John S. Yordy

Phase II Trial of Cetuximab, Gemcitabine, and Oxaliplatin Followed by Chemoradiation With Cetuximab for Locally Advanced (T4) Pancreatic Adenocarcinoma: Correlation of Smad4(Dpc4) Immunostaining With Pattern of Disease Progression

PURPOSE This phase II trial was designed to assess the efficacy and safety of cetuximab, gemcitabine, and oxaliplatin followed by cetuximab, capecitabine, and radiation therapy in locally advanced pancreatic cancer (LAPC). PATIENTS AND METHODS Treatment-naive eligible patients (n = 69) received intravenous gemcitabine (1,000 mg/m(2)) and oxaliplatin (100 mg/m(2)) every 2 weeks for four doses, followed by radiation (50.4 Gy to the gross tumor only) with concurrent capecitabine (825 mg/m(2) twice daily on radiation treatment days). Cetuximab (500 mg/m(2)) was started on day 1 of chemotherapy and was continued every 2 weeks during chemotherapy and chemoradiotherapy. Diagnostic cytology specimens were immunostained for Smad4(Dpc4) expression. RESULTS Median overall survival time was 19.2 months (95% CI, 14.2 to 24.2 months), and 1-year, 2-year, and 4-year actuarial overall survival rates were 66.0%, 25.02%, and 11.3%, respectively. Acneiform rash correlated with improved survival (P = .001), but initial CA19-9, borderline resectable initial stage, and surgical resection (n = 7) did not. The 1-year and 2-year radiographic local progression rates were 22.8% and 61.0%, respectively. The worst acute toxic effects were GI toxicity (32% and 10% for grades 2 and 3, respectively); fatigue (26% and 6% for grades 2 and 3, respectively); sensory neuropathy (9% and 1% for grades 2 and 3, respectively); and acneiform rash (54% and 3% for grades 2 and 3, respectively). Smad4(Dpc4) expression correlated with a local rather than a distant dominant pattern of disease progression (P = .016). CONCLUSION This regimen appears effective and has acceptable toxicity. The primary end point (1-year overall survival rate > 45%) was met, with encouraging survival duration. Smad4(Dpc4) immunostaining correlated with the pattern of disease progression. Prospective validation of Smad4(Dpc4) expression in cytology specimens as a predictive biomarker is warranted and may lead to personalized treatment strategies for patients with localized pancreatic cancer.

Proteomic Profiling Identifies Dysregulated Pathways in Small Cell Lung Cancer and Novel Therapeutic Targets Including PARP1

UNLABELLED Small cell lung cancer (SCLC) is an aggressive malignancy distinct from non-small cell lung cancer (NSCLC) in its metastatic potential and treatment response. Using an integrative proteomic and transcriptomic analysis, we investigated molecular differences contributing to the distinct clinical behavior of SCLCs and NSCLCs. SCLCs showed lower levels of several receptor tyrosine kinases and decreased activation of phosphoinositide 3-kinase (PI3K) and Ras/mitogen-activated protein (MAP)/extracellular signal-regulated kinase (ERK) kinase (MEK) pathways but significantly increased levels of E2F1-regulated factors including enhancer of zeste homolog 2 (EZH2), thymidylate synthase, apoptosis mediators, and DNA repair proteins. In addition, PARP1, a DNA repair protein and E2F1 co-activator, was highly expressed at the mRNA and protein levels in SCLCs. SCLC growth was inhibited by PARP1 and EZH2 knockdown. Furthermore, SCLC was significantly more sensitive to PARP inhibitors than were NSCLCs, and PARP inhibition downregulated key components of the DNA repair machinery and enhanced the efficacy of chemotherapy. SIGNIFICANCE SCLC is a highly lethal cancer with a 5-year survival rate of less than 10%. To date, no molecularly targeted agents have prolonged survival in patients with SCLCs. As a step toward identifying new targets, we systematically profiled SCLCs with a focus on therapeutically relevant signaling pathways. Our data reveal fundamental differences in the patterns of pathway activation in SCLCs and NSCLCs and identify several potential therapeutic targets for SCLCs, including PARP1 and EZH2. On the basis of these results, clinical studies evaluating PARP and EZH2 inhibition, together with chemotherapy or other agents, warrant further investigation.

TP53 Disruptive Mutations Lead to Head and Neck Cancer Treatment Failure through Inhibition of Radiation-Induced Senescence

Purpose: Mortality of patients with head and neck squamous cell carcinoma (HNSCC) is primarily driven by tumor cell radioresistance leading to locoregional recurrence (LRR). In this study, we use a classification of TP53 mutation (disruptive vs. nondisruptive) and examine impact on clinical outcomes and radiation sensitivity. Experimental Design: Seventy-four patients with HNSCC treated with surgery and postoperative radiation and 38 HNSCC cell lines were assembled; for each, TP53 was sequenced and the in vitro radioresistance measured using clonogenic assays. p53 protein expression was inhibited using short hairpin RNA (shRNA) and overexpressed using a retrovirus. Radiation-induced apoptosis, mitotic cell death, senescence, and reactive oxygen species (ROS) assays were carried out. The effect of the drug metformin on overcoming mutant p53-associated radiation resistance was examined in vitro as well as in vivo, using an orthotopic xenograft model. Results: Mutant TP53 alone was not predictive of LRR; however, disruptive TP53 mutation strongly predicted LRR (P = 0.03). Cell lines with disruptive mutations were significantly more radioresistant (P < 0.05). Expression of disruptive TP53 mutations significantly decreased radiation-induced senescence, as measured by SA-β-gal staining, p21 expression, and release of ROS. The mitochondrial agent metformin potentiated the effects of radiation in the presence of a disruptive TP53 mutation partially via senescence. Examination of our patient cohort showed that LRR was decreased in patients taking metformin. Conclusions: Disruptive TP53 mutations in HNSCC tumors predicts for LRR, because of increased radioresistance via the inhibition of senescence. Metformin can serve as a radiosensitizer for HNSCC with disruptive TP53, presaging the possibility of personalizing HNSCC treatment. Clin Cancer Res; 18(1); 290–300. ©2011 AACR.

Isolated central nervous system progression on Crizotinib: An Achilles heel of non-small cell lung cancer with EML4-ALK translocation?

Advanced non-small lung cancer (NSCLC) remains almost uniformly lethal with marginal long-term survival despite efforts to target specific oncogenic addiction pathways that may drive these tumors with small molecularly targeted agents and biologics. The EML4-ALK fusion gene encodes a chimeric tyrosine kinase that activates the Ras signaling pathway, and this fusion protein is found in approximately 5% of NSCLC. Targeting EML4-ALK with Crizotinib in this subset of NSCLC has documented therapeutic efficacy, but the vast majority of patients eventually develop recurrent disease that is often refractory to further treatments. We present the clinicopathologic features of three patients with metastatic NSCLC harboring the EML4-ALK translocation that developed isolated central nervous system (CNS) metastases in the presence of good disease control elsewhere in the body. These cases suggest a differential response of NSCLC to Crizotinib in the brain in comparison to other sites of disease, and are consistent with a previous report of poor CNS penetration of Crizotinib. Results of ongoing clinical trials will clarify whether the CNS is a major sanctuary site for EML4-ALK positive NSCLC being treated with Crizotinib. While understanding molecular mechanisms of resistance is critical to overcome therapeutic resistance, understanding physiologic mechanisms of resistance through analyzing anatomic patterns of failure may be equally crucial to improve long-term survival for patients with EML4-ALK translocation positive NSCLC.

DNA repair biomarker profiling of head and neck cancer: Ku80 expression predicts locoregional failure and death following radiotherapy

Purpose: Radiotherapy plays an integral role in the treatment of head and neck squamous cell carcinoma (HNSCC). Although proteins involved in DNA repair may predict HNSCC response to radiotherapy, none has been validated in this context. We examined whether differential expression of double-strand DNA break (DSB) repair proteins in HNSCC, the chief mediators of DNA repair following irradiation, predict for treatment outcomes. Experimental Design: Archival HNSCC tumor specimens (n = 89) were assembled onto a tissue microarray and stained with antibodies raised against 38 biomarkers. The biomarker set was enriched for proteins involved in DSB repair, in addition to established mechanistic markers of radioresistance. Staining was correlated with treatment response and survival alongside established clinical and pathologic covariates. Results were validated in an independent intramural cohort (n = 34). Results: Ku80, a key mediator of DSB repair, correlated most closely with clinical outcomes. Ku80 was overexpressed in half of all tumors, and its expression was independent of all other covariates examined. Ku80 overexpression was an independent predictor for both locoregional failure and mortality following radiotherapy (P < 0.01). The predictive power of Ku80 overexpression was confined largely to HPV-negative HNSCC, where it conferred a nine-fold greater risk of death at two years. Conclusions: Ku80 overexpression is a common feature of HNSCC, and is a candidate DNA repair-related biomarker for radiation treatment failure and death, particularly in patients with high-risk HPV-negative disease. It is a promising, mechanistically rational biomarker to select individual HPV-negative HNSCC patients for strategies to intensify treatment. Clin Cancer Res; 17(7); 2035–43. ©2011 AACR.

Role of 53BP1 in the regulation of DNA double-strand break repair pathway choice

The p53-binding protein 1 (53BP1) is a well-known DNA damage response (DDR) factor, which is recruited to nuclear structures at the site of DNA damage and forms readily visualized ionizing radiation (IR) induced foci. Depletion of 53BP1 results in cell cycle arrest in G2/M phase as well as genomic instability in human as well as mouse cells. Within the DNA damage response mechanism, 53BP1 is classified as an adaptor/mediator, required for processing of the DNA damage response signal and as a platform for recruitment of other repair factors. More recently, specific 53BP1 contributions to DSB repair pathway choice have been recognized and are being characterized. In this review, we have summarized recent advances in understanding the role of 53BP1 in regulating DNA DSBs repair pathway choice, variable diversity joining [V(D)J] recombination and class-switch recombination (CSR).

Evolutionary action score of TP53 identifies high-risk mutations associated with decreased survival and increased distant metastases in head and neck cancer

TP53 is the most frequently altered gene in head and neck squamous cell carcinoma, with mutations occurring in over two-thirds of cases, but the prognostic significance of these mutations remains elusive. In the current study, we evaluated a novel computational approach termed evolutionary action (EAp53) to stratify patients with tumors harboring TP53 mutations as high or low risk, and validated this system in both in vivo and in vitro models. Patients with high-risk TP53 mutations had the poorest survival outcomes and the shortest time to the development of distant metastases. Tumor cells expressing high-risk TP53 mutations were more invasive and tumorigenic and they exhibited a higher incidence of lung metastases. We also documented an association between the presence of high-risk mutations and decreased expression of TP53 target genes, highlighting key cellular pathways that are likely to be dysregulated by this subset of p53 mutations that confer particularly aggressive tumor behavior. Overall, our work validated EAp53 as a novel computational tool that may be useful in clinical prognosis of tumors harboring p53 mutations.

A contour-guided deformable image registration algorithm for adaptive radiotherapy.

In adaptive radiotherapy, deformable image registration is often conducted between the planning CT and treatment CT (or cone beam CT) to generate a deformation vector field (DVF) for dose accumulation and contour propagation. The auto-propagated contours on the treatment CT may contain relatively large errors, especially in low-contrast regions. A clinicians inspection and editing of the propagated contours are frequently needed. The edited contours are able to meet the clinical requirement for adaptive therapy; however, the DVF is still inaccurate and inconsistent with the edited contours. The purpose of this work is to develop a contour-guided deformable image registration (CG-DIR) algorithm to improve the accuracy and consistency of the DVF for adaptive radiotherapy. Incorporation of the edited contours into the registration algorithm is realized by regularizing the objective function of the original demons algorithm with a term of intensity matching between the delineated structures set pairs. The CG-DIR algorithm is implemented on computer graphics processing units (GPUs) by following the original GPU-based demons algorithm computation framework (Gu et al 2010 Phys Med Biol. 55 207-219). The performance of CG-DIR is evaluated on five clinical head-and-neck and one pelvic cancer patient data. It is found that compared with the original demons, CG-DIR improves the accuracy and consistency of the DVF, while retaining similar high computational efficiency.

Commissioning and initial stereotactic ablative radiotherapy experience with Vero

The purpose of this study is to describe the comprehensive commissioning process and initial clinical performance of the Vero linear accelerator, a new radiotherapy device recently installed at UT Southwestern Medical Center specifically developed for delivery of image‐guided stereotactic ablative radiotherapy (SABR). The Vero system utilizes a ring gantry to integrate a beam delivery platform with image guidance systems. The ring is capable of rotating ± 60° about the vertical axis to facilitate noncoplanar beam arrangements ideal for SABR delivery. The beam delivery platform consists of a 6 MV C‐band linac with a 60 leaf MLC projecting a maximum field size of 15×15 cm2 at isocenter. The Vero planning and delivery systems support a range of treatment techniques, including fixed beam conformal, dynamic conformal arcs, fixed gantry IMRT in either SMLC (step‐and‐shoot) or DMLC (dynamic) delivery, and hybrid arcs, which combines dynamic conformal arcs and fixed beam IMRT delivery. The accelerator and treatment head are mounted on a gimbal mechanism that allows the linac and MLC to pivot in two dimensions for tumor tracking. Two orthogonal kV imaging subsystems built into the ring facilitate both stereoscopic and volumetric (CBCT) image guidance. The system is also equipped with an always‐active electronic portal imaging device (EPID). We present our commissioning process and initial clinical experience focusing on SABR applications with the Vero, including: (1) beam data acquisition; (2) dosimetric commissioning of the treatment planning system, including evaluation of a Monte Carlo algorithm in a specially‐designed anthropomorphic thorax phantom; (3) validation using the Radiological Physics Center thorax, head and neck (IMRT), and spine credentialing phantoms; (4) end‐to‐end evaluation of IGRT localization accuracy; (5) ongoing system performance, including isocenter stability; and (6) clinical SABR applications. PACS number: 87.53.Ly

Tumor-Selective, Futile Redox Cycle-Induced Bystander Effects Elicited by NQO1 Bioactivatable Radiosensitizing Drugs in Triple-Negative Breast Cancers

AIMS β-Lapachone (β-lap), a novel radiosensitizer with potent antitumor efficacy alone, selectively kills solid cancers that over-express NAD(P)H quinone oxidoreductase 1 (NQO1). Since breast or other solid cancers have heterogeneous NQO1 expression, therapies that reduce the resistance (e.g., NQO1(low)) of tumor cells will have significant clinical advantages. We tested whether NQO1-proficient (NQO1(+)) cells generated sufficient hydrogen peroxide (H2O2) after β-lap treatment to elicit bystander effects, DNA damage, and cell death in neighboring NQO1(low) cells. RESULTS β-Lap showed NQO1-dependent efficacy against two triple-negative breast cancer (TNBC) xenografts. NQO1 expression variations in human breast cancer patient samples were noted, where ~60% cancers over-expressed NQO1, with little or no expression in associated normal tissue. Differential DNA damage and lethality were noted in NQO1(+) versus NQO1-deficient (NQO1(-)) TNBC cells and xenografts after β-lap treatment. β-Lap-treated NQO1(+) cells died by programmed necrosis, whereas co-cultured NQO1(-) TNBC cells exhibited DNA damage and caspase-dependent apoptosis. NQO1 inhibition (dicoumarol) or H2O2 scavenging (catalase [CAT]) blocked all responses. Only NQO1(-) cells neighboring NQO1(+) TNBC cells responded to β-lap in vitro, and bystander effects correlated well with H2O2 diffusion. Bystander effects in NQO1(-) cells in vivo within mixed 50:50 co-cultured xenografts were dramatic and depended on NQO1(+) cells. However, normal human cells in vitro or in vivo did not show bystander effects, due to elevated endogenous CAT levels. Innovation and Conclusions: NQO1-dependent bystander effects elicited by NQO1 bioactivatable drugs (β-lap or deoxynyboquinone [DNQ]) likely contribute to their efficacies, killing NQO1(+) solid cancer cells and eliminating surrounding heterogeneous NQO1(low) cancer cells. Normal cells/tissue are protected by low NQO1:CAT ratios.