Cyclin E (CCNE1) has been proposed as a biomarker of sensitivity to adavosertib, a Wee1 kinase inhibitor, and a mechanism of resistance to HER2-targeted therapy.
Copy number and genomic sequencing ...data from The Cancer Genome Atlas and MD Anderson Cancer Center databases were analyzed to assess ERBB2 and CCNE1 expression. Molecular characteristics of tumors and patient-derived xenografts (PDX) were assessed by next-generation sequencing, whole-exome sequencing, fluorescent in situ hybridization, and IHC. In vitro, CCNE1 was overexpressed or knocked down in HER2+ cell lines to evaluate drug combination efficacy. In vivo, NSG mice bearing PDXs were subjected to combinatorial therapy with various treatment regimens, followed by tumor growth assessment. Pharmacodynamic markers in PDXs were characterized by IHC and reverse-phase protein array.
Among several ERBB2-amplified cancers, CCNE1 co-amplification was identified (gastric 37%, endometroid 43%, and ovarian serous adenocarcinoma 41%). We hypothesized that adavosertib may enhance activity of HER2 antibody-drug conjugate trastuzumab deruxtecan (T-DXd). In vitro, sensitivity to T-DXd was decreased by cyclin E overexpression and increased by knockdown, and adavosertib was synergistic with topoisomerase I inhibitor DXd. In vivo, the T-DXd + adavosertib combination significantly increased γH2AX and antitumor activity in HER2 low, cyclin E amplified gastroesophageal cancer PDX models and prolonged event-free survival (EFS) in a HER2-overexpressing gastroesophageal cancer model. T-DXd + adavosertib treatment also increased EFS in other HER2-expressing tumor types, including a T-DXd-treated colon cancer model.
We provide rationale for combining T-DXd with adavosertib in HER2-expressing cancers, especially with co-occuring CCNE1 amplifications. See related commentary by Rolfo et al., p. 4317.
The RET proto-oncogene, a tyrosine kinase receptor, is widely known for its essential role in cell survival. Germ line missense mutations, which give rise to constitutively active oncogenic RET, were ...found to cause multiple endocrine neoplasia type 2, a dominant inherited cancer syndrome that affects neuroendocrine organs. However, the mechanisms by which RET promotes cell survival and prevents cell death remain elusive. We demonstrate that in addition to cytoplasmic localization, RET is localized in the nucleus and functions as a tyrosine-threonine dual specificity kinase. Knockdown of RET by shRNA in medullary thyroid cancer-derived cells stimulated expression of activating transcription factor 4 (ATF4), a master transcription factor for stress-induced apoptosis, through activation of its target proapoptotic genes NOXA and PUMA. RET knockdown also increased sensitivity to cisplatin-induced apoptosis. We observed that RET physically interacted with and phosphorylated ATF4 at tyrosine and threonine residues. Indeed, RET kinase activity was required to inhibit the ATF4-dependent activation of the NOXA gene because the site-specific substitution mutations that block threonine phosphorylation increased ATF4 stability and activated its targets NOXA and PUMA. Moreover, chromatin immunoprecipitation assays revealed that ATF4 occupancy increased at the NOXA promoter in TT cells treated with tyrosine kinase inhibitors or the ATF4 inducer eeyarestatin as well as in RET-depleted TT cells. Together these findings reveal RET as a novel dual kinase with nuclear localization and provide mechanisms by which RET represses the proapoptotic genes through direct interaction with and phosphorylation-dependent inactivation of ATF4 during the pathogenesis of medullary thyroid cancer.
Background: Activating mutations of the receptor tyrosine kinase RET are associated with oncogenic function in medullary thyroid cancer.
Results: RET is a dual specificity kinase, phosphorylates ATF4, and inhibits expression of the ATF4 target proapoptotic genes.
Conclusion: RET prevents apoptosis through inhibition of ATF4 activity.
Significance: Simultaneous targeting of RET and ATF4 may provide clinical benefit in cancers with RET abnormalities.
Molecular alterations in the PI3K/AKT pathway occur frequently in hormone receptor-positive breast tumors. Patients with ER-positive, HER2-negative metastatic breast cancer are often treated with ...CDK4/6 inhibitors such as palbociclib in combination with endocrine therapy. Although this is an effective regimen, most patients ultimately progress. The purpose of this study was identifying synthetic lethality partners that can enhance palbociclib's antitumor efficacy in the presence of PIK3CA/AKT1 mutations. We utilized a barcoded shRNA library to determine critical targets for survival in isogenic MCF7 cells with PIK3CA/AKT1 mutations. We demonstrated that the efficacy of palbociclib is reduced in the presence of PIK3CA/AKT1 mutations. We also identified that the downregulation of discoidin domain receptor 1 (DDR1) is synthetically lethal with palbociclib. DDR1 knockdown and DDR1 pharmacological inhibitor decreased cell growth and inhibited cell cycle progression in all cell lines, while enhanced the sensitivity of PIK3CA/AKT1 mutant cells to palbociclib. Combined treatment of palbociclib and 7rh further induced cell cycle arrest in PIK3CA/AKT1 mutant cell lines. In vivo, 7rh significantly enhanced palbociclib's antitumor efficacy. Our data indicates that DDR1 inhibition can augment cell cycle suppressive effect of palbociclib and could be effective strategy for targeted therapy of ER-positive, HER2-negative breast cancers with PI3K pathway activation.
Zanidatamab is a bispecific human epidermal growth factor receptor 2 (HER2)-targeted antibody that has demonstrated antitumor activity in a broad range of HER2-amplified/expressing solid tumors. We ...determined the antitumor activity of zanidatamab in patient-derived xenograft (PDX) models developed from pretreatment or postprogression biopsies on the first-in-human zanidatamab phase I study (NCT02892123). Of 36 tumors implanted, 19 PDX models were established (52.7% take rate) from 17 patients. Established PDXs represented a broad range of HER2-expressing cancers, and in vivo testing demonstrated an association between antitumor activity in PDXs and matched patients in 7 of 8 co-clinical models tested. We also identified amplification of MET as a potential mechanism of acquired resistance to zanidatamab and demonstrated that MET inhibitors have single-agent activity and can enhance zanidatamab activity in vitro and in vivo. These findings provide evidence that PDXs can be developed from pretreatment biopsies in clinical trials and may provide insight into mechanisms of resistance.
We demonstrate that PDXs can be developed from pretreatment and postprogression biopsies in clinical trials and may represent a powerful preclinical tool. We identified amplification of MET as a potential mechanism of acquired resistance to the HER2 inhibitor zanidatamab and MET inhibitors alone and in combination as a therapeutic strategy. This article is featured in Selected Articles from This Issue, p. 695.
Abstract Hypoxia and adenosine are known to upregulate angiogenesis; however, the role of peroxisome proliferator-activated receptor alpha (PPARα) in angiogenesis is controversial. Using transgenic ...Tg(fli-1:EGFP) zebrafish embryos, interactions of PPARα and adenosine receptors in angiogenesis were evaluated under hypoxic conditions. Epifluorescent microscopy was used to assess angiogenesis by counting the number of intersegmental (ISV) and dorsal longitudinal anastomotic vessel (DLAV) at 28 h post-fertilization (hpf). Hypoxia (6 h) stimulated angiogenesis as the number of ISV and DLAV increased by 18-fold (p < 0.01) and 100 ± 8% (p < 0.001), respectively, at 28 hpf. Under normoxic and hypoxic conditions, WY-14643 (10 μM), a PPARα activator, stimulated angiogenesis at 28 hpf, while MK-886 (0.5 μM), an antagonist of PPARα, attenuated these effects. Compared to normoxic condition, adenosine receptor activation with NECA (10 μM) promoted angiogenesis more effectively under hypoxic conditions. Involvement of A2B receptor was implied in hypoxia-induced angiogenesis as MRS-1706 (10 nM), a selective A2B antagonist attenuated NECA (10 μM)-induced angiogenesis. NECA- or WY-14643-induced angiogenesis was also inhibited by miconazole (0.1 μM), an inhibitor of epoxygenase dependent production of eicosatrienoic acid (EET) epoxide. Thus, we conclude that: activation of PPARα promoted angiogenesis just as activation of A2B receptors through an epoxide dependent mechanism.
Hypoxia (HYP) stimulates angiogenesis (ANG) by promoting expression of many pro‐angiogenic genes, including AR. The role of PPARα in ANG is controversial. A2BR‐induced activation is known to reduce ...PPARα‐mediated activity but the effect of PPARα activation on A2BR is unknown. Here, we tested that PPARα attenuates adenosine(AD)‐mediated HYP –induced ANG. Using zebrafish embryo, ANG was measured by counting intersegmental vessel (ISV) and dorsal longitudinal anastomotic vessels (DLAV). Under normoxic condition, WY‐14643 (WY, 10 μM), a PPARα agonist, increased ISV (P<0.01) at 28 hpf but NECA (10 μM), a non selective AR agonist showed no effect. Combined exposure of WY 10 or 100 μM or NECA (10μM) decreased ISV at 28 hpf (p<0.01). However, WY (10 μM) +NECA increased DLAV (P<0.001), whereas, WY (100 μM)+NECA decreased DLAV (P<0.05) at 48 hpf. HYP alone increased ANG (72±19%) at 48 hpf. WY(10 μM) showed no effect on ISV but increased DLAV (P<0.05). At 28 hpf WY (10 μM)+NECA decreased ISV (P<0.001). Our data show that activation of PPARα and AR individually promote HYP‐induced ANG, but combined activation inhibits AD‐mediated HYP‐induced ANG.
The role of peroxisome proliferator‐activated receptor alpha (PPARα) in angiogenesis is controversial. However, many studies showed that epoxyeisatrienoic acids (EETs), downstream mediators of ...adenosine A2B receptor (A2BR) and PPARα ligand are proangiogenic. Using the Zebrafish (Danio rerio) embryo, we here propose that EET‐induced PPARα activation is involved in A2BR‐mediated angiogenesis under hypoxic condition. Angiogenesis was evaluated at 24, 48, 72 hpf using alkaline phosphatase activity (APA). Under normoxic conditions, APA increased in a time‐dependent manner at 24, 48 (132 %) and 72 hpf (200 %). Clofibrate (10–200 μM) or fenofibrate (10–50 μM); PPARα ligands, NECA (0.1–100 μM), a non selective adenosine receptor agonist, or hydralazine (0.5–1mM), inhibitor of degradation of HIF‐1α did not affect angiogenesis at any of the time points. Compared to normoxia, hypoxia for 24 h reduced angiogenesis by 85 % (p<0.01), but not at 48 or 72 hpf. Using transgenic Tg(fli‐1:EGFP) embryos and epifluorescence microscopy, angiogenesis increased under normoxic conditions in response to clofibrate (50 μM), NECA (10 μM) or hydralazine (0.5 mM), but not to fenofibrate. Epifluorescence microscopy but not APA data demonstrate the possible involvement of PPARα in angiogenesis in the Zebrafish embryo model. However, APA data do not support hypoxia‐induced angiogenesis in Zebrafish embryo.
PDF
