Cytometry by time-of-flight (CyTOF) simultaneously measures multiple cellular proteins at the single-cell level and is used to assess intertumor and intratumor heterogeneity. This approach may be used to investigate the variability of individual tumor responses to treatments. Herein, we stratified lung tumor subpopulations based on AXL signaling as a potential targeting strategy. Integrative transcriptome analyses were used to investigate how TP-0903, an AXL kinase inhibitor, influences redundant oncogenic pathways in metastatic lung cancer cells. CyTOF profiling revealed that AXL inhibition suppressed SMAD4/TGFb signaling and induced JAK1-STAT3 signaling to compensate for the loss of AXL. Interestingly, high JAK1-STAT3 was associated with increased levels of AXL in treatment-na€ ve tumors. Tumors with high AXL, TGFb, and JAK1 signaling concomitantly displayed CD133-mediated cancer stemness and hybrid epithelialto-mesenchymal transition features in advanced-stage patients, suggesting greater potential for distant dissemination. Diffusion pseudotime analysis revealed cell-fate trajectories among four different categories that were linked to clinicopathologic features for each patient. Patient-derived organoids (PDO) obtained from tumors with high AXL and JAK1 were sensitive to TP-0903 and ruxolitinib (JAK inhibitor) treatments, supporting the CyTOF findings. This study shows that single-cell proteomic profiling of treatment-na€ ve lung tumors, coupled with ex vivo testing of PDOs, identifies continuous AXL, TGFb, and JAK1-STAT3 signal activation in select tumors that may be targeted by combined AXL-JAK1 inhibition.Significance: Single-cell proteomic profiling of clinical samples may facilitate the optimal selection of novel drug targets, interpretation of early-phase clinical trial data, and development of predictive biomarkers valuable for patient stratification.

Highlights d We investigate the relationship of patient age and ICB therapy biomarkers d Favorable ICB biomarkers are generally more prevalent in elderly patients d The CAMA web application provides a multi-omics atlas of aging in cancer

No abstract

Type 4 cyclic AMP (cAMP) phosphodiesterase (PDE4) inhibitors, a class of compounds in clinical development that activate cAMP-mediated signaling by inhibiting cAMP catabolism, offer a feasible means by which to potentiate glucocorticoid-mediated apoptosis in lymphoid malignancies such as B-cell chronic lymphocytic leukemia (B-CLL). In this study, we show that PDE4 inhibitors up-regulate glucocorticoid receptor (GRa) transcript levels in B-CLL cells but not T-CLL cells or Sezary cells or normal circulating T cells, B cells, monocytes, or neutrophils. Because GRa transcript half-life does not vary in CLL cells treated with the prototypic PDE4 inhibitor rolipram, the 4-fold increase in GRa mRNA levels observed within 4 h of rolipram treatment seems to result from an increase in GRa transcription. Rolipram treatment increases levels of transcripts derived from the1A3 promoter to a greater extent than the 1B promoter.Treatment of B-CLL cells with two other PDE4 inhibitors currently in clinical development also augments GR transcript levels and glucocorticoid-mediated apoptosis. Washout studies show that simultaneous treatment with both drug classes irreversibly augments apoptosis over the same time frame that GR upregulation occurs. Although treatment of B-CLL cells with glucocorticoids reduces basal GRa transcript levels in a dose-related manner, cotreatment with rolipram maintained GRa transcript levels above baseline. Our results suggest that as a result of their unusual sensitivity to PDE4 inhibitor^mediated up-regulation of GRa expression, treatment of B-CLL patients with combined PDE4 inhibitor/glucocorticoid therapy may be of therapeutic benefit in this disease.Glucocorticoids are an important component of standard therapy for several lymphoid malignancies, including multiple myeloma, acute lymphoblastic leukemia, and diffuse large B-cell lymphoma. As early studies in patients with B-cell chronic lymphocytic leukemia (B-CLL) showed that addition of prednisone to chlorambucil augmented response rate but not median survival, glucocorticoids are not generally a standard component of initial therapy for patients with B-CLL (1, 2). Nonetheless, two studies of high-dose glucocorticoid therapy have suggested that glucocorticoids can be of clinical benefit to a subset of patients with treatment-refractory B-CLL (3, 4).Despite frequent responses to glucocorticoid treatment, monotherapy with glucocorticoids is not curative in any lymphoid malignancy, but the mechanisms underlying clinical glucocorticoid resistance remain controversial. Structural alterations in the glucocorticoid receptor (GR) are commonly identified in lymphoid cell lines that have been selected for glucocorticoid resistance by prolonged culture in dexamethasone, but comparable alterations in primary malignant lymphoid cells have been only infrequently reported (5 -9). A detailed analysis of treated B-CLL patients failed to identify abnormalities in either the DNA-or steroid-binding domains of leukemic GRs (10). Nonstructural modifications of ...

Aggressive tumors of epithelial origin shed cells that intravasate and become circulating tumor cells (CTC). The CTCs that are able to survive the stresses encountered in the bloodstream can then seed metastases. We demonstrated previously that CTCs isolated from the blood of prostate cancer patients display specific nanomechanical phenotypes characteristic of cell endurance and invasiveness and patient sensitivity to androgen deprivation therapy. Here we report that patient-isolated CTCs are nanomechanically distinct from cells randomly shed from the tumor, with high adhesion as the most distinguishing biophysical marker. CTCs uniquely coisolated with macrophage-like cells bearing the markers of tumor-associated macrophages (TAM). The presence of these immune cells was indicative of a survival-promoting phenotype of “mechanical fitness” in CTCs based on high softness and high adhesion as determined by atomic force microscopy. Correlations between enumeration of macrophages and mechanical fitness of CTCs were strong in patients before the start of hormonal therapy. Single-cell proteomic analysis and nanomechanical phenotyping of tumor cell–macrophage cocultures revealed that macrophages promoted epithelial–mesenchymal plasticity in prostate cancer cells, manifesting in their mechanical fitness. The resulting softness and adhesiveness of the mechanically fit CTCs confer resistance to shear stress and enable protective cell clustering. These findings suggest that selected tumor cells are coached by TAMs and accompanied by them to acquire intermediate epithelial/mesenchymal status, thereby facilitating survival during the critical early stage leading to metastasis. Significance: The interaction between macrophages and circulating tumor cells increases the capacity of tumor cells to initiate metastasis and may constitute a new set of blood-based targets for pharmacologic intervention.