The issue of compliance in a research environment in which investigators are subject to disciplinary action if they fail to ensure that patients adhere precisely to the intense monitoring mandates of a clinical trial is explored.
Regulations and ethical principles require that investigators seek consent and that patients participate in experimental studies only under circumstances that minimize the possibility of undue pressure and/or enticements. In recent years, there has been a rapid rise in the monitoring requirements of early-phase trials accompanied by an increasing emphasis on assuring “investigator” compliance with the protocol. It is actually, however, the patient who must comply with the requirements of the study. If there is divergence from the protocol, investigators may be reported to regulatory bodies or agencies. Whereas the investigative community is expected to be vigilant about ensuring that patients participate in studies voluntarily and that their consent is procured without duress, it is also required to guarantee that complex protocols, which entail multiple procedures, be followed exactly by participants who suffer from the complications of advanced cancer. We explore the issue of compliance in a research environment in which investigators are subject to disciplinary action if they fail to ensure that patients adhere precisely to the intense monitoring mandates of a clinical trial.
Compliance; Clinical trials
non-small cell lung cancer; trastuzumab; lapatinib; bevacizumab; HER2
In 31 solid tumor patients treated with the demethylating agent decitabine, we performed tumor biopsies before and after the first cycle of decitabine and used immunohistochemistry (IHC) to assess whether decitabine increased expression of various membrane transporters. Resistance to chemotherapy may arise due to promoter methylation/downregulation of expression of transporters required for drug uptake, and decitabine can reverse resistance in vitro. The endocytosis regulator RhoA, the folate carriers FOLR1 and RFC1, and the glucose transporter GLUT4 were assessed.
Pre-decitabine RhoA was higher in patients who had received their last therapy >3 months previously than in patients with more recent prior therapy (P = 0.02), and varied inversely with global DNA methylation as assessed by LINE1 methylation (r = −0.58, P = 0.006). Tumor RhoA scores increased with decitabine (P = 0.03), and RFC1 also increased in patients with pre-decitabine scores ≤150 (P = 0.004). Change in LINE1 methylation with decitabine did not correlate significantly with change in IHC scores for any transporter assessed. We also assessed methylation of the RFC1 gene (alias SLC19A1). SLC19A1 methylation correlated with tumor LINE1 methylation (r = 0.45, P = 0.02). There was a small (statistically insignificant) decrease in SLC19A1 methylation with decitabine, and there was a trend towards change in SLC19A1 methylation with decitabine correlating with change in LINE1 methylation (r = 0.47, P <0.15). While SLC19A1 methylation did not correlate with RFC1 scores, there was a trend towards an inverse correlation between change in SLC19A1 methylation and change in RFC1 expression (r = −0.45, P = 0.19).
In conclusion, after decitabine administration, there was increased expression of some (but not other) transporters that may play a role in chemotherapy uptake. Larger patient numbers will be needed to define the extent to which this increased expression is associated with changes in DNA methylation.
Decitabine; RhoA; RFC1; FOLR1; GLUT4; LINE1 methylation; Promoter methylation
During a clinical trial of the tyrosine kinase inhibitor dasatinib for advanced non–small cell lung cancer (NSCLC) one patient responded dramatically and remains cancer-free 4 years later. A comprehensive analysis of his tumor revealed a previously undescribed, kinase inactivating BRAF mutation (Y472CBRAF); no inactivating BRAF mutations were found in the non-responding tumors taken from other patients. Cells transfected with Y472CBRAF exhibited CRAF, MEK, and ERK activation – characteristics identical to signaling changes that occur with previously known kinase inactivating BRAF mutants. Dasatinib selectively induced senescence in NSCLC cells with inactivating BRAF mutations. Transfection of other NSCLC cells with these BRAF mutations also increased these cells’ dasatinib sensitivity, whereas transfection with an activating BRAF mutation led to their increased dasatinib resistance. The sensitivity induced by Y472CBRAF was reversed by the introduction of a BRAF mutation that impairs RAF dimerization. Dasatinib inhibited CRAF modestly, but concurrently induced RAF dimerization resulting in ERK activation in NSCLC cells with kinase inactivating BRAF mutations. The sensitivity of NSCLC with kinase impaired BRAF to dasatinib suggested synthetic lethality of BRAF and a dasatinib target. Inhibiting BRAF in NSCLC cells expressing wild-type BRAF likewise enhanced these cells’ dasatinib sensitivity. Thus, the patient’s BRAF mutation was likely responsible for his tumor’s marked response to dasatinib, suggesting that tumors bearing kinase impaired BRAF mutations may be exquisitely sensitive to dasatinib. Moreover, the potential synthetic lethality of combination therapy including dasatinib and BRAF inhibitors may lead to additional therapeutic options against cancers with wild-type BRAF.
The proliferation of nuclear technology in the politically volatile Middle East greatly increases the likelihood of a catastrophic nuclear war. It is widely accepted, while not openly declared, that Israel has nuclear weapons, and that Iran has enriched enough nuclear material to build them. The medical consequences of a nuclear exchange between Iran and Israel in the near future are envisioned, with a focus on the distribution of casualties in urban environments.
Model estimates of nuclear war casualties employed ESRI's ArcGIS 9.3, blast and prompt radiation were calculated using the Defense Nuclear Agency's WE program, and fallout radiation was calculated using the Defense Threat Reduction Agency's (DTRA's) Hazard Prediction and Assessment Capability (HPAC) V404SP4, as well as custom GIS and database software applications. Further development for thermal burn casualties was based on Brode, as modified by Binninger, to calculate thermal fluence. ESRI ArcGISTM programs were used to calculate affected populations from the Oak Ridge National Laboratory's LandScanTM 2007 Global Population Dataset for areas affected by thermal, blast and radiation data.
Trauma, thermal burn, and radiation casualties were thus estimated on a geographic basis for three Israeli and eighteen Iranian cities. Nuclear weapon detonations in the densely populated cities of Iran and Israel will result in an unprecedented millions of numbers of dead, with millions of injured suffering without adequate medical care, a broad base of lingering mental health issues, a devastating loss of municipal infrastructure, long-term disruption of economic, educational, and other essential social activity, and a breakdown in law and order.
This will cause a very limited medical response initially for survivors in Iran and Israel. Strategic use of surviving medical response and collaboration with international relief could be expedited by the predicted casualty distributions and locations. The consequences for health management of thermal burn and radiation patients is the worst, as burn patients require enormous resources to treat, and there will be little to no familiarity with the treatment of radiation victims. Any rational analysis of a nuclear war between Iran and Israel reveals the utterly unacceptable outcomes for either nation.
Nuclear war; Mass casualty; Radiation; Thermal burns; Trauma; Iran; Israel
Platinum resistance is a major limitation in the treatment of advanced non–small-cell lung cancer (NSCLC). Reduced intracellular drug accumulation is one of the most consistently identified features of platinum-resistant cell lines, but clinical data are limited. We assessed the effects of tissue platinum concentrations on response and survival in NSCLC.
Patients and Methods
We measured total platinum concentrations by flameless atomic absorption spectrophotometry in 44 archived fresh-frozen NSCLC specimens from patients who underwent surgical resection after neoadjuvant platinum-based chemotherapy. Tissue platinum concentration was correlated with percent reduction in tumor size on post- versus prechemotherapy computed tomography scans. The relationship between tissue platinum concentration and survival was assessed by univariate and multicovariate Cox proportional hazards regression model analysis and Kaplan-Meier analysis.
Tissue platinum concentration correlated significantly with percent reduction in tumor size (P < .001). The same correlations were seen with cisplatin, carboplatin, and all histology subgroups. Furthermore, there was no significant impact of potential variables such as number of cycles and time lapse from last chemotherapy on platinum concentration. Patients with higher platinum concentration had longer time to recurrence (P = .034), progression-free survival (P = .018), and overall survival (P = .005) in the multicovariate Cox model analysis after adjusting for number of cycles.
This clinical study established a relationship between tissue platinum concentration and response in NSCLC. It suggests that reduced platinum accumulation might be an important mechanism of platinum resistance in the clinical setting. Further studies investigating factors that modulate intracellular platinum concentration are warranted.
We adopted a two-stage study design to screen 927 single nucleotide polymorphisms (SNPs) located in 73 apoptotic-pathway genes in a case-control study and then performed a fast-track validation of the significant SNPs in a replication population to identify sequence variations in the apoptotic pathway modulating lung cancer risk. Fifty-five SNPs showed significant associations in the discovery population comprised of 661 lung cancer cases and 959 controls. Six of these SNPs located in three genes (Bcl-2, CASP9 and ANKS1B) were validated in a replication population with 1154 cases and 1373 controls. Additive model was the best-fitting model for five SNPs (rs1462129 and rs255102 of Bcl-2, rs6685648 of CASP9 and rs1549102, rs11110099 of ANKS1B) and recessive model was the best fit for one SNP (rs10745877 of ANKS1B). In the analysis of joint effects with subjects carrying no unfavorable genotypes as the reference group, those carrying one, two, and three or more unfavorable genotypes had an odds ratio (OR) of 2.22 [95% confidence interval (CI) = 1.08–4.57, P = 0.03], 2.70 (95% CI = 1.33–5.49; P = 0.006) and 4.13 (95% CI = 2.00–8.57; P = 0.0001), respectively (P for trend = 6.05E-06). The joint effect of unfavorable genotypes was also validated in the replication population. The SNPs identified are located in or near key genes known to play important roles in apoptosis regulation, supporting the strong biological relevance of our findings. Future studies are needed to identify the causal SNPs and elucidate the underlying molecular mechanisms.
Folate receptor alpha (FRα) and reduced folate carrier-1 (RFC1) regulate uptake of folate molecules inside the cell. FRα is a potential biomarker of tumors response to antifolate chemotherapy and a target for therapy using humanized monocloncal antibody. Information on the protein expression of these receptors in non–small cell lung carcinoma (NSCLC) is limited.
Material and Methods
Expressions of FRα and RFC1 were examined by IHC in 320 surgically resected NSCLC (202 adenocarcinomas and 118 squamous cell carcinomas) tissue specimens and correlated with patients’ clinicopathologic characteristics. FOLR1 mRNA expression was examined using publicly available microarray datasets. FRα expression was correlated with thymidylate synthase (TS) and p53 expression in NSCLCs, and with EGFR and KRAS mutations in adenocarcinomas.
NSCLC overexpressed FRα and RFC1. In a multivariate analysis, lung adenocarcinomas were more likely to express FRα in the cytoplasm (odds ratio [OR] = 4.39; P<0.0001) and membrane (OR = 5.34; P<0.0001) of malignant cells than squamous cell carcinomas. Tumors from never-smokers were more likely to express cytoplasmic (OR = 3.35; P<0.03) and membrane (OR = 3.60; P=0.0005) FRα than those from smokers. In adenocarcinoma, EGFR mutations correlated with higher expression of membrane FRα and FOLR1 gene expressions. High levels of FRα expression was detected in 42 NSCLC advanced metastatic tumor tissues.
FRα and RFC1 proteins are overexpressed in NSCLC tumor tissues. The high levels of FRα in lung adenocarcinomas may be associated to these tumors’ better responses to antifolate chemotherapy and represents a potential novel target for this tumor type.
non–small cell lung carcinoma; EGFR; membrane transporter; FRα; FRC1
For processes that follow first order kinetics, exponential decay nonlinear regression analysis (EDNRA) may delineate curve characteristics and suggest processes affecting curve shape. We conducted a preliminary feasibility assessment of EDNRA of patient survival curves.
EDNRA was performed on Kaplan-Meier overall survival (OS) and time-to-relapse (TTR) curves for 323 patients with resected NSCLC and on OS and progression-free survival (PFS) curves from selected publications.
Results and Conclusions
In our resected patients, TTR curves were triphasic with a “cured” fraction of 60.7% (half-life [t1/2] >100,000 months), a rapidly-relapsing group (7.4%, t1/2=5.9 months) and a slowly-relapsing group (31.9%, t1/2=23.6 months). OS was uniphasic (t1/2=74.3 months), suggesting an impact of co-morbidities; hence, tumor molecular characteristics would more likely predict TTR than OS. Of 172 published curves analyzed, 72 (42%) were uniphasic, 92 (53%) were biphasic, 8 (5%) were triphasic. With first-line chemotherapy in advanced NSCLC, 87.5% of curves from 2-3 drug regimens were uniphasic vs only 20% of those with best supportive care or 1 drug (p<0.001). 54% of curves from 2-3 drug regimens had convex rapid-decay phases vs 0% with fewer agents (p<0.001). Curve convexities suggest that discontinuing chemotherapy after 3-6 cycles “synchronizes” patient progression and death. With postoperative adjuvant chemotherapy, the PFS rapid-decay phase accounted for a smaller proportion of the population than in controls (p=0.02) with no significant difference in rapid-decay t1/2, suggesting adjuvant chemotherapy may move a subpopulation of patients with sensitive tumors from the relapsing group to the cured group, with minimal impact on time to relapse for a larger group of patients with resistant tumors. In untreated patients, the proportion of patients in the rapid-decay phase increased (p=0.04) while rapid-decay t1/2 decreased (p=0.0004) with increasing stage, suggesting that higher stage may be associated with tumor cells that both grow more rapidly and have a higher probability of surviving metastatic processes than in early stage tumors.
This preliminary assessment of EDNRA suggests that it may be worth exploring this approach further using more sophisticated, statistically rigorous nonlinear modelling approaches. Using such approaches to supplement standard survival analyses could suggest or support specific testable hypotheses.
exponential decay; nonlinear regression analysis; non-small cell lung cancer Running Title: Nonlinear regression of NSCLC patient survival time
Some studies (but not others) suggested high doses are beneficial in small cell lung cancer (SCLC). We hypothesized dose-response curve (DRC) shape reflects resistance mechanisms.
We reviewed published SCLC clinical trialss and converted response rates into estimated mean tumor cell kill, assuming killing is proportional to reduction in tumor volume. Mean % cell survival was plotted vs planned dose-intensity. Nonlinear and linear meta-regression analyses (weighted according to the number of patients in each study) were used to assess DRC characteristics.
Although associations between dose and cell survival were not statistically significant, DRCs sloped downward for 5 of 7 agents across all doses and for all 7 when lowest doses were excluded. Maximum mean cell kill across all drugs and doses was approximately 90%, suggesting there may be a maximum achievable tumor cell kill irrespective of number of agents or drug doses.
Downward DRC slopes suggest that maintaining relatively high doses may possibly maximize palliation, although the associations between dose and slope did not achieve statistical significance, and slopes for most drugs tended to be shallow. DRC flattening at higher doses would preclude cure, and would suggest that “saturable passive resistance” (deficiency of factors required for cell killing) limits maximum achievable cell kill. An example of factors that could flatten the dose-response curve at higher doses and lead to saturable passive resistance would be presence of quiescent, non-cycling cells.
small cell; dose-response; resistance; quiescence
While chemotherapy provides useful palliation, advanced lung cancer remains incurable since those tumors that are initially sensitive to therapy rapidly develop acquired resistance. Resistance may arise from impaired drug delivery, extracellular factors, decreased drug uptake into tumor cells, increased drug efflux, drug inactivation by detoxifying factors, decreased drug activation or binding to target, altered target, increased damage repair, tolerance of damage, decreased proapoptotic factors, increased antiapoptotic factors, or altered cell cycling or transcription factors. Factors for which there is now substantial clinical evidence of a link to small cell lung cancer (SCLC) resistance to chemotherapy include MRP (for platinum-based combination chemotherapy) and MDR1/P-gp (for non-platinum agents). SPECT MIBI and Tc-TF scanning appears to predict chemotherapy benefit in SCLC. In non-small cell lung cancer (NSCLC), the strongest clinical evidence is for taxane resistance with elevated expression or mutation of class III β-tubulin (and possibly α tubulin), platinum resistance and expression of ERCC1 or BCRP, gemcitabine resistance and RRM1 expression, and resistance to several agents and COX-2 expression (although COX-2 inhibitors have had minimal impact on drug efficacy clinically). Tumors expressing high BRCA1 may have increased resistance to platinums but increased sensitivity to taxanes. Limited early clinical data suggest that chemotherapy resistance in NSCLC may also be increased with decreased expression of cyclin B1 or of Eg5, or with increased expression of ICAM, matrilysin, osteopontin, DDH, survivin, PCDGF, caveolin-1, p21WAF1/CIP1, or 14-3-3sigma, and that IGF-1R inhibitors may increase efficacy of chemotherapy, particularly in squamous cell carcinomas. Equivocal data (with some positive studies but other negative studies) suggest that NSCLC tumors with some EGFR mutations may have increased sensitivity to chemotherapy, while K-ras mutations and expression of GST-pi, RB or p27kip1 may possibly confer resistance. While limited clinical data suggest that p53 mutations are associated with resistance to platinum-based therapies in NSCLC, data on p53 IHC positivity are equivocal. To date, resistance-modulating strategies have generally not proven clinically useful in lung cancer, although small randomized trials suggest a modest benefit of verapamil and related agents in NSCLC.
lung cancer; chemotherapy; resistance
Platinum-based regimens are the standard chemotherapy for patients with advanced non–small-cell lung cancer (NSCLC). DNA repair capacity (DRC) in tumor cells plays an important role in resistance to platinum-based drugs. We have previously reported that efficient DRC, as assessed by an in vitro lymphocyte-based assay, was a determinant of poor survival in patients with NSCLC in a relatively small data set. In this larger independent study of 591 patients with NSCLC, we further evaluated whether DRC in peripheral lymphocytes predicts survival of patients with NSCLC who receive platinum-based chemotherapy.
Patients and Methods
All patients were recruited at The University of Texas MD Anderson Cancer Center and donated blood samples before the start of any chemotherapy. We measured DRC in cultured T lymphocytes by using the host-cell reactivation assay, and we assessed associations between DRC in peripheral lymphocytes and survival of patients with NSCLC who were treated with first-line platinum-based chemotherapy.
We found an inverse association between DRC in peripheral lymphocytes and patient survival. Compared with patients in the low tertile of DRC, patients with NSCLC in the high tertile of DRC had significantly worse overall and 3-year survival (adjusted hazard ratio [HR], 1.33; 95% CI, 1.04 to 1.71; P = .023; and HR, 1.35; 95% CI, 1.04 to 1.76; P = .025, respectively). This trend was more pronounced in patients with early-stage tumors, adenocarcinoma, or squamous cell carcinoma.
We confirmed that DRC in peripheral lymphocytes is an independent predictor of survival for patients with NSCLC treated with platinum-based chemotherapy.
ATM gene mutations have been implicated in many human cancers. However, the role of ATM polymorphisms in lung carcinogenesis is largely unexplored. We conducted a case-control analysis of 556 Caucasian non-small-cell lung cancer (NSCLC) patients and 556 controls frequency-matched on age, gender and smoking status. We genotyped 11 single nucleotide polymorphisms of the ATM gene and found that compared with the wild-type allele-containing genotypes, the homozygous variant genotypes of ATM08 (rs227060) and ATM10 (rs170548) were associated with elevated NSCLC risk with ORs of 1.55 (95% CI: 1.02–2.35) and 1.51 (0.99–2.31), respectively. ATM haplotypes and diplotypes were inferred using the Expectation-Maximization algorithm. Haplotype H5 was significantly associated with reduced NSCLC risk in former smokers with an OR of 0.47 (0.25–0.96) compared with the common H1 haplotype. Compared with the H1–H2 diplotype, H2–H2 and H3–H4 diplotypes were associated with increased NSCLC risk with ORs of 1.58 (0.99–2.54) and 2.29 (1.05–5.00), respectively. We then evaluated genotype–phenotype correlation in the control group using the comet assay to determine DNA damage and DNA repair capacity. Compared with individuals with at least 1 wild-type allele, the homozygous variant carriers of either ATM08 or ATM10 exhibited significantly increased DNA damage as evidenced by a higher mean value of the radiation-induced olive tail moment (ATM08: 4.86 ± 2.43 vs. 3.79 ± 1.51, p = 0.04; ATM10: 5.14 ± 2.37 vs. 3.79 ± 1.54, p = 0.01). Our study presents the first epidemiologic evidence that ATM genetic variants may affect NSCLC predisposition, and that the risk-conferring variants might act through down-regulating the functions of ATM in DNA repair activity upon genetic insults such as ionizing radiation.
ATM; polymorphism; haplotype; diplotype; NSCLC
We evaluated the ability of histopathologic response criteria to predict overall survival (OS) and disease-free survival (DFS) in patients with surgically resected non-small cell lung cancer (NSCLC) treated with or without neoadjuvant chemotherapy.
Tissue specimens from 358 patients with NSCLC were evaluated by pathologists blinded to the patient treatment and outcome. The surgical specimens were reviewed for various histopathologic features in the tumor including percentage of residual viable tumor cells, necrosis, and fibrosis. The relationship between the histopathologic findings and OS was assessed.
The percentage of residual viable tumor cells and surgical pathologic stage were associated with OS and DFS in 192 patients with NSCLC receiving neoadjuvant chemotherapy in multivariate analysis (p = 0.005 and p = 0.01, respectively). There was no association of OS or DFS with percentage of viable tumor cells in 166 patients with NSCLC who did not receive neoadjuvant chemotherapy (p = 0.31 and p = 0.45, respectively). Long-term OS and DFS were significantly prolonged in patients who had ≤10% viable tumor compared with patients with >10% viable tumor cells (5 years OS, 85% versus 40%, p < 0.0001 and 5 years DFS, 78% versus 35%, p < 0.001).
The percentages of residual viable tumor cells predict OS and DFS in patients with resected NSCLC after neoadjuvant chemotherapy even when controlled for pathologic stage. Histopathologic assessment of resected specimens after neoadjuvant chemotherapy could potentially have a role in addition to pathologic stage in assessing prognosis, chemotherapy response, and the need for additional adjuvant therapies.
Lung cancer; Neoadjuvant chemotherapy; Histopathology
The magnitude of benefit is variable for advanced non-small cell lung cancer (NSCLC) patients receiving platinum-based chemotherapy. The purpose of this study is to determine whether genetic variations in the transforming growth factor-beta (TGF-β) pathway are associated with clinical outcomes in NSCLC patients receiving first-line platinum-based chemotherapy. Five hundred and ninety-eight advanced-stage NSCLC patients who received first-line platinum-based chemotherapy with or without radiotherapy were recruited at the MD Anderson Cancer Center between 1995 and 2007. DNA from blood was genotyped for 227 single nucleotide polymorphisms (SNPs) in 23 TGF-β pathway-related genes to evaluate their associations with overall survival. In individual SNP analysis, 22 variants were significantly associated with overall survival, of which the strongest associations were found for BMP2:rs235756 [hazard ratio (HR) = 1.45; 95% confidence interval (CI), 1.11–1.90] and SMAD3:rs4776342 (HR = 1.25; 95% CI, 1.06–1.47). Fifteen and 18 genetic loci displayed treatment-specific associations for chemotherapy and chemoradiation, respectively, identifying a majority of the cases who would be predicted to respond favorably to a specific treatment regimen. BMP2:rs235753 and a haplotype in SMAD3 were associated with overall survival for both treatment modalities. Cumulative effect analysis showed that multiple risk genotypes had a significant dose-dependent effect on overall survival (Ptrend = 2.44 x 10−15). Survival tree analysis identified subgroups of patients with dramatically different median survival times of 45.39 versus 13.55 months and 18.02 versus 5.89 months for high- and low- risk populations when treated with chemoradiation and chemotherapy, respectively. These results suggest that genetic variations in the TGF-β pathway are potential predictors of overall survival in NSCLC patients treated with platinum-based chemotherapy with or without radiation.
Interindividual variation in genetic background may influence the response to chemotherapy and overall survival for patients with advanced-stage non–small cell lung cancer (NSCLC).
To identify genetic variants associated with poor overall survival in these patients, we conducted a genome-wide scan of 307 260 single-nucleotide polymorphisms (SNPs) in 327 advanced-stage NSCLC patients who received platinum-based chemotherapy with or without radiation at the University of Texas MD Anderson Cancer Center (the discovery population). A fast-track replication was performed for 315 patients from the Mayo Clinic followed by a second validation at the University of Pittsburgh in 420 patients enrolled in the Spanish Lung Cancer Group PLATAX clinical trial. A pooled analysis combining the Mayo Clinic and PLATAX populations or all three populations was also used to validate the results. We assessed the association of each SNP with overall survival by multivariable Cox proportional hazard regression analysis. All statistical tests were two-sided.
SNP rs1878022 in the chemokine-like receptor 1 (CMKLR1) was statistically significantly associated with poor overall survival in the MD Anderson discovery population (hazard ratio [HR] of death = 1.59, 95% confidence interval [CI] = 1.32 to 1.92, P = 1.42 × 10−6), in the PLATAX clinical trial (HR of death = 1.23, 95% CI = 1.00 to 1.51, P = .05), in the pooled Mayo Clinic and PLATAX validation (HR of death = 1.22, 95% CI = 1.06 to 1.40, P = .005), and in pooled analysis of all three populations (HR of death = 1.33, 95% CI = 1.19 to 1.48, P = 5.13 × 10−7). Carrying a variant genotype of rs10937823 was associated with decreased overall survival (HR of death = 1.82, 95% CI = 1.42 to 2.33, P = 1.73 × 10−6) in the pooled MD Anderson and Mayo Clinic populations but not in the PLATAX trial patient population (HR of death = 0.96, 95% CI = 0.69 to 1.35).
These results have the potential to contribute to the future development of personalized chemotherapy treatments for individual NSCLC patients.
Tumor suppressor gene TUSC2/FUS1 (TUSC2) is frequently inactivated early in lung cancer development. TUSC2 mediates apoptosis in cancer cells but not normal cells by upregulation of the intrinsic apoptotic pathway. No drug strategies currently exist targeting loss-of–function genetic abnormalities. We report the first in-human systemic gene therapy clinical trial of tumor suppressor gene TUSC2.
Patients with recurrent and/or metastatic lung cancer previously treated with platinum-based chemotherapy were treated with escalating doses of intravenous N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTAP):cholesterol nanoparticles encapsulating a TUSC2 expression plasmid (DOTAP:chol-TUSC2) every 3 weeks.
Thirty-one patients were treated at 6 dose levels (range 0.01 to 0.09 milligrams per kilogram). The MTD was determined to be 0.06 mg/kg. Five patients achieved stable disease (2.6–10.8 months, including 2 minor responses). One patient had a metabolic response on positron emission tomography (PET) imaging. RT-PCR analysis detected TUSC2 plasmid expression in 7 of 8 post-treatment tumor specimens but not in pretreatment specimens and peripheral blood lymphocyte controls. Proximity ligation assay, performed on paired biopsies from 3 patients, demonstrated low background TUSC2 protein staining in pretreatment tissues compared with intense (10–25 fold increase) TUSC2 protein staining in post-treatment tissues. RT-PCR gene expression profiling analysis of apoptotic pathway genes in two patients with high post-treatment levels of TUSC2 mRNA and protein showed significant post-treatment changes in the intrinsic apoptotic pathway. Twenty-nine genes of the 82 tested in the apoptosis array were identified by Igenuity Pathway Analysis to be significantly altered post-treatment in both patients (Pearson correlation coefficient 0.519; p<0.01).
DOTAP:chol-TUSC2 can be safely administered intravenously in lung cancer patients and results in uptake of the gene by human primary and metastatic tumors, transgene and gene product expression, specific alterations in TUSC2-regulated pathways, and anti-tumor effects (to our knowledge for the first time for systemic DOTAP:cholesterol nanoparticle gene therapy).
Non-small cell lung cancer (NSCLC) is still the leading cause of cancer-related deaths. The effect of the PI3K/PTEN/AKT/mTOR signaling pathway on cancer treatment, including NSCLC, has been well documented. In this study, we analyzed associations between genetic variations within this pathway and clinical outcomes following platinum-based chemotherapy in 168 patients with stage IIIB (wet) or stage IV NSCLC. Sixteen tagging SNPs in five core genes (PIK3CA, PTEN, AKT1, AKT2, and FRAP1) of this pathway and identified SNPs associated with development of toxicity and disease progression. We observed significantly increased toxicity for patients with PIK3CA:rs2699887 (OR: 3.86, 95% CI: 1.08 – 13.82). In contrast, a SNP in PTEN was associated with significantly reduced risk for chemotherapeutic toxicity (OR: 0.44, 95% CI: 0.20 - 0.95). We identified three SNPs in AKT1 resulting in significantly decreased risks of distant progression in patients carrying at least one variant allele with HRs of 0.66 (95% CI: 0.45 - 0.97), 0.52 (95% CI: 0.35 - 0.77), and 0.62 (95% CI: 0.42 - 0.91) for rs3803304, rs2498804, and rs1130214, respectively. Furthermore, these same variants conferred nearly two-fold increased progression-free survival times. The current study provides evidence that genetic variations within the PI3K/PTEN/AKT/mTOR signaling pathway are associated with variation in clinical outcomes of NSCLC patients. With further validation, our findings may provide additional biomarkers for customized treatment of platinum-based chemotherapy for NSCLC.
lung cancer; chemotherapy; platinum-agents; AKT; clinical outcomes
Src family kinases (SFKs) promote cancer progression and are commonly expressed in non–small-cell lung cancer (NSCLC), but the clinical effects of SFK inhibition in NSCLC are unknown. We conducted a phase II trial of the SFK inhibitor dasatinib for advanced NSCLC. We tested the hypotheses that the activation of epidermal growth factor receptor (EGFR) or SFK or modulation of serum cytokines may predict a response to dasatinib.
Patients and Methods
Patients received dasatinib as first-line therapy. Response was measured by tumor size on computed tomography scans and by metabolic activity on positron emission tomography scans. Tissue samples taken before patients received dasatinib were tested for EGFR and Kras mutation and phosphorylated SFK expression.
Thirty-four patients were enrolled. The overall disease control rate (partial responses plus stable disease) for dasatinib was 43%. One patient had a partial response to therapy. Eleven patients (32%) had a metabolic response to dasatinib. SFK activation and EGFR and Kras mutations in tumor tissue did not predict response to dasatinib. Significant toxicities included fatigue and dyspnea. The presence of a pleural effusion before dasatanib therapy predicted the development of a clinically significant effusion during therapy.
Dasatinib as a single agent had modest clinical activity that was lower than that generally observed in patients with NSCLC who receive chemotherapy. Pleural effusion was an expected and problematic toxicity that was successfully treated with steroids, diuretics, and dose interruptions. Marked activity in one patient and prolonged stable disease in four others suggested a potential subpopulation of patients with dasatinib-sensitive NSCLC.
To understand the role of Nrf2 and Keap1 in NSCLC, we studied their expression in a large series of tumors with annotated clinicopathologic data, including response to platinum-based adjuvant chemotherapy.
We determined the immunohistochemical expression of nuclear Nrf2 and cytoplasmic Keap1 in 304 NSCLCs and its association with patients’ clinicopathologic characteristics, and in 89 tumors from patients who received neoadjuvant (n=26) or adjuvant platinum-based chemotherapy (n=63). We evaluated NFE2L2 and KEAP1 mutations in 31 tumor specimens.
We detected nuclear Nrf2 expression in 26% of NSCLCs; it was significantly more common in squamous cell carcinomas (38%) than in adenocarcinomas (18%; P<0.0001). Low or absent Keap1 expression was detected in 56% of NSCLCs; it was significantly more common in adenocarcinomas (62%) than in squamous cell carcinomas (46%; P=0.0057). In NSCLC, mutations of NFE2L2 and KEAP1 were very uncommon (2 of 29 and 1 of 31 cases, respectively). In multivariate analysis, Nrf2 expression was associated with worse overall survival (P=0.0139; HR=1.75) in NSCLC patients, and low or absent Keap1 expression was associated with worse overall survival (P=0.0181; HR=2.09) in squamous cell carcinoma. In univariate analysis, nuclear Nrf2 expression was associated with worse recurrence-free survival in squamous cell carcinoma patients who received adjuvant treatment (P=0.0410; HR=3.37).
Increased expression of Nrf2 and decreased expression of Keap1 are common abnormalities in NSCLC and are associated with a poor outcome. Nuclear expression of Nrf2 in malignant lung cancer cells may play a role in resistance to platinum-based treatment in squamous cell carcinoma.
Nrf2; Keap1; NSCLC
Treatment of non-small cell lung cancer (NSCLC) with radiotherapy or chemoradiotherapy is often accompanied by the development of esophagitis and pneumonitis. Identifying patients who might be at increased risk for normal tissue toxicity would help in determination of the optimal radiation dose to avoid these events. We profiled 59 single nucleotide polymorphisms (SNPs) from 37 inflammation-related genes in 173 NSCLC patients with stage IIIA/IIIB (dry) disease who were treated with definitive radiation or chemoradiation. For esophagitis risk, nine SNPs were associated with a 1.5- to 4-fold increase in risk, including three PTGS2 (COX2) variants: rs20417 (HR:1.93, 95% CI:1.10–3.39), rs5275 (HR:1.58, 95% CI:1.09–2.27), and rs689470 (HR:3.38, 95% CI:1.09–10.49). Significantly increased risk of pneumonitis was observed for patients with genetic variation in the proinflammatory genes IL1A, IL8, TNF, TNFRSF1B, and MIF. In contrast, NOS3:rs1799983 displayed a protective effect with a 45% reduction in pneumonitis risk (HR:0.55, 95% CI:0.31–0.96). Pneumonitis risk was also modulated by polymorphisms in anti-inflammatory genes, including genetic variation in IL13. rs20541 and rs180925 each resulted in increased risk (HR:2.95, 95% CI:1.14–7.63 and HR:3.23, 95% CI:1.03–10.18, respectively). The cumulative effect of these SNPs on risk was dose-dependent, as evidenced by a significantly increased risk of either toxicity with an increasing number of risk genotypes (P<0.001). These results suggest that genetic variations among inflammation pathway genes may modulate the development of radiation-induced toxicity and, ultimately, help in identifying patients who are at an increased likelihood for such events.
Deciduous molars, extracted from a total of 310 children aged between 3 and 10 years who resided in urban, suburban, and rural environments in Northern Ireland, were analysed for lead by atomic absorption spectrophotometry.
Tooth lead content increased with age and was influenced by the nature of the donor's environment. Lead concentrations were greatest in the teeth of children in the urban group and least in the rural group, with suburban specimens intermediate.
Deciduous teeth, available in quantity due to the high prevalence of dental caries, can conveniently be used as indicators of the past exposure of children to lead contamination. The analytical method is suitable for use on a sufficient scale to enable comparisons to be made between communities.
Mouse knockout technology provides a powerful means of elucidating gene function in vivo, and a publicly available genome-wide collection of mouse knockouts would be significantly enabling for biomedical discovery. To date, published knockouts exist for only about 10% of mouse genes. Furthermore, many of these are limited in utility because they have not been made or phenotyped in standardized ways, and many are not freely available to researchers. It is time to harness new technologies and efficiencies of production to mount a high-throughput international effort to produce and phenotype knockouts for all mouse genes, and place these resources into the public domain.