Squamous Cell Transformation of Primary Lung Adenocarcinoma in a Patient With EML4-ALK Fusion Variant 5 Refractory to ALK Inhibitors.

Histologic transformation from adenocarcinoma to squamous cell carcinoma in lung cancer has not been reported as a mechanism of resistance to ALK inhibition. This report describes the clinical course of a female former light smoker with metastatic lung adenocarcinoma whose tumor underwent histologic transformation from a well-differentiated lung adenocarcinoma to a well-differentiated lung squamous cell carcinoma in the same location at the left mainstem bronchus while maintaining the ALK fusion oncogene without any resistance mutations. After experiencing disease progression while on crizotinib, the patient participated in clinical trials that provided early access to the novel ALK inhibitors ceritinib and alectinib before they were commercially available. Tumor recurrence occurred at the primary and metastatic central nervous system sites (ie, brain and spine). At tumor progression, liquid biopsy and tumor genomic profiling of plasma cell-free DNA next-generation sequencing (NGS) provided an accurate diagnosis with a short turnaround time compared with the tissue-based targeted capture NGS. The patient received several courses of radiation primarily to the brain and spine during her disease course. Her disease did not respond to the immune checkpoint inhibitor nivolumab, and she died on home hospice approximately 4 years after diagnosis. This case supports the importance of both histopathologic assessment and comprehensive genomic profiling in selecting appropriate treatment for patients with refractory, metastatic, ALK oncogene-driven non-small cell lung cancer. Use of symptom-directed radiation in tandem with ALK inhibitors contributed to the disease and symptomatic control and prolonged survival in this patient.

Clinical utility of tumor genomic profiling in patients with high plasma circulating tumor DNA burden or metabolically active tumors.

BACKGROUND: This retrospective study was undertaken to determine if the plasma circulating tumor DNA (ctDNA) level and tumor biological features in patients with advanced solid tumors affected the detection of genomic alterations (GAs) by a plasma ctDNA assay. METHOD: Cell-free DNA (cfDNA) extracted from frozen plasma (N = 35) or fresh whole blood (N = 90) samples were subjected to a 62-gene hybrid capture-based next-generation sequencing assay FoundationACT. Concordance was analyzed for 51 matched FoundationACT and FoundationOne (tissue) cases. The maximum somatic allele frequency (MSAF) was used to estimate the amount of tumor fraction of cfDNA in each sample. The detection of GAs was correlated with the amount of cfDNA, MSAF, total tumor anatomic burden (dimensional sum), and total tumor metabolic burden (SUVmax sum) of the largest ten tumor lesions on PET/CT scans. RESULTS: FoundationACT detected GAs in 69 of 81 (85%) cases with MSAF > 0. Forty-two of 51 (82%) cases had ≥ 1 concordance GAs matched with FoundationOne, and 22 (52%) matched to the National Comprehensive Cancer Network (NCCN)-recommended molecular targets. FoundationACT also detected 8 unique molecular targets, which changed the therapy in 7 (88%) patients who did not have tumor rebiopsy or sufficient tumor DNA for genomic profiling assay. In all samples (N = 81), GAs were detected in plasma cfDNA from cancer patients with high MSAF quantity (P = 0.0006) or high tumor metabolic burden (P = 0.0006) regardless of cfDNA quantity (P = 0.2362). CONCLUSION: This study supports the utility of using plasma-based genomic assays in cancer patients with high plasma MSAF level or high tumor metabolic burden.

Severe nivolumab-induced pneumonitis preceding durable clinical remission in a patient with refractory, metastatic lung squamous cell cancer: a case report.

BACKGROUND: Programmed cell death 1 (PD-1) and its ligand 1 (PD-L1) inhibitors have quickly become standard of care for patients with advanced non-small cell lung cancer and increasing numbers of other cancer types. In this report, we discuss the clinical history, pathological evaluation, and genomic findings in a patient with metastatic lung squamous cell cancer (SCC) who developed severe nivolumab-induced pneumonitis preceding durable clinical remission after three doses of nivolumab. CASE PRESENTATION: A patient with chemotherapy-refractory, metastatic lung SCC developed symptomatic pneumonitis by week 4 after nivolumab treatment, concurrently with onset of a potent antitumor response. Despite discontinuation of nivolumab after three doses and the use of high dose oral corticosteroids for grade 3 pneumonitis, continued tumor response to a complete remission by 3 months was evident by radiographic assessment. At the time of this submission, the patient has remained in clinical remission for 14 months. High PD-L1 expression by immunohistochemistry staining was seen in intra-alveolar macrophages and viable tumor cells in the pneumonitis and recurrent tumor specimens, respectively. Tumor genomic profiling by FoundationOne targeted exome sequencing revealed a very high tumor mutation burden (TMB) corresponding to 95-96 percentile in lung SCC, i.e., 87.4-91.0 and 82.9 mut/Mb, respectively, in pre- and post-nivolumab tumor specimens. Except for one, the 13 functional genomic alterations remained the same in the diagnostic, recurrent, and post-treatment, relapsed tumor specimens, suggesting that nivolumab reset the patient's immune system against one or more preexisting tumor-associated antigens (TAAs). One potential TAA candidate is telomerase reverse transcriptase (TERT) in which an oncogenic promoter -146C>T mutation was detected. Human leukocyte antigen (HLA) typing revealed HLA-A*0201 homozygosity, which is the prevalent HLA class I allele that has been used to develop universal cancer vaccine targeting TERT-derived peptides. CONCLUSIONS: Nivolumab could quickly reset and sustain host immunity against preexisting TAA(s) in this chemotherapy-refractory lung SCC patient. Further mechanistic studies are needed to characterize the effective immune cells and define the HLA-restricted TAA(s) and the specific T cell receptor clones responsible for the potent antitumor effect, with the aim of developing precision immunotherapy with improved effectiveness and safety.

Bridging tumor genomics to patient outcomes through an integrated patient-derived xenograft platform.

New approaches to optimization of cancer drug development in the laboratory and the clinic will be required to fully achieve the goal of individualized, precision cancer therapy. Improved preclinical models that more closely reflect the now recognized genomic complexity of human cancers are needed. Here we describe a collaborative research project that integrates core resources of The Jackson Laboratory Basic Science Cancer Center with genomics and clinical research facilities at the UC Davis Comprehensive Cancer Center to establish a clinically and genomically annotated patient-derived xenograft (PDX) platform designed to enhance new drug development and strategies for targeted therapies. Advanced stage non-small-cell lung cancer (NSCLC) was selected for initial studies because of emergence of a number of "druggable" molecular targets, and recent recognition of substantial inter- and intrapatient tumor heterogeneity. Additionally, clonal evolution after targeted therapy interventions make this tumor type ideal for investigation of this platform. Using the immunodeficient NOD scid gamma mouse, > 200 NSCLC tumor biopsies have been xenotransplanted. During the annotation process, patient tumors and subsequent PDXs are compared at multiple levels, including histomorphology, clinically applicable molecular biomarkers, global gene expression patterns, gene copy number variations, and DNA/chromosomal alterations. NSCLC PDXs are grouped into panels of interest according to oncogene subtype and/or histologic subtype. Multiregimen drug testing, paired with next-generation sequencing before and after therapy and timed tumor pharmacodynamics enables determination of efficacy, signaling pathway alterations, and mechanisms of sensitivity-resistance in individual models. This approach should facilitate derivation of new therapeutic strategies and the transition to individualized therapy.

Percutaneous lung biopsy: an ordering clinician's guide to current practice.

The discovery of a pulmonary nodule or mass usually leads to a clinical assessment of patient risk factors for malignancy and an imaging workup. The latter may include appropriately timed follow-up, computed tomography with the possible addition of a computed tomographic nodule enhancement study, positron emission tomography, and/or percutaneous biopsy of the lung lesion or associated lymph nodes or metastatic lesions. This article reviews the rationale for the imaging workup of lung lesions and the details of percutaneous needle biopsy, including risks, technical feasibility, techniques, needle systems, cytological/pathological evaluation, accuracy, complications, and complication management.

Oxidation of chylomicron remnant-like particles inhibits their uptake by THP-1 macrophages by apolipoprotein E-dependent processes.

The influence of the oxidative state of chylomicron remnants (CMR) on the mechanisms of their uptake and induction of lipid accumulation by macrophages derived from the human monocyte cell line, THP-1, during foam cell formation was investigated using chylomicron-remnant-like particles (CRLPs) at 3 different levels of oxidation. The oxidative state of CRLPs was varied by exposure to CuSO(4) (oxCRLPs) or incorporation of the antioxidant, probucol (pCRLPs) into the particles. oxCRLPs caused significantly less accumulation of triacylglycerol in the macrophages than CRLPs, and their rate of uptake was lower, while pCRLPs caused more lipid accumulation and were taken up faster. Uptake of all 3 types of particles was inhibited to a similar extent when entry via the low density lipoprotein (LDL) receptor related protein (80-90%), LDL receptor (-30-40%), CD36 (-40%) and phagocytosis (-35-40%) was blocked using lactoferrin, excess LDL, anti-CD36 and cytochalasin D, respectively, but blocking scavenger receptors-A or -B1 using poly inosinic acid or excess HDL had no effect. These findings show that oxidation of CRLPs lowers their rate of uptake and induction of lipid accumulation in macrophages. However, oxidation does not change the main pathways of internalisation of CRLPs into THP-1 macrophages, which occur mainly via the LRP with some contribution from the LDLr, while CD36 and phagocytosis have only a minor role, regardless of the oxidative state of the particles. Thus, the effects of CMR oxidation on foam cell formation contrast sharply with those of LDL oxidation and this may be important in the role of dietary oxidized lipids and antioxidants in modulating atherosclerosis.

Right-sided superior vena cava draining into the left atrium: a rare anomaly of systemic venous return.

The most commonly encountered systemic thoracic venous anomaly is a persistent left superior vena cava that drains into the right atrium via the coronary sinus. A much rarer systemic venous anomaly is that of isolated anomalous drainage of a normally positioned right superior vena cava (RSVC) into the left atrium (LA). This has been reported in approximately 20 patients with the diagnosis usually being made by cardiac catheterization. We report the case of a toddler with asymptomatic hypoxemia resulting from anomalous drainage of a normal RSVC into his LA. This was diagnosed non-invasively by contrast-enhanced chest CT.

Efflux of lipid from macrophages after induction of lipid accumulation by chylomicron remnants.

The fate of cholesterol and triacylglycerol taken up and accumulated by macrophages after exposure to chylomicron remnants was investigated using macrophages derived from the human monocyte cell line THP-1 and chylomicron remnant-like particles containing human apolipoprotein (apo) E (CRLPs) as the experimental model. In THP-1 macrophages lipid loaded with CRLPs and incubated with various cholesterol acceptors for 24 h, the mass of cholesterol and cholesteryl ester found in the cells was not changed by HDL, HDL3 or lipid-free ApoA-I, although it was decreased by 38% by ApoA-I-phosphatidylcholine vesicles (ApoA-I-PC). After loading of the macrophages with [3H]cholesterol-labelled CRLPs, only about 5% of the label was effluxed in 24 h in the absence of cholesterol acceptors, and this increased to about 10% with ApoA-I or PC only, and to about 30% with apoA-I-PC. In similar experiments with [3H]triolein, only about 4% of the labelled triacylglycerol taken up by the cells was released into the medium in 24 h, and a large (>60%) and consistent proportion of the intracellular radioactivity remained associated with the triacylglycerol throughout this period. These results suggest that cholesterol and triacylglycerol derived from chylomicron remnants are not readily cleared from macrophages, and this is likely to contribute to the atherogenicity of the remnant lipoproteins.

Differential effects of low-density lipoprotein and chylomicron remnants on lipid accumulation in human macrophages.

The effects of low-density lipoprotein (LDL) and chylomicron remnants on lipid accumulation in human monocyte-derived macrophages (HMDMs) and in macrophages derived from the human monocyte cell line THP-1 were compared. The HMDMs or THP-1 macrophages were incubated with LDL, oxidized LDL (oxLDL), chylomicron remnant-like particles (CMR-LPs), or oxidized CMR-LPs (oxCMR-LPs), and the amount and type of lipid accumulated were determined. As expected, the lipid content of both cell types was increased markedly by oxLDL but not LDL, and this was due to a rise in cholesterol, cholesteryl ester (CE), and triacylglycerol (TG) levels. In contrast, both CMR-LPs and oxCMR-LPs caused a considerable increase in cellular lipid in HMDMs and THP-1 macrophages, but in this case there was a greater rise in the TG than in the cholesterol or CE content. Lipid accumulation in response to oxLDL, CMR-LPs, and oxCMR-LPs was prevented by the ACAT inhibitor CI976 in HMDMs but not in THP-1 macrophages, where TG levels remained markedly elevated. The rate of incorporation of [(3)H]oleate into CE and TG in THP-1 macrophages was increased by oxLDL, CMR-LPs, and oxCMR-LPs, but incorporation into TG was increased to a greater extent with CMR-LPs and oxCMR-LPs compared with oxLDL. These results demonstrate that both CMR-LPs and oxCMR-LPs cause lipid accumulation in human macrophages comparable to that seen with oxLDL and that oxidation of the remnant particles does not enhance this effect. They also demonstrate that a greater proportion of the lipid accumulated in response to CMR-LPs compared with oxLDL is TG rather than cholesterol or CE and that this is associated with a higher rate of TG synthesis. This study, therefore, provides further evidence to suggest that chylomicron remnants have a role in foam cell formation that is distinct from that of oxLDL.

Protection of chylomicron remnants from oxidation by incorporation of probucol into the particles enhances their uptake by human macrophages and increases lipid accumulation in the cells.

The effects of protection of chylomicron remnants from oxidation on their uptake and induction of lipid accumulation in macrophages were investigated using chylomicron remnant-like particles (CRLPs) containing the lipophilic antioxidant drug, probucol, and macrophages derived from the human monocyte cell line, THP-1. The total lipid content of THP-1 macrophages was markedly higher (x2.2) after 48 h of incubation of THP-1 macrophages with CRLPs containing probucol (pCRLPs) when compared to CRLPs without probucol, and this was because of increases in triacylglycerol (x2.3) and cholesterol (x1.8) levels, while cholesteryl ester concentrations were not significantly changed. Determination of the uptake of CRLPs and pCRLPs by the cells using particles labelled with the fluorescent probe 1,1'-dioctadecyl-3,3,3'3'-tetramethylindo-carbocyanine perchlorate showed that pCRLPs are taken up at a faster rate than CRLPs. The synthesis of triacylglycerol, as measured by the incorporation of [(3)H]oleate and [(3)H]glycerol, was also increased in macrophages incubated with pCRLPs as compared to CRLPs without probucol, but phospholipid and cholesteryl ester formation from [(3)H]oleate was unaffected. In addition, no differences between the effects of CRLPs and pCRLPs on the expression of mRNA for a range of genes believed to be involved in lipoprotein uptake, intracellular lipid metabolism and the efflux of cholesterol from macrophages was detected. These results suggest that antioxidants carried in chylomicron remnants enhance lipid accumulation in macrophages by increasing the rate of uptake of the particles and raising the intracellular synthesis of triacylglycerol, but not cholesteryl ester, and that these effects are brought about by changes at the post-transcriptional level. Antioxidants carried in chylomicron remnants therefore may promote the development of atherosclerosis, and this is likely to be particularly important in conditions where clearance of remnants from the circulation is delayed.

Incorporation of lycopene into chylomicron remnant-like particles enhances their induction of lipid accumulation in macrophages.

Lipid accumulation in macrophages exposed to chylomicron remnant-like particles containing the dietary antioxidant lycopene was investigated. After incubation with THP-1 macrophages (48h), chylomicron remnant-like particles containing lycopene (lycCRLPs) as compared to those without (CRLPs) caused significantly more lipid accumulation in the cells, and this was due to increases in both the triacylglycerol (+100%) and cholesterol (+62%) content. In addition, expression of mRNA for diacylglycerol acyltransferase (DGAT), a key enzyme in triacylglycerol synthesis, was significantly decreased by lycCRLPs, but not CRLPs. These findings suggest that lycopene from the diet may promote, rather than retard, lipid accumulation in macrophages during its transport in the blood in chylomicron remnants.