Prospective Evaluation of Circulating Tumor DNA using Next Generation Sequencing as a Biomarker during Neoadjuvant Chemotherapy in Localized Pancreatic Cancer.

OBJECTIVE: In this prospective study, we aim to characterize the prognostic value of circulating tumor DNA (ctDNA) by next-generation-sequencing (NGS) in patients undergoing neoadjuvant chemotherapy (NAC) for pancreatic ductal adenocarcinoma (PDAC). SUMMARY BACKGROUND DATA: Circulating tumor DNA is a promising blood-based biomarker that is prognostic in several malignancies. Detection of ctDNA by NGS may provide insights regarding the mutational profiles in PDAC to help guide clinical decisions for patients in a potentially curative setting. However, the utility of ctDNA as a biomarker in localized PDAC remains unclear. METHODS: Patients with localized PDAC were enrolled in a prospective study at Northwestern Medicine between October 2020 and October 2022. Blood samples were collected to perform targeted tumor agnostic NGS utilizing the Tempus x|F 105 gene panel at three timepoints: pre-therapy (at diagnosis), post-NAC, and after local therapy, including surgery. The relationship between ctDNA detection and CA19-9, and the prognostic significance of ctDNA detection were analyzed. RESULTS: 56 patients were included in the analysis. ctDNA was detectable in 48% at diagnosis, 33% post-NAC, and 41% after local therapy. After completion of NAC, patients with detectable ctDNA had higher CA19-9 levels versus those without (78.4 vs. 30.0, P=0.02). The presence of baseline ctDNA was associated with a CA19-9 response; those without ctDNA had a significant CA19-9 response following NAC (109.0 U/mL vs. 31.5 U/mL; P=0.01), while those with ctDNA present at diagnosis did not (198.1 U/mL vs. 113.8 U/mL; P=0.77). In patients treated with NAC, the presence of KRAS ctDNA at diagnosis was associated with and independently predicted worse progression-free-survival. CONCLUSION: This report demonstrates the prognostic value of ctDNA analysis with NGS in localized PDAC. NGS ctDNA is a biomarker of treatment response to NAC. KRAS ctDNA at diagnosis independently predicts worse survival in patients treated with NAC.

Comparison of perioperative and histopathologic outcomes among neoadjuvant treatment strategies for locoregional gastric cancer.

BACKGROUND AND OBJECTIVES: Neoadjuvant chemotherapy (NAC) and chemoradiation (NCRT) have demonstrated improved survival for gastric cancer. However, the optimal neoadjuvant treatment remains unclear. We sought to evaluate perioperative and histopathologic outcomes among neoadjuvant treatments for locoregional gastric cancer. METHODS: The National Cancer Database queried patients who received NAC or NCRT followed by resection for T2-T4 and/or node-positive gastric cancer (2006-2018). Logistic and Poisson regression assessed perioperative (30-day readmission, 30- and 90-day mortality, length of stay [LOS]) and histopathologic outcomes (pathologic complete response [PCR], margin status, and negative pathologic lymph nodes [ypN0]). Kaplan-Meier methods and Cox regression assessed overall survival (OS). RESULTS: Of 9831 patients, 4221 (42.9%) received NAC and 5610 (57.1%) NCRT. There were no differences in perioperative outcomes, apart from patients treated with NCRT exhibiting increased LOS (incidence rate ratio 1.09, 95% confidence interval [CI] 1.03-1.16). Patients who received NCRT were more likely to achieve PCR, margin-negative resection, and ypN0 (all p < 0.05). Median OS was 36.8 months for NAC and 33.6 months for NCRT (p < 0.001). NCRT independently predicted worse OS (vs. NAC, hazard ratio 1.10, 95% CI 1.03-1.18). CONCLUSION: NCRT was associated with better histologic tumor response although NAC was associated with improved OS. Better understanding prognostication through histologic assessment following neoadjuvant therapy is needed.

Weight loss during neoadjuvant chemotherapy impacts perioperative outcomes in patients undergoing surgery for pancreatic cancer.

BACKGROUND: While use of neoadjuvant chemotherapy (NAC) in pancreatic adenocarcinoma (PDAC) downstages cancers to be eligible for resection, weight loss during the neoadjuvant period due to cancer progression, gastric outlet obstruction, or neoadjuvant therapy itself is an area of concern. The goal of this study is to determine the effect of weight loss during NAC on perioperative outcomes of pancreatectomies. METHODS: The NSQIP database 2014-2019 was utilized to study patients who received NAC for PDAC and underwent significant weight loss, defined as at least 10 % body weight loss in the six months prior to surgery. Univariate and multivariate analyses were conducted using Fisher's Exact Test, Pearson's Chi-squared Test, and logistic regression. RESULTS: Of the 5590 PDAC patients who received NAC, 913 (16%) experienced significant weight loss. Patients who experienced significant weight loss were more likely to experience at least one complication compared to those who did not undergo weight loss (42.2% vs. 38.7%, p = 0.023). Those who had significant weight loss were more likely to undergo unplanned intubation postoperatively (3.8% vs 2.2 %, p = 0.004), have postoperative ventilator need >48 h (3.7% vs 1.8%, p < 0.001), have postoperative septic shock (3.9% vs 1.8 %, p < 0.001), and undergo reoperation (6.0% vs 4.3%, p = 0.027). However, there were no differences for pancreatic fistula (7.7% vs 9.3 %, p = 0.15), readmission rates (15% vs 15 %, p = 0.7), or 30-day mortality (1.5% vs 1.2%, p = 0.5). Utilizing logistic regression, BMI (OR: 1.05, p = 0.032), significant weight loss (OR = 1.18, p = 0.025), sex (OR = 1.26 with female baseline, p < 0.001), history of COPD (OR = 1.39, p = 0.012), hypertensive medication use (OR = 1.18, p = 0.004), and pancreatic radiotherapy (OR = 1.16, p = 0.010) were independent preoperative predictors of a post-operative complication. CONCLUSIONS: Nutritional measures to stabilize weight during NAC should be considered to decrease post-pancreatectomy complications.

The differential effect of neoadjuvant chemotherapy and chemoradiation on nodal downstaging in pancreatic adenocarcinoma.

BACKGROUND/OBJECTIVES: Neoadjuvant chemotherapy (NCT) and chemoradiotherapy (NCRT) enhance resectability in patients with pancreatic adenocarcinoma (PDAC). This study compares the effect of NCT and NCRT on lymph nodal downstaging and survival. METHODS: The 2004-2016 National Cancer Database Pancreas Participant User File was used to identify patients who underwent surgery for PDAC. Fisher's exact, Wilcoxon rank-sum, multivariate logistic regression, and log-rank were used. Downstaging was defined as clinically node-positive patients who demonstrated node-negativity on pathology. RESULTS: Of 42,545 patients meeting criteria, 3311 received NCT and 1511 received NCRT. After surgery for clinically node-positive disease, 23.3% of NCT patients and 41.3% of NCRT patients demonstrated nodal downstaging. Younger age and lower tumor grade independently predicted downstaging. Downstaging after neoadjuvant therapy was associated with improved survival versus no nodal treatment response (29.8 vs. 22.8 months, p < 0.001). Downstaging by NCT was associated with improved overall survival versus downstaging by NCRT (37.5 vs. 26.6 months, p = 0.001). No survival difference existed between those with no nodal response after NCT or NCRT (p = 0.101). CONCLUSIONS: Although nodal downstaging is more likely post-NCRT, survival is superior in those downstaged post-NCT. Overall survival is determined by the systemic burden of disease. Post-therapy histologic analysis may be less prognostic post-NCRT.

The inaccuracies of gastric adenocarcinoma clinical staging and its predictive factors.

INTRODUCTION: Accurate clinical staging (CS) of gastric adenocarcinoma is important to guide treatment planning. Our objectives were to (1) assess clinical to pathologic stage migration patterns for patients with gastric adenocarcinoma, (2) identify factors associated with inaccurate CS, and (3) evaluate the association of understaging with survival. METHODS: The National Cancer Database was queried for patients who underwent upfront resection for stage I-III gastric adenocarcinoma. Multivariable logistic regression was used to detect factors associated with inaccurate understaging. Kaplan-Meier analyses and cox proportional hazards regression were performed to assess overall survival (OS) for patients with inaccurate CS. RESULTS: Of 14 425 analyzed patients, 5781 (40.1%) patients were inaccurately staged. Factors associated with understaging included treatment at a Comprehensive Community Cancer Program, presence of lymphovascular invasion, moderate to poor differentiation, large tumor size, and T2 disease. Based on overall CS, median OS was 51.0 months for accurately staged patients and 29.5 months for understaged patients (<0.001). CONCLUSION: Clinical T-category, large tumor size, and worse histologic features lead to inaccurate CS for gastric adenocarcinoma, impacting OS. Improvements to staging parameters and diagnostic modalities focusing on these factors may improve prognostication.

Identification of high-risk germline variants for the development of pancreatic cancer: Common characteristics and potential guidance to screening guidelines.

Pancreatic cancer (PC) is a product of a variety of environmental and genetic factors. Recent work has highlighted the influence of hereditary syndromes on pancreatic cancer incidence. The purpose of this review is to identify the high-risk syndromes, common variants, and risks associated with PC. The study also elucidates common characteristics of patients with these mutations, which is used to recommend potential changes to current screening protocols for greater screening efficacy. We analyzed 8 syndromes and their respective variants: Hereditary Breast and Ovarian Cancer (BRCA1/2), Familial Atypical Multiple Mole Melanoma Syndrome (CDKN2A), Peutz-Jeghers Syndrome (STK11), Lynch Syndrome (PMS2, MLH1, MSH2, MSH6, EPCAM), Ataxia Telangiectasia (ATM), Li-Fraumeni Syndrome (TP53), Fanconi Anemia (PALB2), and Hereditary Pancreatitis (PRSS1, SPINK1, CFTR). Of 587 studies evaluated, 79 studies fit into our inclusion criteria. Information from each study was analyzed to draw conclusions on these variants as well as their association with pancreatic cancer. Information from this review is intended to improve precision medicine and improve criteria for screening.

The role of oncologic resection and enucleation for small pancreatic neuroendocrine tumors.

BACKGROUND: Surgical management of small pancreatic neuroendocrine tumors (PNETs) is variable. Patients may undergo formal oncologic resection, encompassing regional lymphadenectomy, or enucleation. This study's aim was to understand if enucleation is adequate treatment for PNETs <2 cm METHODS: The US National Cancer Database (NCDB) from 2004 to 2016 was used to identify patients who underwent oncologic resection or enucleation for PNETs <2 cm. Fisher's exact test, log-rank, and logistic regression were used. RESULTS: Of 4083 patients, 75.6% underwent oncologic resection with a median (range) number of 8 (0-99) lymph nodes examined, and 24.1% underwent enucleation. Five-year overall survival rate was 89.7% in node-negative patients versus 82.1% in node-positive patients (p < 0.001). No survival difference existed between patients who underwent enucleation versus oncologic resection (5-yr OS of 88.5% vs 88.2%, p = 0.064). According to AJCC classification, 3776 patients were clinically-staged with evidence of node-negative disease. Of these, 75.1% underwent oncologic resection, of which 9.9% had node-positive disease after resection. Tumor grade and size independently predicted nodal upstaging after oncologic resection. CONCLUSION: One-tenth of patients with clinically node-negative disease were node-positive after surgery. Although this was not reflected in overall survival, patients who receive enucleation with higher grade and larger size may benefit from enhanced surveillance for locoregional recurrence.

Liver resection for metastatic disease after y90 radioembolization: a case series with long-term follow-up.

INTRODUCTION: There are only few reports of liver resections for metastatic disease in patients previously treated with Y-90 radioembolization (RE), and long-term outcome data are sparse. We reviewed our center's experience in patients undergoing hepatectomy after hepatic RE. METHODS: A retrospective chart review of patients undergoing RE from 2004 to 2011 was performed. Demographic, clinicopathologic, operative, and long-term outcomes variables were collected. Independent pathologic review of tumor necrosis and normal liver tissue grading of fibrosis and inflammation after resection was performed. Data are expressed as medians and ranges. RESULTS: RE was delivered to 106 patients with primary and metastatic disease of the liver, of whom 9 patients (6 males, 3 females, median age 54 (47-76) years) with metastatic disease ultimately underwent resection. RE was previously administered to the right liver in five, the left liver in one, and to the whole liver in three. Two patients had a second RE performed before resection. Six of the nine patients had previously received several infusions of cytotoxic therapy. The operations occurred at a median of 115 (56-245) days after RE and included right lobectomy (n = 5), left lobectomy (n = 1), left-lateral sectionectomy (n = 1), and bilobar wedge resections (n = 2). Extrahepatic sites were resected in three patients. Median blood loss was 900 (range 250-3600) ml. Grade 3 or higher complications occurred in seven cases (78 %). Follow-up was complete all nine patients. Three patients (33 %) died within 30 days of resection. All those surviving the operative period had disease recurrence (time to recurrence: 202 [range 54-315] days), and all have since died (overall survival: 584 [range 127-1230] days). Review of resected specimens demonstrated median tumor necrosis of 70 % (range 20-90 %). In nontumor-bearing liver, fibrosis grade (0-4) and inflammation score (0-4) was 2 or less in all specimens. CONCLUSIONS: In this small cohort of highly selected and heavily pretreated patients, long-term survival in patients undergoing resection after RE appears possible, but the operations may carry substantial risks-highlighting the importance of careful patient selection for these resections. The etiology of morbidity and mortality is likely multifactorial and additional reports that include long-term outcomes will be necessary to identify more clearly the impact of RE on postoperative complications and death.

Characterization of human T lymphocytes engineered to express interleukin-15 and herpes simplex virus-thymidine kinase.

INTRODUCTION: Preclinical studies have demonstrated that tumor-reactive T cells expressing the interleukin (IL)-15 transgene had enhanced activity. Gene therapy strategies using IL-15 should include a safety mechanism in anticipation of possible adverse effects because IL-15 overexpression has been implicated in autoimmune disorders and may be involved in the pathogenesis of some leukemias. We developed a retroviral vector carrying both IL-15 and the herpes simplex virus-thymidine kinase (HSV-TK) suicide gene and characterized its application in the transduction of human T lymphocytes. METHODS: A retroviral vector carrying IL-15 and HSV-TK genes was optimized for the transduction of human T lymphocytes. IL-15 production was measured by enzyme-linked immunosorbent assay. Thymidine incorporation and cell viability assays were used to assess the efficacy of the HSV-TK suicide gene. Genetically modified tumor-infiltrating lymphocytes (TILs) were assayed for survival after withdrawal from exogenous IL-2. The activity and specificity of retrovirally transduced TILs were assessed using tumor coculture assays. RESULTS: Human T cells transduced with the IL-15 HSV-TK vector exhibited thymidine uptake in the absence of exogenous cytokine support and survived in culture for up to 80 d without IL-2. IL-15 HSV-TK-transduced T cells were efficiently killed by ganciclovir at concentrations as low as 0.1 µM. TILs transduced with the IL-15 HSV-TK vector retained specific recognition of HLA-A2+, MART1+ melanomas, even after withdrawal of IL-2. CONCLUSIONS: Human T lymphocytes genetically modified with the IL-15 HSV-TK retroviral vector retained the ability to recognize tumor antigen while gaining the ability to secrete IL-15 and prolong their own survival. IL-15 HSV-TK-transduced T cells expressed HSV-TK and could be efficiently eliminated by ganciclovir.

Lentiviral vector design for optimal T cell receptor gene expression in the transduction of peripheral blood lymphocytes and tumor-infiltrating lymphocytes.

Lentiviral vectors containing promoters of distinct origins, that is, strong viral promoters (cytomegalovirus [CMV] and murine stem cell virus [MSCV]), a cellular promoter (phosphoglycerate kinase [PGK]), and two composite promoters (CAG [a composite promoter sequence comprised of the CMV enhancer and portions of the chicken beta-actin promoter and the rabbit beta-globin gene] and SV40/CD43), were used to evaluate green fluorescent protein (GFP) reporter gene expression in human primary peripheral blood lymphocytes (PBLs) and tumor-infiltrating lymphocytes (TILs). In PBLs, vectors containing the MSCV promoter were found to be optimal for expression in both minimally stimulated and highly activated lymphocytes. The stability of gene expression was monitored for up to 7 weeks in culture and the MSCV promoter-containing vector was found to be comparable to the cellular PGK promoter-containing vector. The MSCV promoter-containing lentiviral vector was also the most active in transduced TILs and these cells retained biological activity as measured by antimelanoma antigen reactivity. Using the knowledge gained in comparing individual promoters, a series of two-gene-containing lentiviral vectors was constructed in an attempt to produce the alpha and beta chains of antitumor antigen T cell receptors (TCRs). Dual-promoter or internal ribosome entry site (IRES)-containing vector designs were evaluated and found to be unable to produce both chains of the TCR in amounts that led to significant biological activity. In contrast, if the alpha and beta chains were linked by a 2A ribosomal skip peptide, both proper TCR chain pairing and biologically activity were observed. This paper emphasizes the need to optimize both promoter function and protein synthesis in constructs that require stoichiometric production of multiple protein subunits.

T-cell receptor gene therapy of established tumors in a murine melanoma model.

Adoptive cell transfer therapy using tumor-infiltrating lymphocytes for patients with metastatic melanoma has demonstrated significant objective response rates. One major limitation of these current therapies is the frequent inability to isolate tumor-reactive lymphocytes for treatment. Genetic engineering of peripheral blood lymphocytes with retroviral vectors encoding tumor antigen-specific T-cell receptors (TCRs) bypasses this restriction. To evaluate the efficacy of TCR gene therapy, a murine treatment model was developed. A retroviral vector was constructed encoding the pmel-1 TCR genes targeting the B16 melanoma antigen, gp100. Transduction of C57BL/6 lymphocytes resulted in efficient pmel-1 TCR expression. Lymphocytes transduced with this retrovirus specifically recognized gp100-pulsed target cells as measured by interferon-gamma secretion assays. Upon transfer into B16 tumor-bearing mice, the genetically engineered lymphocytes significantly slowed tumor development. The effectiveness of tumor treatment was directly correlated with the number of TCR-engineered T cells administered. These results demonstrated that TCR gene therapy targeting a native tumor antigen significantly delayed the growth of established tumors. When C57BL/6 lymphocytes were added to antigen-reactive pmel-1 T cells, a reduction in the ability of pmel-1 T cell to treat B16 melanomas was seen, suggesting that untransduced cells may be deleterious to TCR gene therapy. This model may be a powerful tool for evaluating future TCR gene transfer-based strategies.

Treatment of oligometastases after successful immunotherapy.

Local destruction of individual metastases by any of a number of effective modalities fails as a treatment for most patients with disseminated cancer because of the presence of either undetected micrometastases or simply too many lesions. The availability of a systemic therapy that could reduce the number of metastases to a manageable few would dramatically increase the utility of surgical metastasectomy or other locally ablative measures. Interleukin-2-based immunotherapy can serve exactly this function in some patients with renal cancer or melanoma. We review the effectiveness of surgery in treating limited relapses or residual disease in patients who have responded to systemic immunotherapy. These data indicate that a surprising percentage of such patients can enjoy durable disease-free survival after surgical removal of their oligometastases, and, for a significant minority, it appears to be curative.