Although most lung cancers are a result of smoking, approximately 25% of lung cancer cases worldwide are not attributable to tobacco use, accounting for over 300,000 deaths each year. Striking differences in the epidemiological, clinical and molecular characteristics of lung cancers arising in never smokers versus smokers have been identified, suggesting that they are separate entities. This Review summarizes our current knowledge of this unique and poorly understood disease.

To site-specifically incorporate an unnatural amino acid (UAA) into target proteins in Escherichia coli, we use a suppressor plasmid that provides an engineered suppressor tRNA and an aminoacyl-tRNA synthetase (aaRS) specific for the UAA of interest. The continuous drive to further improve UAA incorporation efficiency in E. coli has resulted in several generations of suppressor plasmids. Here we describe a new, highly efficient suppressor plasmid, pUltra, harboring a single copy each of the tRNA and aaRS expression cassettes that exhibits higher suppression activity than its predecessors. This system is able to efficiently incorporate up to three UAAs within the same protein at levels up to 30% of the level of wild-type protein expression. Its unique origin of replication (CloDF13) and antibiotic resistance marker (spectinomycin) allow pUltra to be used in conjunction with the previously reported pEVOL suppressor plasmid, each encoding a distinct tRNA/aaRS pair, to simultaneously insert two different UAAs into the same protein. We demonstrate the utility of this system by efficiently incorporating two bio-orthogonal UAAs containing keto and azido side chains into ketosteroid isomerase and subsequently derivatizing these amino acid residues with two distinct fluorophores, capable of Förster resonance energy transfer interaction. Finally, because of its minimal composition, two different tRNA/aaRS pairs were encoded in pUltra, allowing the generation of a single plasmid capable of dual suppression. The high suppression efficiency and the ability to harbor multiple tRNA/aaRS pairs make pUltra a useful system for conducting single- and multiple-UAA mutagenesis in E. coli.

The adoptive transfer of autologous T cells engineered to express a chimeric antigen receptor (CAR) has emerged as a promising cancer therapy. Despite impressive clinical efficacy, the general application of current CAR–T-cell therapy is limited by serious treatment-related toxicities. One approach to improve the safety of CAR-T cells involves making their activation and proliferation dependent upon adaptor molecules that mediate formation of the immunological synapse between the target cancer cell and T-cell. Here, we describe the design and synthesis of structurally defined semisynthetic adaptors we refer to as “switch” molecules, in which anti-CD19 and anti-CD22 antibody fragments are site-specifically modified with FITC using genetically encoded noncanonical amino acids. This approach allows the precise control over the geometry and stoichiometry of complex formation between CD19- or CD22-expressing cancer cells and a “universal” anti-FITC–directed CAR-T cell. Optimization of this CAR–switch combination results in potent, dose-dependent in vivo antitumor activity in xenograft models. The advantage of being able to titrate CAR–T-cell in vivo activity was further evidenced by reduced in vivo toxicity and the elimination of persistent B-cell aplasia in immune-competent mice. The ability to control CAR-T cell and cancer cell interactions using intermediate switch molecules may expand the scope of engineered T-cell therapy to solid tumors, as well as indications beyond cancer therapy.

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Global cancer statistics indicate that lung cancer is responsible for over 1 million deaths each year (1). Approximately 80%-85% of all lung cancers are non-small cell lung cancer (NSCLC), which include squamous cell carcinoma, adenocarcinoma, and large-cell carcinoma. Small cell lung cancer (SCLC) represents approximately 15%-20% of cases, and the incidence of SCLC has been decreasing over the last several years in the United States (2). Current treatment options include surgical resection, platinum-based doublet chemotherapy (a combination of two drugs that includes carboplatin or cisplatin as the backbone), and radiation therapy alone or in combination. Unfortunately, despite these therapies, the disease is rarely curable and prognosis is poor, with an overall 5-year survival rate of only 15% (3). This article will review recent advances and future perspectives in the development of novel strategies for the treatment of lung cancer, with an emphasis on how these can be derived from our understanding of the molecular pathogenesis of the disease. Historical perspectiveIn the mid-1800s, lung cancer was a rare disease, representing only 1% of all cancers seen at autopsy. By the early 1900s, the incidence of malignant lung tumors had begun to rise, and although most lung cancers occurred in men, a steady increase in women was observed beginning in the 1960s. While the link between cigarette smoking and lung cancer was suspected by clinicians in the 1930s, the cause of the dramatic increase was not well established until landmark epidemiologic studies in the 1950s provided evidence for a strong causal association between smoking and lung cancer (4). Surgical therapy for lung cancer began in the 1930s, with the first successful pneumonectomy for lung cancer reported by Evarts Graham in 1933 (5). The prognostic significance of nodal metastases was subsequently recognized, and surgical mediastinal lymph node sampling became an important aspect of staging. Current surgical approaches use lobectomy or, if necessary, pneumonectomy for curative treatment, while wedge resections are reserved for patients with severely impaired lung function. Likewise, preoperative evaluation and postoperative care have improved, resulting in operative mortality rates of less than 5%. With the invention of megavoltage linear accelerators, radical radiotherapy for potential cure of lung cancer was introduced in the 1950s. There have been significant technologic developments in radiation therapy planning and delivery such that currently, approximately 15% of patients with early-stage disease can be cured with radiotherapy alone. Nitrogen mustard was the first chemotherapy used for the treatment of lung cancer in the 1940s. Pilot studies from the late 1970s and early 1980s identified other drugs with activity against lung cancer, particularly drug combinations using cisplatin with a vinca alkaloid or etoposide (6). Over the next two decades, several prospective, randomized studies were conducted to evaluate various chemotherapy regimens (6, 7)...