Efficacy evaluation of anticancer agents in single-arm clinical trials: analysis of review reports from Pharmaceuticals and Medical Devices Agency.

BACKGROUND: Comparative studies often cannot be conducted for cancer types with a small patient population. We reviewed the efficacy evaluations of new drug approvals in Japan based on the results of single-arm clinical trials. METHODS: We reviewed review reports on anticancer agents approved in Japan between 2006 and 2019 that are publicly available on the website of the Pharmaceuticals and Medical Devices Agency, Japan. RESULTS: The number of single-arm trial-based approvals increased, with a total of 43 drugs approved during the study period. Compared with comparative trial-based approvals, single-arm trial-based approvals had a tendency toward more biomarker-related indications (37.2% vs 22.6%, p = .053), as well as more approvals for hematological malignancies, orphan designation, and response-related outcomes as the primary endpoint. Only 13 of the pivotal trials of single-arm trial-based approvals had a predefined threshold for efficacy based on the same target population as the pivotal trial, and nearly half of the trials did not have an appropriate predefined threshold for efficacy. In particular, the efficacy thresholds for clinical trials for 4 molecular targeted agents were set based on the results of the nonbiomarker-selected population. CONCLUSIONS: Evidence on standard cancer therapies for rare molecular subtypes is lacking. External control data from registries might support the efficacy evaluations of new drugs for newly established rare molecular subtypes.

Relationship between the immune microenvironment of different locations in a primary tumour and clinical outcomes of oesophageal squamous cell carcinoma.

BACKGROUND: Tumour microenvironments can differ according to intratumoural locations. We investigated the immune status at different locations in primary tumours and its clinical significance in oesophageal squamous cell carcinoma (ESCC). METHODS: The number of CD8+ tumour-infiltrating immune cells (TIICs) and PD-1+ TIICs, and PD-L1 expression on tumour cells (PD-L1TC) were immunohistochemically examined in the surface (Surf), centre (Cent) and invasive front (Inv) of tumours surgically resected from 192 patients with ESCC. RESULTS: The PD-L1+ rate was lower in Inv than in Cent (12.0% vs. 18.2%, P = 0.012), although the numbers of CD8+ TIICs and PD-1+ TIICs were comparable among intratumoural locations. High numbers of CD8+ and PD-1+ TIICs and positive PD-L1TC were related to better overall survival (OS) only in Surf and Cent (CD8: P = 0.012 in Surf, 0.018 in Cent, and 0.165 in Inv; PD-1: P = 0.028 in Surf, 0.021 in Cent, and 0.208 in Inv; and PD-L1: 0.044 in Surf, 0.026 in Cent, and 0.718 in Inv). Positive PD-L1TC in Surf and/or Cent but not in Inv demonstrated a strong tendency toward better OS (P = 0.053). CONCLUSIONS: Immune microenvironments according to the intratumoural location have different effects on the survival of patients with ESCC.

Risk factors for febrile neutropenia in neoadjuvant docetaxel, cisplatin, and 5-fluorouracil chemotherapy for esophageal cancer.

PURPOSE: The aim of the present study was to evaluate the incidence and explore the risk factors of febrile neutropenia (FN) in patients with esophageal cancer receiving neoadjuvant docetaxel, cisplatin, and 5-fluorouracil (DCF) chemotherapy in real-world settings. METHODS: We retrospectively reviewed clinical data of 128 consecutive patients with esophageal cancer. Specifically, these patients underwent neoadjuvant DCF chemotherapy with prophylactic antibiotic administration at our institution between July 2009 and January 2015. Two FN-related endpoints were set as follows: definite FN (dFN) defined as grade 4 neutropenia at the onset of fever and clinically suspected FN (csFN), which included both patients with dFN and patients without grade 4 neutropenia but with deteriorating grade 3 neutropenia at the onset of fever who were clinically diagnosed with FN for which they underwent treatment. The risk factors for dFN and csFN were evaluated based on patients' characteristics. RESULTS: A total of 72 (56.3%) patients developed grade 3 or grade 4 neutropenia and 26 (20.3%) developed csFN including 14 (10.9%) with dFN. Multivariate analysis revealed that older age (OR 3.57, CI 1.27-10.1, P = 0.016) and living alone (OR 5.17, 95% CI 1.26-21.3, P = 0.023) were statistically significant risk factors for csFN development. As to dFN, no statistically significant risk factors were identified. CONCLUSIONS: Older age and living alone are significant risk factors for developing FN, and thus, particularly for patients with risk factors for FN, G-CSF should be considered instead of prophylactic antibiotics with careful observation.

The Tumor Microenvironment of Bladder Cancer.

Bladder cancer has been well known as immunotherapy-responsive disease as intravesical therapy with BCG has been the standard of care for non-muscle invasive disease for several decades. In addition, immune checkpoint inhibitors have dramatically changed the treatment of metastatic bladder cancer. However, only a small fraction of patients with bladder cancer can benefit from these therapies. As immunotherapies act on the tumor microenvironment, understanding it is essential to expand the efficacy of modern treatments. The bladder cancer microenvironment consists of various components including tumor cells, immune cells, and other stromal cells, affecting each other via immune checkpoint molecules, cytokines, and chemokines. The development of an antitumor immune response depends on tumor antigen recognition by antigen presenting cells and priming and recruitment of effector T cells. Accumulated evidence shows that these processes are impacted by multiple types of immune cells in the tumor microenvironment including regulatory T cells, tumor-associated macrophages, and myeloid derived suppressor cells. In addition, recent advances in genomic profiling have shed light on the relationship between molecular subtypes and the tumor microenvironment. Finally, emerging evidence has shown that multiple factors can impact the tumor microenvironment in bladder cancer, including tumor-oncogenic signaling, patient genetics, and the commensal microbiome.

Review of early endoscopic findings in patients with local recurrence after definitive chemoradiotherapy for esophageal squamous cell carcinoma.

BACKGROUNDS: Local recurrence after definitive chemoradiotherapy, if diagnosed early, can be cured by salvage endoscopic therapy, which allows organ preservation and contributes to maintaining patient quality of life. This study aimed to investigate early endoscopic findings of local recurrence in post-definitive chemoradiotherapy patients. METHODS: Between January 2008 and June 2012, 17 esophageal squamous cell carcinoma patients with no metastasis but local recurrence after definitive chemoradiotherapy were enrolled. We attempted to find endoscopic hallmarks suggestive of local recurrence by comparing pre- and post-local recurrence diagnostic images. The influence of follow-up schedule on chosen salvage therapy type was also investigated. RESULTS: Endoscopic local recurrence findings included eight submucosal tumors, five ulcers, and four erosions. Upon review of prior images, findings suggestive of local recurrence were detected in seven patients, including six submucosal tumors and one erosion, all of which were smaller than 10 mm. These lesions had changed morphologically at local recurrence diagnosis: three submucosal tumors had become larger and three submucosal tumors and one erosion had changed to ulcers. Of 12 patients with cT1 at local recurrence, four (33%) underwent follow-up endoscopy within 1 month of local recurrence findings and 11 patients (92%) were treated with salvage endoscopic therapy. CONCLUSIONS: Endoscopists should be aware that SMTs or erosions, even those smaller than 10 mm, can indicate local recurrence after complete response to definitive chemoradiotherapy. Follow-up endoscopy should be performed within 1-2 months if findings suggestive of local recurrence are observed on prior endoscopy, even when biopsy results are negative.

Pathological tumor regression grade of metastatic tumors in lymph node predicts prognosis in esophageal cancer patients.

Tumor regression grade of the primary tumor (TRG-PT) and residual lymph node metastasis have been pathologically determined in esophageal squamous cell carcinoma (ESCC) patients who had received neoadjuvant chemotherapy (nCT) followed by surgery; however, TRG of the metastatic tumor involving lymph nodes (LN) has not yet been determined. The aim of the present study was to clarify the impact of TRG on the prognosis of ESCC patients. ESCC patients (n = 110) who had received nCT followed by surgery were enrolled. Dissected LN were classified into 2 categories: plausible positive metastatic LN (pp-MLN) where viable and/or degenerated ESCC cells and/or tissue modifications were observed, and non-metastatic LN (non-MLN) where neither of them was observed. We defined nCT-effective rate (CER) as the ratio of the number of pp-MLN that showed tumor regression to the total number of pp-MLN, and divided CER into low-CER (LCER; ≥0% and <50%) and high-CER (HCER; ≥50% and ≤100%). Relationships between CER and clinicopathological factors including prognosis were then examined. Multivariate analyses of 110 patients indicated that ypT3-4 (P = .023, HR; 2.551), positive venous infiltration (P = .006, HR; 3.526), and LCER (P = .033, HR; 1.922) were independently associated with shorter recurrence-free survival (RFS). Multivariate analyses of 43 patients with grade 0 TRG-PT showed that ypT3-4 (P = .033, HR; 3.397) and LCER (P = .008, HR; 3.543) were independently associated with shorter RFS. This study showed that CER was one of the prognostic factors for ESCC patients who had received nCT followed by surgery.

Salvage endoscopic resection (ER) after chemoradiotherapy for esophageal squamous cell carcinoma: What are the risk factors for recurrence after salvage ER?

BACKGROUND AND AIM: Salvage endoscopic resection (ER) is among the curative treatments for superficial local failure after chemoradiotherapy (CRT) for esophageal squamous cell carcinoma (ESCC). The present study aimed to clarify risk factors for recurrence after salvage ER. METHODS: This study enrolled consecutive ESCC patients treated with salvage ER for local failure after CRT between 1998 and 2013. Recurrences after salvage ER included locoregional recurrences and distant metastases. Multivariate analysis was carried out on clinicopathological parameters to identify risk factors for post-salvage ER recurrence. RESULTS: Of the 72 patients enrolled in this study, 37/8/23/4 patients had been staged before CRT as cT1/T2/T3/T4 and 44/28 patients as cN0/N1, respectively, and local failures detected before salvage ER were residual lesions after CRT in 19 and local recurrences in 53 patients. Resected specimens were classified as pT1a (M) in 45 and pT1b (SM) in 27 patients. During the median 45-month follow up (range, 3-175 months) after salvage ER, 27 (38%) patients developed recurrence with a 3-year recurrence-free survival rate of 48.9% (95% confidence interval [CI], 36.5-60.3). Multivariate analysis showed that residual lesions after CRT (HR, 2.55; 95% CI, 1.32-4.94) and lesions with a submucosal tumor (SMT)-like appearance before salvage ER (HR, 2.08; 95% CI, 1.04-4.18) were significantly associated with post-salvage ER recurrence. CONCLUSIONS: Clinical findings (e.g. residual tumors found immediately after CRT and macroscopic SMT-like appearance before salvage ER) were shown to be significant risk factors for post-salvage ER recurrence.

Concordance between PIK3CA mutations in endoscopic biopsy and surgically resected specimens of esophageal squamous cell carcinoma.

BACKGROUND: PIK3CA mutations are expected to be potential therapeutic targets for esophageal squamous cell carcinoma (ESCC). We aimed to clarify the concordance between PIK3CA mutations detected in endoscopic biopsy specimens and corresponding surgically resected specimens. METHODS: We examined five hotspot mutations in the PIK3CA gene (E542K, E545K, E546K, H1047R, and H1047L) in formalin-fixed and paraffin-embedded tissue sections of paired endoscopic biopsy and surgically resected specimens from 181 patients undergoing curative resection for ESCC between 2000 and 2011 using a Luminex technology-based multiplex gene mutation detection kit. RESULTS: Mutation analyses were successfully performed for both endoscopic biopsy and surgically resected specimens in all the cases. A PIK3CA mutation was detected in either type of specimen in 13 cases (7.2%, 95% confidence interval: 3.9-12.0). The overall concordance rate, positive predictive value, and negative predictive value were 98.3% (178/181), 90.9% (10/11), and 98.8% (168/170), respectively. Among patients with a PIK3CA mutation detected in both types of specimens, the concordance between PIK3CA mutation genotypes was 100%. There were three cases with a discordant mutation status between the types of specimens (PIK3CA mutation in surgically resected specimen and wild-type in biopsy specimen in two cases, and the opposite pattern in one case), suggesting possible intratumoral heterogeneity in the PIK3CA mutation status. CONCLUSIONS: The PIK3CA mutation status was highly concordant between endoscopic biopsy and surgically resected specimens from the same patient, suggesting that endoscopic biopsy specimens can be clinically used to detect PIK3CA mutations in patients with ESCC.

Salvage photodynamic therapy for local failure after chemoradiotherapy for esophageal squamous cell carcinoma.

BACKGROUND AND AIMS: Photodynamic therapy (PDT) is a less-invasive salvage treatment option for local failure at the primary site after chemoradiotherapy (CRT) for esophageal squamous cell carcinoma. The objective of this study was to clarify the long-term outcomes and prognostic factors of salvage PDT. METHODS: One hundred thirteen consecutive patients treated in our institution with PDT for local failure limited to within T2 without any metastases after definitive CRT performed between 1998 and 2008 were retrospectively enrolled. The complete response rate, adverse events, and survival outcomes were assessed and prognostic factors were investigated using a multivariate analysis. RESULTS: The complete response rate was 58.4% (95% confidence interval [CI], 49.3%-67.5%). The progression-free survival (PFS) and the overall survival (OS) rates at 5 years after salvage PDT were 22.1% (95% CI, 14.3%-30.0%) and 35.9% (95% CI, 26.7%-45.1%). N0 before CRT was significantly associated with OS (hazard ratio [HR], 0.54; 95% CI, 0.33-0.91, P = .020), whereas the impact of T1 or T2 before CRT on PFS (HR, 0.63; 95% CI, 0.38-1.04, P = .068) and that of a longer period between CRT and PDT on OS (HR, 0.64; 95% CI, 0.39-1.05, P = .078) were marginal. The treatment-related death rate was 1.8%. CONCLUSIONS: Salvage PDT was found to have a superior outcome and a satisfactory safety profile. An earlier clinical stage before CRT and a longer interval between CRT and PDT may be associated with a longer survival period.

Prognostic significance of tumor regression grade for patients with esophageal squamous cell carcinoma after neoadjuvant chemotherapy followed by surgery.

BACKGROUND AND OBJECTIVES: To clarify prognostic factors for the patients with esophageal squamous cell carcinoma (ESCC) through an assessment of surgically resected specimens modified by neoadjuvant chemotherapy (nCT). METHODS: We retrospectively reviewed the clinicopathological data of 143 consecutive patients with ESCC who underwent nCT followed by surgery between 2008 and 2012 at our institution and conducted survival analysis. The tumor regression grade (TRG) was classified based on the proportion of residual tumor cells in the area where the tumor was thought to have existed before nCT as follows: Grade 0 (no therapeutic effect), Grade 1a (residual tumor cells ≥2/3), Grade 1b (1/3≤ residual tumor cells <2/3), Grade 2 (residual tumor cells <1/3), and Grade 3 (no residual tumor). RESULTS: The 3-year OS and RFS of patients with tumor regression grade 0/1a/1b-3 were 53.6%/73.3%/88.6% and 37.7%/60.5%/83.8%, respectively. A multivariate analysis demonstrated that TRG was an independent predictor of OS (TRG 1a-3: HR, 0.46; 95%CI, 0.23-0.89), in addition to venous invasion, and of RFS (TRG 1a-3: HR, 0.49; 95%CI, 0.28-0.84), in addition to ypT factor, and venous invasion. CONCLUSIONS: TRG is a critical prognostic factor in patients with ESCC who had undergone nCT followed by surgery. J. Surg. Oncol. 2016;113:390-396. © 2016 Wiley Periodicals, Inc.

[Molecular Subtypes of Gastric Cancer].

Gastric cancer has been classified based on the pathological characteristics including microscopic configuration and growth pattern. Although these classifications have been used in studies investigating prognosis and recurrence pattern, they are not considered for decisions regarding the therapeutic strategy. In the ToGA study, trastuzumab, an anti-HER2 monoclonal antibody, demonstrated clinical efficacy for gastric cancer with HER2 overexpression or HER2 gene amplification. Based on these findings of the ToGA study, the definition of HER2-positive gastric cancer was established. Thereafter, several molecular targeted agents, including agents targeting other receptor tyrosine kinases, have been investigated in gastric cancer. However, to date no biomarker, except HER2, has been established. Based on the recent technological development in the field of gene analysis, a comprehensive molecular evaluation of gastric cancer was performed as part of The Cancer Genome Atlas (TCGA) project, and a new molecular classification was proposed that divided gastric cancer into the following 4 subtypes: tumors positive for Epstein-Barr virus, microsatellite instability tumors, genomically stable tumors, and tumors with chromosomal instability. Each subtype has specific molecular alterations including gene mutation and amplification, DNA methylation, and protein overexpression. Additionally, some subtypes were suggested to be correlated with the clinicopathological characteristics or as targets of some molecular targeted agents that are currently under development. The new molecular classification is expected to be a roadmap for patient stratification and clinical trials on molecular targeted therapies in gastric cancer.

Batf3+ DCs and the 4-1BB/4-1BBL axis are required at the effector phase in the tumor microenvironment for PD-1/PD-L1 blockade efficacy.

The cellular source of positive signals that reinvigorate T cells within the tumor microenvironment (TME) for the therapeutic efficacy of programmed death-1 (PD-1)/programmed death-ligand 1 (PD-L1) blockade has not been clearly defined. We now show that Batf3-lineage dendritic cells (DCs) are essential in this process. Flow cytometric analysis, gene-targeted mice, and blocking antibody studies revealed that 4-1BBL is a major positive co-stimulatory signal provided by these DCs within the TME that translates to CD8+ T cell functional reinvigoration and tumor regression. Immunofluorescence and spatial transcriptomics on human tumor samples revealed clustering of Batf3+ DCs and CD8+ T cells, which correlates with anti-PD-1 efficacy. In addition, proximity to Batf3+ DCs within the TME is associated with CD8+ T cell transcriptional states linked to anti-PD-1 response. Our results demonstrate that Batf3+ DCs within the TME are critical for PD-1/PD-L1 blockade efficacy and indicate a major role for the 4-1BB/4-1BB ligand (4-1BBL) axis during this process.

[Timely colonoscopy leads to faster identification of bleeding diverticulum].

We conducted a retrospective study of the efficacy of a polyethylene glycol purge before colonoscopic examination in 110 patients with colonic diverticular bleeding. The patients' data were assessed for the timing of colonoscopy and the methods used to stop bleeding. The rate at which bleeding diverticula were identified was markedly higher when a purge was used than when it was not (28.2% vs. 12.0%, p=0.11). In addition, the identification rate was significantly higher when colonoscopic examination was performed within 18 hours of the final hematochezia than when it was performed after 18 hours (40.5% vs. 10.5%, p<0.01). These findings suggest that patients with diverticular bleeding should undergo colonoscopy following an orally administered colonic purge, providing their condition permits. Furthermore, colonoscopy should be performed within 18 hours of the final hematochezia in order to improve identification of the bleeding diverticulum.

[A case of hepatic artery aneurysm that had formed asymptomatically and penetrated into the duodenum].

A 41-year-old man was admitted to our hospital with gastrointestinal bleeding. Esophagogastroduodenoscopy revealed a submucosal protrusion with erosion in the duodenal bulb which was thought to be the bleeding source. Dynamic CT scan, ultrasonography and angiography of the abdomen revealed a hepatic artery aneurysm and a dilated celiac artery that dissected from its origin. Although we considered percutaneous transcatheter arterial embolization with metallic coils, we chose surgical resection and vascular reconstruction to prevent hepatic ischemia resulting from interruption of collateral circulation. On the 8(th) day, hepatic artery aneurysmectomy and revascularization with a great saphenous vein was carried out without any severe complication. The pathological specimen demonstrated segmental arterial mediolysis.

Wilms tumor reveals DNA repair gene hyperexpression is linked to lack of tumor immune infiltration.

BACKGROUND: A T cell-rich tumor microenvironment has been associated with improved clinical outcome and response to immune checkpoint blockade therapies in several adult cancers. Understanding the mechanisms for lack of immune cell infiltration in tumors is critical for expanding immunotherapy efficacy. To gain new insights into the mechanisms of poor tumor immunogenicity, we turned to pediatric cancers, which are generally unresponsive to checkpoint blockade. METHODS: RNA sequencing and clinical data were obtained for Wilms tumor, rhabdoid tumor, osteosarcoma, and neuroblastoma from the Therapeutically Applicable Research to Generate Effective Treatments (TARGET) database, and adult cancers from The Cancer Genome Atlas (TCGA). Using an 18-gene tumor inflammation signature (TIS) representing activated CD8+ T cells, we identified genes inversely correlated with the signature. Based on these results, adult tumors were also analyzed, and immunofluorescence was performed on metastatic melanoma samples to assess the MSH2 relationship to anti-programmed cell death protein-1 (PD-1) efficacy. RESULTS: Among the four pediatric cancers, we observed the lowest TIS scores in Wilms tumor. TIS scores were lower in Wilms tumors compared with matched normal kidney tissues, arguing for loss of endogenous T cell infiltration. Pathway analysis of genes upregulated in Wilms tumor and anti-correlated with TIS revealed activated pathways involved DNA repair. The majority of adult tumors in TCGA also showed high DNA repair scores associated with low TIS. Melanoma samples from an independent cohort revealed an inverse correlation between MSH2+ tumor cells and CD8+ T cells. Additionally, melanomas with high MSH2+ tumor cell numbers were largely non-responders to anti-PD-1 therapy. CONCLUSIONS: Increased tumor expression of DNA repair genes is associated with a less robust immune response in Wilms tumor and the majority of TCGA tumor types. Surprisingly, the negative relationship between DNA repair score and TIS remained strong across TCGA when correcting for mutation count, indicating a potential role for DNA repair genes outside of preventing the accumulation of mutations. While loss of DNA repair machinery has been associated with carcinogenesis and mutational antigen generation, our results suggest that hyperexpression of DNA repair genes might be prohibitive for antitumor immunity, arguing for pharmacologic targeting of DNA repair as a potential therapeutic strategy.

Pembrolizumab Monotherapy for Previously Untreated Advanced Hepatocellular Carcinoma: Data from the Open-Label, Phase II KEYNOTE-224 Trial.

PURPOSE: KEYNOTE-224 cohort 1 demonstrated that pembrolizumab was efficacious and tolerable in patients with advanced hepatocellular carcinoma (HCC) previously treated with sorafenib. We report results from KEYNOTE-224 (NCT02702414) cohort 2, which enrolled patients with advanced HCC and no prior systemic therapy. PATIENTS AND METHODS: KEYNOTE-224 was an open-label, multicountry phase II trial. Eligible patients in cohort 2 had advanced HCC not amenable or refractory to locoregional therapy and not previously treated with systemic therapy. Patients received pembrolizumab 200 mg intravenously every 3 weeks for ≤2 years. Primary endpoint was objective response rate (ORR) by central imaging review per RECIST v1.1. Secondary endpoints included duration of response (DOR), disease control rate (DCR), time to progression (TTP), progression-free survival (PFS), overall survival (OS), and safety/tolerability. RESULTS: Between September 4, 2018, and February 20, 2019, 51 patients were allocated in cohort 2. The median time from the first dose to data cutoff (January 19, 2021) was 27 months (range, 23-29). ORR was 16% [95% confidence interval (CI), 7-29] and was similar across key subgroups. Median DOR was 16 months (range, 3-24+), and DCR was 57%. The median PFS was 4 months (95% CI, 2-8), and median TTP was 4 months (95% CI, 3-9). Median OS was 17 months (95% CI, 8-23). Grade ≥3 treatment-related adverse events occurred in 16% of patients. CONCLUSIONS: In patients with advanced HCC with no prior systemic therapy, pembrolizumab provided durable antitumor activity, promising OS, and had a safety profile consistent with previous observations. These findings support further evaluation of pembrolizumab-based regimens for HCC.

[Advanced gallbladder cancer that showed complete response to gemcitabine plus S-1 chemotherapy].

A 57-year-old man with advanced gallbladder cancer and accompanying hepatic, colonic and duodenal invasion and para-aortic lymph node metastasis was referred to our hospital. Gemcitabine plus S-1 administration was chosen. Gemcitabine was administered intravenously at a dose of 1000 mg/m(2) on days 1 and 15, and repeated every 4 weeks. S-1 was administered orally at a dose of 40 mg/m(2) b.i.d. on days 1-14. Chemotherapy was effective for the primary gallbladder tumor and lymph node metastasis. The primary tumor and metastatic lymph nodes were shown to have disappeared by a FDG-PET CT study after 10 courses of chemotherapy. Informed consent was obtained prior to performing surgery of the primary lesion. Pathological examination showed fibrosis and a small focus of residual cancer in the resected gallbladder. Complete resection was achieved as all the margins were negative. The findings suggest that gemcitabine plus S-1 treatment may be effective against advanced gallbladder cancer.

Immune cell and tumor cell-derived CXCL10 is indicative of immunotherapy response in metastatic melanoma.

A T cell-inflamed tumor microenvironment is characterized by the accumulation and local activation of CD8+ T cells and Bat3-lineage dendritic cells, which together are associated with clinical response to anti-programmed cell death protein 1 (anti-PD-1)-based immunotherapy. Preclinical models have demonstrated a crucial role for the chemokine CXCL10 in the recruitment of effector CD8+ T cells into the tumor site, and a chemokine gene signature is also seen in T cell-inflamed tumors from patients. However, the cellular source of CXCL10 in human solid tumors is not known. To identify the cellular source of CXCL10 we analyzed 22 pretreatment biopsy samples of melanoma metastases from patients who subsequently underwent checkpoint blockade immunotherapy. We stained for CD45+ and Sox10+ cells with multiparameter immunofluorescence staining, and RNA in situ hybridization technology was used in concert to identify CXCL10 transcripts. The results were correlated with the expression levels of CXCL10 transcripts from bulk RNA sequencing and the best overall response to immune checkpoint inhibition (anti-PD-1 alone or with anti-CTLA-4) in the same patients. We identified CD45+ cells as the major cellular source for CXCL10 in human melanoma metastases, with additional CXCL10 production seen by Sox10+ cells. Up to 90% of CD45+ cells and up to 69% of Sox10+ cells produced CXCL10 transcripts. The CXCL10 staining result was consistent with the level of CXCL10 expression determined by bulk RNA sequencing. The percentages of CD45+ CXCL10+ cells and Sox10+ CXCL10+ cells independently predicted response (p<0.001). The average number of transcripts per cell correlated with the CD45+ cell infiltrate (R=0.37). Immune cells and melanoma cells produce CXCL10 in human melanoma metastases. Intratumoral CXCL10 is a positive prognostic factor for response to immunotherapy, and the RNAscope technique is achievable using paraffin tissue. Strategies that support effector T cell recruitment via induction of CXCL10 should be considered as a mechanism-based intervention to expand immunotherapy efficacy.

Severe COVID-19 infection is associated with aberrant cytokine production by infected lung epithelial cells rather than by systemic immune dysfunction.

The mechanisms explaining progression to severe COVID-19 remain poorly understood. It has been proposed that immune system dysregulation/over-stimulation may be implicated, but it is not clear how such processes would lead to respiratory failure. We performed comprehensive multiparameter immune monitoring in a tightly controlled cohort of 128 COVID-19 patients, and used the ratio of oxygen saturation to fraction of inspired oxygen (SpO2 / FiO2) as a physiologic measure of disease severity. Machine learning algorithms integrating 139 parameters identified IL-6 and CCL2 as two factors predictive of severe disease, consistent with the therapeutic benefit observed with anti-IL6-R antibody treatment. However, transcripts encoding these cytokines were not detected among circulating immune cells. Rather, in situ analysis of lung specimens using RNAscope and immunofluorescent staining revealed that elevated IL-6 and CCL2 were dominantly produced by infected lung type II pneumocytes. Severe disease was not associated with higher viral load, deficient antibody responses, or dysfunctional T cell responses. These results refine our understanding of severe COVID-19 pathophysiology, indicating that aberrant cytokine production by infected lung epithelial cells is a major driver of immunopathology. We propose that these factors cause local immune regulation towards the benefit of the virus.

Complete response of renal cell carcinoma vena cava tumor thrombus to neoadjuvant immunotherapy.

BACKGROUND: Clinically localized renal cell carcinoma is treated primarily with surgery followed by observation or adjuvant sunitinib in selected high-risk patients. The checkpoint inhibitor immunotherapeutic agents nivolumab and ipilimumab have recently shown a survival benefit in the first-line metastatic setting. To date, there have been no reports on the response of localized renal cancer to modern immunotherapy. We report a remarkable response of an advanced tumor thrombus to combined immunotherapy which facilitated curative-intent resection of the non-responding primary renal tumor. We characterized the tumor microenvironment within the responding and non-responding tumors. CASE PRESENTATION: A 54-year-old female was diagnosed with a locally advanced clear cell renal cell carcinoma with a level IV tumor thrombus of the vena cava. She was initially deemed unfit for surgical resection due to poor performance status. She underwent neoadjuvant immunotherapy with nivolumab and ipilimumab with a complete response of the vena cava and renal vein tumor thrombus, but had stable disease within her renal mass. She underwent complete surgical resection with negative margins and remains disease-free longer than 1 year after her diagnosis with no further systemic therapy. Notably, pathologic analysis showed a complete response within the vena cava and renal vein, but substantial viable cancer remained in the kidney. Multichannel immunofluorescence was performed and showed marked infiltration of immune cells including CD8+ T cells and Batf3+ dendritic cells in the thrombus, while the residual renal tumor showed a non-T cell-inflamed phenotype. CONCLUSIONS: Preoperative immunotherapy with nivolumab and ipilimumab for locally advanced clear cell renal cancer resulted in a complete response of an extensive vena cava tumor thrombus, which enabled curative-intent resection of a non-responding primary tumor. If validated in larger cohorts, preoperative immunotherapy for locally advanced renal cell carcinoma may ultimately impact surgical planning and long-term prognosis.