Richard E. Cutler

A phase I/II study of carfilzomib 2–10-min infusion in patients with advanced solid tumors

AbstractPurposenTolerability, pharmacokinetics (PK), pharmacodynamics, and antitumor activity of carfilzomib, a selective proteasome inhibitor, administered twice weekly by 2–10-min intravenous (IV) infusion on days 1, 2, 8, 9, 15, and 16 in 28-day cycles, were assessed in patients with advanced solid tumors in this phase I/II study.nMethodsAdult patients with solid tumors progressing after ≥1 prior therapies were enrolled. The dose was 20xa0mg/m2 in week 1 of cycle 1 and 20, 27, or 36xa0mg/m2 thereafter. The maximum tolerated dose or protocol-defined maximum planned dose (MPD) identified during dose escalation was administered to an expansion cohort and to patients with small cell lung, non-small cell lung, ovarian, and renal cancer in phase II tumor-specific cohorts.ResultsFourteen patients received carfilzomib during dose escalation. The single dose-limiting toxicity at 20/36xa0mg/m2 was grade 3 fatigue, establishing the MPD as the expansion and phase II dose. Sixty-five additional patients received carfilzomib at the MPD. Adverse events included fatigue, nausea, anorexia, and dyspnea. Carfilzomib PK was dose proportional with a half-life <1xa0h. All doses resulted in at least 80xa0% proteasome inhibition in blood. Partial responses occurred in two patients in phase I, with 21.5xa0% stable disease after four cycles in evaluable patients in the expansion and phase II cohorts.ConclusionCarfilzomib 20/36xa0mg/m2 was well tolerated when administered twice weekly by 2–10-min IV infusion. At this dose and infusion rate, carfilzomib inhibited the proteasome in blood but demonstrated limited antitumor activity in patients with advanced solid tumors.n

The levels of mutant K-RAS and mutant N-RAS are rapidly reduced in a Beclin1 / ATG5 -dependent fashion by the irreversible ERBB1/2/4 inhibitor neratinib

ABSTRACT The FDA approved irreversible inhibitor of ERBB1/2/4, neratinib, was recently shown to rapidly down-regulate the expression of ERBB1/2/4 as well as the levels of c-MET and mutant K-RAS via autophagic degradation. In the present studies, in a dose-dependent fashion, neratinib reduced the expression levels of mutant K-RAS or of mutant N-RAS, which was augmented in an additive to greater than additive fashion by the HDAC inhibitors sodium valproate and AR42. Neratinib could reduce PDGFRα levels in GBM cells, that was enhanced by sodium valproate. Knock down of Beclin1 or of ATG5 prevented neratinib and neratinib combined with sodium valproate / AR42 from reducing the expression of mutant N-RAS in established PDX and fresh PDX models of ovarian cancer and melanoma, respectively. Neratinib and the drug combinations caused the co-localization of mutant RAS proteins and ERBB2 with Beclin1 and cathepsin B. The drug combination activated the AMP-dependent protein kinase that was causal in enhancing HMG Co A reductase phosphorylation. Collectively, our data reinforce the concept that the irreversible ERBB1/2/4 inhibitor neratinib has the potential for use in the treatment of tumors expressing mutant RAS proteins.

Abstract PD2-08: Neratinib + fulvestrant in ERBB2-mutant, HER2–non-amplified, estrogen receptor (ER)-positive, metastatic breast cancer (MBC): Preliminary analysis from the phase II SUMMIT trial

Background: Somatic mutations in ERBB2 are a new class of oncogenic drivers in HER2–non amplified MBC. Neratinib is an irreversible pan-HER tyrosine kinase inhibitor that inhibits the growth of ERBB2-mutant breast tumors in preclinical models and has encouraging single-agent clinical activity in patients (pts) with ERBB2-mutant, HER2–non amplified MBC. Bi-directional signaling between HER2 and ER may limit the effectiveness of endocrine and HER2 directed therapy, if each is given alone, in ER+ MBC with ERBB2 amplifications/mutations. Preclinical data suggest that dual blockade of ER and HER2 signaling results in enhanced anti-tumor activity in ER+ HER2+ MBC. SUMMIT, a multicenter multi-histology phase II 9basket9 trial, is investigating the efficacy of neratinib monotherapy (in ER+ and ER– pts) and neratinib + fulvestrant (ER+ pts only) in ERBB2-mutant MBC. Methods: MBC pts with ERBB2 mutations documented by local testing were eligible and received oral neratinib 240 mg qd. Pts with ER+ MBC received fulvestrant 500 mg, a selective ER degrader, in addition to neratinib on d1 & 15 of month 1 then on d1 q4w. Patients received high dose loperamide prophylaxis during cycle 1. Primary endpoint is objective response rate (ORR) at 8w, defined using RECIST 1.1 and/or modified PERCIST assessments. Secondary endpoints include ORR, clinical benefit rate (CBR), progression free survival (PFS), and safety. Mutation profiling and central confirmation of ERBB2 mutation(s) from available fresh or archival tumor tissues and plasma DNA were performed retrospectively by next-generation sequencing (MSK-IMPACT). Clinicaltrials.gov: NCT01953926. Results: As of 23 Sep 2016, 35 efficacy-evaluable ERBB2-mutant MBC pts received neratinib, either as monotherapy (n=24) or in combination with fulvestrant (n=11). Efficacy findings are shown in the table. The overall safety profile of neratinib + fulvestrant was similar to that previously reported with neratinib monotherapy. Grade 3 diarrhea rate was 24% with neratinib monotherapy and 18% with neratinib + fulvestrant. Conclusions: Encouraging clinical activity has been observed with neratinib + fulvestrant in heavily pretreated pts with ERBB2-mutant, ER+ MBC. Clinical efficacy in the ER+ MBC cohort met pre-specified efficacy requirements; a confirmatory trial of neratinib + fulvestrant for targeting ERBB2 mutations in ER+ MBC is warranted. The safety profile of neratinib was acceptable and diarrhea was manageable with loperamide prophylaxis. Citation Format: Hyman D, Piha-Paul S, Saura C, Arteaga C, Mayer I, Shapiro G, Loi S, Lalani A, Xu F, Cutler R, Butturini A, Bryce R, Meric-Bernstam F, Baselga J, Solit D. Neratinib + fulvestrant in ERBB2-mutant, HER2–non-amplified, estrogen receptor (ER)-positive, metastatic breast cancer (MBC): Preliminary analysis from the phase II SUMMIT trial [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr PD2-08.

The irreversible ERBB1/2/4 inhibitor neratinib interacts with the PARP1 inhibitor niraparib to kill ovarian cancer cells

ABSTRACT The irreversible ERBB1/2/4 inhibitor neratinib has been shown to rapidly down-regulate the expression of ERBB1/2/4 as well as the levels of c-MET, PDGFRα and mutant RAS proteins via autophagic degradation. Neratinib interacted in an additive to synergistic fashion with the approved PARP1 inhibitor niraparib to kill ovarian cancer cells. Neratinib and niraparib caused the ATM-dependent activation of AMPK which in turn was required to cause mTOR inactivation, ULK-1 activation and ATG13 phosphorylation. The drug combination initially increased autophagosome levels followed later by autolysosome levels. Preventing autophagosome formation by expressing activated mTOR or knocking down of Beclin1, or knock down of the autolysosome protein cathepsin B, reduced drug combination lethality. The drug combination caused an endoplasmic reticulum stress response as judged by enhanced eIF2α phosphorylation that was responsible for reducing MCL-1 and BCL-XL levels and increasing ATG5 and Beclin1 expression. Knock down of BIM, but not of BAX or BAK, reduced cell killing. Expression of activated MEK1 prevented the drug combination increasing BIM expression and reduced cell killing. Downstream of the mitochondrion, drug lethality was partially reduced by knock down of AIF, but expression of dominant negative caspase 9 was not protective. Our data demonstrate that neratinib and niraparib interact to kill ovarian cancer cells through convergent DNA damage and endoplasmic reticulum stress signaling. Cell killing required the induction of autophagy and was cathepsin B and AIF -dependent, and effector caspase independent.

Neratinib is effective in breast tumors bearing both amplification and mutation of ERBB2 (HER2)

Metastatic breast cancer patients with coexistent HER2 mutation and amplification respond to neratinib. Neratinib for resistant metastatic breast cancer Breast cancers with amplification or mutation in the epidermal growth factor receptor (EGFR) family member HER2 are usually treated with targeted inhibitors, but resistance is common. Amplification and mutation of HER2 are generally considered mutually exclusive occurrences in treatment-naïve patients. However, Cocco et al. discovered a small proportion of treatment-naïve and, more often, previously treated patients with metastatic breast cancer in which HER2 amplification and mutation were coincident. It is not yet clear why, but these co-amplified/mutant cells were resistant to currently approved HER2-specific and HER2/EGFR-specific inhibitors but were sensitive to the new pan-EGFR inhibitor neratinib. Neratinib, which inhibits EGFR and HER2, as well as HER3 and HER4, was more effective at blocking the activity of the EGFR pathway and other receptor tyrosine kinases, common modes of resistance in HER2-driven tumors. Patients and mice bearing their tumor cells showed improved survival and even tumor regression on neratinib, suggesting that this may be a treatment option for certain breast cancer patients. Mutations in ERBB2, the gene encoding epidermal growth factor receptor (EGFR) family member HER2, are common in and drive the growth of “HER2-negative” (not ERBB2 amplified) tumors but are rare in “HER2-positive” (ERBB2 amplified) breast cancer. We analyzed DNA-sequencing data from HER2-positive patients and used cell lines and a patient-derived xenograft model to test the consequence of HER2 mutations on the efficacy of anti-HER2 agents such as trastuzumab, lapatinib, and neratinib, an irreversible pan-EGFR inhibitor. HER2 mutations were present in ~7% of HER2-positive tumors, all of which were metastatic but not all were previously treated. Compared to HER2 amplification alone, in both patients and cultured cell lines, the co-occurrence of HER2 mutation and amplification was associated with poor response to trastuzumab and lapatinib, the standard-of-care anti-HER2 agents. In mice, xenografts established from a patient whose HER2-positive tumor acquired a D769Y mutation in HER2 after progression on trastuzumab-based therapy were resistant to trastuzumab or lapatinib but were sensitive to neratinib. Clinical data revealed that six heavily pretreated patients with tumors bearing coincident HER2 amplification and mutation subsequently exhibited a statistically significant response to neratinib monotherapy. Thus, these findings indicate that coincident HER2 mutation reduces the efficacy of therapies commonly used to treat HER2-positive breast cancer, particularly in metastatic and previously HER2 inhibitor–treated patients, as well as potentially in patients scheduled for first-line treatment. Therefore, we propose that clinical studies testing the efficacy of neratinib are warranted selectively in breast cancer patients whose tumors carry both amplification and mutation of ERBB2/HER2.

Abstract PD2-05: Inhibition of mutant HER2 results in synthetic lethality when combined with ER antagonists in ER+/HER2 mutant human breast cancer cells

Background: Human epidermal growth factor receptor 2 (ERBB2 or commonly known as HER2) missense mutations have been reported in 2-4% of breast cancers and occur primarily in the absence of HER2 gene amplification. Based on TCGA, approximately 60% of these tumors are hormone-dependent and express estrogen receptor (ER) a. Among ER+ breast cancers with HER2 missense mutations, more than 80% are in the HER2 kinase domain. We examined herein whether ER+/HER2 mutant breast cancer cells are resistant to anti-estrogen therapies and, thus, whether they should be treated with combined ER and HER2 inhibitors. Methods: Three common HER2 activating mutations (G309A, L755S, V777L) and wild type (WT) HER2 were incorporated into ER+ MCF7 cells using AAV-mediated homologous recombination. The isogenic incorporation of a heterozygous mutation more accurately represents primary human tumors as compared to transfection and overexpression of exogenous vectors. We examined cell viability and ER transcriptional activity, using an ERE-luciferase reporter, in response to estrogen deprivation and treatment with fulvestrant (a selective ER downregulator) and neratinib (an irreversible, pan-HER tyrosine kinase inhibitor), either alone or in combination. Signaling downstream mutant HER2 was examined by immunoblot analysis. In vivo anti-tumor efficacy of fulvestrant ± neratinib is currently being assessed in ovariectomized athymic mice bearing MCF7/HER2V777L xenografts. Results: MCF7 cells containing HER2 kinase missense mutations (L755S and V777L), but not cells with HER2WT or an extracellular domain mutation (G309A), were able to proliferate exponentially in estrogen-free medium. MCF7/HER2L755S and MCF7/HER2V777L were also resistant to 1 mM fulvestrant, despite fulvestrant9s ability to downregulate ER in these cells. Additionally, MCF7/HER2L755S and MCF7/HER2V777L showed increased levels of pERK and p70S6K. Treatment with 200 nM neratinib potently inhibited growth of MCF7/HER2L755S and MCF7/HER2V777L in estrogen-free conditions and resensitized them to fulvestrant while partially downregulating HER2 levels. Addition of 1 nM estradiol markedly rescued all three HER2 mutant cells from neratinib-induced cell death suggesting that the inhibition of both ER and mutant HER2 is required for tumor cell apoptosis. Using ERE-luc reporter assays, neratinib did not inhibit basal or estrogen-induced ER transcriptional activity or ERα Ser118 phosphorylation, thus not supporting HER2 mutation-to-ER crosstalk in these genetically engineered cells. This result also suggests that the ER and HER2 mutant pathways can operate independently and it is the dual pathway inhibition that results in synthetic lethality. Conclusions: These data suggest that, in ER+ breast cancers and similar to HER2 gene amplification, HER2 kinase domain mutations induce resistance to antiestrogen therapies. Therefore, we propose simultaneous therapeutic targeting of both ER and HER2 signaling pathways is required for maximal inhibition of ER+ breast cancers also harboring HER2 activating mutations, as is currently being investigated in the phase II SUMMIT trial (NCT01953926). Citation Format: Croessmann S, Zabransky DJ, Cutler, Jr. RE, Lalani AS, Park BH, Arteaga CL. Inhibition of mutant HER2 results in synthetic lethality when combined with ER antagonists in ER+/HER2 mutant human breast cancer cells [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr PD2-05.