Genotoxic Damage Activates the AMPK-α1 Isoform in the Nucleus via Ca2+/CaMKK2 Signaling to Enhance Tumor Cell Survival

2017 American Association for Cancer Research. Many genotoxic cancer treatments activate AMP-activated protein kinase (AMPK), but the mechanisms of AMPK activation in response to DNA damage, and its downstream consequences, have been unclear. In this study, etoposide activates the a1 but not the a2 isoform of AMPK, primarily within the nucleus. AMPK activation is independent of ataxia-telangiectasia mutated (ATM), a DNA damage-activated kinase, and the principal upstream kinase for AMPK, LKB1, but correlates with increased nuclear Ca2þ and requires the Ca2þ/calmodulin-dependent kinase, CaMKK2. Intriguingly, Ca2þ-dependent activation of AMPK in two different LKB1-null cancer cell lines caused G1-phase cell-cycle arrest, and enhanced cell viability/ survival after etoposide treatment, with both effects being abolished by knockout of AMPK-a1 and a2. The CDK4/6 inhibitor palbociclib also caused G1 arrest in G361 but not HeLa cells and, consistent with this, enhanced cell survival after etoposide treatment only in G361 cells. These results suggest that AMPK activation protects cells against etoposide by limiting entry into S-phase, where cells would be more vulnerable to genotoxic stress. Implications: These results reveal that the a1 isoform of AMPK promotes tumorigenesis by protecting cells against genotoxic stress, which may explain findings that the gene encoding AMPK-a1 (but not -a2) is amplified in some human cancers. Furthermore, a1-selective inhibitors might enhance the anticancer effects of genotoxic-based therapies

Targeting AMP-activated kinase impacts hepatocellular cancer stem cells induced by long-term treatment with sorafenib

Hepatocellular carcinoma (HCC) is the third leading cause of cancer death worldwide. HCC treatment is hindered by the frequent emergence of chemoresistance to the multikinase inhibitor sorafenib, which has been related to the presence of cancer stem cells (CSCs) that self‐renew and often escape therapy. The key metabolic sensor AMP‐activated kinase (AMPK) has recently been recognized as a tumour growth regulator. In this study, we aimed to elucidate the role of AMPK in the development of a stem cell phenotype in HCC cells. To this end, we enriched the CSC population in HCC cell lines that showed increased expression of drug resistance (ALDH1A1, ABCB1A) and stem cell (CD133, Nanog, Oct4, alpha fetoprotein) markers and demonstrated their stemness phenotype. These cells were refractory to sorafenib‐induced cell death. We report that sorafenib‐resistant cells had lower levels of total and phosphorylated AMPK as well as its downstream substrate, ACC, compared with the parental cells. Interestingly, AMPK knockdown with siRNA or inhibition with dorsomorphin increased the expression of stem cell markers in parental cells and blocked sorafenib‐induced cell death. Conversely, the upregulation of AMPK, either by transfection or by pharmacological activation with A‐769662, decreased the expression of ALDH1A1, ABCB1A, CD133, Nanog, Oct4, and alpha fetoprotein, and restored sensitivity to sorafenib. Analysis of the underlying mechanism points to hypoxia‐inducible factor HIF‐1α as a regulator of stemness. In vivo studies in a xenograft mouse model demonstrated that stem‐like cells have greater tumourigenic capacity. AMPK activation reduced xenograft tumour growth and decreased the expression of stem cell markers. Taken together, these results indicate that AMPK may serve as a novel target to overcome chemoresistance in HCC

AMPK Causes Cell Cycle Arrest in LKB1-deficient Cells via Activation of CAMKK2

The AMP-activated protein kinase (AMPK) is activated by phosphorylation at Thr172, either by the tumor suppressor kinase LKB1 or by an alternate pathway involving the Ca(2+)/calmodulin-dependent kinase, CAMKK2. Increases in AMP:ATP and ADP:ATP ratios, signifying energy deficit, promote allosteric activation and net Thr172 phosphorylation mediated by LKB1, so that the LKB1-AMPK pathway acts as an energy sensor. Many tumor cells carry loss-of-function mutations in the STK11 gene encoding LKB1, but LKB1 re-expression in these cells causes cell cycle arrest. Therefore, it was investigated as to whether arrest by LKB1 is caused by activation of AMPK or of one of the AMPK-related kinases, which are also dependent on LKB1 but are not activated by CAMKK2. In three LKB1-null tumor cell lines, treatment with the Ca(2+) ionophore A23187 caused a G1-arrest that correlated with AMPK activation and Thr172 phosphorylation. In G361 cells, expression of a truncated, CAMKK2 mutant also caused G1-arrest similar to that caused by expression of LKB1, while expression of a dominant negative AMPK mutant, or a double knockout of both AMPK-α subunits, also prevented the cell cycle arrest caused by A23187. These mechanistic findings confirm that AMPK activation triggers cell cycle arrest, and also suggest that the rapid proliferation of LKB1-null tumor cells is due to lack of the restraining influence of AMPK. However, cell cycle arrest can be restored by re-expressing LKB1 or a constitutively active CAMKK2, or by pharmacological agents that increase intracellular Ca(2+) and thus activate endogenous CAMKK2. IMPLICATIONS: Evidence here reveals that the rapid growth and proliferation of cancer cells lacking the tumor suppressor LKB1 is due to reduced activity of AMPK, and suggests a therapeutic approach by which this block might be circumvented

SEOM clinical guidelines in early stage breast cancer (2018)

Breast cancer is the most common cancer in women in our country and it is usually diagnosed in the early and potentially curable stages. Nevertheless, around 20–30% of patients will relapse despite appropriate locoregional and systemic therapies. A better knowledge of this disease is improving our ability to select the most appropriate therapy for each patient with a recent diagnosis of an early stage breast cancer, minimizing unnecessary toxicities and improving long-term efficacy

A retrospective, multicenter study of the efficacy of lapatinib plus trastuzumab in HER2-positive metastatic breast cancer patients previously treated with trastuzumab, lapatinib, or both: the Trastyvere study

[Purpose]: To evaluate the efficacy and safety of lapatinib (L) and trastuzumab (T) combination in HER2-positive metastatic breast cancer (MBC) patients previously treated with T and/or L.[Materials and methods]: We conducted a retrospective, post-authorized, multicenter study including patients with HER2-positive MBC or locally advanced breast cancer (ABC) treated with the combination of L–T. Concomitant endocrine therapy, as well as brain metastasis and/or prior exposure to L, were allowed.[Results]: One hundred and fifteen patients from 14 institutions were included. The median age was 59.8 years. The median number of prior T regimens in the advanced setting was 3 and 73 patients had received a prior L regimen. The clinical benefit rate (CBR) was 34.8% (95% CI 26.1–43.5). Among other efficacy endpoints, the overall response rate was 21.7%, and median progression-free survival (PFS) and overall survival were 3.9 and 21.6 months, respectively. Heavily pretreated and ≥ 3 metastatic organ patients showed lower CBR and PFS than patients with a low number of previous regimens and < 3 metastatic organs. Moreover, CBR did not significantly change in L-pretreated compared with L-naïve patients (31.5% versus 40.5% for L-pretreated versus L-naïve). Grade 3/4 adverse events were reported in 19 patients (16.5%).[Conclusion]: The combination of L–T is an effective and well-tolerated regimen in heavily pretreated patients and remains active among patients progressing on prior L-based therapy. Our study suggests that the L–T regimen is a safe and active chemotherapy-free option for MBC patients previously treated with T and/or L.This work was supported by GlaxoSmithKline plc (GSK) through a contract with Medica Scientia Innovation Research (MedSIR), an academic research organization focused on independent clinical research development

Time course and management of key adverse events during the randomized phase III SOLAR-1 study of PI3K inhibitor alpelisib plus fulvestrant in patients with HR-positive advanced breast cancer.

Background Alpelisib (α-selective phosphatidylinositol 3-kinase inhibitor) plus fulvestrant is approved in multiple countries for men and postmenopausal women with PIK3CA-mutated, hormone receptor-positive, human epidermal growth factor receptor 2-negative advanced breast cancer following progression on or after endocrine therapy. A detailed understanding of alpelisib's safety profile should inform adverse event (AE) management and enhance patient care. Patients and methods AEs in the phase III SOLAR-1 trial were assessed in patients with and without PIK3CA mutations. The impact of protocol-specified AE-management recommendations was evaluated, including an amendment to optimize hyperglycemia and rash management. Results Patients were randomly assigned to receive fulvestrant plus alpelisib (n = 284) or placebo (n = 287). The most common grade 3/4 AEs with alpelisib were hyperglycemia (grade 3, 32.7%; grade 4, 3.9%), rash (grade 3, 9.9%), and diarrhea (grade 3, 6.7%). Median time to onset of grade ≥3 toxicity was 15 days (hyperglycemia, based on fasting plasma glucose), 13 days (rash), and 139 days (diarrhea). Metformin alone or in combination with other antidiabetic agents was used by most patients (87.1%) with hyperglycemia. Preventive anti-rash medication resulted in lower incidence (any grade, 26.7% versus 64.1%) and severity of rash (grade 3, 11.6% versus 22.7%) versus no preventative medication. Discontinuations due to grade ≥3 AEs were lower following more-detailed AE management guidelines (7.9% versus 18.1% previously). Patients with PIK3CA mutations had a median alpelisib dose intensity of 248 mg/day. Median progression-free survival with alpelisib was 12.5 and 9.6 months for alpelisib dose intensities of ≥248 mg/day and Conclusions Hyperglycemia and rash occurred early during alpelisib treatment, while diarrhea occurred at a later time point. Early identification, prevention, and intervention, including concomitant medications and alpelisib dose modifications, resulted in less severe toxicities. Reductions in treatment discontinuations and improved progression-free survival at higher alpelisib dose intensities support the need for optimal AE management. ClinicalTrials.gov Id NCT02437318

Alpelisib plus fulvestrant for PIK3CA-mutated, hormone receptor-positive, human epidermal growth factor receptor-2–negative advanced breast cancer: final overall survival results from SOLAR-1

Background Activation of the phosphatidylinositol-3-kinase (PI3K) pathway via PIK3CA mutations occurs in 28%-46% of hormone receptor-positive (HR+), human epidermal growth factor receptor-2-negative (HER2−) advanced breast cancers (ABCs) and is associated with poor prognosis. The SOLAR-1 trial showed that the addition of alpelisib to fulvestrant treatment provided statistically significant and clinically meaningful progression-free survival (PFS) benefit in PIK3CA-mutated, HR+, HER2− ABC. Patients and methods Men and postmenopausal women with HR+, HER2− ABC whose disease progressed on or after aromatase inhibitor (AI) were randomized 1 : 1 to receive alpelisib (300 mg/day) plus fulvestrant (500 mg every 28 days and once on day 15) or placebo plus fulvestrant. Overall survival (OS) in the PIK3CA-mutant cohort was evaluated by Kaplan–Meier methodology and a one-sided stratified log-rank test was carried out with an O'Brien–Fleming efficacy boundary of P ≤ 0.0161. Results In the PIK3CA-mutated cohort (n = 341), median OS [95% confidence interval (CI)] was 39.3 months (34.1-44.9) for alpelisib-fulvestrant and 31.4 months (26.8-41.3) for placebo-fulvestrant [hazard ratio (HR) = 0.86 (95% CI, 0.64-1.15; P = 0.15)]. OS results did not cross the prespecified efficacy boundary. Median OS (95% CI) in patients with lung and/or liver metastases was 37.2 months (28.7-43.6) and 22.8 months (19.0-26.8) in the alpelisib-fulvestrant and placebo-fulvestrant arms, respectively [HR = 0.68 (0.46-1.00)]. Median times to chemotherapy (95% CI) for the alpelisib-fulvestrant and placebo-fulvestrant arms were 23.3 months (15.2-28.4) and 14.8 months (10.5-22.6), respectively [HR = 0.72 (0.54-0.95)]. No new safety signals were observed with longer follow-up. Conclusions Although the analysis did not cross the prespecified boundary for statistical significance, there was a 7.9-month numeric improvement in median OS when alpelisib was added to fulvestrant treatment of patients with PIK3CA-mutated, HR+, HER2− ABC. Overall, these results further support the statistically significant prolongation of PFS observed with alpelisib plus fulvestrant in this population, which has a poor prognosis due to a PIK3CA mutation. ClinicalTrials.gov Id NCT02437318

LBA3_PRAlpelisib (ALP) + fulvestrant (FUL) for advanced breast cancer (ABC): Results of the phase III SOLAR-1 trial

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