Therapeutic targeting of Chk1 in NSCLC stem cells during chemotherapy

Cancer stem cell (SC) chemoresistance may be responsible for the poor clinical outcome of non-small-cell lung cancer (NSCLC) patients. In order to identify the molecular events that contribute to NSCLC chemoresistance, we investigated the DNA damage response in SCs derived from NSCLC patients. We found that after exposure to chemotherapeutic drugs NSCLC-SCs undergo cell cycle arrest, thus allowing DNA damage repair and subsequent cell survival. Activation of the DNA damage checkpoint protein kinase (Chk) 1 was the earliest and most significant event detected in NSCLC-SCs treated with chemotherapy, independently of their p53 status. In contrast, a weak Chk1 activation was found in differentiated NSCLC cells, corresponding to an increased sensitivity to chemotherapeutic drugs as compared with their undifferentiated counterparts. The use of Chk1 inhibitors in combination with chemotherapy dramatically reduced NSCLC-SC survival in vitro by inducing premature cell cycle progression and mitotic catastrophe. Consistently, the co-administration of the Chk1 inhibitor AZD7762 and chemotherapy abrogated tumor growth in vivo, whereas chemotherapy alone was scarcely effective. Such increased efficacy in the combined use of Chk1 inhibitors and chemotherapy was associated with a significant reduction of NSCLC-SCs in mouse xenografts. Taken together, these observations support the clinical evaluation of Chk1 inhibitors in combination with chemotherapy for a more effective treatment of NSCLC. \uc2\ua9 2012 Macmillan Publishers Limited. All rights reserved

The Hippo transducer TAZ as a biomarker of pathological complete response in HER2-positive breast cancer patients treated with trastuzumab-based neoadjuvant therapy

Activation of the Hippo transducer TAZ is emerging as a novel oncogenic route in breast cancer and it has been associated with breast cancer stem cells. Additionally, TAZ expression has been linked with HER-2 positivity. We investigated the association between TAZ expression and pathological complete response in HER2-positive breast cancer patients treated with trastuzumab-based neoadjuvant therapy.TAZ was assessed in diagnostic core biopsies by immunohistochemistry. To categorize samples with low TAZ and samples with high TAZ we generated a score by combining staining intensity and cellular localization. The pathological complete response rate was 78.6% in patients with low TAZ tumors and 57.6% in patients with high TAZ tumors (p=0.082). In HER2-enriched tumors there was no signi cant association between TAZ and pathological complete response, whereas in the luminal B subtype the pathological complete response rate was 82.4% in tumors with low TAZ and 44.4% in tumors with high TAZ (p=0.035). This association remained statistically signi cant when restricting our analysis to triple-positive tumors with expression of both estrogen receptor and progesterone receptor 65 50% (p=0.035). Results from this exploratory study suggest that the TAZ score ef ciently predicts pathological complete response in Luminal B, HER2-positive breast cancer patients who received neoadjuvant chemotherapy and trastuzumab

Body mass index modifies the relationship between \u3b3-H2AX, a DNA damage biomarker, and pathological complete response in triple-negative breast cancer

Background: Body mass index (BMI) is largely investigated as a prognostic and predictive factor in triple-negative breast cancer (TNBC). Overweight and obesity are linked to a variety of pathways regulating tumor-promoting functions, including the DNA damage response (DDR). The DDR physiologically safeguards genome integrity but, in a neoplastic background, it is aberrantly engaged and protects cancer cells from chemotherapy. We herein verified the role of BMI on a previously assessed association between DDR biomarkers and pathological complete response (pCR) in TNBC patients treated with neoadjuvant chemotherapy (NACT). Methods: In this retrospective analysis 54 TNBC patients treated with NACT were included. The relationship between DDR biomarkers, namely phosphorylated H2A Histone Family Member X (\u3b3-H2AX) and phosphorylated checkpoint kinase 1 (pChk1), and pCR was reconsidered in light of BMI data. The Pearson\u2019s Chi-squared test of independence (2-tailed) and the Fisher Exact test were employed to assess the relationship between clinical-molecular variables and pCR. Uni- and multivariate logistic regression models were used to identify variables impacting pCR. Internal validation was carried out. Results: We observed a significant association between elevated levels of the two DDR biomarkers and pCR in patients with BMI < 25 (p = 0.009 and p = 0.022 for \u3b3-H2AX and pChk1, respectively), but not in their heavier counterpart. Results regarding \u3b3-H2AX were confirmed in uni- and multivariate models and, again, for leaner patients only (\u3b3-H2AXhigh vs \u3b3-H2AXlow: OR 10.83, 95% CI: 1.79\u201365.55, p = 0.009). The consistency of this finding was confirmed upon internal validation. Conclusions: The predictive significance of \u3b3-H2AX varies according to BMI status. Indeed, elevated levels of \u3b3-H2AX seemed associated with lower pCR rate only in leaner patients, whereas differences in pCR rate according to \u3b3-H2AX levels were not appreciable in heavier patients. Larger investigations are warranted concerning the potential role of BMI as effect modifier of the relationship between DDR-related biomarkers and clinical outcomes in TNBC

CHK1-targeted therapy to deplete DNA replication- stressed, p53-deficient, hyperdiploid colorectal cancer stem cells

Objective Cancer stem cells (CSCs) are responsible for tumour formation and spreading, and their targeting is required for tumour eradication. There are limited therapeutic options for advanced colorectal cancer (CRC), particularly for tumours carrying RAS-activating mutations. The aim of this study was to identify novel CSC-targeting strategies. Design To discover potential therapeutics to be clinically investigated as single agent, we performed a screening with a panel of FDA-approved or investigational drugs on primary CRC cells enriched for CSCs (CRC-SCs) isolated from 27 patients. Candidate predictive biomarkers of efficacy were identified by integrating genomic, reverse-phase protein microarray (RPPA) and cytogenetic analyses, and validated by immunostainings. DNA replication stress (RS) was increased by employing DNA replication-perturbing or polyploidising agents. Results The drug-library screening led to the identification of LY2606368 as a potent anti-CSC agent acting in vitro and in vivo in tumour cells from a considerable number of patients ( 3c36%). By inhibiting checkpoint kinase (CHK)1, LY2606368 affected DNA replication in most CRC-SCs, including RAS-mutated ones, forcing them into premature, lethal mitoses. Parallel genomic, RPPA and cytogenetic analyses indicated that CRC-SCs sensitive to LY2606368 displayed signs of ongoing RS response, including the phosphorylation of RPA32 and ataxia telangiectasia mutated serine/threonine kinase (ATM). This was associated with mutation(s) in TP53 and hyperdiploidy, and made these CRC-SCs exquisitely dependent on CHK1 function. Accordingly, experimental increase of RS sensitised resistant CRC-SCs to LY2606368. Conclusions LY2606368 selectively eliminates replication-stressed, p53-deficient and hyperdiploid CRC-SCs independently of RAS mutational status. These results provide a strong rationale for biomarker-driven clinical trials with LY2606368 in patients with CRC

\u201cTriple positive\u201d early breast cancer: an observational multicenter retrospective analysis of outcome

We recently found that trastuzumab benefit may be lower in a small subset of early breast cancer (BC) patients (pts) with tumors expressing high levels of both hormonal receptors (HRs), i.e. triple positive (TP). To better investigate the role of HRs in HER2 positive BC, we retrospectively identified 872 TP BC pts treated with adjuvant chemotherapy alone (cohort A-366 pts), or plus trastuzumab (cohort B-506 pts). Relapse-free-survival (RFS) and breast-cancer-specific-survival (BCSS) were evaluated. Trastuzumab improved RFS and BCSS in all the subsets analyzed, but the effect on BCSS in tumors expressing both HRs in >30% of cells (TP30), and even on RFS in tumors with both HRs expressed in >50% of cells (TP50) was not significant. Distinct patterns of relapse were observed in TP50 and no-TP50 tumors, the former showing low and constant risk in the first 5 years, a late increase beyond 5 years and modest trastuzumab effect. Trastuzumab effect tended to disappear in pts whose tumors expressed ER in >50% of cells. Multivariate analysis of RFS confirmed a significant interaction between trastuzumab and ER expression, with benefit confined to pts whose tumors expressed ER in 6450% of cells. Our data suggest that the pattern of relapse of TP tumors with high HRs is similar to that of "luminal", HER2 negative tumors, without clear benefit from adjuvant trastuzumab, which remains the standard treatment even in TP tumors. Confirmatory findings on the extent to which quantitative expression of HRs may impact clinical behavior of HER2 positive BC are warranted

Association of kidney disease measures with risk of renal function worsening in patients with type 1 diabetes

Background: Albuminuria has been classically considered a marker of kidney damage progression in diabetic patients and it is routinely assessed to monitor kidney function. However, the role of a mild GFR reduction on the development of stage 653 CKD has been less explored in type 1 diabetes mellitus (T1DM) patients. Aim of the present study was to evaluate the prognostic role of kidney disease measures, namely albuminuria and reduced GFR, on the development of stage 653 CKD in a large cohort of patients affected by T1DM. Methods: A total of 4284 patients affected by T1DM followed-up at 76 diabetes centers participating to the Italian Association of Clinical Diabetologists (Associazione Medici Diabetologi, AMD) initiative constitutes the study population. Urinary albumin excretion (ACR) and estimated GFR (eGFR) were retrieved and analyzed. The incidence of stage 653 CKD (eGFR < 60 mL/min/1.73 m2) or eGFR reduction > 30% from baseline was evaluated. Results: The mean estimated GFR was 98 \ub1 17 mL/min/1.73m2 and the proportion of patients with albuminuria was 15.3% (n = 654) at baseline. About 8% (n = 337) of patients developed one of the two renal endpoints during the 4-year follow-up period. Age, albuminuria (micro or macro) and baseline eGFR < 90 ml/min/m2 were independent risk factors for stage 653 CKD and renal function worsening. When compared to patients with eGFR > 90 ml/min/1.73m2 and normoalbuminuria, those with albuminuria at baseline had a 1.69 greater risk of reaching stage 3 CKD, while patients with mild eGFR reduction (i.e. eGFR between 90 and 60 mL/min/1.73 m2) show a 3.81 greater risk that rose to 8.24 for those patients with albuminuria and mild eGFR reduction at baseline. Conclusions: Albuminuria and eGFR reduction represent independent risk factors for incident stage 653 CKD in T1DM patients. The simultaneous occurrence of reduced eGFR and albuminuria have a synergistic effect on renal function worsening

No evidence of association between prothrombotic gene polymorphisms and the development of acute myocardial infarction at a young age

Background : we investigated the association between 9 polymorphisms of genes encoding hemostasis factors and myocardial infarction in a large sample of young patients chosen because they have less coronary atherosclerosis than older patients, and thus their disease is more likely to be related to a genetic predisposition to a prothrombotic state Methods and Results : this nationwide case-control study involved 1210 patients who had survived a first myocardial infarction at an age of 45 years who underwent coronary arteriography in 125 coronary care units and 1210 healthy subjects matched for age, sex, and geographical origin. None of the 9 polymorphisms of genes encoding proteins involved in coagulation (G-455A -fibrinogen: OR, 1.0; CI, 0.8 to 1.2; G1691A factor V: OR, 1.1; CI, 0.6 to 2.1; G20210A factor II: OR, 1.0; CI, 0.5 to 1.9; and G10976A factor VII: OR, 1.0; CI, 0.8 to 1.3), platelet function (C807T glycoprotein Ia: OR, 1.1; CI, 0.9 to 1.3; and C1565T glycoprotein IIIa: OR, 0.9; CI, 0.8 to 1.2), fibrinolysis (G185T factor XIII: OR, 1.2; CI, 0.9 to 1.6; and 4G/5G plasminogen activator inhibitor type 1: OR, 0.9; CI, 0.7 to 1.2), or homocysteine metabolism (C677T methylenetetrahydrofolate reductase: OR, 0.9; CI, 0.8 to 1.1) were associated with an increased or decreased risk of myocardial infarction Conclusions : this study provides no evidence supporting an association between 9 polymorphisms of genes encoding proteins involved in hemostasis and the occurrence of premature myocardial infarction or protection against it