Symptomatic Pneumonitis With Durvalumab After Concurrent Chemoradiotherapy in Unresectable Stage III NSCLC.

Introduction: In the placebo-controlled, phase 3 PACIFIC trial, durvalumab significantly prolonged progression-free survival (PFS) (p < 0.0001) and overall survival (OS) (p = 0.00251) in patients with unresectable stage III NSCLC and no progression after platinum-based concurrent chemoradiotherapy (cCRT). Pneumonitis or radiation pneumonitis (PRP) was common in both arms. We report exploratory analyses evaluating the association of symptomatic (grade ≥2) PRP (G2+PRP) with baseline factors and clinical outcomes. Methods: Patients with WHO performance status of 0 or 1 were randomized (2:1) to 12 months of durvalumab or placebo, 1 to 42 days after cCRT. Associations between baseline factors and on-study G2+PRP in durvalumab-treated patients were investigated using univariate and multivariate logistic regression. PFS and OS were analyzed using Cox proportional hazards models adjusted for time-dependent G2+PRP plus covariates for randomization stratification factors without and with additional baseline factors. Results: On-study G2+PRP occurred in 94 of 475 (19.8%) and 33 of 234 patients (14.1%) on durvalumab and placebo, respectively (median follow-up, 25.2 mo); grade greater than or equal to 3 PRP was uncommon (4.6% and 4.7%, respectively). Time to onset and resolution of G2+PRP was similar with durvalumab and placebo. Univariate and multivariate analyses identified patients treated in Asia, those with stage IIIA disease, those with performance status of 1, and those who had not received induction chemotherapy as having a higher risk of G2+PRP. PFS and OS benefit favoring durvalumab versus placebo was maintained regardless of time-dependent G2+PRP. Conclusions: Factors associated with higher risk of G2+PRP with durvalumab after cCRT were identified. Clinical benefit was maintained regardless of on-study G2+PRP, suggesting the risk of this event should not deter the use of durvalumab in eligible patients with unresectable stage III NSCLC.

Differential roles of normal and lung cancer-associated fibroblasts in microvascular network formation.

Perfusable microvascular networks offer promising three-dimensional in vitro models to study normal and compromised vascular tissues as well as phenomena such as cancer cell metastasis. Engineering of these microvascular networks generally involves the use of endothelial cells stabilized by fibroblasts to generate robust and stable vasculature. However, fibroblasts are highly heterogenous and may contribute variably to the microvascular structure. Here, we study the effect of normal and cancer-associated lung fibroblasts on the formation and function of perfusable microvascular networks. We examine the influence of cancer-associated fibroblasts on microvascular networks when cultured in direct (juxtacrine) and indirect (paracrine) contacts with endothelial cells, discovering a generative inhibition of microvasculature in juxtacrine co-cultures and a functional inhibition in paracrine co-cultures. Furthermore, we probed the secreted factors differential between cancer-associated fibroblasts and normal human lung fibroblasts, identifying several cytokines putatively influencing the resulting microvasculature morphology and functionality. These findings suggest the potential contribution of cancer-associated fibroblasts in aberrant microvasculature associated with tumors and the plausible application of such in vitro platforms in identifying new therapeutic targets and/or agents that can prevent formation of aberrant vascular structures.

Transcriptional and epigenetic regulators of human CD8+ T cell function identified through orthogonal CRISPR screens.

Clinical response to adoptive T cell therapies is associated with the transcriptional and epigenetic state of the cell product. Thus, discovery of regulators of T cell gene networks and their corresponding phenotypes has potential to improve T cell therapies. Here we developed pooled, epigenetic CRISPR screening approaches to systematically profile the effects of activating or repressing 120 transcriptional and epigenetic regulators on human CD8+ T cell state. We found that BATF3 overexpression promoted specific features of memory T cells and attenuated gene programs associated with cytotoxicity, regulatory T cell function, and exhaustion. Upon chronic antigen stimulation, BATF3 overexpression countered phenotypic and epigenetic signatures of T cell exhaustion. Moreover, BATF3 enhanced the potency of CAR T cells in both in vitro and in vivo tumor models and programmed a transcriptional profile that correlates with positive clinical response to adoptive T cell therapy. Finally, we performed CRISPR knockout screens that defined cofactors and downstream mediators of the BATF3 gene network.

Survivin Dendritic Cell Vaccine Safely Induces Immune Responses and Is Associated with Durable Disease Control after Autologous Transplant in Patients with Myeloma.

PURPOSE: We investigated whether a dendritic cell (DC) vaccine transduced with an adenoviral vector encoded with full-length survivin (Ad-S), with mutations neutralizing its antiapoptotic function, could safely generate an immune response and deepen clinical responses when administered before and after autologous stem cell transplant (ASCT) for multiple myeloma. PATIENTS AND METHODS: This phase I first-in-human trial (NCT02851056) evaluated the safety of DC:Ad-S in newly diagnosed multiple myeloma not having achieved complete response with induction, given 7 to 30 days prior to stem cell collection and 20 to 34 days after ASCT. Anti-survivin antibodies and CD4+ and CD8+ specific T cells were quantified. RESULTS: A total of 14 patients were treated and 13 included in the primary efficacy analysis. No serious adverse events were attributed to DC:Ad-S vaccine. Detectable anti-survivin antibodies increased from baseline in 9 of 13 (69%) patients, and 11 of 13 (85%) mounted either a cellular or humoral immune response to survivin. Seven patients had an improved clinical response at day +90, all of whom had mounted an immune response, and 6 of 7 patients remain event-free at a median follow-up of 4.2 years. Estimated progression-free survival at 4 years is 71% (95% confidence interval, 41-88). CONCLUSIONS: Two doses of DC:Ad-S, one given immediately before and another after ASCT, were feasible and safe. A high frequency of vaccine-specific immune responses was seen in combination with durable clinical outcomes, supporting ongoing investigation into the potential of this approach. See related commentary by Dhodapkar, p. 4524.

A phase Ib/II study of galunisertib in combination with nivolumab in solid tumors and non-small cell lung cancer.

BACKGROUND: In this phase Ib/II open-label study, tumor immune suppression was targeted in patients with advanced refractory solid tumors and patients with recurrent/refractory non-small cell lung cancer (NSCLC) using galunisertib with nivolumab. METHODS: Eligible patients were ≥ 18 years old, had an Eastern Cooperative Oncology Group performance status ≤ 1, and were treatment-naive for anti-programmed cell death-1, its ligand, or transforming growth factor ß receptor 1 kinase inhibitors. Phase Ib was an open-label, dose-escalation assessment of the safety and tolerability of galunisertib with nivolumab in patients with advanced refractory solid tumors. Phase II evaluated the safety of galunisertib with nivolumab in NSCLC patients who had received prior platinum-based treatment but were immuno-oncology agent-naive. RESULTS: This trial was conducted between October 2015 and August 2020. No dose-limiting toxicities were observed in phase I. In the phase II NSCLC cohort (n = 25), patients received 150 mg twice daily galunisertib (14 days on/14 days off dosing schedule for all phases) plus nivolumab at 3 mg/kg (intravenously every 2 weeks). In this phase, the most frequent treatment-related adverse events (AEs) were pruritus (n = 9, 36%), fatigue (n = 8, 32%), and decreased appetite (n = 7, 28%). No grade 4 or 5 treatment-related AEs were observed. Six (24%) patients had confirmed partial response (PR) and 4 (16%) had stable disease; 1 additional patient had confirmed PR after initial pseudo-progression. The median duration of response was 7.43 months (95% confidence interval [CI]: 3.75, NR). Among the 7 responders, including the delayed responder, 1 had high PD-L1 expression (≥ 50%). The median progression-free survival was 5.26 months (95% CI: 1.77, 9.20) and the median overall survival was 11.99 months (95% CI: 8.15, NR). Interferon gamma response genes were induced post-treatment and cell adhesion genes were repressed, although the association of these observations with tumor response and clinical outcomes was not statistically powered due to limited samples available. CONCLUSIONS: The study met its primary endpoint as galunisertib combined with nivolumab was well tolerated. Preliminary efficacy was observed in a subset of patients in the Phase 2 NSCLC cohort. TRIAL REGISTRATION: Trial registered with ClinicalTrials.gov (NCT02423343; 22.04.2015).

A phase I/IB trial of binimetinib in combination with erlotinib in NSCLC harboring activating KRAS or EGFR mutations.

BACKGROUND: Activating mutations in EGFR or KRAS are highly prevalent in NSCLC, share activation of the MAPK pathway and may be amenable to combination therapy to prevent negative feedback activation. METHODS: In this phase 1/1B trial, we tested the combination of binimetinib and erlotinib in patients with advanced NSCLC with at least 1 prior line of treatment (unless with activating EGFR mutation which could be treatment-naïve). A subsequent phase 1B expansion accrued patients with either EGFR- or KRAS-mutation using the recommended phase 2 dose (RP2D) from Phase 1. The primary objective was to evaluate the safety of binimetinib plus erlotinib and establish the RP2D. RESULTS: 43 patients enrolled (dose-escalation = 23; expansion = 20). 17 harbored EGFR mutation and 22 had KRAS mutation. The RP2D was erlotinib 100 mg daily and binimetinib 15 mg BID × 5 days/week. Common AEs across all doses included diarrhea (69.8%), rash (44.2%), fatigue (32.6%), and nausea (32.6%), and were primarily grade 1/2. Among KRAS mutant patients, 1 (5%) had confirmed partial response and 8 (36%) achieved stable disease as best overall response. Among EGFR mutant patients, 9 were TKI-naïve with 8 (89%) having partial response, and 8 were TKI-pretreated with no partial responses and 1 (13%) stable disease as best overall response. CONCLUSIONS: Binimetinib plus erlotinib demonstrated a manageable safety profile and modest efficacy including one confirmed objective response in a KRAS mutant patient. While clinical utility of this specific combination was limited, these results support development of combinations using novel small molecule inhibitors of RAS, selective EGFR- and other MAPK pathway inhibitors, many of which have improved therapeutic indices. CLINICAL TRIAL REGISTRATION: NCT01859026.

Brief Report: Safety and Antitumor Activity of Durvalumab Plus Tremelimumab in Programmed Cell Death-(Ligand)1-Monotherapy Pretreated, Advanced NSCLC: Results From a Phase 1b Clinical Trial.

INTRODUCTION: Although first-line immunotherapy approaches are standard, in patients with non-small cell lung cancer (NSCLC) previously treated with programmed cell death protein-1 or programmed death-(ligand)1 (PD-[L]1) inhibitors, the activity of combined CTLA-4 plus PD-(L)1 inhibition is unknown. This phase 1b study evaluated the safety and efficacy of durvalumab plus tremelimumab in adults with advanced NSCLC who received anti-PD-(L)1 monotherapy as their most recent line of therapy. METHODS: Patients with PD-(L)1-relapsed or refractory NSCLC were enrolled between October 25, 2013, and September 17, 2019. Durvalumab 20 mg/kg plus tremelimumab 1 mg/kg was administered intravenously every 4 weeks for four doses, followed by up to nine doses of durvalumab monotherapy every 4 weeks for up to 12 months of treatment or disease progression. Primary end points included safety and objective response rate (ORR) on the basis of Response Evaluation Criteria in Solid Tumors version 1.1 (RECIST v1.1) per blinded independent central review; secondary end points were ORR on the basis of RECIST v1.1 per investigator; duration of response, disease control, and progression-free survival on the basis of RECIST v1.1 per blinded independent central review and investigator; and overall survival. CLINICALTRIALS: gov identifier: NCT02000947. RESULTS: PD-(L)1-refractory (n = 38) and PD-(L)1-relapsed (n = 40) patients were treated. The most common treatment-related adverse events were fatigue (26.3%, PD-(L)1-refractory patients) and diarrhea (27.5%, PD-(L)1-relapsed patients). Grade 3 to 4 treatment-related adverse events occurred in 22 patients. Median follow-up duration was 43.6 months for PD-(L)1-refractory patients and 41.2 months for PD-(L)1-relapsed patients. The ORR was 5.3% for PD-(L)1-refractory patients (one complete response, one partial response) and 0% for PD-(L)1-relapsed patients. CONCLUSIONS: Durvalumab plus tremelimumab had a manageable safety profile, but the combination did not have efficacy after PD-(L)1 treatment failure.

Orthogonal CRISPR screens to identify transcriptional and epigenetic regulators of human CD8 T cell function.

The clinical response to adoptive T cell therapies is strongly associated with transcriptional and epigenetic state. Thus, technologies to discover regulators of T cell gene networks and their corresponding phenotypes have great potential to improve the efficacy of T cell therapies. We developed pooled CRISPR screening approaches with compact epigenome editors to systematically profile the effects of activation and repression of 120 transcription factors and epigenetic modifiers on human CD8+ T cell state. These screens nominated known and novel regulators of T cell phenotypes with BATF3 emerging as a high confidence gene in both screens. We found that BATF3 overexpression promoted specific features of memory T cells such as increased IL7R expression and glycolytic capacity, while attenuating gene programs associated with cytotoxicity, regulatory T cell function, and T cell exhaustion. In the context of chronic antigen stimulation, BATF3 overexpression countered phenotypic and epigenetic signatures of T cell exhaustion. CAR T cells overexpressing BATF3 significantly outperformed control CAR T cells in both in vitro and in vivo tumor models. Moreover, we found that BATF3 programmed a transcriptional profile that correlated with positive clinical response to adoptive T cell therapy. Finally, we performed CRISPR knockout screens with and without BATF3 overexpression to define co-factors and downstream factors of BATF3, as well as other therapeutic targets. These screens pointed to a model where BATF3 interacts with JUNB and IRF4 to regulate gene expression and illuminated several other novel targets for further investigation.

Combination IFNß and Membrane-Stable CD40L Maximize Tumor Dendritic Cell Activation and Lymph Node Trafficking to Elicit Systemic T-cell Immunity.

Oncolytic virus therapies induce the direct killing of tumor cells and activation of conventional dendritic cells (cDC); however, cDC activation has not been optimized with current therapies. We evaluated the adenoviral delivery of engineered membrane-stable CD40L (MEM40) and IFNß to locally activate cDCs in mouse tumor models. Combined tumor MEM40 and IFNß expression induced the highest cDC activation coupled with increased lymph node migration, increased systemic antitumor CD8+ T-cell responses, and regression of established tumors in a cDC1-dependent manner. MEM40 + IFNß combined with checkpoint inhibitors led to effective control of distant tumors and lung metastases. An oncolytic adenovirus (MEM-288) expressing MEM40 + IFNß  in phase I clinical testing induced cancer cell loss concomitant with enhanced T-cell infiltration and increased systemic presence of tumor T-cell clonotypes in non-small cell lung cancer (NSCLC) patients. This approach to simultaneously target two major DC-activating pathways has the potential to significantly affect the solid tumor immunotherapy landscape.

Brief Report: Durvalumab After Chemoradiotherapy in Unresectable Stage III EGFR-Mutant NSCLC: A Post Hoc Subgroup Analysis From PACIFIC.

INTRODUCTION: Consolidation durvalumab (the "PACIFIC regimen") is standard of care for patients with unresectable stage III NSCLC who have not progressed after chemoradiotherapy, on the basis of data from the phase 3 placebo-controlled PACIFIC study (NCT02125461). Nevertheless, the benefit of immunotherapy in patients with stage III EGFR-mutant (EGFRm) NSCLC is not well characterized. Here, we report a post hoc exploratory efficacy and safety analysis from a subgroup of patients with EGFRm NSCLC from the PACIFIC. METHODS: Patients with stage III unresectable NSCLC and no progression after more than or equal to two cycles of platinum-based concurrent chemoradiotherapy were randomized (2:1) to receive durvalumab (10 mg/kg intravenously every 2 weeks [wk], for up to 1 y) or placebo; stratified by age, sex, and smoking history. Enrollment was not restricted by oncogenic driver gene mutation status or programmed death-ligand 1 expression. Patients with NSCLC with an EGFR mutation, determined by local testing only, were included in this subgroup analysis. The primary end points were progression-free survival (PFS; assessed by blinded independent central review) and overall survival (OS). Secondary end points included objective response rate and safety. Statistical analyses for the subgroup of patients with EGFRm NSCLC were post hoc and considered exploratory. RESULTS: Of 713 patients randomized, 35 had locally confirmed EGFRm NSCLC (durvalumab, n = 24; placebo, n = 11). At data cutoff (January 11, 2021), median duration of follow-up for survival was 42.7 months (range: 3.7-74.3 mo) for all randomized patients in the subgroup. Median PFS was 11.2 months (95% confidence interval [CI]: 7.3-20.7) with durvalumab versus 10.9 months (95% CI: 1.9-not evaluable [NE]) with placebo; hazard ratio = 0.91 (95% CI: 0.39-2.13). Median OS was 46.8 months (95% CI: 29.9-NE) with durvalumab versus 43.0 months (95% CI: 14.9-NE) with placebo; hazard ratio = 1.02 (95% CI: 0.39-2.63). The safety profile of durvalumab was generally consistent with the overall population and known profile for durvalumab. CONCLUSIONS: PFS and OS outcomes with durvalumab were similar to placebo for patients with EGFRm tumors, with wide CIs. These data should be interpreted with caution owing to small patient numbers and lack of a prospective study that evaluates clinical outcomes by tumor biomarker status. Further research to determine the optimal treatment for unresectable stage III EGFRm NSCLC is warranted.

Dose-Limiting Pulmonary Toxicity in a Phase 1/2 Study of Radiation and Chemotherapy with Ipilimumab Followed by Nivolumab for Patients With Stage 3 Unresectable Non-Small Cell Lung Cancer.

PURPOSE: We hypothesized that concurrent ipilimumab with chemoradiationtherapy (chemoRT) followed by maintenance nivolumab would be safe for patients with unresectable stage III non-small cell lung cancer (NSCLC). We aimed to assess the safety (phase 1) and the 12-month progression-free survival (PFS) (phase 2) in a multi-institution prospective trial. METHODS AND MATERIALS: Eligible patients had unresectable stage III NSCLC. The treatment included platinum doublet chemotherapy with concurrent thoracic radiation therapy to 60 Gy in 30 fractions and ipilimumab (1 mg/kg) delivered during weeks 1 and 4. After chemoRT, maintenance nivolumab (480 mg) was given every 4 weeks for up to 12 cycles. Adverse events (AEs) were assessed according to the Common Terminology Criteria for Adverse Events, version 5.0. Survival analyses were performed with Kaplan Meier (KM) methods and log-rank tests. RESULTS: The trial was discontinued early after enrolling 19 patients without proceeding to the phase 2 component because of unacceptable toxicity. Sixteen patients (84%) had grade ≥3 (G3+) possible treatment-related toxicity, most commonly pulmonary AEs (n = 8, 42%). Fourteen patients (74%) discontinued study therapy early because of AEs (n = 12, 63%) or patient choice (n = 2, 11%). Eleven patients (58%) experienced G2+ pulmonary toxicity with median time to onset 4.1 months (95% CI 2.6-not reached [NR]), and 12-month freedom from G2+ pulmonary toxicity 37% (95% CI, 16-59). Five patients had G5 AEs, including 3 with G5 pulmonary AEs (1 respiratory failure with pneumonitis and pulmonary embolism, 1 pneumonia/chronic obstructive pulmonary disease exacerbation, 1 pulmonary fibrosis). Despite toxicities, the median PFS was 19.2 months (95% CI 6.1-NR) and the median overall survival was NR (95% CI 6.1-NR) with median follow-up of 30.1 months by the reverse KM method. CONCLUSIONS: Concurrent ipilimumab with chemoRT for unresectable stage III NSCLC is associated with pulmonary toxicity that may limit opportunities for improved outcomes. Future studies aiming to incorporate ipilimumab or other anti-CTLA4 therapies into management of unresectable stage III NSCLC should consider careful measures to minimize toxicity risk.

High Yield of Pleural Cell-Free DNA for Diagnosis of Oncogenic Mutations in Lung Adenocarcinoma.

BACKGROUND: Pleural cytology is currently used to assess targetable mutations in patients with advanced lung adenocarcinoma. However, it is fraught with low diagnostic yield. RESEARCH QUESTION: Can pleural cell-free DNA (cfDNA) be used to assess targetable mutations in lung adenocarcinoma patients with malignant pleural effusions (MPE)? STUDY DESIGN AND METHODS: Patients with lung adenocarcinoma MPE were recruited prospectively between January 2017 and September 2021. Oncogenic mutations were assessed by treating providers using pleural fluid cytology or lung cancer biopsies. Pleural and plasma cfDNA were used to assess the mutations using next-generation sequencing (NGS). RESULTS: Fifty-four pleural fluid samples were collected from 42 patients. The diagnostic yield to detect oncogenic mutations for pleural cfDNA, pleural cytology, biopsy, and plasma cfDNA was 49/54 (90.7%), 16/33 (48.5%), 22/25 (88%), and 24/32 (75%), respectively, P < .001. The agreement of mutations in positive samples between pleural cfDNA and pleural cytology was 100%, whereas the agreement of pleural cfDNA with biopsies was 89.4%. The median concentration (interquartile range) of pleural cfDNA was higher than plasma: 28,444 (4,957-67,051) vs 2,966.5 (2,167-5,025) copies of amplifiable DNA per mL, P < .01. Median of 5 mL (interquartile range, 4.5-5) of pleural fluid supernatant was adequate for cfDNA testing. INTERPRETATION: The diagnostic yield of pleural cfDNA NGS for oncogenic mutations in lung adenocarcinoma patients is comparable to tumor biopsies and higher than pleural cytology and plasma cfDNA. The pleural cfDNA can be longitudinally collected, can be readily incorporated in clinical workflow, and may decrease the need for additional biopsies.

Patient-Reported Distress and Clinical Outcomes with Immuno-Oncology Agents in Metastatic Non-Small Cell Lung Cancer (mNSCLC): A Real-World Retrospective Cohort Study.

OBJECTIVES: There are limited real-world data about patient-reported outcomes with immunotherapies (IO) in metastatic non-small cell lung cancer (mNSCLC). We describe patient-reported distress and clinical outcomes with IO-based treatments or cytotoxic chemotherapies (Chemo). METHODS: We conducted a single-institution retrospective chart review of adults with mNSCLC treated at Duke from 03/2015 to 06/2020. At each visit, patients self-reported their distress level and sources of distress using the NCCN Distress Thermometer (DT) and its 39-item Problem List. We abstracted demographic, clinical, distress, and investigator assessed-clinical response data, then analyzed these using descriptive statistics and generalized estimating equations. RESULTS: Data from 152 patients were analyzed in four groups: Chemo alone, IO + Chemo, single agent IO, dual agent IO. Distress was worse before treatment start in all groups, and the odds of actionable distress (DT score > 4) decreased by 10 % per month. The most frequent sources of distress were physical symptoms (e.g., fatigue, pain), which remained high longitudinally. Patients receiving IO had higher clinical response rates and a lower rate of unplanned healthcare encounters compared to patients treated with Chemo alone. Only one-third of all patients were seen by palliative care. CONCLUSIONS: This single-center, real-world evidence study demonstrates that patients with mNSCLC experience significant distress prior to starting first-line treatment. IO treatment was associated with higher clinical benefit rates and lower healthcare utilization compared to chemotherapy. Symptom distress persists over time, highlighting potential unmet palliative and supportive care needs in mNSCLC care in the IO treatment era.

Anti-4-1BB immunotherapy enhances systemic immune effects of radiotherapy to induce B and T cell-dependent anti-tumor immune activation and improve tumor control at unirradiated sites.

Radiation therapy (RT) can prime and boost systemic anti-tumor effects via STING activation, resulting in enhanced tumor antigen presentation and antigen recognition by T cells. It is increasingly recognized that optimal anti-tumor immune responses benefit from coordinated cellular (T cell) and humoral (B cell) responses. However, the nature and functional relevance of the RT-induced immune response are controversial, beyond STING signaling, and agonistic interventions are lacking. Here, we show that B and CD4+ T cell accumulation at tumor beds in response to RT precedes the arrival of CD8+ T cells, and both cell types are absolutely required for abrogated tumor growth in non-irradiated tumors. Further, RT induces increased expression of 4-1BB (CD137) in both T and B cells; both in preclinical models and in a cohort of patients with small cell lung cancer treated with thoracic RT. Accordingly, the combination of RT and anti-41BB therapy leads to increased immune cell infiltration in the tumor microenvironment and significant abscopal effects. Thus, 4-1BB therapy enhances radiation-induced tumor-specific immune responses via coordinated B and T cell responses, thereby preventing malignant progression at unirradiated tumor sites. These findings provide a rationale for combining RT and 4-1bb therapy in future clinical trials.

Peroxynitrite in the tumor microenvironment changes the profile of antigens allowing escape from cancer immunotherapy.

Cancer immunotherapy often depends on recognition of peptide epitopes by cytotoxic T lymphocytes (CTLs). The tumor microenvironment (TME) is enriched for peroxynitrite (PNT), a potent oxidant produced by infiltrating myeloid cells and some tumor cells. We demonstrate that PNT alters the profile of MHC class I bound peptides presented on tumor cells. Only CTLs specific for PNT-resistant peptides have a strong antitumor effect in vivo, whereas CTLs specific for PNT-sensitive peptides are not effective. Therapeutic targeting of PNT in mice reduces resistance of tumor cells to CTLs. Melanoma patients with low PNT activity in their tumors demonstrate a better clinical response to immunotherapy than patients with high PNT activity. Our data suggest that intratumoral PNT activity should be considered for the design of neoantigen-based therapy and also may be an important immunotherapeutic target.

IGF-binding proteins secreted by cancer-associated fibroblasts induce context-dependent drug sensitization of lung cancer cells.

Cancer-associated fibroblasts (CAFs) in the tumor microenvironment are often linked to drug resistance. Here, we found that coculture with CAFs or culture in CAF-conditioned medium unexpectedly induced drug sensitivity in certain lung cancer cell lines. Gene expression and secretome analyses of CAFs and normal lung-associated fibroblasts (NAFs) revealed differential abundance of insulin-like growth factors (IGFs) and IGF-binding proteins (IGFBPs), which promoted or inhibited, respectively, signaling by the receptor IGF1R and the kinase FAK. Similar drug sensitization was seen in gefitinib-resistant, EGFR-mutant PC9GR lung cancer cells treated with recombinant IGFBPs. Conversely, drug sensitivity was decreased by recombinant IGFs or conditioned medium from CAFs in which IGFBP5 or IGFBP6 was silenced. Phosphoproteomics and receptor tyrosine kinase (RTK) array analyses indicated that exposure of PC9GR cells to CAF-conditioned medium also inhibited compensatory IGF1R and FAK signaling induced by the EGFR inhibitor osimertinib. Combined small-molecule inhibition of IGF1R and FAK phenocopied the CAF-mediated effects in culture and increased the antitumor effect of osimertinib in mice. Cells that were osimertinib resistant and had MET amplification or showed epithelial-to-mesenchymal transition also displayed residual sensitivity to IGFBPs. Thus, CAFs promote or reduce drug resistance in a context-dependent manner, and deciphering the relationship between the differential content of CAF secretomes and the signaling dependencies of the tumor may reveal effective combination treatment strategies.

Characterizing the Shifting Real-World Treatment Landscape by PD-L1 Testing Status and Expression Level in Advanced Non-Small Cell Lung Cancer.

INTRODUCTION: Contemporary real-world data on advanced non-small cell lung cancer (aNSCLC) treatment patterns across programmed cell death-ligand 1 (PD-L1) expression levels and testing status are limited. METHODS: A retrospective cohort was selected of adults newly diagnosed with aNSCLC between January 1, 2018, and July 31, 2021, who initiated first-line treatments, which were described by PD-L1 status and expression levels (≥ 50%, 1-49%, < 1%). Treatment received before and after PD-L1 test results were described for patients initiating first-line treatment before PD-L1 results. For patients who initiated chemotherapy alone before PD-L1 results, the probability of receiving immune checkpoint inhibitors (ICIs) after PD-L1 results was estimated by PD-L1 level and associated factors were explored. RESULTS: Among 12,202 patients with aNSCLC initiating first-line treatment [54.7% male, mean (standard deviation) age 69.2 (9.4) years], the most common therapies were ICI-based regimens across PD-L1 levels, and chemotherapy alone among PD-L1-untested patients. Use of chemotherapy alone decreased between 2018 and 2019 and stabilized thereafter, accounting for 21-29% of first-line treatments across PD-L1 levels and 48% of untested patients in 2021. Of 1468 patients initiating first-line treatment before PD-L1 results, treatments remained unchanged in most patients after PD-L1 results. Among patients initiating chemotherapy alone before PD-L1 results, the probability of receiving ICIs within 45 days after test results was 40.5% [95% confidence interval (CI) 31.6-48.3%], 28.6% (95% CI 20.3-36.0%), and 22.9% (95% CI 16.9-28.4%) at PD-L1 ≥ 50%, 1-49%, and < 1%, respectively. CONCLUSION: While ICI-based regimens accounted for most first-line treatments across PD-L1 levels, chemotherapy alone was initiated in > 20% of patients tested for PD-L1 and 48% of untested patients in 2021. Patients who initiated chemotherapy alone had a low probability of receiving ICIs after PD-L1 test results. These results highlight the need for understanding the role and timing of PD-L1 test results for informing treatment decisions for patients with aNSCLC.

Mistic: An open-source multiplexed image t-SNE viewer.

Understanding the complex ecology of a tumor tissue and the spatiotemporal relationships between its cellular and microenvironment components is becoming a key component of translational research, especially in immuno-oncology. The generation and analysis of multiplexed images from patient samples is of paramount importance to facilitate this understanding. Here, we present Mistic, an open-source multiplexed image t-SNE viewer that enables the simultaneous viewing of multiple 2D images rendered using multiple layout options to provide an overall visual preview of the entire dataset. In particular, the positions of the images can be t-SNE or UMAP coordinates. This grouped view of all images allows an exploratory understanding of the specific expression pattern of a given biomarker or collection of biomarkers across all images, helps to identify images expressing a particular phenotype, and can help select images for subsequent downstream analysis. Currently, there is no freely available tool to generate such image t-SNEs.

A Phase 2 Clinical Trial of Combination Nivolumab, Ipilimumab, and Paclitaxel in Patients With Untreated Metastatic NSCLC: The OPTIMAL Trial.

Introduction: Most patients with advanced NSCLC will experience disease progression and death within 2 years. Novel approaches are needed to improve outcomes. Methods: We conducted an open-label, nonrandomized, phase 2 trial in patients with treatment-naive, advanced NSCLC to assess the safety and efficacy of nivolumab 360 mg every 3 weeks, ipilimumab 1 mg/kg every 6 weeks, and four to six cycles of paclitaxel 80 mg/m2 on days 1 and 8 of every 21-day treatment. The primary end point of the study was median progression-free survival (PFS), with secondary end points of safety, objective response rate, and median overall survival (OS). Results: A total of 46 patients underwent consent and received treatment. The median age was 66 (range: 48-82) years, most had adenocarcinoma (63%), and 50% (23) had programmed death-ligand 1 greater than or equal to 1%. The median follow-up on the study as of October 2021 was 19 months. The primary end point of median PFS was 9.4 months (95% confidence interval [CI]: 5.9-16.6) in all patients regardless of programmed death-ligand 1 expression. The objective response rate for patients in the study was 47.8% (95% CI: 33.4-62.3). The 12-month OS rate was 69.5% (95% CI: 53%-81%), and median OS was not yet reached. Treatment-related grade greater than or equal to 3 adverse events was found in 54.3% of the patients. Conclusions: The toxicity observed was consistent with other reported chemo-immunotherapeutic combinations and was manageable. The primary end point of exceeding median PFS of 9 months was achieved with nivolumab, ipilimumab, and weekly paclitaxel and should be evaluated further in a randomized trial.

Patient-derived micro-organospheres enable clinical precision oncology.

Patient-derived xenografts (PDXs) and patient-derived organoids (PDOs) have been shown to model clinical response to cancer therapy. However, it remains challenging to use these models to guide timely clinical decisions for cancer patients. Here, we used droplet emulsion microfluidics with temperature control and dead-volume minimization to rapidly generate thousands of micro-organospheres (MOSs) from low-volume patient tissues, which serve as an ideal patient-derived model for clinical precision oncology. A clinical study of recently diagnosed metastatic colorectal cancer (CRC) patients using an MOS-based precision oncology pipeline reliably assessed tumor drug response within 14 days, a timeline suitable for guiding treatment decisions in the clinic. Furthermore, MOSs capture original stromal cells and allow T cell penetration, providing a clinical assay for testing immuno-oncology (IO) therapies such as PD-1 blockade, bispecific antibodies, and T cell therapies on patient tumors.