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Following resection of colorectal liver metastasis, most patients have disease recurrence, most commonly intrahepatic. Although the role of resection in colorectal liver metastasis is well-established, there have been limited investigations assessing the benefit of repeat hepatic resection compared with systemic treatment alone for intrahepatic recurrence.A retrospective single-institution cohort study of patients with recurrent colorectal liver metastasis following curative-intent hepatectomy was performed from 2003 to 2019. The oncologic outcomes, including post-recurrence overall survival, were evaluated using Kaplan-Meier and Cox proportional hazards modeling. Patients undergoing repeat hepatic resection were propensity-matched with patients receiving systemic treatment alone based on relevant clinicopathologic variables.There were 338 patients treated with hepatic resection for colorectal liver metastasis over the study period. Liver recurrence was observed in 147 (43%) patients at a median time of 10 months from prior resection, with a median post-recurrence overall survival of 29 months. There were 37 patients managed with repeat hepatic resection; 33 (89%) received perioperative chemotherapy. On propensity matching, there were no significant clinicopathologic differences between 37 patients having repeat hepatic resection and 37 patients treated with systemic treatment alone. Repeat hepatic resection was independently associated with improved 5-year post-recurrence overall survival compared with systemic treatment alone (median overall survival 41 vs 35 months, 5-year overall survival 19% vs 3%, P = .048).Disease characteristics of patients with intrahepatic recurrence of colorectal liver metastasis, specifically the number of liver lesions and size of the largest lesion, are most predictive of survival and response to systemic therapy. Patients who recur with oligometastatic liver disease experience improved outcomes and derive benefit from curative-intent repeat hepatic resection with integrated perioperative systemic therapy.

Circulating hybrid cells (CHCs) are a novel, rare cell population that harbor tumor and immune cell phenotypes and genotypes and are detectible in peripheral blood. Several recent reports implicated CHCs in the metastatic cascade and found their enumeration to provide better prognostic value than conventionally-defined circulating tumor cells (CTCs). However, methods for isolation and enrichment of CHCs are not well-studied or established. Here, we developed an ultrasensitive, antigen-independent platform leveraging the principles of magnetic levitation for the detection and isolation of disseminated neoplastic CHCs. For the first time, we demonstrate that CHCs can be magnetically focused to different levitation heights, under various paramagnetic conditions using a static levitation system, and we quantified the biophysical properties of CHCs (i.e., levitation heights). In addition, we investigated whether magnetic levitation approach can be combined with the affinity-based strategies to enrich CHCs under the magnetic field. Using clinical samples from breast and colorectal cancer patients, we demonstrated that neoplastic cells can be sorted with a magnetic levitation-based sorting device, without relying on any surface markers. Overall, we demonstrated the feasibility of the magnetic levitation method for unbiased enrichment of rare neoplastic-immune hybrid cells from peripheral blood specimens from cancer patients. This approach can be expanded to more clinical samples and cancer types to unprecedentedly explore the biology of rare neoplastic cells and develop metastasis-tailored therapies broadly impacting personalized and precision clinical treatments.

Cellular circulating biomarkers from the primary tumor such as circulating tumor cells (CTCs) and circulating hybrid cells (CHCs) have been described to harbor tumor-like phenotype and genotype. CHCs are present in higher numbers than CTCs supporting their translational potential. Methods for isolation of CHCs do not exist and are restricted to low-throughput, time consuming, and biased methodologies. We report the development of a label-free dielectrophoretic microfluidic platform facilitating enrichment of CHCs in a high-throughput and rapid fashion by depleting healthy peripheral blood mononuclear cells (PBMCs). We demonstrated up to 96.5% depletion of PBMCs resulting in 18.6-fold enrichment of cancer cells. In PBMCs from pancreatic adenocarcinoma patients, the platform enriched neoplastic cells identified by their KRAS mutant status using droplet digital PCR with one hour of processing. Enrichment was achieved in 75% of the clinical samples analyzed, establishing this approach as a promising way to non-invasively analyze tumor cells from patients., Montoya Mira & Sapre et al. design a microfluidic device utilizing dielectrophoresis for the enrichment of circulating neoplastic cells that possess both epithelial and immune-like characteristics. They optimized the device using in vitro models and then applied it to clinical samples for clinically relevant mutation analyses.

BACKGROUND Intrahepatic cholangiocarcinoma (ICC) is a rare cancer. Patients in rural areas may face reduced access to advanced treatments often only available at referral centers. We evaluated the association of referral center treatment with treatment patterns, outcomes, and geography in patients with ICC. METHODS We queried the Oregon State Cancer Registry for ICC between 1997 and 2016, collecting clinicopathologic, demographic, and oncologic data. Patients were classified by treatment at a referral center or non-referral center. 'Crowfly' distance to the nearest referral center (DRC) was calculated. Outcomes were evaluated using Kaplan-Meier, Cox proportional hazards modeling, and logistic regression. RESULTS Over 20 years, 740 patients with ICC had a median age of 66 years. Slightly more than half (n = 424, 57%) were non-referral center treated and 316 (43%) were referral center treated. Referral center treatment increased over time (odds ratio [OR] 1.03/year, p

Resected colorectal liver metastases (CRLM) frequently recur intrahepatically. Selection criteria for repeat hepatectomy of recurrent CRLM are ill-defined.We performed an institutional review of patients with recurrent CRLM undergoing repeat hepatectomy from 2003 to 19. Post-recurrence overall (rOS) and recurrence-free survival (RFS) were analyzed with Cox proportional hazards modeling.n = 147 experienced recurrent CRLM; 11% (n = 38) received repeat hepatectomy of which there was one Clavien-Dindo IIIa complication. Median rOS was 41 months; median RFS was 9 months. Improved rOS and RFS were independently associated with additional post-operative chemotherapy after repeat hepatectomy (HR 0.35 and 0.34, respectively); poor rOS with recurrent CRLM3 cm (HR 4.4) and12 months from first hepatectomy to recurrence (HR 4.8); poor RFS with ≥3 recurrence liver metastases (HR 2.8) (All P 0.05).Repeat hepatectomy for recurrent CRLM can be performed safely. Worse survival following repeat hepatectomy is independently associated with3 cm and ≥3 liver lesions at recurrence, and12 months to recurrence. Additional post-operative chemotherapy after repeat hepatectomy is associated with improved outcomes.

Gastrointestinal stromal tumors (GIST) commonly recur following curative-intent resection. Patients with recurrent GIST display heterogeneous outcomes with limited prognostic tools. We investigated factors associated with post-recurrence survival (PRS) and progression-free survival (PFS). We performed a review of our institutional cancer registry from 2003 to 2018 for patients with GIST. Clinicopathologic and outcome data were collected. The disease-free interval (DFI) was calculated from the end of curative-intent oncologic therapy until recurrence. Outcomes were evaluated using Kaplan–Meier and Cox proportional hazards modeling. Overall, 254 patients underwent resection of primary, non-metastatic GIST, with 81 (32%) recurrences. The median age was 58 years and more than half of the patients with recurrence (n = 44; 54%) received adjuvant imatinib. Recurrence was most common in the liver (n = 34, 42%), peritoneum (n = 31, 38%), or liver plus peritoneum (n = 10, 12%). The median DFI was 14 months (interquartile range 2–26 months); 51 (63%) patients had a DFI ≤24 months and 30 (37%) had a DFI > 24 months. The median post-recurrence follow-up was 46 months. Compared with a DFI ≤24 months, patients with a DFI >24 months had increased 10-year PRS (77% vs. 41%, p 24 months was independently associated with increased PRS (hazard ratio [HR] 0.24, p

INTRODUCTION. Patients developing metastatic gastrointestinal stromal tumors (mGIST) have heterogenous disease biology and oncologic outcomes; prognostic factors are incompletely characterized. We sought to evaluate predictors of 10-year metastatic survivorship in the era of tyrosine kinase inhibitor (TKI) therapy. METHODS. We reviewed patients with mGIST treated at our Comprehensive Cancer Center from 2003 to 2019, including only patients with either mortality or 10 years of follow-up. Ten-year survivorship was evaluated with logistic regression. RESULTS. We identified 109 patients with a median age of 57 years at mGIST diagnosis. Synchronous disease was present in 57% (n = 62) of patients; liver (n = 48, 44%), peritoneum (n = 40, 37%), and liver + peritoneum (n = 18, 17%) were the most common sites. Forty-six (42%) patients were 10-year mGIST survivors. Following mGIST diagnosis, radiographic progression occurred within 2 years in 53% (n = 58) of patients, 2–5 years in 16% (n = 17), and 5–10 years in 16% (n = 17), with median survival of 32, 76, and 173 months, respectively. Seventeen (16%) patients had not progressed by 10 years. Fifty-two (47%) patients underwent metastasectomy, which was associated with improved progression-free survival (hazard ratio 0.63, p = 0.04). In patients experiencing progression, factors independently associated with 10-year survivorship were age (odds ratio [OR] 0.96, p = 0.03) and time to progression (OR 1.71/year, p < 0.001). CONCLUSIONS. Ten-year survivorship is achievable in mGIST in the era of TKIs and is associated with younger age and longer time to first progression, while metastasectomy is associated with longer time to first progression. The role of metastasectomy in the management of patients with disease progression receiving TKI therapy merits further study.

Multiple tumor foci (MTF) in intrahepatic cholangiocarcinoma (ICC), including satellitosis and true multifocality, is a known negative prognostic factor and can inform pre-operative decision-making. Lack of standardized pre-operative liver staging practices may contribute to undiagnosed MTF and poor outcomes. We sought to investigate the sensitivity of different cross-sectional imaging modalities for MTF at our institution. We identified n = 52 patients with ICC who underwent curative-intent resection from 2004 to 2017 in a multidisciplinary hepato-pancreato-biliary cancer program. Timing and modality of pre-operative imaging were recorded. Blinded review of imaging was performed and modalities were evaluated for false-negative rate (FNR) in detecting MTF, satellitosis, and true multifocality. Forty-one (79%) patients underwent CT and 20 (38%) underwent MRI prior to hepatectomy. MTF was pre-operatively identified in six (12%) patients. An additional seven patients had MTF discovered on final surgical pathology, despite a median interval from CT/MRI to surgery of 20 days. On blinded review the FNR of MRI compared to CT for multifocality was 0% vs. 38%, 50% vs 80% for satellitosis, and 22% vs 46% for MTF as a whole. CT is inadequate for pre-operative diagnosis of MTF in resectable ICC, even when performed within 30 days of hepatectomy. We recommend liver-protocol MRI as the standard pre-operative imaging modality in non-metastatic ICC.

Colorectal liver metastasis (CRLM) is a leading cause of morbidity and mortality in patients with colorectal cancer. Hepatic arterial infusion (HAI) chemotherapy has been demonstrated to improve survival in patients with resected CRLM and to facilitate conversion of technically unresectable disease.Between 2016 and 2018, n = 22 HAI pumps were placed for CRLM. All patients received systemic chemotherapy concurrently with HAI floxuridine/dexamethasone. Overall survival (OS) and progression-free survival (PFS) were assessed using the Kaplan-Meier method.HAI pumps were placed in seven patients with completely resected CRLM and 15 patients with unresectable disease. Twenty-one patients received HAI floxuridine with a median of 5 total HAI cycles (interquartile range: 4-7). Biliary sclerosis was the most common HAI-related complication (n = 5, 24%). Of the 13 patients treated to convert unresectable CRLM, 3 (23%) underwent hepatic resection with curative intent after a median of 7 HAI cycles (range: 4-10). For all HAI patients, the mean OS was 26.7 months from CRLM diagnosis, while the median PFS and hepatic PFS from pump placement were 9 and 13 months, respectively.Concomitant HAI and systemic therapy can be utilized at multidisciplinary programs for patients with advanced CRLM, both in the adjuvant setting and to facilitate conversion of unresectable disease.

Preoperative chemotherapy (POC) is often employed for patients with resectable colorectal liver metastasis (CRLM). The time to resection (TTR) following the end of chemotherapy may impact oncologic outcomes; this phenomenon has not been studied in CRLM.We queried our institutional cancer database for patients with resected CRLM after POC from 2003 to 2019. TTR was calculated from date of last cytotoxic chemotherapy. Kaplan-Meier analysis and multivariable Cox proportional hazards modeling were used to analyze recurrence-free survival (RFS) and overall survival (OS).We identified n = 187 patients. One hundred twenty-four (66%) patients had a TTR of2 months, while 63 (33%) had a TTR of ≥2 months. Median follow-up was 36 months. On Kaplan-Meier analysis, patients with TTR ≥ 2 months had shorter RFS (median 11 vs. 17 months, p = 0.002) and OS (median 44 vs. 62 months, p 0.001). On multivariable analysis, TTR ≥ 2 months was independently associated with worse RFS (hazard ratio [HR] = 1.54, 95% confidence interval [CI] = 1.06-2.22, p = 0.02) and OS (HR = 1.75, 95% CI = 1.11-2.77, p = 0.01).TTR ≥ 2 months following POC is independently associated with worse oncologic outcomes in patients with resectable CRLM. We therefore recommend consideration for hepatic resection of CRLM within this window whenever feasible.

Background Management of elderly patients with solitary hepatocellular carcinoma (sHCC) is challenging with perceived clinicopathologic differences driving treatment options. We sought to determine factors predictive of disease control and survival after hepatic resection of sHCC in elderly patients. Methods We identified n = 45 elderly patients (³≥65 yo) with sHCC treated with hepatic resection alone from our prospective database from 2003–16. Clinicopathologic data were analyzed and survival was assessed from the time of hepatic resection. Results The median age was 75-years-old. Less than half of patients (47%) had viral hepatitis. At resection, the median Child-Pugh score was A6, median tumor size 5 cm, and mean AFP of 1050 (ng/mL). Major hepatectomy was performed in 23 patients (51%) with R0 resection achieved in 96%. Two patients (4%) had Grade III complications with no mortalities at 30 days and one death (2%) at 90-days. After R0 resection 44% (n = 20) had intrahepatic recurrence at a median of 32 months (95% CI: 15–46) with 20% (n = 9) developing extrahepatic recurrence at a median of 78 months (95% CI: 78-.). The median survival was 72 months (95% CI: 30–108 months). For patients with at least 3 years of follow-up, the 1-, 3-, and 5-year overall survival was 74%, 59%, and 50%, respectively. Mortality was associated with higher AFP and lower Prognostic Nutritional Index (PNI). Conclusion Carefully selected elderly patients with sHCC appear to have unique disease that is amenable to hepatic resection with low morbidity and mortality with excellent overall and recurrence-free survival.

BACKGROUND Gallbladder cancer (GBC) is often incidentally diagnosed after cholecystectomy. Intra-operative biliary tract violations (BTV) have been recently associated with development of peritoneal disease (PD). The degree of BTV may be associated with PD risk, but has not been previously investigated. METHODS We reviewed patients with initially non-metastatic GBC treated at our institution from 2003 to 2018. Patients were grouped based on degree of BTV during their treatment: major (e.g., cholecystotomy with bile spillage, n = 27, 29%), minor (e.g., intra-operative cholangiogram, n = 18, 19%), and no violations (n = 48, 55%). Overall survival (OS) and peritoneal disease-free survival (PDFS) were evaluated with Kaplan-Meier and Cox proportional hazards modeling. RESULTS Ninety-three patients were identified; the median age was 64 years (range 31-87 years). Seventy-six (82%) were incidentally diagnosed. The median follow-up was 23 months; 20 (22%) patients developed PD. The 3-year PDFS for patients with major, minor, and no BTV was 52%, 83%, and 98%, respectively (major vs. none: p < 0.001; minor vs. none: p < 0.01). BTV was not associated with 5-year OS (HR 1.53, p = 0.16). CONCLUSION Increasing degree of BTV is associated with higher risk of peritoneal carcinomatosis in patients with GBC and should be considered during preoperative risk stratification. Reporting biliary tract violations during cholecystectomy is encouraged.

Background Levels of circulating hybrid cells (CHCs), a newly identified circulating tumor cell (CTC), correlate with disease stage and progression in cancer. We investigated their utility to risk-stratify patients with clinically N0 (cN0) oral cavity squamous cell carcinoma (OCSCC), and to identify patients with occult cervical lymph node metastases (pN+). Methods We analyzed peripheral blood samples for CHCs with co-expression of cytokeratin (tumor) and CD45 (leukocyte) from 22 patients with cN0 OCSCC using immunofluorescence microscopy, then correlated levels with pathologic lymph node status. Results CHC levels exceeded CTCs and correlated with the presence of both clinically overt (p = 0.002) and occult nodal metastases (p = 0.006). Conclusions For evaluated cN0 OCSCC patients, those with cN0 → pN+ status harbored elevated CHC levels compared to patients without occult disease. Our findings highlight a promising blood-based biologic assay with potential utility to determine the necessity of surgical neck dissection for staging and treatment.

Metastatic progression defines the final stages of tumor evolution and underlies the majority of cancer-related deaths. The heterogeneity in disseminated tumor cell populations capable of seeding and growing in distant organ sites contributes to the development of treatment resistant disease. We recently reported the identification of a novel tumor-derived cell population, circulating hybrid cells (CHCs), harboring attributes from both macrophages and neoplastic cells, including functional characteristics important to metastatic spread. These disseminated hybrids outnumber conventionally defined circulating tumor cells (CTCs) in cancer patients. It is unknown if CHCs represent a generalized cancer mechanism for cell dissemination, or if this population is relevant to the metastatic cascade. Herein, we detect CHCs in the peripheral blood of patients with cancer in myriad disease sites encompassing epithelial and non-epithelial malignancies. Further, we demonstrate that in vivo-derived hybrid cells harbor tumor-initiating capacity in murine cancer models and that CHCs from human breast cancer patients express stem cell antigens, features consistent with the ability to seed and grow at metastatic sites. Finally, we reveal heterogeneity of CHC phenotypes reflect key tumor features, including oncogenic mutations and functional protein expression. Importantly, this novel population of disseminated neoplastic cells opens a new area in cancer biology and renewed opportunity for battling metastatic disease.Simple SummaryThere is an incomplete understanding of circulating neoplastic cell populations and the fundamental mechanisms that drive dissemination, immune evasion, and growth —all critical information to more effectively prevent and treat cancer progression. A novel disseminated tumor cell population, circulating hybrid cells, are detected across many cancer types and carry functional tumor-initiating properties. Additionally, circulating hybrid cells are found at significantly higher levels than conventionally defined circulating tumor cells. Our study demonstrates that neoplastic hybrid cells harbor phenotypic and genetic characteristics of tumor and immune cells, display stem features, and are a generalizable phenomenon in solid tumors. Circulating hybrid cells therefore have relevance as a novel biomarker and open a new field of study in malignancy.

Background Imatinib resistance is associated with a poor prognosis in patients with gastrointestinal stromal tumors. Although novel tyrosine kinase inhibitors have improved outcomes in imatinib-resistant gastrointestinal stromal tumors, the role of resection remains unclear. We sought to investigate factors predictive of overall and progression-free survival in patients with imatinib-resistant gastrointestinal stromal tumors. Methods A query of our prospectively maintained Comprehensive Cancer Center registry was performed from 2003 to 2019 for patients with imatinib-resistant gastrointestinal stromal tumors. Clinicopathologic characteristics and medical and surgical treatments were collected; overall survival and progression-free survival after imatinib-resistance were analyzed with Kaplan-Meier and Cox proportional hazards modeling. Results A total of 84 patients developed imatinib resistance at a median age of 59 years. Median time to imatinib resistance after diagnosis and overall survival after imatinib resistance was 50 and 51 months, respectively. After being diagnosed with imatinib resistance, 17 (20%) patients underwent resection. On multivariable analysis, resection after imatinib resistance was independently associated with improved progression-free survival (hazard ratio 0.50; P = .027) but not overall survival (hazard ratio 0.62; P = .215). Similar findings were found on subgroup analysis of patients treated with second-line sunitinib (n = 71). Conclusion Long-term survival can be achieved in patients who develop imatinib-resistant gastrointestinal stromal tumors. Surgical resection of imatinib-resistant gastrointestinal stromal tumors is associated with improved progression-free survival and should be considered in selected patients.

Variant hepatic arterial anatomy (vHAA) is thought to occur in 20-30% of patients. Hepatic arterial infusion (HAI) pump placement for liver cancers requires thorough hepatic artery dissection; we sought to compare vHAA identified during pump placement with established dogma.Between 2016 and 2020, n = 30 patients received a HAI pump. Intra-operatively identified vHAA was characterized and compared with published data.vHAA was identified in 60% (n = 18) of patients, significantly higher than 19% (3671 of 19013) in the largest published series (P 0.001). The most common variations were accessory left (n = 12; 40%) and replaced right (n = 6; 20%) hepatic arteries; six (20%) had ≥2 variants. Pre-operative imaging correctly identified 67% of variant hepatic arteries.Meticulous operative dissection of the hepatic arterial tree reveals vHAA not captured by imaging or cadaveric dissection. vHAA likely has a higher prevalence than previously reported and should be addressed to optimize therapeutic efficacy of HAI pump therapy.

Background The impact of neoadjuvant chemotherapy (NAC) on overall and recurrence-free survival (OS, RFS) in resectable intrahepatic cholangiocarcinoma (ICC) is poorly characterized. We sought to investigate the association of NAC with oncologic outcomes in ICC. Methods We identified n = 52 patients with ICC undergoing hepatectomy from 2004 to 2017. Oncologic outcomes were analyzed using Kaplan-Meier and multivariate Cox proportional hazard modeling. Results The median patient age was 64-years. NAC was administered in ten (19%) patients, most commonly with gemcitabine-cisplatin (n = 8, 80%). Median RFS and OS were 15 months. and 49 months, respectively. Controlling for stage and margins, NAC was independently associated with improved OS (HR 0.16, P = 0.01) but not RFS (HR 0.54, P = 0.27). NAC was not associated with major post-operative complications (P = 0.25) or R1 margins (P = 0.58). Conclusion NAC in ICC may hold oncologic benefits beyond downstaging borderline resectable disease, such as identifying patients with favorable biology who are more likely to benefit from resection.

Multiple tumor foci (MTF) in intrahepatic cholangiocarcinoma (ICC), including satellitosis and true multifocality, is a known negative prognostic factor and can inform pre-operative decision-making. Lack of standardized pre-operative liver staging practices may contribute to undiagnosed MTF and poor outcomes. We sought to investigate the sensitivity of different cross-sectional imaging modalities for MTF at our institution.We identified n = 52 patients with ICC who underwent curative-intent resection from 2004 to 2017 in a multidisciplinary hepato-pancreato-biliary cancer program. Timing and modality of pre-operative imaging were recorded. Blinded review of imaging was performed and modalities were evaluated for false-negative rate (FNR) in detecting MTF, satellitosis, and true multifocality.Forty-one (79%) patients underwent CT and 20 (38%) underwent MRI prior to hepatectomy. MTF was pre-operatively identified in six (12%) patients. An additional seven patients had MTF discovered on final surgical pathology, despite a median interval from CT/MRI to surgery of 20 days. On blinded review the FNR of MRI compared to CT for multifocality was 0% vs. 38%, 50% vs 80% for satellitosis, and 22% vs 46% for MTF as a whole.CT is inadequate for pre-operative diagnosis of MTF in resectable ICC, even when performed within 30 days of hepatectomy. We recommend liver-protocol MRI as the standard pre-operative imaging modality in non-metastatic ICC.

Introduction Hepatic artery infusion pump (HAIP) chemotherapy is a specialized therapy for patients with unresectable colorectal liver metastases (uCRLM). Its effectiveness was demonstrated from a high volume center, with uncertainty regarding the feasibility and safety at other centers. Therefore, we sought to assess the safety and feasibility of HAIP for the management of uCRLM at other centers. Methods We conducted a multicenter retrospective cohort study of patients with uCRLM treated with HAIP from January 2003 to December 2017 at six North American centers initiating the HAIP program. Outcomes included the safety and feasibility of HAIP chemotherapy. Results We identified 154 patients with HAIP insertion and the median age of 54 (48-61) years. The burden of disease was >10 intra-hepatic metastatic foci in 59 (38.3%) patients. Patients received at least one cycle of systemic chemotherapy before HAIP insertion. Major complications occurred in 7 (4.6%) patients during their hospitalization and 13 (8.4%) patients developed biliary sclerosis during follow-up. A total of 148 patients (96.1%) received at least one-dose of HAIP chemotherapy with a median of 5 (4-7) cycles. 78 patients (56.5%) had a complete or partial response and 12 (7.8%) received a curative liver resection. Conclusion HAIP programs can be safely and effectively initiated in previously inexperienced centers with good response.

Colorectal cancer (CRC) ranks second in cancer deaths worldwide and presents multiple management challenges, one of which is identifying high risk stage II disease that may benefit from adjuvant therapy. Molecular biomarkers, such as ones that identify stem cell activity, could better stratify high-risk cohorts for additional treatment. To identify possible biomarkers of high-risk disease in early-stage CRC, a discovery set (n = 66) of advanced-stage tumors were immunostained with antibodies to stemness proteins (CD166, CD44, CD26, and LGR5) and then digitally analyzed. Using a second validation cohort (n = 54) of primary CRC tumors, we analyzed protein and gene expression of CD166 across disease stages, and extended our analyses to CD166-associated genes (LGR5, ASCL2, BMI1, POSTN, and VIM) by qRT-PCR. Stage III and metastatic CRC tumors highly expressed stem cell-associated proteins, CD166, CD44, and LGR5. When evaluated across stages, CD166 protein expression was elevated in advanced-stage compared to early-stage tumors. Notably, a small subset of stage I and II cancers harbored elevated CD166 protein expression, which correlated with development of recurrent cancer or adenomatous polyps. Gene expression analyses of CD166-associated molecules revealed elevated ASCL2 in primary tumors from patients who recurred. We identified a protein signature prognostic of aggressive disease in early stage CRC. Stem cell-associated protein and gene expression identified a subset of early-stage tumors associated with cancer recurrence and/or subsequent adenoma formation. Signatures for stemness offer promising fingerprints for stratifying early-stage patients at high risk of recurrence.

Gastrointestinal cancers account for more cancer-related deaths than any other organ system, owing in part to difficulties in early detection, treatment response assessment, and post-treatment surveillance. Circulating biomarkers hold the promise for noninvasive liquid biopsy platforms to overcome these obstacles. Although tumors shed detectable levels of degraded genetic material and cellular debris into peripheral blood, identifying reproducible and clinically relevant information from these analytes (eg, cell-free nucleotides, exosomes, proteins) has proven difficult. Cell-based circulating biomarkers also present challenges, but have multiple advantages including allowing for a more comprehensive tumor analysis, and communicating the risk of metastatic spread. Circulating tumor cells have dominated the cancer cell biomarker field with robust evidence in extraintestinal cancers; however, establishing their clinical utility beyond that of prognostication in colorectal and pancreatic cancers has remained elusive. Recently identified novel populations of tumor-derived cells bring renewed potential to this area of investigation. Cancer-associated macrophage-like cells, immune cells with phagocytosed tumor material, also show utility in prognostication and assessing treatment responsiveness. In addition, circulating hybrid cells are the result of tumor–macrophage fusion, with mounting evidence for a role in the metastatic cascade. Because of their relative abundance in circulation, circulating hybrid cells have great potential as a liquid biomarker for early detection, prognostication, and surveillance. In all, the power of the cell reaches beyond enumeration by providing a cellular source of tumor DNA, RNA, and protein, which can be harnessed to impact overall survival. Keywords: Fusion Hybrid, CAML, CHC, Liquid Biopsy, Macrophage

TPS500 Background: The majority of patients with intrahepatic cholangiocarcinoma (ICC) present with advanced liver dominant disease rendering them unresectable. The current standard of care supports palliative treatment of unresectable ICC with gemcitabine plus cisplatin. Alternatively, continuous liver-directed therapy using a surgically implanted hepatic arterial infusion (HAI) pump allows for maximal treatment of liver tumors with limited systemic side-effects and can facilitate conversion to a resectable status. No prospective clinical trial has utilized FOLFIRINOX in combination with HAI floxuridine-dexamethasone for ICC. Methods: HELIX-ICC is a first-line, single-center, single-arm, phase II clinical trial enrolling patients with liver-dominant unresectable or multifocal ICC. Only patients with microsatellite stable cancer and no prior liver radiotherapy will be included. Patients are administered dose-modified FOLFIRINOX systemically for 4 cycles over 8 weeks. Those with evidence of disease control on restaging imaging and laparoscopy will proceed to HAI pump placement. Treatment continues with two 28-day cycles of combined HAI floxuridine (1.08 mg/kg) with dexamethasone for 14 days and systemic dose-modified FOLFIRI starting on day 15. The primary objectives are to evaluate the safety and efficacy (disease control rate [DCR] at 6 months) of this novel treatment approach that utilizes our institutional dose-reduced HAI floxuridine protocol in combination with modified systemic regimens to facilitate control of liver disease and maximize patient quality of life (QoL). HELIX-ICC includes many exploratory analyses through longitudinal collection of blood samples, liver biopsies (including an end of trial research biopsy), and QoL metrics (Table). The study is open to n = 30 with an initial safety run-in of 6 patients, of which 4 have enrolled at the time of submission. This study is designed to allow for drop out of 9 patients, with total accrual of 21 patients to protocol completion achieving 80.2% power at 0.05 significance to detect a 25% increase in DCR at 6 months. Clinical trial information: NCT04251715. [Table: see text]

Hepatic resection for metastatic GIST (mGIST) is often performed with either curative-intent or for tyrosine kinase inhibitor (TKI)-resistant lesions. The efficacy of hepatectomy for treatment-resistant lesions (TRL) is uncertain.We reviewed patients with liver-mGIST treated from 2003 to 2018. Oncologic outcomes including overall (OS), post-operative progression-free survival (PFS), and post-progression OS were evaluated using Kaplan-Meier and Cox proportional hazards modeling.We identified n = 91 patients; 31 (34%) underwent curative-intent hepatectomy, 60 (66%) were initially managed with TKI alone, and 17 (19%) had resection of a TRL. The median follow-up for resected patients was 102 months (range 5-209 months) with 23 (25%) managed with a major hepatectomy. Patients having curative-intent hepatectomy had 72% 10-year OS following diagnosis of liver-mGIST, compared with 58% (P = 0.50) for TRL resection and 41% (P = 0.01) for non-resected patients. Curative-intent hepatectomy (HR 0.39, P = 0.03) and age (HR 1.04, P = 0.004) were independently associated with 10-year OS, but not TRL resection. TRL resection was not associated with improved post-progression OS compared to second-line TKI therapy (HR 0.61, P = 0.21).Curative-intent hepatectomy is associated with improved OS in liver-mGIST. The oncologic benefit of resecting treatment-resistant liver-mGIST compared to second-line TKI therapy alone remains unclear in the era of multi-line TKI therapy.

Surgical site infection is a relatively common and devastating complication following pediatric orthopedic surgery. Many infections have been determined to be the result of settled airborne particles on surgical equipment and the sterile field. Fiberglass casts are commonly used orthopedic fixation devices before and after surgery; however, fiberglass casting material is expelled during the removal process and represents an uninvestigated area for the possibility of cast saw dust as a source of airborne bacterial contamination in an operating room setting. This study evaluates the prevalence and distribution of microbiota on 90 pediatric casts by collecting and culturing fiberglass cast material from 90 pediatric casts. Bacterial identification was performed using a Bruker Biotyper Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry device. 81 out of 90 casts (90%) showed evidence of microbial contamination. Isolated species were very diverse and ranged from normal skin flora to opportunistic pathogens. The 5 most commonly isolated organisms were Acinetobacter pittii, Enterobacter cloacae, Micrococcus luteus, Staphylococcus epidermidis, and Staphylococcus hominis. Further investigation is required to determine if casting material is truly a cause of surgical site infection.

The identification of a disseminated tumor cell generated by heterotypic cell fusion, a circulating hybrid cell (CHCs) is a biologically relevant analyte of disease across myriad of cancers. Detected at elevated levels in peripheral blood, this novel population can be leveraged to understand mechanisms driving metastatic progression and predict metastatic spread. Our long term goal is to leverage evolving knowledge of mechanisms of metastatic progression to identify biologically relevant tumor markers for clinical disease assessment strategies. We therefore detected and delineated the role of CHCs as a measure of disease across a multitude of solid tumors. Human blood specimens were collected and analyzed with IRB approved protocols. Peripheral blood was obtained from patients with cancer in 14 different organ sites: ampullary adenocarcinoma, breast adenocarcinoma, cholangiocarcinoma, colon adenocarcinoma, esophageal cancer, high grade glioma (pediatric & adult), head and neck squamous cell carcinoma, pancreatic ductal adenocarcinoma, pancreatic neuroendocrine tumor, prostate adenocarcinoma, rectal adenocarcinoma and uveal melanoma and healthy subjects. Peripheral blood mononuclear cells were isolated and prepared for flow cytometry or immunofluorescence microscopy analyses with antibodies to CD45 and panCytokeratin, EpCAM, NKI Beteb, chromogranin A (CHGA)/synaptophysin (SYP) or glial fibrillary acid protein (GFAP). Discrete cellular populations including CHCs and circulating tumor cells (CTCs) were identified by protein expression. Cyclic immunofluorescence (cycIF) techniques were performed on a subset of specimens, for indepth interrogation of phenotypic heterogeneity in both the tumor and among CHCs. CHCs, defined by co-expression of CD45+ and a tumor associated protein were detected in all subjects across all disease sites at statistically significantly higher numbers than in controls samples using both detection paradigms. Furthermore, higher levels of CHCs compared with CTCs (that were CD45-) across disease sites. CycIF analysis of tumor tissue and dissemnated tumor cells from a triple negative breast cancer subject revealed a heterogeneous population of CHCs as well as complex tumor ecosystem. The phenotypic diversity of CHCs included CD44 and androgen receptor (AR) expression, which aligned with those detected in the proliferative tumor compartment. AR expression in triple negative breast cancer is known to correlate with treatment resistance and indeed, our subject rapidly succumb to disease. Taken together, these data suggest that tumors disseminate a heterogeneous population of CHCs that reflect the overall phenotype of viable tumor, supporting the value of phenotypically profiling CHCs for liquid biopsy to anticipate disease evolution and treatment resistance. This study established an isolation and detection platform for measuring CHCs in myriad of cancers and methods of phenotypic profiling CHC heterogeneity to identify discrete subpopulations. Citation Format: Matthew S. Dietz, Thomas Sutton, Brett Walker, Young Hwan Chang, Koei Chin, Melissa H. Wong. A novel disseminated tumor cell identified in myriad cancer harbors tumor initiating properties [abstract]. In: Proceedings of the AACR Virtual Special Conference on Tumor Heterogeneity: From Single Cells to Clinical Impact; 2020 Sep 17-18. Philadelphia (PA): AACR; Cancer Res 2020;80(21 Suppl):Abstract nr PO-102.

Pancreatic adenocarcinoma (PDAC) remains one of the most lethal malignancies in the United States and tumor heterogeneity has major implications on the effectiveness of systemic therapies. Comprehensive tumor analysis at the single cell level offers an in-depth examination of neoplastic cells with metastatic potential and supports development of individualized treatment strategies to ultimately improve survival. Hybrid cells are a novel cell population derived from immune-cancer cell fusion within solid tumors and disseminate into peripheral blood, where they can be detected as circulating hybrid cells (CHCs) based on their dual expression of tumor and immune cell epitopes. Compared to conventionally-defined circulating tumor cells (CTCs), CHCs are found in patients at higher levels which allows for more robust phenotypic analyses of tumor cell features and collection for downstream multi-omic profiling. To investigate heterogeneity among disseminated CHCs in patients with PDAC, we assessed for tumor cell features using immunohistochemical analyses of single cells from peripheral blood mononuclear cell smears and by single cell RNA sequencing (scRNA-seq) from FACS-isolated cells. Downstream image analyses was performed Zeiss Zen blue software and scRNA-seq was analyzed using the Seurat analysis package. We determined that PDAC patients harbored heterogeneous populations of CHCs with differential protein expression that mirrored expression in corresponding primary tumors. Further, scRNA-seq analyses identified CHC populations with varying degrees of retained tumor identity, as well as subsets derived from different immune cell lineages. Unsupervised clustering of CHCs with tumor-dominate phenotypes demonstrated high differential expression of cancer related genes with known implications in PDAC tumors and treatment resistant pathways. Inference copy number variation analysis revealed chromosomal amplifications of 1q and 19q along with deletions of 6p, 12q, and 22q, characteristic of PDAC tumors. Additionally, a myriad of KRAS mutations were identified in patient CHCs but absent in normal leukocytes. As the product of tumor and immune cell fusion, CHCs provide a rich source for assessing PDAC heterogeneity through genetic and protein expression analyses. Ultimately, CHCs have great potential as a liquid biopsy in pancreatic cancer which could be harnessed to guide therapy. Citation Format: Brett Scott Walker, Sidharth Sengupta, Michael Parappilly, Aysegul Ors, Jared Fischer, Thomas Sutton, Brett Sheppard, Skye Mayo, Melissa Wong. Harnessing the heterogeneity of circulating hybrid cells in pancreatic adenocarcinoma [abstract]. In: Proceedings of the AACR Virtual Special Conference on Tumor Heterogeneity: From Single Cells to Clinical Impact; 2020 Sep 17-18. Philadelphia (PA): AACR; Cancer Res 2020;80(21 Suppl):Abstract nr PO-014.

Topical haemostatic agents have become an essential tool to assist with the control of bleeding during surgery as well as to facilitate wound closure. The imaging appearance of these agents can overlap that of abscess or tumour. Knowledge of the appearance of these various agents on ultrasound and CT is crucial to avoid misdiagnosing pathology, potentially resulting in unnecessary interventional procedures.

Background Infections caused by community-associated strains of methicillin-resistant Staphylococcus aureus (CA-MRSA) are associated with a marked and prolonged host inflammatory response. In a sepsis simulation model, we tested whether the anesthetic ketamine inhibits the macrophage TNF response to antibiotic-exposed CA-MRSA bacteria via its antagonism of N-methyl-D-aspartate (NMDA) receptors. RAW264.7 cells were stimulated for 18 hrs with 105 to 107 CFU/mL inocula of either of two prototypical CA-MRSA isolates, USA300 strain LAC and USA400 strain MW2, in the presence of either vancomycin or daptomycin. One hour before bacterial stimulation, ketamine was added with or without MK-801 (dizocilpine, a chemically unrelated non-competitive NMDA receptor antagonist), APV (D-2-amino-5-phosphono-valerate, a competitive NMDA receptor antagonist), NMDA, or combinations of these agents. Supernatants were collected and assayed for TNF concentration by ELISA. Results RAW264.7 cells exposed to either LAC or MW2 in the presence of daptomycin secreted less TNF than in the presence of vancomycin. The addition of ketamine inhibited macrophage TNF secretion after stimulation with either of the CA-MRSA isolates (LAC, MW2) in the presence of either antibiotic. The NMDA inhibitors, MK-801 and APV, also suppressed macrophage TNF secretion after stimulation with either of the antibiotic-exposed CA-MRSA isolates, and the effect was not additive or synergistic with ketamine. The addition of NMDA substrate augmented TNF secretion in response to the CA-MRSA bacteria, and the addition of APV suppressed the effect of NMDA in a dose-dependent fashion. Conclusions Ketamine inhibits TNF secretion by MRSA-stimulated RAW264.7 macrophages and the mechanism likely involves NMDA receptor antagonism. These findings may have therapeutic significance in MRSA sepsis.