Craig D. Peacock

A crucial requirement for Hedgehog signaling in small cell lung cancer

Small-cell lung cancer (SCLC) is an aggressive neuroendocrine subtype of lung cancer for which there is no effective treatment. Using a mouse model in which deletion of Rb1 and Trp53 in the lung epithelium of adult mice induces SCLC, we found that the Hedgehog signaling pathway is activated in SCLC cells independently of the lung microenvironment. Constitutive activation of the Hedgehog signaling molecule Smoothened (Smo) promoted the clonogenicity of human SCLC in vitro and the initiation and progression of mouse SCLC in vivo. Reciprocally, deletion of Smo in Rb1 and Trp53-mutant lung epithelial cells strongly suppressed SCLC initiation and progression in mice. Furthermore, pharmacological blockade of Hedgehog signaling inhibited the growth of mouse and human SCLC, most notably following chemotherapy. These findings show a crucial cell-intrinsic role for Hedgehog signaling in the development and maintenance of SCLC and identify Hedgehog pathway inhibition as a therapeutic strategy to slow the progression of disease and delay cancer recurrence in individuals with SCLC.

Self-Renewal of Acute Lymphocytic Leukemia Cells Is Limited by the Hedgehog Pathway Inhibitors Cyclopamine and IPI-926

Conserved embryonic signaling pathways such as Hedgehog (Hh), Wingless and Notch have been implicated in the pathogenesis of several malignancies. Recent data suggests that Hh signaling plays a role in normal B-cell development, and we hypothesized that Hh signaling may be important in precursor B-cell acute lymphocytic leukemia (B-ALL). We found that the expression of Hh pathway components was common in human B-ALL cell lines and clinical samples. Moreover, pathway activity could be modulated by Hh ligand or several pathway inhibitors including cyclopamine and the novel SMOOTHENED (SMO) inhibitor IPI-926. The inhibition of pathway activity primarily impacted highly clonogenic B-ALL cells expressing aldehyde dehydrogenase (ALDH) by limiting their self-renewal potential both in vitro and in vivo. These data demonstrate that Hh pathway activation is common in B-ALL and represents a novel therapeutic target regulating self-renewal and persistence of the malignant clone.

Cancer stem cells in lung cancer: Evidence and controversies.

The cancer stem cell (CSC) model is based on a myriad of experimental and clinical observations suggesting that the malignant phenotype is sustained by a subset of cells characterized by the capacity for self‐renewal, differentiation and innate resistance to chemotherapy and radiation. CSC may be responsible for disease recurrence after definitive therapy and may therefore be functionally synonymous with minimal residual disease. Similar to other solid tumours, several putative surface markers for lung CSC have been identified, including CD133 and CD44. In addition, expression and/or activity of the cytoplasmic enzyme aldehyde dehydrogenase ALDH and capacity of cells to exclude membrane permeable dyes (known as the ‘side population’) correlate with stem‐like function in vitro and in vivo. Embryonic stem cell pathways such as Hedgehog, Notch and WNT may also be active in lung cancers stem cells and therefore may be therapeutically targetable for maintenance therapy in patients achieving a complete response to surgery, radiotherapy or chemotherapy. This paper will review the evidence regarding the existence and function of lung CSC in the context of the experimental and clinical evidence and discuss some ongoing controversies regarding this model.

Small Cell Lung Cancer: Can Recent Advances in Biology and Molecular Biology Be Translated into Improved Outcomes?

Paul A. Bunn Jr., MD, John D. Minna, MD, Alexander Augustyn, PhD, Adi F. Gazdar, MD, Youcef Ouadah, BS, Mark A. Krasnow, MD, PhD, Anton Berns, PhD, Elisabeth Brambilla, MD, Natasha Rekhtman, MD, PhD, Pierre P. Massion, MD, Matthew Niederst, PhD, Martin Peifer, PhD, Jun Yokota, MD, Ramaswamy Govindan, MD, John T. Poirier, PhD, Lauren A. Byers, MD, Murry W. Wynes, PhD, David G. McFadden, MD, PhD, David MacPherson, PhD, Christine L. Hann, MD, PhD, Anna F. Farago, MD, PhD, Caroline Dive, PhD, Beverly A. Teicher, PhD, Craig D. Peacock, PhD, Jane E. Johnson, PhD, Melanie H. Cobb, PhD, Hans-Guido Wendel, MD, David Spigel, MD, Julien Sage, PhD, Ping Yang, MD, PhD, M. Catherine Pietanza, MD, Lee M. Krug, MD, John Heymach, MD, PhD, Peter Ujhazy, MD, PhD, Caicun Zhou, MD, PhD, Koichi Goto, MD, Afshin Dowlati, MD, Camilla Laulund Christensen, PhD, Keunchil Park, MD, PhD, Lawrence H. Einhorn, MD, Martin J. Edelman, MD, Giuseppe Giaccone, MD, PhD, David E. Gerber, MD, Ravi Salgia, MD, PhD, Taofeek Owonikoko, MD, PhD, Shakun Malik, MD, Niki Karachaliou, MD, David R. Gandara, MD, Ben J. Slotman, MD, PhD, Fiona Blackhall, MD, PhD, Glenwood Goss, MD, FRCPC, Roman Thomas, MD, Charles M. Rudin, MD, PhD, Fred R. Hirsch, MD, PhD*

A genetic basis for the variation in the vulnerability of cancer to DNA damage

Radiotherapy is not currently informed by the genetic composition of an individual patients tumour. To identify genetic features regulating survival after DNA damage, here we conduct large-scale profiling of cellular survival after exposure to radiation in a diverse collection of 533 genetically annotated human tumour cell lines. We show that sensitivity to radiation is characterized by significant variation across and within lineages. We combine results from our platform with genomic features to identify parameters that predict radiation sensitivity. We identify somatic copy number alterations, gene mutations and the basal expression of individual genes and gene sets that correlate with the radiation survival, revealing new insights into the genetic basis of tumour cellular response to DNA damage. These results demonstrate the diversity of tumour cellular response to ionizing radiation and establish multiple lines of evidence that new genetic features regulating cellular response after DNA damage can be identified.

Next-Generation Sequence Analysis of Cancer Xenograft Models

Next-generation sequencing (NGS) studies in cancer are limited by the amount, quality and purity of tissue samples. In this situation, primary xenografts have proven useful preclinical models. However, the presence of mouse-derived stromal cells represents a technical challenge to their use in NGS studies. We examined this problem in an established primary xenograft model of small cell lung cancer (SCLC), a malignancy often diagnosed from small biopsy or needle aspirate samples. Using an in silico strategy that assign reads according to species-of-origin, we prospectively compared NGS data from primary xenograft models with matched cell lines and with published datasets. We show here that low-coverage whole-genome analysis demonstrated remarkable concordance between published genome data and internal controls, despite the presence of mouse genomic DNA. Exome capture sequencing revealed that this enrichment procedure was highly species-specific, with less than 4% of reads aligning to the mouse genome. Human-specific expression profiling with RNA-Seq replicated array-based gene expression experiments, whereas mouse-specific transcript profiles correlated with published datasets from human cancer stroma. We conclude that primary xenografts represent a useful platform for complex NGS analysis in cancer research for tumours with limited sample resources, or those with prominent stromal cell populations.

A Histologic Basis for the Efficacy of SBRT to the lung

Purpose: Stereotactic body radiation therapy (SBRT) is the standard of care for medically inoperable patients with early‐stage NSCLC. However, NSCLC is composed of several histological subtypes and the impact of this heterogeneity on SBRT treatments has yet to be established. Methods: We analyzed 740 patients with early‐stage NSCLC treated definitively with SBRT from 2003 through 2015. We calculated cumulative incidence curves using the competing risk method and identified predictors of local failure using Fine and Gray regression. Results: Overall, 72 patients had a local failure, with a cumulative incidence of local failure at 3 years of 11.8%. On univariate analysis, squamous histological subtype, younger age, fewer medical comorbidities, higher body mass index, higher positron emission tomography standardized uptake value, central tumors, and lower radiation dose were associated with an increased risk for local failure. On multivariable analysis, squamous histological subtype (hazard ratio = 2.4 p = 0.008) was the strongest predictor of local failure. Patients with squamous cancers fail SBRT at a significantly higher rate than do those with adenocarcinomas or NSCLC not otherwise specified, with 3‐year cumulative rates of local failure of 18.9% (95% confidence interval [CI]: 12.7–25.1), 8.7% (95% CI: 4.6–12.8), and 4.1% (95% CI: 0–9.6), respectively. Conclusion: Our results demonstrate an increased rate of local failure in patients with squamous cell carcinoma. Standard approaches for radiotherapy that demonstrate efficacy for a population may not achieve optimal results for individual patients. Establishing the differential dose effect of SBRT across histological groups is likely to improve efficacy and inform ongoing and future studies that aim to expand indications for SBRT.

Integration of Hedgehog and mutant FLT3 signaling in myeloid leukemia

Activation of the Hedgehog pathway drives FLT3-mutated leukemia, and dual pathway inhibition effectively inhibits tumor growth. Hedgehog to the rescue Acute myeloid leukemia is generally difficult to treat, and the presence of internal tandem duplication in a gene called FLT3 (FLT3-ITD) is associated with a particularly poor prognosis. Lim et al. discovered that patients with FLT3-ITD leukemia also have increased activity of the Hedgehog protein signaling pathway. Experiments in mouse models of FLT3-ITD confirmed the functional role of Hedgehog in the development of leukemia and showed that combined treatment targeting FLT3 and Hedgehog is effective as a therapeutic strategy in this setting. FMS-like tyrosine kinase 3 (FLT3) internal tandem duplication (ITD) mutations resulting in constitutive kinase activity are common in acute myeloid leukemia (AML) and carry a poor prognosis. Several agents targeting FLT3 have been developed, but their limited clinical activity suggests that the inhibition of other factors contributing to the malignant phenotype is required. We examined gene expression data sets as well as primary specimens and found that the expression of GLI2, a major effector of the Hedgehog (Hh) signaling pathway, was increased in FLT3-ITD compared to wild-type FLT3 AML. To examine the functional role of the Hh pathway, we studied mice in which Flt3-ITD expression results in an indolent myeloproliferative state and found that constitutive Hh signaling accelerated the development of AML by enhancing signal transducer and activator of transcription 5 (STAT5) signaling and the proliferation of bone marrow myeloid progenitors. Furthermore, combined FLT3 and Hh pathway inhibition limited leukemic growth in vitro and in vivo, and this approach may serve as a therapeutic strategy for FLT3-ITD AML.

Radiotherapy in the Era of Precision Medicine

Current predictors of radiation response are largely limited to clinical and histopathologic parameters, and extensive systematic analyses of the correlation between radiation sensitivity and genomic parameters remain lacking. In the era of precision medicine, the lack of -omic determinants of radiation response has hindered the personalization of radiation delivery to the unique characteristics of each patient׳s cancer and impeded the discovery of new therapies that can be administered concurrently with radiation therapy. The cataloging of the -omic determinants of radiation sensitivity of cancer has great potential in enhancing efficacy and limiting toxicity in the context of a new approach to precision radiotherapy. Herein, we review concepts and data that contribute to the delineation of the radiogenomic landscape of cancer.

Case study: patient-derived clear cell adenocarcinoma xenograft model longitudinally predicts treatment response

There has been little progress in the use of patient-derived xenografts (PDX) to guide individual therapeutic strategies. In part, this can be attributed to the operational challenges of effecting successful engraftment and testing multiple candidate drugs in a clinically workable timeframe. It also remains unclear whether the ancestral tumor will evolve along similar evolutionary trajectories in its human and rodent hosts in response to similar selective pressures (i.e., drugs). Herein, we combine a metastatic clear cell adenocarcinoma PDX with a timely 3 mouse x 1 drug experimental design, followed by a co-clinical trial to longitudinally guide a patient’s care. Using this approach, we accurately predict response to first- and second-line therapies in so far as tumor response in mice correlated with the patient’s clinical response to first-line therapy (gemcitabine/nivolumab), development of resistance and response to second-line therapy (paclitaxel/neratinib) before these events were observed in the patient. Treatment resistance to first-line therapy in the PDX is coincident with biologically relevant changes in gene and gene set expression, including upregulation of phase I/II drug metabolism (CYP2C18, UGT2A, and ATP2A1) and DNA interstrand cross-link repair (i.e., XPA, FANCE, FANCG, and FANCL) genes. A total of 5.3% of our engrafted PDX collection is established within 2 weeks of implantation, suggesting our experimental designs can be broadened to other cancers. These findings could have significant implications for PDX-based avatars of aggressive human cancers.