Recurrence patterns after complex multimodality therapy and hepatic arterial infusion for colorectal liver metastases: A reflection of biology and technique.

BACKGROUND AND METHODS: We characterized colorectal liver metastasis recurrence and survival patterns after surgical resection and intraoperative ablation ± hepatic arterial infusion pump (HAIP) placement. We estimated patterns of recurrence and survival in patients undergoing contemporary multimodal treatments. Between 2017 and 2021, patient, tumor characteristics, and recurrence data were collected. Primary outcomes included recurrence patterns and survival data based on operative intervention. RESULTS: There were 184 patients who underwent hepatectomy and intraoperative ablation. Sixty patients (32.6%) underwent HAIP placement. A total of 513 metastases were ablated, median total of 2 ablations per patient. Median time to recurrence was 31 [22-40] months. Recurrence patterns included tumor at ablative margin on first scheduled postoperative imaging (8, 4.3%), local tumor recurrence at ablative site (69, 37.5%), and non-ablated liver tumor recurrence (38, 20.6%). In patients who underwent HAIP placement, the rate of liver recurrence was reduced (45% vs 70.9%, p = 0.0001). Median overall survival was 64 [41-58] months and prolonged survival was associated with HAIP treatment (85 [66-109] vs 60 [51-70] months. CONCLUSIONS AND DISCUSSION: Hepatic recurrence is common and combination of intraoperative ablation and HAIP treatments were associated with prolonged survival. These data may reflect patient selection however, future work will clarify preoperative tumor and patient characteristics that may better predict recurrence expectations.

Tumor and Peritoneum-Associated Macrophage Gene Signature as a Novel Molecular Biomarker in Gastric Cancer.

A spectrum of immune states resulting from tumor resident macrophages and T-lymphocytes in the solid tumor microenvironment correlates with patient outcomes. We hypothesized that in gastric cancer (GC), macrophages in a polarized immunosuppressive transcriptional state would be prognostic of poor survival. We derived transcriptomic signatures for M2 (M2TS, MRC1; MS4A4A; CD36; CCL13; CCL18; CCL23; SLC38A6; FGL2; FN1; MAF) and M1 (M1TS, CCR7; IL2RA; CXCL11; CCL19; CXCL10; PLA1A; PTX3) macrophages, and cytolytic T-lymphocytes (CTLTS, GZMA; GZMB; GZMH; GZMM; PRF1). Primary GC in a TCGA stomach cancer dataset was evaluated for signature expressions, and a log-rank test determined overall survival (OS) and the disease-free interval (DFI). In 341 TCGA GC entries, high M2TS expression was associated with histological types and later stages. Low M2TS expression was associated with significantly better 5-year OS and DFI. We validated M2TS in prospectively collected peritoneal fluid of a GC patient cohort (n = 28). Single-cell RNA sequencing was used for signature expression in CD68+CD163+ cells and the log-rank test compared OS. GC patients with high M2TS in CD68+CD163+ cells in their peritoneal fluid had significantly worse OS than those with low expression. Multivariate analyses confirmed M2TS was significantly and independently associated with survival. As an independent predictor of poor survival, M2TS may be prognostic in primary tumors and peritoneal fluid of GC patients.

Different Methods of Minimally Invasive Esophagojejunostomy After Total Gastrectomy for Gastric Cancer: Outcomes from Two Experienced Centers.

BACKGROUND: Esophagojejunostomy after minimally invasive total gastrectomy (MITG) for gastric cancer (GC) is technically challenging. Failure of the esophagojejunal anastomosis can lead to significant morbidity, leading to short- and long-term quality of life (QoL) impairment or mortality. The optimal reconstruction method following MITG remains controversial. We evaluated outcomes of minimally invasive esophagojejunostomy after laparoscopic or robotic total gastrectomies. METHODS: We retrospectively reviewed MITG patients between 2015 and 2020 at two high-volume centers in China and the United States. Eligible patients were divided into groups by different reconstruction methods. We compared clinicopathologic characteristics, postoperative outcomes, including complication rates, overall survival rate (OS), disease-free survival rate (DFS), and patient-reported QoL. RESULTS: GC patients (n = 105) were divided into intracorporeal esophagojejunostomy (IEJ, n = 60) and extracorporeal esophagojejunostomy (EEJ, n = 45) groups. EEJ had higher incidence of wound infection (8.3% vs 13.3%, P = 0.044) and pneumonia (21.7% vs 40.0%, P = 0.042) than IEJ. The linear stapler (LS) group was inferior to the circular stapler (CS) group in reflux [50.0 (11.1-77.8) vs 44.4 (0.0-66.7), P = 0.041] and diarrhea [33.3 (0.0-66.7) vs 0.0 (0.0-66.7), P = 0.045] while LS was better than CS for dysphagia [22.2 (0.0-33.3) vs 11.1 (0.0-33.3), P = 0.049] and eating restrictions [33.3 (16.7-58.3) vs 41.7 (16.7-66.7), P = 0.029] at 1 year. OS and DFS did not differ significantly between LS and CS. CONCLUSIONS: IEJ anastomosis generated better results than EEJ. LS was associated with a better patient eating experience, but more diarrhea and reflux compared with CS. Clinical and patient-reported outcomes show the superiority of IEJ with the LS reconstruction method in MITG for GC.

Layilin Anchors Regulatory T Cells in Skin.

Regulatory T cells (Tregs) reside in nonlymphoid tissues where they carry out unique functions. The molecular mechanisms responsible for Treg accumulation and maintenance in these tissues are relatively unknown. Using an unbiased discovery approach, we identified LAYN (layilin), a C-type lectin-like receptor, to be preferentially and highly expressed on a subset of activated Tregs in healthy and diseased human skin. Expression of layilin on Tregs was induced by TCR-mediated activation in the presence of IL-2 or TGF-ß. Mice with a conditional deletion of layilin in Tregs had reduced accumulation of these cells in tumors. However, these animals somewhat paradoxically had enhanced immune regulation in the tumor microenvironment, resulting in increased tumor growth. Mechanistically, layilin expression on Tregs had a minimal effect on their activation and suppressive capacity in vitro. However, expression of this molecule resulted in a cumulative anchoring effect on Treg dynamic motility in vivo. Taken together, our results suggest a model whereby layilin facilitates Treg adhesion in skin and, in doing so, limits their suppressive capacity. These findings uncover a unique mechanism whereby reduced Treg motility acts to limit immune regulation in nonlymphoid organs and may help guide strategies to exploit this phenomenon for therapeutic benefit.

Tumor Immune Profiling-Based Neoadjuvant Immunotherapy for Locally Advanced Melanoma.

BACKGROUND: The frequency of "exhausted" or checkpoint-positive (PD-1+CTLA-4+) cytotoxic lymphocytes (Tex) in the tumor microenvironment is associated with response to anti-PD-1 therapy in metastatic melanoma. The current study determined whether pretreatment Tex cells in locally advanced melanoma predicted response to neoadjuvant anti-PD-1 blockade. METHODS: Pretreatment tumor samples from 17 patients with locally advanced melanoma underwent flow cytometric analysis of pretreatment Tex and regulatory T cell frequency. Patients who met the criteria for neoadjuvant checkpoint blockade were treated with either PD-1 monotherapy or PD-1/CTLA-4 combination therapy. Best overall response was evaluated by response evaluation criteria in solid tumors version 1.1, with recurrence-free survival (RFS) calculated by the Kaplan-Meier test. The incidence and severity of adverse events were tabulated by clinicians using the National Cancer Institute Common Terminology Criteria for Adverse Events version 4. RESULTS: Of the neoadjuvant treated patients, 10 received anti-PD-1 monotherapy and 7 received anti-CTLA-4/PD-1 combination therapy. Of these 17 patients, 12 achieved a complete response, 4 achieved partial responses, and 1 exhibited stable disease. Surgery was subsequently performed for 11 of the 17 patients, and 8 attained a complete pathologic response. Median RFS and overall survival (OS) were not reached. Immune-related adverse events comprised four grade 3 or 4 events, including pneumonitis, transaminitis, and anaphylaxis. CONCLUSION: The results showed high rates of objective response, RFS, and OS for patients undergoing immune profile-directed neoadjuvant immunotherapy for locally advanced melanoma. Furthermore, the study showed that treatment stratification based upon Tex frequency can potentially limit the adverse events associated with combination immunotherapy. These data merit further investigation with a larger validation study.

A novel stratification of mesenteric mass involvement as a predictor of challenging mesenteric lymph node dissection by minimally invasive approach for ileal neuroendocrine tumors.

BACKGROUND AND OBJECTIVES: We classified the extent of mesenteric mass (MM) involvement that predicts challenging mesenteric lymph node dissection (mLND) by minimally invasive surgery (MIS) for ileal neuroendocrine tumors (i-NETs). METHODS: Patients who underwent surgery for i-NETs were retrospectively reviewed. MM involvement was classified as region-0: no MM; region-1: >2 cm from the origins of the ileocolic artery/vein; region-2: ≤2 cm from the origins; and region-3: more proximal superior mesenteric artery/vein. Logistic regression analysis was used to evaluate the predictive value of MM regions for gross positive mesenteric margin (mR2) and/or conversion among the MIS cohort. The open surgery cohort was used as a reference for mR2 rates. RESULTS: Of 108 patients, 83 patients (77%) underwent MIS. MMs in region-2 and region-3 were independent risk factors for mR2 and/or conversion (odds ratio [95% confidence interval]: 4.25 [1.17-16.4] and 8.51 × 107 [11.0-], respectively, against regions-0 and 1]. mR2 rates of MIS and open surgery cohorts per region did not differ significantly (4% and 7% for regions-0 and 1; 17% and 25% for region-2; and 100% and 83% for region-3). CONCLUSIONS: The novel stratification of MM regions was predictive of challenging mLND by MIS. Surgeons should have a low threshold for conversion for MMs in proximal regions.

Prognostic impact of a large mesenteric mass >2 cm in ileal neuroendocrine tumors.

BACKGROUND AND OBJECTIVES: Ileal neuroendocrine tumors (i-NETs) frequently metastasize to mesenteric lymph nodes and the liver. Regional lymphadenopathy is associated with desmoplasia of the mesentery forming a large mesenteric mass (LMM). Although the latest American Joint Committee on Cancer TNM staging (8th edition) defined LMM >2 cm as N2, the prognostic impact of LMM is ill-defined. We evaluated whether LMM is prognostic for patients with i-NETs. METHODS: This single-institution, retrospective cohort study included 106 patients who underwent resection of i-NETs between 2007 and 2018. Overall survival (OS) and liver progression-free survival (LPFS) were compared between patients with and without LMM. RESULTS: LMM was present in 66 patients (62%) and was not associated with the presence or absence of liver metastasis (P = .969) or the extent of liver involvement (P = .938). OS and LPFS differed significantly between patients with and without LMM (5-year OS rates of 64.8% and 92.9%, respectively, P = .011; 3-year LPFS rates of 45.3% and 67.5%, respectively, P = .025). In multivariate analysis, LMM was an independent prognostic factor for both OS (hazard ratio: 4.69, 95% confidence interval: 1.63-17.6) and LPFS (1.99, 1.08-3.88). CONCLUSION: LMM >2 cm is prognostic for OS and LPFS and represents aggressive tumor biology.

Applications of Liquid Biopsy for Surgical Patients With Cancer: A Review.

Importance: Liquid biopsy is an emerging tool with the potential to change oncologic care practices. Optimal clinical applications for its use are currently undefined for surgical patients. Observations: Liquid biopsy analytes such as circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) have been the most clinically studied assays and were initially limited to advanced-stage disease. In the metastatic setting, CTCs and ctDNA levels are prognostic. Although their levels correlate with treatment response, CTC-guided systemic regimen switches for nonresponders have not been shown to improve clinical outcomes. ctDNA genomic profiling has succeeded, and there are now multiple plasma-based assays approved by the US Food and Drug Administration that can detect actionable mutations to guide systemic therapy. Technological advancements in assay sensitivity have expanded the use of ctDNA to early-stage and resectable disease, allowing for detection of minimal residual disease. Postoperative ctDNA levels are a strong predictor of disease recurrence, and ctDNA detection often precedes serum carcinoembryonic antigen elevation and radiographic changes. However, its use for surveillance has not been shown to improve clinical outcomes. A promising application of ctDNA is for adjuvant therapy escalation and de-escalation. A phase 2 clinical trial demonstrated that treatment de-escalation for patients with high-risk stage II colorectal cancer and negative postoperative ctDNA had similar recurrence-free survival as patients receiving standard-of-care chemotherapy. These results suggest that ctDNA may help select patients who will benefit from adjuvant chemotherapy, and multiple clinical trials are actively underway. Conclusions and Relevance: Although uncertainties regarding the optimal use of liquid biopsy remain, it has the potential to significantly improve care for patients with cancer at all stages of disease. It is critical that surgeons understand how to use and interpret these assays, and they should be active participants in clinical trials to advance the field.

Diffuse-Type Histology Is Prognostic for All Siewert Types of Gastroesophageal Adenocarcinoma.

PURPOSE: The optimal treatment for gastroesophageal junction adenocarcinoma (GEJA) remains controversial. We evaluated the treatment patterns and outcomes of patients with locally advanced GEJA according to the histological type. MATERIALS AND METHODS: We conducted a single-institution retrospective cohort study of patients with locally advanced GEJA who underwent curative-intent surgical resection between 2010 and 2020. Perioperative therapies as well as clinicopathologic, surgical, and survival data were collected. The results of endoscopy and histopathological examinations were assessed for Siewert and Lauren classifications. RESULTS: Among the 58 patients included in this study, 44 (76%) were clinical stage III, and all received neoadjuvant therapy (72% chemoradiation, 41% chemotherapy, 14% both chemoradiation and chemotherapy). Tumor locations were evenly distributed by Siewert Classification (33% Siewert-I, 40% Siewert-II, and 28% Siewert-III). Esophagogastrectomy (EG) was performed for 47 (81%) patients and total gastrectomy (TG) for 11 (19%) patients. All TG patients received D2 lymphadenectomy compared to 10 (21%) EG patients. Histopathological examination showed the presence of 64% intestinal-type and 36% diffuse-type histology. The frequencies of diffuse-type histology were similar among Siewert groups (37% Siewert-I, 36% Siewert-II, and 33% Siewert-III). Regardless of Siewert type and compared to intestinal-type, diffuse histology was associated with increased intraabdominal recurrence rates (P=0.03) and decreased overall survival (hazard ratio, 2.33; P=0.02). With a median follow-up of 31.2 months, 29 (50%) patients had a recurrence, and the median overall survival was 50.5 months. CONCLUSIONS: Present in equal proportions among Siewert types of esophageal and gastric cancer, a diffuse-type histology was associated with high intraabdominal recurrence rates and poor survival. Histopathological evaluation should be considered in addition to anatomic location in the determination of multimodal GEJA treatment strategies.

Defining a New Classification System for the Surgical Management of Neuroendocrine Tumor Liver Metastases.

Although rarely curative, hepatic cytoreduction of neuroendocrine tumor liver metastases (NETLM) is associated with improved symptom control and prolonged survival. Preoperative 68Ga DOTATATE and gadoxetic acid-enhanced liver MRI can improve characterization of hepatic disease extent to improve surgical clearance, and resection of the primary tumor is associated with improved survival regardless of whether the liver metastases are treated. As parenchymal-sparing surgical techniques and the lowering of the debulking threshold have expanded the numbers of eligible NETLM patients for hepatic cytoreduction, we propose a new classification system to help guide surgical management. A multimodal approach that includes surgery, liver-directed therapies, and systemic therapies has improved outcomes and increased longevity for patients with well-differentiated metastatic NET.

Liver-Directed Therapy in Neuroendocrine Neoplasms Metastatic to Both Liver and Bone.

Bone metastases from gastroenteropancreatic neuroendocrine neoplasms (GEPNENs) have been associated with poor prognosis, but it is unclear whether patients with concurrent bone metastases who receive liver-directed therapy (LDT) would derive survival benefit. The California Cancer Registry dataset, merged with data from the California Office of Statewide Health Planning and Development, was used to perform a retrospective study of GEPNENs metastatic to both liver and bone between 2000 and 2012. A total of 203 patients were identified. Of these, 14.8% underwent LDT after bone metastasis diagnosis, 22.1% received LDT prior to that diagnosis, and 63.1% never received LDT. The median overall survival from the time of bone metastasis diagnosis was significantly longer in those that received LDT after diagnosis when compared with those that never received LDT (p = 0.005) and was not significantly different from the median overall survival of those that had received LDT prior to diagnosis (p = 0.256). LDT may still be associated with improved survival even after a diagnosis of bone metastasis.

Indole-3-carbinol downregulation of telomerase gene expression requires the inhibition of estrogen receptor-alpha and Sp1 transcription factor interactions within the hTERT promoter and mediates the G1 cell cycle arrest of human breast cancer cells.

Indole-3-carbinol (I3C), a naturally occurring hydrolysis product of glucobrassicin from cruciferous vegetables such as broccoli, cabbage and Brussels sprouts, is an anticancer phytochemical that triggers complementary sets of antiproliferative pathways to induce a cell cycle arrest of estrogen-responsive MCF7 breast cancer cells. I3C strongly downregulated transcript expression of the catalytic subunit of the human telomerase (hTERT) gene, which correlated with the dose-dependent indole-mediated G(1) cell cycle arrest without altering the transcript levels of the RNA template (hTR) for telomerase elongation. Exogenous expression of hTERT driven by a constitutive promoter prevented the I3C-induced cell cycle arrest and rescued the I3C inhibition of telomerase enzymatic activity and activation of cellular senescence. Time course studies showed that I3C downregulated expression of estrogen receptor-alpha (ERα) and cyclin-dependent kinase-6 transcripts levels (which is regulated through the Sp1 transcription factor) prior to the downregulation of hTERT suggesting a mechanistic link. Chromatin immunoprecipitation assays demonstrated that I3C disrupted endogenous interactions of both ERα and Sp1 with an estrogen response element-Sp1 composite element within the hTERT promoter. I3C inhibited 17ß-estradiol stimulated hTERT expression and stimulated the production of threonine-phosphorylated Sp1, which inhibits Sp1-DNA interactions. Exogenous expression of both ERα and Sp1, but not either alone, in MCF7 cells blocked the I3C-mediated downregulation of hTERT expression. These results demonstrate that I3C disrupts the combined ERα- and Sp1-driven transcription of hTERT gene expression, which plays a significant role in the I3C-induced cell cycle arrest of human breast cancer cells.

Stress and the Surgical Resident in the COVID-19 Pandemic.

OBJECTIVE: The COVID-19 pandemic has drastically transformed the healthcare community and medical education across the United States. The aim of this study was to evaluate the impact of COVID-19 on the surgical resident training experience, assess possible sources of stress or anxiety among surgery residents, and examine how patterns of anxiety vary by resident rank. DESIGN: We developed and disseminated a survey, which included the Generalized Anxiety Disorder 7-Item Scale (GAD-7), to all general and integrated plastic surgery residents in their clinical years of training at the University of California, San Francisco. Statistical analysis of the survey responses was performed using the Kruskal-Wallis or Wilcoxon rank sum test. Post-hoc analysis was performed using the Bonferroni-corrected Dunn test. Survey data were combined with aggregated duty hour information and operative case numbers from select hospitals for March and April of 2019 (historical baseline) and 2020. RESULTS: The overall survey response rate was 73.7% (n = 73). With an estimated operative volume reduction of 63.3% for general surgery cases, over 90% of residents expressed concern about the decline in operative exposure. While the senior residents tended to work more shifts, they were not more likely to have higher risk perception scores for contracting COVID-19 nor higher anxiety levels about the possibility of contracting COVID-19. They were, however, significantly more likely to have high GAD-7 scores (≥ 10) when compared to interns (z = -2.82, p-adj = 0.014). Overall, residents were more concerned about the general health of loved ones than about their own risk of contracting COVID-19 (U = 3897.5, p < 0.01). CONCLUSIONS: While the work-related experiences of residents varied across a number of factors during the pandemic, residents tended to report similar sources of anxiety. Moving forward, surgical residency training programs will need to develop ways to optimize available surgical experiences and address the unique resident anxieties that an infectious pandemic presents.

Single-cell analyses identify circulating anti-tumor CD8 T cells and markers for their enrichment.

The ability to monitor anti-tumor CD8+ T cell responses in the blood has tremendous therapeutic potential. Here, we used paired single-cell RNA and TCR sequencing to detect and characterize "tumor-matching" (TM) CD8+ T cells in the blood of mice with MC38 tumors or melanoma patients using the TCR as a molecular barcode. TM cells showed increased activation compared with nonmatching T cells in blood and were less exhausted than matching cells in tumors. Importantly, PD-1, which has been used to identify putative circulating anti-tumor CD8+ T cells, showed poor sensitivity for identifying TM cells. By leveraging the transcriptome, we identified candidate cell surface markers for TM cells in mice and patients and validated NKG2D, CD39, and CX3CR1 in mice. These data show that the TCR can be used to identify tumor-relevant cells for characterization, reveal unique transcriptional properties of TM cells, and develop marker panels for tracking and analysis of these cells.

Layilin augments integrin activation to promote antitumor immunity.

Tumor-infiltrating CD8+ T cells mediate antitumor immune responses. However, the mechanisms by which T cells remain poised to kill cancer cells despite expressing high levels of inhibitory receptors are unknown. Here, we report that layilin, a C-type lectin domain-containing membrane glycoprotein, is selectively expressed on highly activated, clonally expanded, but phenotypically exhausted CD8+ T cells in human melanoma. Lineage-specific deletion of layilin on murine CD8+ T cells reduced their accumulation in tumors and increased tumor growth in vivo. Congruently, gene editing of LAYN in human CD8+ T cells reduced direct tumor cell killing ex vivo. On a molecular level, layilin colocalized with integrin αLß2 (LFA-1) on T cells, and cross-linking layilin promoted the activated state of this integrin. Accordingly, LAYN deletion resulted in attenuated LFA-1-dependent cellular adhesion. Collectively, our results identify layilin as part of a molecular pathway in which exhausted or "dysfunctional" CD8+ T cells enhance cellular adhesiveness to maintain their cytotoxic potential.

Integrating Robotic Technology Into Resident Training: Challenges and Recommendations From the Front Lines.

PURPOSE: To develop recommendations for improving the integration of robotic technology into today's apprentice-based resident training. METHOD: During a national meeting in 2017, 24 robotic surgeons were interviewed about their experiences integrating robotic technology into resident training. Qualitative thematic analysis of interview notes and recordings revealed themes related to challenges and recommendations. RESULTS: Four themes emerged, each corresponding to a general recommendation for integrating robotic technology into training. The first, surgical techniques versus tools, contrasts faculty's sequential mastery-surgical techniques first, then the robotic tool-with residents' simultaneous learning. The recommendation is to create separate learning opportunities for focused skill acquisition. The second theme, timing of exposure to the robotic tool, describes trainees' initial focus on tool use for basic surgical steps. The recommendation is to increase access to basic robotic cases. The third theme covers the relationship of laparoscopic and robotic surgery. The recommendation is to emphasize similar and dissimilar features during all minimally invasive surgical cases. The fourth theme, use of the dual console (which enables two consoles to operate the robot, the primary determines the secondary's functionality), highlights the unique teaching opportunities this console creates. The recommendation is for surgeons to give verbal guidance so residents completely understand surgical techniques. CONCLUSIONS: Surgical educators should consider technique versus tool, timing of exposure to the tool, overlapping and varying features of robotic and laparoscopic surgery, and use of the dual console as they develop curricula to ensure thorough acquisition and synthesis of all elements of robotic surgery.

Regulatory T cells use arginase 2 to enhance their metabolic fitness in tissues.

Distinct subsets of Tregs reside in nonlymphoid tissues where they mediate unique functions. To interrogate the biology of tissue Tregs in human health and disease, we phenotypically and functionally compared healthy skin Tregs with those in peripheral blood, inflamed psoriatic skin, and metastatic melanoma. The mitochondrial enzyme, arginase 2 (ARG2), was preferentially expressed in Tregs in healthy skin, increased in Tregs in metastatic melanoma, and reduced in Tregs from psoriatic skin. ARG2 enhanced Treg suppressive capacity in vitro and conferred a selective advantage for accumulation in inflamed tissues in vivo. CRISPR-mediated deletion of this gene in primary human Tregs was sufficient to skew away from a tissue Treg transcriptional signature. Notably, the inhibition of ARG2 increased mTOR signaling, whereas the overexpression of this enzyme suppressed it. Taken together, our results suggest that Tregs express ARG2 in human tissues to both regulate inflammation and enhance their metabolic fitness.