Meta-analysis of tumor- and T cell-intrinsic mechanisms of sensitization to checkpoint inhibition.

Checkpoint inhibitors (CPIs) augment adaptive immunity. Systematic pan-tumor analyses may reveal the relative importance of tumor-cell-intrinsic and microenvironmental features underpinning CPI sensitization. Here, we collated whole-exome and transcriptomic data for >1,000 CPI-treated patients across seven tumor types, utilizing standardized bioinformatics workflows and clinical outcome criteria to validate multivariable predictors of CPI sensitization. Clonal tumor mutation burden (TMB) was the strongest predictor of CPI response, followed by total TMB and CXCL9 expression. Subclonal TMB, somatic copy alteration burden, and histocompatibility leukocyte antigen (HLA) evolutionary divergence failed to attain pan-cancer significance. Dinucleotide variants were identified as a source of immunogenic epitopes associated with radical amino acid substitutions and enhanced peptide hydrophobicity/immunogenicity. Copy-number analysis revealed two additional determinants of CPI outcome supported by prior functional evidence: 9q34 (TRAF2) loss associated with response and CCND1 amplification associated with resistance. Finally, single-cell RNA sequencing (RNA-seq) of clonal neoantigen-reactive CD8 tumor-infiltrating lymphocytes (TILs), combined with bulk RNA-seq analysis of CPI-responding tumors, identified CCR5 and CXCL13 as T-cell-intrinsic markers of CPI sensitivity.

To study the survival analysis and recurrence pattern in women treated for breast cancer: Retrospective study, a tertiary cancer center experience from Sub-Himalayan Region of India.

BACKGROUND: Breast cancer is a leading health concern in India, comprising 25% of female cancers with significant mortality. This study was conducted at the Cancer Research Institute in the Northern Sub-Himalayan region of India from 2016 to 2021, evaluated 674 breast cancer cases to analyze factors that influence recurrence. METHODOLOGY: Retrospective clinical audit assessing patients' survival outcomes using Kaplan-Meier curves and Cox proportional hazard regression. Factors including age, molecular subtype, TNM staging, and treatment modalities were evaluated. RESULTS: Notable findings include a high occurrence of breast cancer in young patients (24.48% ≤ 40 years) and varying recurrence rates among molecular subtypes with human epidermal growth factor receptor 2 neu-enriched (25.24%) and triple­negative breast cancer (22.58%) being the most common. Advanced T and N stages, neoadjuvant chemotherapy, and the number of nodes dissected showed significant associations with higher recurrence rates. CONCLUSION: This study sheds light on survival and recurrence patterns in Northern Sub-Himalayan breast cancer patients, emphasizing the need for tailored treatment strategies, comprehensive follow-up care, with improved understanding of regional outcomes. These findings contribute valuable insights for optimizing patient care and improving survival rates in this region.

Oncobiology and treatment of breast cancer in young women.

Female breast cancer emerged as the leading cancer type in terms of incidence globally in 2020. Although mortality due to breast cancer has improved during the past three decades in many countries, this trend has reversed in women less than 40 years since the past decade. From the biological standpoint, there is consensus among experts regarding the clinically relevant definition of breast cancer in young women (BCYW), with an age cut-off of 40 years. The idea that breast cancer is an aging disease has apparently broken in the case of BCYW due to the young onset and an overall poor outcome of BCYW patients. In general, younger patients exhibit a worse prognosis than older pre- and postmenopausal patients due to the aggressive nature of cancer subtypes, a high percentage of cases with advanced stages at diagnosis, and a high risk of relapse and death in younger patients. Because of clinically and biologically unique features of BCYW, it is suspected to represent a distinct biologic entity. It is unclear why BCYW is more aggressive and has an inferior prognosis with factors that contribute to increased incidence. However, unique developmental features, adiposity and immune components of the mammary gland, hormonal interplay and crosstalk with growth factors, and a host of intrinsic and extrinsic risk factors and cellular regulatory interactions are considered to be the major contributing factors. In the present article, we discuss the status of BCYW oncobiology, therapeutic interventions and considerations, current limitations in fully understanding the basis and underlying cause(s) of BCYW, understudied areas of BCYW research, and postulated advances in the coming years for the field.

Walking Exercise Therapy Effects on Lower Extremity Skeletal Muscle in Peripheral Artery Disease.

Walking exercise is the most effective noninvasive therapy that improves walking ability in peripheral artery disease (PAD). Biologic mechanisms by which exercise improves walking in PAD are unclear. This review summarizes evidence regarding effects of walking exercise on lower extremity skeletal muscle in PAD. In older people without PAD, aerobic exercise improves mitochondrial activity, muscle mass, capillary density, and insulin sensitivity in skeletal muscle. However, walking exercise increases lower extremity ischemia in people with PAD, and therefore, mechanisms by which this exercise improves walking may differ between people with and without PAD. Compared with people without PAD, gastrocnemius muscle in people with PAD has greater mitochondrial impairment, increased reactive oxygen species, and increased fibrosis. In multiple small trials, walking exercise therapy did not consistently improve mitochondrial activity in people with PAD. In one 12-week randomized trial of people with PAD randomized to supervised exercise or control, supervised treadmill exercise increased treadmill walking time from 9.3 to 15.1 minutes, but simultaneously increased the proportion of angular muscle fibers, consistent with muscle denervation (from 7.6% to 15.6%), while angular myofibers did not change in the control group (from 9.1% to 9.1%). These findings suggest an adaptive response to exercise in PAD that includes denervation and reinnervation, an adaptive process observed in skeletal muscle of people without PAD during aging. Small studies have not shown significant effects of exercise on increased capillary density in lower extremity skeletal muscle of participants with PAD, and there are no data showing that exercise improves microcirculatory delivery of oxygen and nutrients in patients with PAD. However, the effects of supervised exercise on increased plasma nitrite abundance after a treadmill walking test in people with PAD may be associated with improved lower extremity skeletal muscle perfusion and may contribute to improved walking performance in response to exercise in people with PAD. Randomized trials with serial, comprehensive measures of muscle biology, and physiology are needed to clarify mechanisms by which walking exercise interventions improve mobility in PAD.

Herpesvirus immunology in solid organ transplant recipients - liver transplant study (HISTORY): a retrospective and prospective observational cohort study.

BACKGROUND: Life-long immunosuppressive treatment after liver transplantation (LT) prevents graft rejection but predisposes the LT recipient to infections. Herpesvirus infections are associated with morbidity and mortality among LT recipients. Among those, especially cytomegalovirus (CMV) and varicella-zoster virus (VZV) pose challenges after LT. The aim of this study is to provide an in-depth characterization of the cellular immune response against CMV and VZV infections in LT recipients and identify potential risk factors for infection. METHODS: The Herpesvirus Infections in Solid Organ Transplant Recipients - Liver Transplant Study (HISTORY) consists of an epidemiological and immunological substudy. The epidemiological substudy is a retrospective observational cohort study that includes all patients who underwent LT in Denmark between 2010 and 2023 (N ≈ 500). Using data from nationwide hospital records and national health registries, the incidence of and clinical risk factors for CMV and VZV infections will be determined. The immunological substudy is an explorative prospective observational cohort study including patients enlisted for LT in Denmark during a 1.5-year period (N > 80). Participants will be followed with scheduled blood samples until 12 months after LT. CMV- and VZV-derived peptides will be predicted for their likelihood to be presented in participants based on their HLA type. Peptide-MHC complexes (pMHC) will be produced to isolate CMV- and VZV-specific T cells from peripheral blood mononuclear cells before and after CMV and VZV infection. Their frequency, T cell receptor sequences, and phenotypic characteristics will be examined, and in a subset of participants, CMV- and VZV-specific T cells will be expanded ex vivo. DISCUSSION: This study will provide novel insight into T cell immunity required for viral control of CMV and VZV and has the potential to develop a prediction model to identify LT recipients at high risk for infection based on a combination of clinical and immunological data. Furthermore, this study has the potential to provide proof-of-concept for adoptive T cell therapy against CMV and VZV. Combined, this study has the potential to reduce the burden and consequence of CMV and VZV infections and improve health and survival in LT recipients. TRIAL REGISTRATION: ClinicalTrials.gov (NCT05532540), registered 8 September 2022.

Herpes Virus Infections in Kidney Transplant Patients (HINT) - a prospective observational cohort study.

BACKGROUND: Kidney transplant recipients receive maintenance immunosuppressive therapy to avoid allograft rejection resulting in increased risk of infections and infection-related morbidity and mortality. Approximately 98% of adults are infected with varicella zoster virus, which upon reactivation causes herpes zoster. The incidence of herpes zoster is higher in kidney transplant recipients than in immunocompetent individuals, and kidney transplant recipients are at increased risk of severe herpes zoster-associated disease. Vaccination with adjuvanted recombinant glycoprotein E subunit herpes zoster vaccine (RZV) prevents herpes zoster in older adults with excellent efficacy (90%), and vaccination of kidney transplant candidates is recommended in Danish and international guidelines. However, the robustness and duration of immune responses after RZV vaccination, as well as the optimal timing of vaccination in relation to transplantation remain unanswered questions. Thus, the aim of this study is to characterize the immune response to RZV vaccination in kidney transplant candidates and recipients at different timepoints before and after transplantation. METHODS: The Herpes Virus Infections in Kidney Transplant Patients (HINT) study is a prospective observational cohort study. The study will include kidney transplant candidates on the waiting list for transplantation (n = 375) and kidney transplant recipients transplanted since January 1, 2019 (n = 500) from all Danish kidney transplant centers who are offered a RZV vaccine as routine care. Participants are followed with repeated blood sampling until 12 months after inclusion. In the case of transplantation or herpes zoster disease, additional blood samples will be collected until 12 months after transplantation. The immune response will be characterized by immunophenotyping and functional characterization of varicella zoster virus-specific T cells, by detection of anti-glycoprotein E antibodies, and by measuring cytokine profiles. DISCUSSION: The study will provide new knowledge on the immune response to RZV vaccination in kidney transplant candidates and recipients and the robustness and duration of the response, potentially enhancing preventive strategies against herpes zoster in a population at increased risk. TRIAL REGISTRATION: ClinicalTrials.gov (NCT05604911).

Impact of Enhanced Recovery Protocols on Short-Term Outcomes in Esophagectomy: A Retrospective Cohort Study from Cancer Research Institute, Uttarakhand, India.

OBJECTIVE: Surgery for esophageal cancer is associated with high mortality and morbidity, especially in low and middle-income countries. The recent enhanced recovery after surgery guidelines for esophagectomy (2018) which attempt to reduce complications and length of stay (LOS) have rarely been validated in these settings. This study aimed to analyse the effect of this protocol on short-term outcomes in our subset of patients. METHODS: A retrospective review was conducted to investigate the outcomes of enhanced recovery protocol (ERP) compared to standard pre-protocol care (PP) in patients who underwent esophagectomy for cancer (31 in ERP vs 61 in PP group) at Cancer Research Institute, Uttarakhand, India. The main outcomes measured were 30-day mortality, morbidity and LOS. Risk assessment was stratified as per Physiological and Operative Severity Score for the Enumeration of Mortality and Morbidity (POSSUM) systems while complications were classified as per the Clavien-Dindo scale. RESULTS: Preoperative clinical characteristics were similar between groups. Though the predicted POSSUM mortality and morbidity were significantly higher in the ERP group (p = 0.007), 30-day morbidity (19.35% vs 42.62%, p = 0.027) as well as median LOS (12 vs 15 days, p < 0.001) was significantly lower in ERP group. The PP group reported 4 deaths within 30 days as compared to none in the ERP group (p = 0.296). Furthermore, the ERP group reported lower occurrence of pulmonary complications (6.4%vs24.6%,p = 0.046), hemodynamic instability (0%vs14.75%,p = 0.026) as well as need for prolonged postoperative ventilation (> 24 h; 0% vs 11.48%, p = 0.004). Both minor and major complications as assessed by the Clavien-Dindo scale were lower in the group ERP though these differences were not statistically significant (0.059). CONCLUSIONS: Implementation of ERP improved short-term outcomes; hence can be strongly recommended in patients undergoing esophagectomy.

Let the Kid Speak: Dynamics of Triadic Medical Interactions Involving Pediatric Patients.

Communication in healthcare represents the complex interplay between multiple individual and contextual factors unfolding over the course of the medical encounter. Despite significant improvements in patient-centered care delivery, studies of health communication typically focus exclusively on clinical interactions between adult patients and their clinicians. Much less is known about non-dyadic interactions, such as pediatric triads involving a child patient and accompanying parent. Understanding the dynamics of triadic pediatric healthcare communication is the first step toward evaluating and ultimately optimizing these healthcare interactions. Thus, we undertook a mixed-method analysis of 28 audio-recorded triadic medical interactions between healthcare providers, pediatric asthma and allergy patients, and their parents to explore the prevalence of various features of these interactions. Our findings point to mechanisms through which healthcare providers and parents may facilitate or hinder children's involvement in their own asthma and allergy care, including interruptions, unclarified technical medical language, the flow of information exchange, and the formation of dyadic conversational partnerships (coalitions) between providers and parents. Our analyses further reveal that children's participation during their medical visits was minimal (13% of the interaction). Providers in our sample elicited input directly from pediatric patients more often than from parents, though the difference was small. Taken together, these findings provide a foundation on which to develop training and communication interventions to ensure that children have a voice in their medical care.

Pattern of nutritional status in node-negative versus node-positive head and neck cancer patients undergoing treatment: a prospective cohort study.

PURPOSE: The aim of this study was to study the nutritional profile of node-negative and node-positive patients undergoing treatment for head and neck squamous cell cancer (HNSCC). METHODS: This prospective cohort study was conducted between 2018 and 2020. Patients diagnosed with HNSCC, planned for treatment, were enrolled after written informed consent. In node-negative (N0) and node-positive (N +) cohorts of patients, nutritional status was determined using anthropometric measures and Subjective Global Assessment (SGA) scale pre-treatment, and during and after treatment. Statistical analysis was performed using SPSS version 22. Data was analyzed using parametric and non-parametric tests, and p value of 0.05 was considered significant. RESULTS: In total, 161 patients were analyzed, 73 N0 and 88 N + cohorts. Pre-treatment, 9.6 to 20.4% patients in N0 and 23.9 to 32.8% patients in N + cohorts were malnourished. Incidence of malnutrition at completion of treatment was 40.8 to 52.5% overall, 20.5 to 41.1% N0, and 39.5 to 62.8% N + . Mean reduction in weight (11.1% ± 7.82 vs 6.26% ± 8.3, p = 0.000), mean reduction in BMI (2.57 ± 1.87 vs 1.29 ± 1.62, p = 0.000), median reduction in MUAC (2 cm vs 1 cm, p = 0.000), and median increase in SGA score (13 vs 6, p = 0.000) were higher in multi-modality as compared to those in a single-modality treatment. Similar findings were noted in N0 and N + cohorts. CONCLUSION: As compared to N0, N + patients had higher burden of malnutrition at diagnosis, and more worsening of nutritional parameters during treatment. More decline in nutritional status was seen in patients receiving multi-modality as compared to single-modality treatment.

Skin substitutes with noncultured autologous skin cell suspension heal porcine full-thickness wounds in a one-stage procedure.

Clinical application of skin substitute is typically a two-stage procedure with application of skin substitute matrix to the wound followed by engraftment of a split-thickness skin graft (STSG). This two-stage procedure requires multiple interventions, increasing the time until the wound is epithelialised. In this study, the feasibility of a one-stage procedure by combining bioengineered collagen-chondroitin-6-sulfate (DS1) or decellularised fetal bovine skin substitute (DS2) with autologous skin cell suspension (ASCS) in a porcine full-thickness wound healing model was evaluated. Twelve full-thickness excisional wounds on the backs of pigs received one of six different treatments: empty; ASCS; DS1 with or without ASCS; DS2 with or without ASCS. The ASCS was prepared using a point-of-care device and was seeded onto the bottom side of DS1, DS2, and empty wounds at 80 000 cells/cm2 . Wound measurements and photographs were taken on days 0, 9, 14, 21, 28, 35, and 42 post-wounding. Histological analysis was performed on samples obtained on days 9, 14, 28, and 42. Wounds in the empty group or with ASCS alone showed increased wound contraction, fibrosis, and myofibroblast density compared with other treatment groups. The addition of ASCS to DS1 or DS2 resulted in a marked increase in re-epithelialisation of wounds at 14 days, from 15 ± 11% to 71 ± 20% (DS1 vs DS1 + ASCS) or 28 ± 14% to 77 ± 26 (DS2 vs DS2 + ASCS) despite different mechanisms of tissue regeneration employed by the DS used. These results suggest that this approach may be a viable one-stage treatment in clinical practice.

Skeletal Muscle Pathology in Peripheral Artery Disease: A Brief Review.

This brief review summarizes current evidence regarding lower extremity peripheral artery disease (PAD) and lower extremity skeletal muscle pathology. Lower extremity ischemia is associated with reduced calf skeletal muscle area and increased calf muscle fat infiltration and fibrosis on computed tomography or magnetic resonance imaging. Even within the same individual, the leg with more severe ischemia has more adverse calf muscle characteristics than the leg with less severe ischemia. More adverse computed tomography-measured calf muscle characteristics, such as reduced calf muscle density, are associated with higher rates of mobility loss in people with PAD. Calf muscle in people with PAD may also have reduced mitochondrial activity compared with those without PAD, although evidence is inconsistent. Muscle biopsy document increased oxidative stress in PAD. Reduced calf muscle perfusion, impaired mitochondrial activity, and smaller myofibers are associated with greater walking impairment in PAD. Preliminary evidence suggests that calf muscle pathology in PAD may be reversible. In a small uncontrolled trial, revascularization improved both the ankle-brachial index and mitochondrial activity, measured by calf muscle phosphocreatine recovery time. A pilot clinical trial showed that cocoa flavanols increased measures of myofiber health, mitochondrial activity, and capillary density while simultaneously improving 6-minute walk distance in PAD. Calf muscle pathological changes are associated with impaired walking performance in people with PAD, and interventions that both increase calf perfusion and improve calf muscle health are promising therapies to improve walking performance in PAD.

Effectiveness of Yoga on Quality of Life of Breast Cancer Patients Undergoing Chemotherapy: A Randomized Clinical Controlled Study.

BACKGROUND: Cancer of breast is most common cancer among women in India and vast majority of countries worldwide. While undergoing chemotherapy for carcinoma management, women encounter side effects, which affects their quality of life (QOL). A randomized controlled study with quantitative research approach and time series design was conducted, to study the effectiveness of yoga on QOL of breast cancer patients undergoing chemotherapy. METHODOLOGY: One hundred breast cancer patients scheduled for 3-weekly, day-care adjuvant chemotherapy (CEF regimen) were enrolled with consecutive sampling technique, into control (n = 52) and experiment (n = 48) groups, by concealed randomization following written informed consent. Baseline data on QOL were collected before first-cycle chemotherapy using the European Organization for Research and Treatment of Cancer QLQ C30. Patients in the experimental group were taught diaphragmatic breathing techniques, systematic relaxation, and alternate nostril breathing, and Joints and Glands neck and shoulder exercises were instructed to practice twice daily at home. They were supervised in practicing these techniques while they received second, third, fourth, fifth, and sixth cycles of chemotherapy in the day-care facility. Participants in the control group received only routine care. All participants received standard post chemotherapy prescription. Data on QOL were collected from all patients during the second, third, fourth, fifth, and sixth cycles of chemotherapy. RESULTS: The analysis revealed that at the baseline (first chemotherapy cycle), breast cancer patients in control and experimental groups were homogeneous in terms of their sociodemographic and clinical variables and QOL score. Yoga practices were effective in improving the QOL of breast cancer patients in the experimental group in the areas of global health status, physical function, role function, and emotional function and decreasing the symptoms of fatigue, insomnia, loss of appetite, and constipation, during the period of chemotherapy. CONCLUSION: Yoga practices comprising of relaxation techniques reduce many side effects and improve the QOL of women undergoing chemotherapy for breast cancer.

miR-145 supports cancer cell survival and shows association with DDR genes, methylation pattern, and epithelial to mesenchymal transition.

BACKGROUND: Despite several reports describing the dual role of miR-145 as an oncogene and a tumor suppressor in cancer, not much has been resolved and understood. METHOD: In this study, the potential targets of miR-145 were identified bio-informatically using different target prediction tools. The identified target genes were validated in vitro by dual luciferase assay. Wound healing and soft agar colony assay assessed cell proliferation and migration. miR-145 expression level was measured quantitatively by RT-PCR at different stages of breast tumor. Western blot was used to verify the role of miR-145 in EMT transition using key marker proteins. RESULT: Wound healing and soft agar colony assays, using miR-145 over-expressing stably transfected MCF7 cells, unraveled its role as a pro-proliferation candidate in cancerous cells. The association between miR-145 over-expression and differential methylation patterns in representative target genes (DR5, BCL2, TP53, RNF8, TIP60, CHK2, and DCR2) supported the inference drawn. These in vitro observations were validated in a representative set of nodal positive tumors of stage 3 and 4 depicting higher miR-145 expression as compared to early stages. Further, the role of miR-145 in epithelial-mesenchymal (EMT) transition found support through the observation of two key markers, Vimentin and ALDL, where a positive correlation with Vimentin protein and a negative correlation with ALDL mRNA expression were observed. CONCLUSION: Our results demonstrate miR-145 as a pro-cancerous candidate, evident from the phenotypes of aggressive cellular proliferation, epithelial to mesenchymal transition, hypermethylation of CpG sites in DDR and apoptotic genes and upregulation of miR-145 in later stages of tumor tissues.

Cancer Pain, Anxiety, and Depression in Admitted Patients at a Tertiary Care Hospital: A Prospective Observational Study.

CONTEXT: Pain is the most common symptom in admitted cancer patients. The association between the severity of cancer pain and distress symptoms such as depression and anxiety is a subject of research. AIMS: The aim is to study the prevalence of pain, anxiety, and depression in admitted cancer patients and determine the association between pain and anxiety and depression at a tertiary cancer care institute. SETTINGS AND DESIGN: This was prospective observational study. SUBJECTS AND METHODS: We enrolled 393 cancer inpatients prospectively after written informed consent. Their disease details, presence, severity, and character of pain were recorded. Numerical Pain Scale was used for pain scores, self-reporting Hospital Anxiety and Depression Scale for anxiety and depression. STATISTICAL ANALYSIS USED: Normal data were analyzed with parametric, nonnormal with nonparametric methods, and categorical with the Chi-square test. RESULTS: The prevalence of moderate-to-severe pain was 41.5%, anxiety 20.3%, and depression 24.8%. Proportion of patients with anxiety and depression was 9.2% and 17.7% in patients with no pain; about 32.8% and 36.7% with severe pain, respectively (P < 0.000). In patients with no depression 6% had anxiety; with depression 44.9% had anxiety (P < 0.000). Odd's ratio to have anxiety and depression was 4.44 (95% confidence interval [CI] 2.0318-9.7024) and 2.92 (95% CI 1.5739-5.4186), respectively, in patients with pain as compared to no pain (P < 0.00). There was a positive correlation between pain, anxiety, and depression scores. CONCLUSIONS: There is strong association between the presence and severity of pain and distress symptoms such as anxiety and depression in admitted cancer patients.

Pyruvate kinase M knockdown-induced signaling via AMP-activated protein kinase promotes mitochondrial biogenesis, autophagy, and cancer cell survival.

Preferential expression of the low-activity (dimeric) M2 isoform of pyruvate kinase (PK) over its constitutively active splice variant M1 isoform is considered critical for aerobic glycolysis in cancer cells. However, our results reported here indicate co-expression of PKM1 and PKM2 and their possible physical interaction in cancer cells. We show that knockdown of either PKM1 or PKM2 differentially affects net PK activity, viability, and cellular ATP levels of the lung carcinoma cell lines H1299 and A549. The stable knockdown of PK isoforms in A549 cells significantly reduced the cellular ATP level, whereas in H1299 cells the level of ATP was unaltered. Interestingly, the PKM1/2 knockdown in H1299 cells activated AMP-activated protein kinase (AMPK) signaling and stimulated mitochondrial biogenesis and autophagy to maintain energy homeostasis. In contrast, knocking down either of the PKM isoforms in A549 cells lacking LKB1, a serine/threonine protein kinase upstream of AMPK, failed to activate AMPK and sustain energy homeostasis and resulted in apoptosis. Moreover, in a similar genetic background of silenced PKM1 or PKM2, the knocking down of AMPKα1/2 catalytic subunit in H1299 cells induced apoptosis. Our findings help explain why previous targeting of PKM2 in cancer cells to control tumor growth has not met with the expected success. We suggest that this lack of success is because of AMPK-mediated energy metabolism rewiring, protecting cancer cell viability. On the basis of our observations, we propose an alternative therapeutic strategy of silencing either of the PKM isoforms along with AMPK in tumors.

Dipeptides catalyze rapid peptide exchange on MHC class I molecules.

Peptide ligand selection by MHC class I molecules, which occurs by iterative optimization, is the centerpiece of immunodominance in antiviral and antitumor immune responses. For its understanding, the molecular mechanisms of peptide binding and dissociation by class I molecules must be elucidated. To this end, we have investigated dipeptides that bind to the F pocket of class I molecules. We find that they accelerate the dissociation of prebound peptides of both low and high affinity, suggesting a mechanism of action for the peptide-exchange chaperone tapasin. Peptide exchange on class I molecules also has practical uses in epitope discovery and T-cell monitoring.

Association of mitochondrial copy number variation and T16189C polymorphism with colorectal cancer in North Indian population.

Globally, colorectal cancer is the third most common type of cancer. Genetic instability leading to cancer development is one of the major causes for development of cancer. Alterations in mitochondrial genome, that is, mutations, single-nucleotide polymorphisms, and copy number variations are known to contribute in cancer development. The aim of our study was to investigate association of mitochondrial T16189C polymorphism and copy number variation with colorectal cancer in North Indian population. DNA isolated from peripheral blood of 126 colorectal cancer patients and 114 healthy North Indian subjects was analyzed for T16189C polymorphism and half of them for mitochondrial copy number variation. Genotyping was done using polymerase chain reaction-restriction fragment length polymorphism, and copy number variation was estimated using real-time polymerase chain reaction, numbers of mitochondrial copies and found to be significantly higher in colorectal cancer patients than healthy controls (88 (58-154), p = 0.001). In the regression analysis, increased mitochondrial copy number variation was associated with risk of colorectal cancer (odds ratio = 2.885, 95% confidence interval = 1.3-6.358). However, T16189C polymorphism was found to be significantly associated with the risk of rectal cancer (odds ratio = 5.213, p = 0.001) and non-significantly with colon cancer (odds ratio = 0.867, p = 0.791). Also, false-positive report probability analysis was done to validate the significant findings. Our results here indicate that mitochondrial copy number variation may be playing an important role in the development of colorectal cancer, and detection of mitochondrial copy number variation can be used as a biomarker for predicting the risk of colorectal cancer in North Indian subjects.

Peptide-independent stabilization of MHC class I molecules breaches cellular quality control.

The intracellular trafficking of major histocompatibility complex class I (MHC-I) proteins is directed by three quality control mechanisms that test for their structural integrity, which is correlated to the binding of high-affinity antigenic peptide ligands. To investigate which molecular features of MHC-I these quality control mechanisms detect, we have followed the hypothesis that suboptimally loaded MHC-I molecules are characterized by their conformational mobility in the F-pocket region of the peptide-binding site. We have created a novel variant of an MHC-I protein, K(b)-Y84C, in which two α-helices in this region are linked by a disulfide bond that mimics the conformational and dynamic effects of bound high-affinity peptide. K(b)-Y84C shows a remarkable increase in the binding affinity to its light chain, beta-2 microglobulin (ß2m), and bypasses all three cellular quality control steps. Our data demonstrate (1) that coupling between peptide and ß2m binding to the MHC-I heavy chain is mediated by conformational dynamics; (2) that the folded conformation of MHC-I, supported by ß2m, plays a decisive role in passing the ER-to-cell-surface transport quality controls; and (3) that ß2m association is also tested by the cell surface quality control that leads to MHC-I endocytosis.

Dipeptides promote folding and peptide binding of MHC class I molecules.

MHC class I molecules bind only those peptides with high affinity that conform to stringent length and sequence requirements. We have now investigated which peptides can aid the in vitro folding of class I molecules, and we find that the dipeptide glycyl-leucine efficiently supports the folding of HLA-A*02:01 and H-2K(b) into a peptide-receptive conformation that rapidly binds high-affinity peptides. Treatment of cells with glycyl-leucine induces accumulation of peptide-receptive H-2K(b) and HLA-A*02:01 at the surface of cells. Other dipeptides with a hydrophobic second amino acid show similar enhancement effects. Our data suggest that the dipeptides bind into the F pocket like the C-terminal amino acids of a high-affinity peptide.