Risk Factors for Lymph Node Metastasis in a Western Series of Patients with Distal Early Gastric Cancer.

Background: Assessment of potential lymph node metastasis is mandatory in the appropriate treatment of early gastric cancers. This study analysed factors associated with lymph node metastasis to identify differences between node-negative and node-positive patients and between T1a and T1b cancers. Methods: The clinicopathological features of 129 early gastric cancer patients who had undergone radical gastrectomy were analysed to identify predictive factors for lymph node metastasis. Results: Lymph node metastasis was detected in 76 (59.0%) patients. Node-positive patients were younger (58.1 ± 11.3 years) than those without metastasis (61.9 ± 9.6 years, p = 0.02). Greater tumour sizes were observed in patients with lymph node metastasis (3.6 ± 1.0 cm) compared to node-negative patients (1.9 ± 0.5 cm, p = 0.00001). Depressed form, ulceration, diffuse histological type, and undifferentiated lesions were more frequent in node-positive patients than in the node-negative group. Tumour size > 3.0 cm showed a correlation with lymph node metastasis in both T1a (p = 0.0001) and T1b (p = 0.006) cancer. The male sex (p = 0.006) had a significant correlation with lymph node metastasis in T1a cancer. Depressed appearance (p = 0.02), ulceration (p = 0.03), differentiation (p = 0.0001), diffuse type (p = 0.0002), and lower third location (p = 0.005) were associated with lymph node metastasis in T1b cancer. Conclusions: Tumour size > 3 cm, undifferentiated lesions, ulceration, diffuse type, lower third location, and submucosal invasion are risk factors for lymph node metastasis in early gastric cancer.

Entero-Cutaneous and Entero-Atmospheric Fistulas: Insights into Management Using Negative Pressure Wound Therapy.

Enteric fistulas are a common problem in gastrointestinal tract surgery and remain associated with significant mortality rates, due to complications such as sepsis, malnutrition, and electrolyte imbalance. The increasingly widespread use of open abdomen techniques for the initial treatment of abdominal sepsis and trauma has led to the observation of so-called entero-atmospheric fistulas. Because of their clinical complexity, the proper management of enteric fistula requires a multidisciplinary team. The main goal of the treatment is the closure of enteric fistula, but also mortality reduction and improvement of patients' quality of life are fundamental. Successful management of patients with enteric fistula requires the establishment of controlled drainage, management of sepsis, prevention of fluid and electrolyte depletion, protection of the skin, and provision of adequate nutrition. Many of these fistulas will heal spontaneously within 4 to 6 weeks of conservative management. If closure is not accomplished after this time point, surgery is indicated. Despite advances in perioperative care and nutritional support, the mortality remains in the range of 15 to 30%. In more recent years, the use of negative pressure wound therapy for the resolution of enteric fistulas improved the outcomes, so patients can be successfully treated with a non-operative approach. In this review, our intent is to highlight the most important aspects of negative pressure wound therapy in the treatment of patients with enterocutaneous or entero-atmospheric fistulas.

Artificial intelligence in the diagnosis and treatment of acute appendicitis: a narrative review.

Artificial intelligence is transforming healthcare. Artificial intelligence can improve patient care by analyzing large amounts of data to help make more informed decisions regarding treatments and enhance medical research through analyzing and interpreting data from clinical trials and research projects to identify subtle but meaningful trends beyond ordinary perception. Artificial intelligence refers to the simulation of human intelligence in computers, where systems of artificial intelligence can perform tasks that require human-like intelligence like speech recognition, visual perception, pattern-recognition, decision-making, and language processing. Artificial intelligence has several subdivisions, including machine learning, natural language processing, computer vision, and robotics. By automating specific routine tasks, artificial intelligence can improve healthcare efficiency. By leveraging machine learning algorithms, the systems of artificial intelligence can offer new opportunities for enhancing both the efficiency and effectiveness of surgical procedures, particularly regarding training of minimally invasive surgery. As artificial intelligence continues to advance, it is likely to play an increasingly significant role in the field of surgical learning. Physicians have assisted to a spreading role of artificial intelligence in the last decade. This involved different medical specialties such as ophthalmology, cardiology, urology, but also abdominal surgery. In addition to improvements in diagnosis, ascertainment of efficacy of treatment and autonomous actions, artificial intelligence has the potential to improve surgeons' ability to better decide if acute surgery is indicated or not. The role of artificial intelligence in the emergency departments has also been investigated. We considered one of the most common condition the emergency surgeons have to face, acute appendicitis, to assess the state of the art of artificial intelligence in this frequent acute disease. The role of artificial intelligence in diagnosis and treatment of acute appendicitis will be discussed in this narrative review.

A phase 1 trial of adoptive transfer of vaccine-primed autologous circulating T cells in ovarian cancer.

We have previously shown that vaccination with tumor-pulsed dendritic cells amplifies neoantigen recognition in ovarian cancer. Here, in a phase 1 clinical study ( NCT01312376 /UPCC26810) including 19 patients, we show that such responses are further reinvigorated by subsequent adoptive transfer of vaccine-primed, ex vivo-expanded autologous peripheral blood T cells. The treatment is safe, and epitope spreading with novel neopeptide reactivities was observed after cell infusion in patients who experienced clinical benefit, suggesting reinvigoration of tumor-sculpting immunity.

Surgical complications of oncological treatments: A narrative review.

Gastrointestinal complications are common in patients undergoing various forms of cancer treatments, including chemotherapy, radiation therapy, and molecular-targeted therapies. Surgical complications of oncologic therapies can occur in the upper gastrointestinal tract, small bowel, colon, and rectum. The mechanisms of action of these therapies are different. Chemotherapy includes cytotoxic drugs, which block the activity of cancer cells by targeting intracellular DNA, RNA, or proteins. Gastrointestinal symptoms are very common during chemotherapy, due to a direct effect on the intestinal mucosa resulting in edema, inflammation, ulceration, and stricture. Serious adverse events have been described as complications of molecular targeted therapies, including bowel perforation, bleeding, and pneumatosis intestinalis, which may require surgical evaluation. Radiotherapy is a local anti-cancer therapy, which uses ionizing radiation to cause inhibition of cell division and ultimately lead to cell death. Complications related to radiotherapy can be both acute and chronic. Ablative therapies, including radiofrequency, laser, microwave, cryoablation, and chemical ablation with acetic acid or ethanol, can cause thermal or chemical injuries to the nearby structures. Treatment of the different gastrointestinal complications should be tailored to the individual patient and based on the underlying pathophysiology of the complication. Furthermore, it is important to know the stage and prognosis of the disease, and a multidisciplinary approach is necessary to personalize the surgical treatment. The purpose of this narrative review is to describe complications related to different oncologic therapies that may require surgical interventions.

Targeting of multiple tumor-associated antigens by individual T cell receptors during successful cancer immunotherapy.

The T cells of the immune system can target tumors and clear solid cancers following tumor-infiltrating lymphocyte (TIL) therapy. We used combinatorial peptide libraries and a proteomic database to reveal the antigen specificities of persistent cancer-specific T cell receptors (TCRs) following successful TIL therapy for stage IV malignant melanoma. Remarkably, individual TCRs could target multiple different tumor types via the HLA A∗02:01-restricted epitopes EAAGIGILTV, LLLGIGILVL, and NLSALGIFST from Melan A, BST2, and IMP2, respectively. Atomic structures of a TCR bound to all three antigens revealed the importance of the shared x-x-x-A/G-I/L-G-I-x-x-x recognition motif. Multi-epitope targeting allows individual T cells to attack cancer in several ways simultaneously. Such "multipronged" T cells exhibited superior recognition of cancer cells compared with conventional T cell recognition of individual epitopes, making them attractive candidates for the development of future immunotherapies.

The spectrum of pneumatosis intestinalis in the adult. A surgical dilemma.

Pneumatosis intestinalis (PI) is a striking radiological diagnosis. Formerly a rare diagnostic finding, it is becoming more frequently diagnosed due to the wider availability and improvement of computed tomography scan imaging. Once associated only with poor outcome, its clinical and prognostic significance nowadays has to be cross-referenced to the nature of the underlying condition. Multiple mechanisms of pathogenesis have been debated and multiple causes have been detected during the years. All this contributes to creating a broad range of clinical and radiological presentations. The management of patients presenting PI is related to the determining cause if it is identified. Otherwise, in particular if an association with portal venous gas and/or pneumoperitoneum is present, the eventual decision between surgery and non-operative management is challenging, even for stable patients, since this clinical condition is traditionally associated to intestinal ischemia and consequently to pending clinical collapse if not treated. Considering the wide variety of origin and outcomes, PI still remains for surgeons a demanding clinical entity. The manuscript is an updated narrative review and gives some suggestions that may help make the decisional process easier, identifying patients who can benefit from surgical intervention and those who can benefit from non-operative management avoiding unnecessary procedures.

Pattern of Distribution of Lymph Node Metastases in Individual Stations in Middle and Lower Gastric Carcinoma.

(1) Background: Lymph node (LN) dissection is the cornerstone of curative treatment of GC. The pattern of distribution of LN metastases is closely related to several factors. The aim of this study is to evaluate the factors determining the distribution of nodal metastases in a population of N+ distal GC patients undergoing gastrectomy and D2 lymphadenectomy. (2) Methods: The medical charts of 162 N+ GC patients who underwent surgical resection over a 15-year period were retrospectively analyzed. Clinical, pathological and anatomical characteristics were evaluated to identify the factors affecting the patterns and prevalence of metastases in individual LN stations. (3) Results: LN metastasis is correlated with the depth of the tumor and to diffuse-type tumors. A higher number of metastatic nodes was documented in patients with middle-third tumors (8.2 ± 7.3 vs. 4.5 ± 5.0 in lower-third tumors, p = 0.0001) and in patients with tumors located on the lesser curve. Station 4 showed the highest rate of metastases (53.1%). Concerning stations 7 to 12, station 8 showed the highest metastasis rate (28.4%). Metastases at stations 1, 2, 4 and 7 to 11 were dominant in middle-third cancer, whereas stations 5 and 6 were dominant in lower-third cancers. Station 4, 5, 6, 10 and 11 metastases were dominant when the cancer was located on the greater curve, whereas stations 1, 2, 7, 8 and 12 were dominant in lesser-curve cancers. (4) Conclusions: The study documented that in patients with distal GC, the distribution of nodal metastases at individual stations is closely related to primary tumor location.

Perforated peptic ulcer (PPU) treatment: an Italian nationwide propensity score-matched cohort study investigating laparoscopic vs open approach.

BACKGROUND: Perforated peptic ulcer (PPU) remain a surgical emergency accounting for 37% of all peptic ulcer-related deaths. Surgery remains the standard of care. The benefits of laparoscopic approach have been well-established even in the elderly. However, because of inconsistent results with specific regard to some technical aspects of such technique surgeons questioned the adoption of laparoscopic approach. This leads to choose the type of approach based on personal experience. The aim of our study was to critically appraise the use of the laparoscopic approach in PPU treatment comparing it with open procedure. METHODS: A retrospective study with propensity score matching analysis of patients underwent surgical procedure for PPU was performed. Patients undergoing PPU repair were divided into: Laparoscopic approach (LapA) and Open approach (OpenA) groups and clinical-pathological features of patients in the both groups were compared. RESULTS: A total of 453 patients underwent PPU simple repair. Among these, a LapA was adopted in 49% (222/453 patients). After propensity score matching, 172 patients were included in each group (the LapA and the OpenA). Analysis demonstrated increased operative times in the OpenA [OpenA: 96.4 ± 37.2 vs LapA 88.47 ± 33 min, p = 0.035], with shorter overall length of stay in the LapA group [OpenA 13 ± 12 vs LapA 10.3 ± 11.4 days p = 0.038]. There was no statistically significant difference in mortality [OpenA 26 (15.1%) vs LapA 18 (10.5%), p = 0.258]. Focusing on morbidity, the overall rate of 30-day postoperative morbidity was significantly lower in the LapA group [OpenA 67 patients (39.0%) vs LapA 37 patients (21.5%) p = 0.002]. When stratified using the Clavien-Dindo classification, the severity of postoperative complications was statistically different only for C-D 1-2. CONCLUSIONS: Based on the present study, we can support that laparoscopic suturing of perforated peptic ulcers, apart from being a safe technique, could provide significant advantages in terms of postoperative complications and hospital stay.

Endoluminal vacuum-assisted therapy as a treatment for anastomotic leakage in colorectal surgery.

Anastomotic leakage (AL) has a wide range of clinical features ranging from radiological only findings to peritonitis and sepsis with multiorgan failure. An early diagnosis of AL is essential in order to establish the most appropriate treatment for this complication. Despite AL continues to be a dreadful compli-cation after colorectal surgery, there has been no consensus on its management. However, based on patient's presentation and timing of the AL, there has been a gradual shift to a more conservative management, keeping surgery as the last option Reoperation for sepsis control is rarely necessary especially in those patients who already have a diverting stoma at the time of the leak. A nonoperative management is usually preferred in these patients. There are several treatment options, also for patients without a stoma who do not require a reoperation for a contained pelvic leak, including recently developed endoscopic procedures, such as clip placement or endoluminal vacuum-assisted therapy. More conservative treatments could be an option in patients who are clinically stable or in presence of a small defect.

Therapeutic strategies in Crohn's disease in an emergency surgical setting.

Crohn's disease (CD) remains a chronic, incurable disorder that presents unique challenges to the surgeon. Multiple factors must be considered to allow development of an appropriate treatment plan. Medical therapy often precedes or complements the surgical management. The indications for operative management of CD include acute and chronic disease complications and failed medical therapy. Elective surgery comes into play when patients are refractory to medical treatment if they have an obstructive phenotype. Toxic colitis, acute obstruction, perforation, acute abscess, or massive hemorrhage represent indications for emergency surgery. These patients are generally in critical conditions and present with intra-abdominal sepsis and a preoperative status of immunosuppression and malnutrition that exposes them to a higher risk of complications and mortality. A multidisciplinary team including surgeons, gastroenterologists, radiologists, nutritional support services, and enterostomal therapists are required for optimal patient care and decision making. Management of each emergency should be individualized based on patient age, disease type and duration, and patient goals of care. Moreover, the recurrent nature of disease mandates that we continue searching for innovative medical therapies and operative techniques that reduce the need to repeat surgical operations. In this review, we aimed to discuss the acute complications of CD and their treatment.

Anastomotic leakage in colorectal cancer surgery.

The safety of colorectal surgery for oncological disease is steadily improving, but anastomotic leakage is still the most feared and devastating complication from both a surgical and oncological point of view. Anastomotic leakage affects the outcome of the surgery, increases the times and costs of hospitalization, and worsens the prognosis in terms of short- and long-term outcomes. Anastomotic leakage has a wide range of clinical features ranging from radiological only finding to peritonitis and sepsis with multi-organ failure. C-reactive protein and procalcitonin have been identified as early predictors of anastomotic leakage starting from postoperative day 2-3, but abdominal-pelvic computed tomography scan is still the gold standard for the diagnosis. Several treatments can be adopted for anastomotic leakage. However, there is not a universally accepted flowchart for the management, which should be individualized based on patient's general condition, anastomotic defect size and location, indication for primary resection and presence of the proximal stoma. Non-operative management is usually preferred in patients who underwent proximal faecal diversion at the initial operation. Laparoscopy can be attempted after minimal invasive surgery and can reduce surgical stress in patients allowing a definitive treatment. Reoperation for sepsis control is rarely necessary in those patients who already have a diverting stoma at the time of the leak, especially in extraperitoneal anastomoses. In patients without a stoma who do not require abdominal reoperation for a contained pelvic leak, there are several treatment options, including laparoscopic diverting ileostomy combined with trans-anal anastomotic tube drainage, percutaneous drainage or recently developed endoscopic procedures, such as stent or clip placement or endoluminal vacuum-assisted therapy. We describe the current approaches to treat this complication, as well as the clinical tests necessary to diagnose and provide an effective therapy.

Anastomotic leakage in rectal cancer surgery: Retrospective analysis of risk factors.

BACKGROUND: Anastomotic leakage (AL) after restorative surgery for rectal cancer (RC) is associated with significant morbidity and mortality. AIM: To ascertain the risk factors by examining cases of AL in rectal surgery in this retrospective cohort study. METHODS: To identify risk factors for AL, a review of 583 patients who underwent rectal resection with a double-stapling colorectal anastomosis between January 2007 and January 2022 was performed. Clinical, demographic and operative features, intraoperative outcomes and oncological characteristics were evaluated. RESULTS: The incidence of AL was 10.4%, with a mean time interval of 6.2 ± 2.1 d. Overall mortality was 0.8%. Mortality was higher in patients with AL (4.9%) than in patients without leak (0.4%, P = 0.009). Poor bowel preparation, blood transfusion, median age, prognostic nutritional index < 40 points, tumor diameter and intraoperative blood loss were identified as risk factors for AL. Location of anastomosis, number of stapler cartridges used to divide the rectum, diameter of circular stapler, level of vascular section, T and N status and stage of disease were also correlated to AL in our patients. The diverting ileostomy did not reduce the leak rate, while the use of the transanastomic tube significantly did. CONCLUSION: Clinical, surgical and pathological factors are associated with an increased risk of AL. It adversely affects the morbidity and mortality of RC patients.

Management of the kidney transplant patient with Cancer: Report from a Multidisciplinary Consensus Conference.

INTRODUCTION: Cancer is the second most common cause of mortality and morbidity in Kidney Transplant Recipients (KTRs). Immunosuppression can influence the efficacy of cancer treatment and modification of the immunosuppressive regimen may restore anti-neoplastic immune responses improving oncologic prognosis. However, patients and transplant physicians are usually reluctant to modify immunosuppression, fearing rejection and potential graft loss. Due to the lack of extensive and recognised data supporting how to manage the immunosuppressive therapy in KTRs, in the context of immunotherapy, chemotherapy, radiotherapy and loco-regional treatments, a Consensus Conference was organised under the auspices of the European Society of Organ Transplantation and the Italian Society of Organ Transplantation. The conference involved a multidisciplinary group of transplant experts in the field across Europe. METHODS: The overall process included a) the formulation of 12 specific questions based on the PICO methodology, b) systematic literature review and summary for experts for each question, c) a two-day conference celebration and the collection of experts' agreements. The conference was articulated in three sessions: "Immunosuppressive therapy and immunotherapy", "Systemic therapy", "Integrated Therapy", while the final experts' agreement was collected with a televoting procedure and defined according to the majority criterion. RESULTS: Twenty-six European experts attended the conference and expressed their vote. A total of 14 statements were finally elaborated and voted. Strong agreement was found for ten statements, moderate agreement for two, moderate disagreement for one and uncertainty for the last one. CONCLUSIONS: The consensus statements provide guidance to transplant physicians caring for kidney transplant recipients with cancer and indicate key aspects that need to be addressed by future clinical research.

Neoantigen-Specific Adoptive Cell Therapies for Cancer: Making T-Cell Products More Personal.

Mutation-derived neoantigens are taking central stage as a determinant in eliciting effective antitumor immune responses following adoptive T-cell therapies. These mutations are patient-specific, and their targeting calls for highly personalized pipelines. The promising clinical outcomes of tumor-infiltrating lymphocyte (TIL) therapy have spurred interest in generating T-cell infusion products that have been selectively enriched in neoantigen (or autologous tumor) reactivity. The implementation of an isolation step, prior to T-cell in vitro expansion and reinfusion, may provide a way to improve the overall response rates achieved to date by adoptive T-cell therapies in metastatic cancer patients. Here we provide an overview of the main technologies [i.e., peptide major histocompatibility complex (pMHC) multimers, cytokine capture, and activation markers] to enrich infiltrating or circulating T-cells in predefined neoantigen specificities (or tumor reactivity). The unique technical and regulatory challenges faced by such highly specialized and patient-specific manufacturing T-cell platforms are also discussed.

A Self-Calibrating IoT Portable Electrochemical Immunosensor for Serum Human Epididymis Protein 4 as a Tumor Biomarker for Ovarian Cancer.

Nowadays, analytical techniques are moving towards the development of smart biosensing strategies for the point-of-care accurate screening of disease biomarkers, such as human epididymis protein 4 (HE4), a recently discovered serum marker for early ovarian cancer diagnosis. In this context, the present work represents the first implementation of a competitive enzyme-labelled magneto-immunoassay exploiting a homemade IoT Wi-Fi cloud-based portable potentiostat for differential pulse voltammetry readout. The electrochemical device was specifically designed to be capable of autonomous calibration and data processing, switching between calibration, and measurement modes: in particular, firstly, a baseline estimation algorithm is applied for correct peak computation, then calibration function is built by interpolating data with a four-parameter logistic function. The calibration function parameters are stored on the cloud for inverse prediction to determine the concentration of unknown samples. Interpolation function calibration and concentration evaluation are performed directly on-board, thus reducing the power consumption. The analytical device was validated in human serum, demonstrating good sensing performance for analysis of HE4 with detection and quantitation limits in human serum of 3.5 and 29.2 pM, respectively, reaching the sensitivity that is required for diagnostic purposes, with high potential for applications as portable and smart diagnostic tool for point-of-care testing.

Peptide Super-Agonist Enhances T-Cell Responses to Melanoma.

Recent immunotherapeutic approaches using adoptive cell therapy, or checkpoint blockade, have demonstrated the powerful anti-cancer potential of CD8 cytotoxic T-lymphocytes (CTL). While these approaches have shown great promise, they are only effective in some patients with some cancers. The potential power, and relative ease, of therapeutic vaccination against tumour associated antigens (TAA) present in different cancers has been a long sought-after approach for harnessing the discriminating sensitivity of CTL to treat cancer and has seen recent renewed interest following cancer vaccination successes using unique tumour neoantigens. Unfortunately, results with TAA-targeted "universal" cancer vaccines (UCV) have been largely disappointing. Infectious disease models have demonstrated that T-cell clonotypes that recognise the same antigen should not be viewed as being equally effective. Extrapolation of this notion to UCV would suggest that the quality of response in terms of the T-cell receptor (TCR) clonotypes induced might be more important than the quantity of the response. Unfortunately, there is little opportunity to assess the effectiveness of individual T-cell clonotypes in vivo. Here, we identified effective, persistent T-cell clonotypes in an HLA A2+ patient following successful tumour infiltrating lymphocyte (TIL) therapy. One such T-cell clone was used to generate super-agonist altered peptide ligands (APLs). Further refinement produced an APL that was capable of inducing T-cells in greater magnitude, and with improved effectiveness, from the blood of all 14 healthy donors tested. Importantly, this APL also induced T-cells from melanoma patient blood that exhibited superior recognition of the patient's own tumour compared to those induced by the natural antigen sequence. These results suggest that use of APL to skew the clonotypic quality of T-cells induced by cancer vaccination could provide a promising avenue in the hunt for the UCV "magic bullet."

Prothrombotic activity of cytokine-activated endothelial cells and shear-activated platelets in the setting of ventricular assist device support.

BACKGROUND: We systematically analyzed the synergistic effect of: (i) cytokine-mediated inflammatory activation of endothelial cells (ECs) with and (ii) shear-mediated platelet activation (SMPA) as a potential contributory mechanism to intraventricular thrombus formation in the setting of left ventricular assist device (LVAD) support. METHODS: Intact and shear-activated human platelets were exposed to non-activated and cytokine-activated ECs. To modulate the level of LVAD-related shear activation, platelets were exposed to shear stress patterns of varying magnitude (30, 50, and 70 dynes/cm2, 10 minutes) via a hemodynamic shearing device. ECs were activated via exposure to inflammatory tumor necrosis factor-α (TNF-α 10 and 100 ng/ml, 24 hours), consistent with inflammatory activation recorded in patients on LVAD circulatory support. RESULTS: Adhesivity of shear-activated platelets to ECs was significantly higher than that of intact/unactivated platelets, regardless of the initial activation level (70 dynes/cm2 shear-activated platelets vs intact platelets: +80%, p < 0.001). Importantly, inflammatory activation of ECs amplified platelet prothrombinase activity progressively with increasing shear stress magnitude and TNF-α concentration: thrombin generation of 70 dynes/cm2 shear-activated platelets was 2.6-fold higher after exposure and adhesion to 100 ng/ml TNF-α‒activated ECs (p < 0.0001). CONCLUSIONS: We demonstrated synergistic effect of SMPA and cytokine-mediated EC inflammatory activation to enhance EC‒platelet adhesion and platelet prothrombotic function. These mechanisms may contribute to intraventricular thrombosis in the setting of mechanical circulatory support.

In Silico and Structural Analyses Demonstrate That Intrinsic Protein Motions Guide T Cell Receptor Complementarity Determining Region Loop Flexibility.

T-cell immunity is controlled by T cell receptor (TCR) binding to peptide major histocompatibility complexes (pMHCs). The nature of the interaction between these two proteins has been the subject of many investigations because of its central role in immunity against pathogens, cancer, in autoimmunity, and during organ transplant rejection. Crystal structures comparing unbound and pMHC-bound TCRs have revealed flexibility at the interaction interface, particularly from the perspective of the TCR. However, crystal structures represent only a snapshot of protein conformation that could be influenced through biologically irrelevant crystal lattice contacts and other factors. Here, we solved the structures of three unbound TCRs from multiple crystals. Superposition of identical TCR structures from different crystals revealed some conformation differences of up to 5 Å in individual complementarity determining region (CDR) loops that are similar to those that have previously been attributed to antigen engagement. We then used a combination of rigidity analysis and simulations of protein motion to reveal the theoretical potential of TCR CDR loop flexibility in unbound state. These simulations of protein motion support the notion that crystal structures may only offer an artifactual indication of TCR flexibility, influenced by crystallization conditions and crystal packing that is inconsistent with the theoretical potential of intrinsic TCR motions.

Sensitive and frequent identification of high avidity neo-epitope specific CD8 + T cells in immunotherapy-naive ovarian cancer.

Immunotherapy directed against private tumor neo-antigens derived from non-synonymous somatic mutations is a promising strategy of personalized cancer immunotherapy. However, feasibility in low mutational load tumor types remains unknown. Comprehensive and deep analysis of circulating and tumor-infiltrating lymphocytes (TILs) for neo-epitope specific CD8+ T cells has allowed prompt identification of oligoclonal and polyfunctional such cells from most immunotherapy-naive patients with advanced epithelial ovarian cancer studied. Neo-epitope recognition is discordant between circulating T cells and TILs, and is more likely to be found among TILs, which display higher functional avidity and unique TCRs with higher predicted affinity than their blood counterparts. Our results imply that identification of neo-epitope specific CD8+ T cells is achievable even in tumors with relatively low number of somatic mutations, and neo-epitope validation in TILs extends opportunities for mutanome-based personalized immunotherapies to such tumors.