prophylaxis of chemotherapy-induced neutropenia in patients with early-stage breast cancer (ESBC) met the primary endpoint of non-inferiority in duration of severe neutropenia (SN; ANC75kg. The safety profiles, including AEs and discontinuations, for Efla and Peg were comparable, and >99% of all patients received full dose of TC on schedule. The majority (67%) of patients with SN experienced a 1 day duration, occurring between Days 7 and 8 after TC. Mean duration of SN for Efla was statistically lower than for Peg (0.24 vs. 0.36 days; p=0.029). The above statistical significance was maintained for Efla after adjusting for demographic and baseline characteristics, namely age, weight, enrolling geographical region, and treatment setting in a multivariate model. Similarly, the incidence of SN for Efla was statistically lower than Peg in Cycle 1 (17.5% vs 24%; relative risk reduction [RRR]=27%; p=0.043). Univariate analysis of the incidence of SN showed a significant risk reduction in favor of Efla (8.6% vs 14.1%; p=0.034) for patients weighing >75kg (p=0.034). Multivariate analysis of SN showed significant odds ratio of SN for age ≥65 years and baseline ANC >6 × 109/L in favor of Efla (OR=0.42 and 0.39, respectively). The incidence of SN in Cycles 2-4 was comparable between treatment groups. Also, the incidence of febrile neutropenia and neutropenic complications was similar with 75kg. Eflapegrastim is a novel, long-acting and potent recombinant human G-CSF which may provide an attractive option in supporting patients at risk for SN-related complications. Citation Format: Lee S Schwartzberg, Gajanan Bhat, Alvaro Restrepo, Osama Hlalah, Inderjit Mehmi, Yong Wha Moon, Seungjae Baek, Shanta Chawla, Francois Lebel, Patrick Wayne Cobb. Pooled efficacy analysis from two phase 3 studies in patients receiving eflapegrastim, a novel, long-acting granulocyte-colony stimulating factor, following TC for early-stage breast cancer [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS9-59.

Background Eflapegrastim, a novel, long‐acting recombinant human granulocyte‐colony stimulating factor (rhG‐CSF), consists of a rhG‐CSF analog conjugated to a human IgG4 Fc fragment via a short polyethylene glycol linker. Preclinical and phase I and II pharmacodynamic and pharmacokinetic data showed increased potency for neutrophil counts for eflapegrastim versus pegfilgrastim. This open‐label phase III trial compared the efficacy and safety of eflapegrastim with pegfilgrastim for reducing the risk of chemotherapy‐induced neutropenia. Materials and Methods Patients with early‐stage breast cancer were randomized 1:1 to fixed‐dose eflapegrastim 13.2 mg (3.6 mg G‐CSF) or standard pegfilgrastim (6 mg G‐CSF) following standard docetaxel plus cyclophosphamide chemotherapy for 4 cycles. The primary objective was to demonstrate the noninferiority of eflapegrastim compared with pegfilgrastim in mean duration of severe neutropenia (DSN; grade 4) in cycle 1. Results Eligible patients were randomized 1:1 to study arms (eflapegrastim, n = 196; pegfilgrastim, n = 210). The incidence of cycle 1 severe neutropenia was 16% (n = 31) for eflapegrastim versus 24% (n = 51) for pegfilgrastim, reducing the relative risk by 35% (p = .034). The difference in mean cycle 1 DSN (−0.148 day) met the primary endpoint of noninferiority (p < .0001) and also showed statistical superiority for eflapegrastim (p = .013). Noninferiority was maintained for the duration of treatment (all cycles, p < .0001), and secondary efficacy endpoints and safety results were also comparable for study arms. Conclusion These results demonstrate noninferiority and comparable safety for eflapegrastim at a lower G‐CSF dose versus pegfilgrastim. The potential for increased potency of eflapegrastim to deliver improved clinical benefit warrants further clinical study in patients at higher risk for CIN. Implications for Practice Chemotherapy‐induced neutropenia (CIN) remains a significant clinical dilemma for oncology patients who are striving to complete their prescribed chemotherapy regimen. In a randomized, phase III trial comparing eflapegrastim to pegfilgrastim in the prevention of CIN, the efficacy of eflapegrastim was noninferior to pegfilgrastim and had comparable safety. Nevertheless, the risk of CIN remains a great concern for patients undergoing chemotherapy, as the condition frequently results in chemotherapy delays, dose reductions, and treatment discontinuations., Myelosuppression, particularly neutropenia, has presented a major challenge in cancer treatment since the introduction of cytotoxic chemotherapy. This article reports the results of a phase III trial that compared the efficacy and safety of eflapegrastim with pegfilgrastim for reducing the risk of chemotherapy‐induced neutropenia.

Background: Eflapegrastim (E) represents the first myeloid growth factor innovation in more than 15 years. A novel, long-acting recombinant human granulocyte-colony stimulating factor (rhG-CSF), E consists of a rhG-CSF analog conjugated to a human IgG4 Fc fragment via a polyethylene glycol linker. Preclinical and Phase I and II pharmacodynamic and pharmacokinetic data showed increased potency for E versus pegfilgrastim (P). Two independent randomized Phase III trials comparing fixed dose E and P (E 3.6 mg G-CSF and P 6.0 mg G-CSF) for the management of chemotherapy-induced neutropenia (CIN) have recently been completed. Both trials met the primary endpoint of non-inferiority for E vs P in Cycle 1 duration of severe neutropenia (P Patients and Methods: Patients with early-stage breast cancer (ESBC) who were candidates for adjuvant or neoadjuvant chemotherapy were randomized 1:1 in two open-label Phase III trials to E 13.2 mg (3.6 mg G-CSF) or standard P (6 mg G-CSF) administered on Day 2 following TC (docetaxel/cyclophosphamide) chemotherapy on Day 1 of each of 4 cycles. Blood for CBC and serum chemistry was collected in every cycle. Safety assessments began with the first dose of TC and continued for one year after the last dose of study drug. AEs and laboratory values were graded according to NCI CTCAE version 4.03. Immunogenicity was assessed from blood samples collected on Day 1 of each cycle, at the end-of-treatment visit, and at 6- and 12-month follow-up visits. Results: A total of 660 patients who received at least one dose of eflapegrastim (n=334) or pegfilgrastim (n=326) were included in this integrated safety analysis. The two treatment groups were well balanced for demographics and baseline disease characteristics. The mean age was 59y, ~40% were aged >65y, ~54% weighed >75kg, and ~80% were treated in the adjuvant setting. Median relative dose intensity for T and C was >99% for both groups. A similar percentage of patients in both treatment groups discontinued treatment due to AEs (4% E vs 6% P), with 2% in each group discontinuing due to AEs related to E or P. Serious AEs were similar in both groups (15% each). Incidence of AEs irrespective of causality were also similar between groups (74% E vs. 72% P). No notable differences between groups were observed in the types of study-drug-related AEs. The majority of study-drug-related AEs occurred with an incidence ≤10% for both E and P. As expected with myeloid growth factors, study-drug-related AEs occurring in >10% in either group were bone pain (E vs P: 33% vs 34%), arthralgia (15% vs 10%), back pain (14% vs 9%), myalgia (14% vs 9%), and headache (11% vs 8%). Incidence of febrile neutropenia and neutropenic complications were similar and less than 5% in each treatment group. No leukocytosis, splenic rupture, or anaphylaxis was reported in any patient receiving E or P. The overall incidence of immunogenicity was similar in both groups and there was no demonstrable impact on clinical safety or efficacy. Conclusions: Two large, randomized Phase III trials (Total n=660) of E vs P administered once-per-cycle showed E at a lower G-CSF dose to be safe and effective for the prophylaxis of CIN in patients with ESBC receiving TC chemotherapy. E is a novel long-acting rhG-CSF with increased potency and similar toxicity to P and may provide an attractive alternative for growth factor support of patients at high risk for CIN-related complications. Citation Format: Lee S Schwartzberg, Gajanan Bhat, Julio Peguero, Richy Agajanian, Jayaram Bharadwaj, Alvaro Restrepo, Osama Hlalah, Inderjit Mehmi, Shanta Chawla, Francois Lebel, Patrick W Cobb. Eflapegrastim, a novel long-acting granulocyte-colony stimulating factor: Integrated safety results in patients with early-stage breast cancer treated with TC chemotherapy [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P2-14-12.

Background: The ABC trials and the WSG Plan B trial demonstrated that certain early stage breast cancer patients can be appropriately treated with docetaxel and cyclophosphamide (TC) rather than doxorubicin and cyclophosphamide (AC); however both of these trials utilized six cycles of TC as the comparator. TC is given for 4 cycles based on the US Oncology 9735 trial. To our knowledge, there is no available data that has compared adverse events or outcomes of four versus six cycles of TC. Methods: This was a single-center, retrospective study evaluating recurrence-free survival, overall survival, and occurrence of toxicities between four and six cycles of docetaxel and cyclophosphamide adjuvant chemotherapy for early stage breast cancer at West Virginia University Cancer Institute. Results: 112 patients were included; 81 received four cycles and 31 received six. Recurrence free survival rates of patients that received four and six cycles of chemotherapy were 94% and 90%, respectively, at three years (P=0.68). Overall survival at three years was 99% versus 97% (P=0.49), respectively. The incidence of peripheral neuropathy requiring treatment was 42% in patients that received six cycles and 19% in patients that received four (P=0.015). Conclusion: The addition of two cycles of docetaxel and cyclophosphamide adjuvant chemotherapy for early stage breast cancer provides no additional recurrence-free or overall survival benefit but does increase the incidence of peripheral neuropathy requiring treatment. Table 1.Baseline Characteristics 4 Cycles (n=81)6 Cycles (n=31)P-valueAge, median years (range)57 (37-75)48 (32-71)0.001Receptor status, n (%) 0.63HR positive, HER2 negative61 (75)25 (81) Triple negative20 (25)6 (19) Cancer stage, n (%) 0.031I39 (48)9 (29) II39 (48)17 (55) III3 (4)5 (16) Lymph node involvement, n (%)24 (30)20 (65)0.001Ki67 (%), median (range)23 (2-99)23.5 (4-99)0.99Tumor grade, n (%) 0.07212 (2)4 (13) 242 (52)12 (39) 337 (46)15 (48) Surgery, n (%) Breast conservation35 (43)14 (45)0.99Unilateral mastectomy32 (40)9 (29)0.38Bilateral mastectomy14 (17)8 (26)0.43 Table 2.Adverse Events 4 Cycles (n=81)6 Cycles (n=31)P-valueDose reductions, n (%)17 (21)8 (26)0.62Dose delay, n (%)3 (4)1 (3)0.99Inpatient admissions, n (%)22 (27)8 (26)0.99Treatment for peripheral neuropathy, n (%)15 (19)13 (42)0.015 Citation Format: Mehmi I, Wight CE, Hill JL. Comparison of overall and recurrence-free survival between four and six cycles of adjuvant docetaxel and cyclophosphamide in early stage breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P1-13-10.

Background Therapeutic options are limited for patients with advanced melanoma that is refractory to PD-1 blockade. This study was performed in this patient population to assess the safety and antitumor activity of CMP-001, a CpG-A TLR9 agonist packaged within a virus-like particle. Methods Patients were eligible for this 2-part, open-label, multicenter, phase 1b study if they had metastatic/unresectable melanoma and stable disease after =12 weeks or progressive disease (PD) on/after anti-PD-1 therapy. Part 1 evaluated CMP-001 plus pembrolizumab dose-escalation and dose-expansion. Part 2 evaluated CMP-001 monotherapy. Accessible lesion(s) were injected intratumorally with CMP-001, at a polysorbate 20 (PS20) concentration of either 0.01% or 0.00167%. The Part 1 primary objective was to identify the recommended phase 2 dose (RP2D) and schedule of CMP-001 plus pembrolizumab, while the Part 2 primary objective was to assess the safety of CMP-001 monotherapy. Secondary objectives for both parts were a preliminary assessment of antitumor activity of CMP-001 plus pembrolizumab and CMP-001 monotherapy, and the overall safety profile and pharmacodynamics of the combination. Results In Part 1 (N=159) and Part 2 (N=40), 93.1% and 80.0% of patients had PD as their last response to prior anti–PD-1 therapy, respectively. The most common treatment-related adverse events (TRAEs; >25%) were flu-like symptoms (Parts 1 and 2) and injection-site reactions (Part 1). Grade 3/4 TRAEs were reported in 36.5% (Part 1) and 22.5% (Part 2) of patients, the most common being hypotension (Part 1: 6.9%; Part 2: 5.0%). No Grade 5 TRAEs were observed. In Part 1, the best objective response rate (ORR; RECIST v1.1) in patients treated with pembrolizumab and CMP-001 (PS20 0.01%) was 23.5% (23/98), while CMP-001 PS20 (0.00167%) resulted in a lower ORR of 11.5% (7/61). Seven additional patients had a delayed response after initial PD (table 1). The median duration of response was >1 year. In the 37 RECIST v1.1 and post-progression responders, the mean regression in injected and noninjected target lesions was 54.7% and 52.7%, respectively. In Part 2, the best ORR with CMP-001 monotherapy was 17.5% (7/40 patients); the response duration was shorter than in Part 1. Intratumoral CMP-001 PS20 0.01% 10 mg was selected as the RP2D. Conclusions Intratumoral CMP-001 was well-tolerated and provided both local and distant responses in patients with advanced melanoma with disease progression on prior PD-1 blockade. CMP-001 monotherapy induced systemic tumor regression in some patients, but duration of response was substantially increased by the addition of pembrolizumab. Acknowledgements This work was supported by Checkmate Pharmaceuticals. Medical writing assistance was provided by Cindy Rigby, PhD, of ApotheCom (San Francisco, CA) and was funded by Checkmate Pharmaceuticals. Trial Registration NCT02680184 Ethics Approval This study was approved by the WCG-WIRB, WIRB approval tracking number 20152597. Consent N/A References N/A

Background In the treatment-naive setting, PD-1 blockade is associated with greater response in T cell-inflamed vs non-T cell-inflamed tumors. CMP-001 is a CpG-A oligonucleotide TLR9 agonist in a virus-like particle that is hypothesized to activate tumor-associated plasmacytoid dendritic cells (pDCs) to secrete type I interferons. Through this activity, CMP-001 may convert the tumor microenvironment to a Th1-like chemokine milieu (eg, increased CXCL10) and induce an antitumor CD8+ T-cell response. We have recently reported that intratumoral injection of CMP-001 + IV pembrolizumab (pembro) had an acceptable safety profile and can reverse PD-1 blockade resistance in patients (pts) with melanoma (Milhem et al, SITC 2019). Regression was observed in injected and uninjected lesions. Herein we report pharmacodynamic and translational data. Methods This 2-part, open-label, multicenter, phase 1b study (NCT02680184) enrolled pts with metastatic/unresectable melanoma and stable disease (SD) or progressive disease (PD) on/after anti−PD-1 therapy. In part 1 (3+3 dose-escalation and dose-expansion), pts received CMP-001 + pembro. In part 2, pts received CMP-001 monotherapy. Determination of safety and clinical activity were the study's main objectives. Prespecified pharmacodynamic and translational studies evaluated serum chemokines and evaluated tumor biopsies using RNA and/or whole exome sequencing and immunohistochemistry for PD-L1 (reported as H-score), CD8, and CD303 (pDC marker). Results As of September 30, 2020, 159 pts (part 1) and 40 pts (part 2) have been treated. A greater median fold increase of serum CXCL10 (a marker of innate immunity, n=40) was observed in responders (R) to CMP-001 + pembro (18.8x) vs nonresponders (NR) after treatment (9.9x in SD; 6.15x in PD; differences were not statistically significant). Preliminary analyses showed that interferon gene expression distinguished R vs NR. Tumor biopsy analyses (part 1, n=139; part 2, n=34) showed that pts with high PD-L1, high CD8+ T cells, or inflamed transcriptional signatures at baseline were less likely to respond to CMP-001 + pembro vs pts without inflammation markers at baseline. Baseline mean PD-L1 expression (H-score) was 8.1 in R (n=10) vs 21.8 in NR (n=49). Posttreatment biopsies generally showed increased PD-L1, CD8+ T cells, and inflamed transcriptional signatures in R vs NR. Neither tumor mutational burden nor baseline pDC density distinguished R vs NR. Conclusions In pts with anti-PD-1 refractory melanoma, intratumoral CMP-001 ± pembro appears to disproportionately induce antitumor responses in noninflamed tumors. Clinical response to CMP-001 ± pembro was associated with induction of markers of both innate and adaptive antitumor immunity. Citation Format: Jason John Luke, Riyue Bao, John M. Kirkwood, Yousef Zakharia, Diwakar Davar, Elizabeth Buchbinder, Theresa Medina, Adil Daud, Antoni Ribas, Jiaxin Niu, Geoffrey Gibney, Kim Margolin, Anthony J. Olszanski, Inderjit Mehmi, Takami Sato, Montaser Shaheen, Aaron Morris, Dmitri Bobilev, Katie Campbell, George Weiner, James E. Wooldridge, Arthur M. Krieg, Mohammed Milhem. CMP-001 demonstrates improved response in noninflamed anti-PD-1 refractory melanoma and response is associated with serum CXCL10 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT032.

TPS5600 Background: KIF18A is a mitotic kinesin motor protein that regulates chromosome positioning during cell division and is overexpressed in a subset of human cancers. TP53 mutant unstable aneuploid cancer cells with chromosomal instability (CIN) features are dependent on KIF18A motor activity to prevent lethal multipolar cell division. Preclinical data demonstrate that treatment with AMG 650; an oral, first in class, selective small molecule inhibitor of KIF18A may be safe and tolerable. We are conducting a first-in-human phase 1 study with AMG 650 in adult subjects with locally advanced or metastatic solid tumors with TP53MUT, triple negative breast cancer (TNBC), high grade serous ovarian cancer (HGSOC) or serous like endometrial cancers and other solid tumors. Methods: In this phase 1, multicentric, dose escalation and dose expansion study we evaluate the safety and tolerability of AMG 650 monotherapy in patients with advanced/metastatic solid tumors (NCT04293094). The main objective is to determine the maximum tolerated dose (MTD) and/or recommended phase 2 dose (RP2D) based on emerging safety, efficacy, and pharmacodynamics (PD) data prior to reaching the MTD. Key inclusion criteria include the presence of measurable disease and diagnosis of advanced/metastatic triple negative breast cancer (TNBC), high-grade serous ovarian cancer (HGSOC), serous-like endometrial cancer or other solid tumors with documented TP53 mutations. In the dose expansion phase, participants with locally advanced or metastatic TNBC or HGSOC will be treated with the preliminary RP2D identified from the dose exploration part of the study. Primary endpoints include the incidence of Dose Limiting Toxicities (DLTs),Treatment-Emergent Adverse Events (TEAEs), Serious Adverse Events (SAEs), Treatment-related Adverse Events and the evaluation of the number of participants who experience a clinically significant change from baseline in vital signs, electrocardiogram and laboratory tests parameters. Secondary endpoints include Objective Response Rate, Duration of Response, Progression-free Survival, Clinical Benefit Rate, Time to Response, Time to Progression, Overall Survival (OS), Maximum Plasma Concentration (Cmax) of AMG 650, Time to Maximum Plasma Concentration (Tmax) of AMG 650 as well as Area Under the Plasma Concentration-time Curve (AUC) Over the Dosing Interval for AMG 650. Continuous monitoring of toxicity is conducted. The study began enrolling pts in March 2020 and is ongoing. For more information, please contact Amgen Medical Information: medinfo@amgen.com Clinical trial information: NCT04293094.

140 Background: Reduced-dose nivolumab in combination with standard-dose ipilimumab was shown to improve overall response rate (58%), progression free survival (11.5 months), and overall survival compared to ipilimumab alone in Checkmate 067. Improved outcomes were accompanied by an increased risk of toxicities with a 59% rate of grade 3-4 toxicities for the combination. Keynote 029 evaluated standard-dose pembrolizumab with reduced-dose ipilimumab with a slightly lower risk of grade 3-4 toxicities at 45%. Here we report outcomes for standard-dose nivolumab combined with reduced-dose ipilimumab in metastatic melanoma. Methods: We conducted a retrospective review of metastatic melanoma patients that received standard-dose nivolumab with reduced-dose ipilimumab at West Virginia University Cancer Institute. Response to therapy and incidence of immune-related adverse events were assessed. Results: From February 2017 to October 2018, 11 patients received the alternative dosing regimen. Only 1 (9%) patient experienced a grade 3-4 toxicity, and 3 (27%) additional patients experienced a grade 1-2 toxicity. The overall response rate was 54%. Conclusions: Standard-dose nivolumab in combination with reduced-dose ipilimumab is better tolerated than reduced-dose nivolumab with standard-dose ipilimumab without a loss in overall response rate.

Background: CMP-001 comprises a CpG-A oligodeoxynucleotide packaged within a virus-like particle. It is designed to activate tumor-associated plasmacytoid dendritic cells via TLR9 inducing an interferon-rich tumor microenvironment and anti-tumor CD8+ T cell responses. Materials and Methods: CMP-001-001 is an ongoing phase Ib trial evaluating intratumoral (IT) CMP-001 in combination with pembrolizumab (administered per label) in subjects with advanced melanoma resistant (either did not respond or progressed) on prior anti-PD-1 monotherapy or in combination. During dose escalation, subjects were enrolled to cohorts of ≥ 3 subjects at CMP-001 doses of 1, 3, 5, 7.5, and 10 mg in two dosing schedules (weekly for 7w, followed by q3w; or weekly for 2w, followed by q3w). CMP-001 was administered IT into an accessible lesion(s), and response assessed in all target lesions (injected and non-injected) by RECIST v1.1. Study therapy was continued until progression, toxicity, investigator decision or withdrawal of consent. Baseline and on-therapy serum was collected for cytokine analysis. Immunohistochemical and RNA-Seq analysis was performed on available pre- and post-treatment tumor biopsies. Results: As of December 31, 2017, 68 subjects have been treated (44 in Escalation and 24 in Expansion). Safety data from 63 subjects demonstrated a manageable acute toxicity profile consisting predominately of fever, N/V, headache, hypotension and rigors. Grade 3/4 related AEs reported in ≥1 subject; hypotension (n=7), anemia (n=2), chills (n=2), hypertension (n=2) and fever (n=2). The Objective Response Rates (ORR) across all dose cohorts on weekly (n=40) and q3week schedules (n=13) were 22.5% (9/40; 95 % CI 11-39%) and 7.7%% (1/13; 95% CI 0-36%) respectively. For subjects dosed weekly at 3 and 5 mg, the ORR was 33.3% (6/18 95% CI 13-59%). Of the 10 responders, 1 progressed (w36), 2 withdrew consent (w13, w25), 7 remain on study with 2 subjects maintaining their response though w72. Regression of non-injected tumors occurred in cutaneous, nodal, hepatic, and splenic metastases. CMP-001 induced TLR9 activation with a median 5.9 fold increase in serum CXCL10 (range of 0.9 - 276.3; mean fold increase of 21.8 with SD=48.8; n=39). Immunohistochemical and RNA-Seq analysis of tumor biopsies revealed increases in tumor-infiltrating CD8 T cells (>5 fold), PD-L1 expression (>3 fold increase in H score), and transcriptional signature of inflammation in 2/4 subjects with analyzable pre-and post-treatment samples. Conclusions: CMP-001 in combination with pembrolizumab resulted in objective, durable tumor responses with tolerable toxicities in subjects with advanced melanoma resistant to prior anti-PD-1 therapy. CMP-001 dosing at 5 mg/weekly has been selected for further evaluation in the ongoing dose expansion phase of this study. Citation Format: Mohammed Milhem, Rene Gonzales, Theresa Medina, John M. Kirkwood, Elizabeth Buchbinder, Inderjit Mehmi, Jiaxin Niu, Montaser Shaheen, Ryan Weight, Kim Margolin, Jason Luke, Aaron Morris, David Mauro, Arthur M. Krieg, Antoni Ribas. Intratumoral toll-like receptor 9 (TLR9) agonist, CMP-001, in combination with pembrolizumab can reverse resistance to PD-1 inhibition in a phase Ib trial in subjects with advanced melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr CT144.

e21552Background: Metastatic melanoma (MM) continues to present a challenge in clinical oncology. Anti-PD-1 antibodies (nivolumab, pembrolizumab) have improved outcomes for a significant number of ...

e12513Background: Eflapegrastim is a novel investigational biologic comprised of recombinant human G-CSF covalently linked to the human immunoglobulin G4FC fragment using proprietary LAPSCOVERY™ te...

Overexpression of the lipogenic enzyme fatty acid synthase (FAS) is a common molecular feature in subsets of sex-steroid-related tumors including endometrium and breast carcinomas that are associated with poor prognosis. Pharmacological inhibition of tumor-associated FAS hyperactivity is under investigation as a chemotherapeutic target. We examined the effects of the mycotoxin cerulenin (a covalent FAS inactivator), and the novel small compound C75 (a slow-binding FAS inhibitor) on estradiol (E2)- and tamoxifen (TAM)-stimulated ER-driven molecular responses in Ishikawa cells, an in vitro model of well-differentiated human endometrial carcinoma. We evaluated the effects of FAS inhibition on E2- and TAM-induced estrogen receptor (ER) transcriptional activity by using transient cotransfection assays with an estrogen-response element reporter construct (ERE-Luciferase). Antiestrogenic effects of cerulenin and C75 were observed by dose-dependent inhibition of E2-stimulated ERE-dependent transcription, whereas FAS inhibitors did not significantly increase the levels of ERE transcriptional activity in the absence of E2. Moreover, pharmacological blockade of FAS activity completely abolished TAM-stimulated ERE activity. To address the reliability of transient transfection assays, the effects of FAS inhibitors on E2-inducible gene products were evaluated. FAS blockade induced a dose-dependent decrease in E2-inducible alkaline phosphatase activity. E2-stimulated accumulation of progesterone receptor (PR) and HER-2/neu oncogene was abolished in the presence of FAS blockers. FAS inhibition also resulted in a marked downregulation of E2-stimulated ERalpha expression, and noticeably impaired E2-induced ERalpha nuclear accumulation. A dose-dependent decrease in cell proliferation and cell viability was observed after FAS blockade. A Cell Death ELISA, detecting DNA fragmentation, demonstrated that FAS inhibitors stimulated apoptosis of Ishikawa cells. The analysis of critical E2- and TAM-related cell cycle proteins revealed an increase of both the expression and the nuclear accumulation of cyclin-dependent kinase inhibitors p21WAF1/CIP1 and p27Kip1 following FAS inhibition. To rule out non-FAS cerulenin- and C75-related effects, we finally monitored ER signaling after silencing of FAS gene expression using the highly sequence-specific mechanism of RNA interference (RNAi). The concentrations of E2 and TAM inducing half-maximal ERE activity (EC50) dramatically increased (100 times) in FAS RNAi-transfected Ishikawa cells. Moreover, depletion of FAS by RNAi also caused loss of ERalpha expression, downregulation of PR, and accumulation of p21WAF1/CIP1 and p27Kip1 in E2-stimulated Ishikawa cells. If chemically stable FAS inhibitors or cell-selective vector systems able to deliver RNAi targeting FAS gene demonstrate systemic anticancer effects in vivo, our results render FAS as a novel target for the prevention and treatment of endometrial carcinoma.

CYR61 (CNN1), a member of the cysteine rich 61/connective tissue growth factor/nephroblastoma overexpressed (CYR61/CTFG/NOV) family of growth regulators (CNN), is a pro-angiogenic factor that mediates diverse roles in development, cell proliferation, and tumorigenesis. We have recently shown that CYR61 is overexpressed in invasive and metastatic human breast cancer cells. Accordingly, elevated levels of CYR61 in breast cancer are associated with more advanced disease. Unfortunately, the exact mechanisms by which CYR61 promotes an aggressive breast cancer phenotype are still largely unknown. This review examines the functional role of CYR61 in breast cancer disease, presenting evidence that CYR61 signaling may play a major role in estrogen- as well as growth factor-dependent breast cancer progression. We also emphasize the functional significance of the molecular connection of CYR61 and its integrin receptor alpha(v)beta(3) enhancing breast cancer aggressiveness. Moreover, we describe experimental evidence that establishes a novel role for CYR61 determining the protection of human breast cancer cells against chemotherapy-induced apoptosis through its interactions with the integrin receptor alpha(v)beta(3). All these findings delineate a new noteworthy function of a CYR61/alpha(v)beta(3) autocrine-paracrine signaling pathway within both angiogenesis and breast cancer progression, which would allow a dual anti-angiogenic and anti-tumor benefit with a single drug.

Heregulin (HRG) is an activator of the erbB2-, erbB3- and erbB4-(erbB-2/3/4) signaling pathway. Transfection of full-length HRG cDNA into the estrogen (E2)-dependent cell line MCF-7 promoted an invasive E2-independent phenotype, as well as persistent activation of the erbB-2/3/4 receptors. Moreover, HRG expression in MCF-7 cells renders the cells sensitive to the topoisomerase II inhibitor doxorubicin (Doxo). In an attempt to dissociate the tumorigenic effect of HRG from the sensitizing effect to chemotherapy, we constructed a structural deletion mutant of HRG. Transfection of the deletion mutant of HRG described in this study (HRG/M) into MCF-7 cells resulted in the dissociation of the tumor-promoting activity of HRG from the sensitization to Doxo, that is, although the cells did not become more aggressive or E2-independent they became more sensitive to Doxo. HRG/M was unable to autophosphorylate the erbB receptors and did not affect the level of MAPK phosphorylation. Furthermore, the intracellular localization of the protein was different from that of the full-length protein. Our data show that the HRG/M sequences are sufficient to sensitize MCF-7 cells to Doxo, and provide evidence that this sensitization is independent of erbB2 activation.

Black cohosh is an increasingly popular alternative to estrogen replacement therapy for the relief of menopausal symptoms, primarily hot flushes. However, an important consideration for long-term therapy is potential toxicity and carcinogenicity. Therefore, we undertook a study to assess the estrogenic activity of black cohosh to examine its safety for those with, or at high risk of developing, breast cancer. Several assays were utilized as listed: RNAse protection assays, which ascertain the regulation of the expression of E2-responsive genes; estrogen-responsive-element (ERE)-luciferase, which determines modulation of the ER function by transactivation of the ERE; the Ishikawa cell system, which has an E2-regulated endogenous alkaline phosphatase; and colony formation of ER-expressing breast cancer cells, which indicates possible progression of early stage breast cancer into a more aggressive state. Black cohosh extracts did not demonstrate estrogenic activity in any of these assay systems. This is an encouraging step in the assessment of the safety of black cohosh for treatment of menopausal hot flushes.

The growth factor heregulin (HRG), expressed in about 30% of breast cancer tumors, activates the erbB-2 receptor via induction of heterodimeric complexes of erbB-2 with erbB-3 or erbB-4. HRG induces tumorigenicity and metastasis of breast cancer cells. Our investigation into whether HRG is a factor likely to promote tumor formation independently of erbB-2 overexpression concludes that blockage of HRG expression suppresses the aggressive phenotype of MDA-MB-231 breast cancer cells by inhibiting cell proliferation, preventing anchorage-independent growth, and suppressing the invasive potential of the cells in vitro. More importantly, we observed a marked reduction in tumor formation, tumor size, and a lack of metastasis in vivo. These studies were achieved by blocking HRG expression in MDA-MB-231 cells using an HRG antisense cDNA. In the search for the mechanism by which blockage of HRG reverts this aggressive phenotype, we discovered that the cells in which HRG is blocked exhibit a marked decrease in erbB activation and a significant reduction in MMP-9 activity, demonstrating a direct causal role in HRG induction of tumorigenicity. Our study is the first report and serves as a proof of the concept that HRG is a key promoter of breast cancer tumorigenicity and metastasis independently of erbB-2 overexpression and should be deemed a potential target in developing therapies for breast cancer.

e19092 Background: Whether a delay in the initiation of systemic chemotherapy for advanced non-small cell lung cancer (NSCLC) can affect overall survival (OS) is not well studied. In this study we ...