Activation of Peripheral Blood Mononuclear Cells and Leptin Secretion: New Potential Role of Interleukin-2 and High Mobility Group Box (HMGB)1.

Activation of innate immunity and low-grade inflammation contributes to hyperglycemia and an onset of Type 2 Diabetes Mellitus (T2DM). Interleukin-2 (IL-2), leptin, High Mobility Group Box-1 (HMGB-1), and increased glucose concentrations are mediators of these processes also by modulating peripheral blood mononuclear cells (PBMCs) response. The aim of this study was to investigate if HMGB-1 and IL-2 turn on PBMCs and their leptin secretion. In isolated human PBMCs and their subpopulations from healthy individuals and naïve T2DM patients, leptin release, pro-inflammatory response and Toll-like Receptors (TLRs) activation was measured. After treatment with IL-2 and HMGB1, NK (Natural Killer) have the highest amount of leptin secretion, whilst NK-T have the maximal release in basal conditions. TLR4 (TAK242) and/or TLR2 (TLR2-IgA) inhibitors decreased leptin secretion after IL-2 and HMGB1 treatment. A further non-significant increase in leptin secretion was reported in PBMCs of naive T2DM patients in response to IL-2 and HMGB-1 stimulation. Finally, hyperglycemia or hyperinsulinemia might stimulate leptin secretion from PBMCs. The amount of leptin released from PBMCs after the different treatments was enough to stimulate the secretion of IL-1ß from monocytes. Targeting leptin sera levels and secretion from PBMCs could represent a new therapeutic strategy to counteract metabolic diseases such as T2DM.

CD16-158-valine chimeric receptor T cells overcome the resistance of KRAS-mutated colorectal carcinoma cells to cetuximab.

KRAS mutations hinder therapeutic efficacy of epidermal growth factor receptor (EGFR)-specific monoclonal antibodies cetuximab and panitumumab-based immunotherapy of EGFR+ cancers. Although cetuximab inhibits KRAS-mutated cancer cell growth in vitro by natural killer (NK) cell-mediated antibody-dependent cellular cytotoxicity (ADCC), KRAS-mutated colorectal carcinoma (CRC) cells escape NK cell immunosurveillance in vivo. To overcome this limitation, we used cetuximab and panitumumab to redirect Fcγ chimeric receptor (CR) T cells against KRAS-mutated HCT116 colorectal cancer (CRC) cells. We compared four polymorphic Fcγ-CR constructs including CD16158F -CR, CD16158V -CR, CD32131H -CR, and CD32131R -CR transduced into T cells by retroviral vectors. Percentages of transduced T cells expressing CD32131H -CR (83.5 ± 9.5) and CD32131R -CR (77.7 ± 13.2) were significantly higher than those expressing with CD16158F -CR (30.3 ± 10.2) and CD16158V -CR (51.7 ± 13.7) (p < 0.003). CD32131R -CR T cells specifically bound soluble cetuximab and panitumumab. However, only CD16158V -CR T cells released high levels of interferon gamma (IFNγ = 1,145.5 pg/ml ±16.5 pg/ml, p < 0.001) and tumor necrosis factor alpha (TNFα = 614 pg/ml ± 21 pg/ml, p < 0.001) upon incubation with cetuximab-opsonized HCT116 cells. Moreover, only CD16158V -CR T cells combined with cetuximab killed HCT116 cells and A549 KRAS-mutated cells in vitro. CD16158V -CR T cells also effectively controlled subcutaneous growth of HCT116 cells in CB17-SCID mice in vivo. Thus, CD16158V -CR T cells combined with cetuximab represent useful reagents to develop innovative EGFR+KRAS-mutated CRC immunotherapies.

In vitro elimination of epidermal growth factor receptor-overexpressing cancer cells by CD32A-chimeric receptor T cells in combination with cetuximab or panitumumab.

Cetuximab and panitumumab bind the human epidermal growth factor receptor (EGFR). Although the chimeric cetuximab (IgG1) triggers antibody-dependent-cellular-cytotoxicity (ADCC) of EGFR positive target cells, panitumumab (a human IgG2) does not. The inability of panitumumab to trigger ADCC reflects the poor binding affinity of human IgG2 Fc for the FcγRIII (CD16) on natural killer (NK) cells. However, both human IgG1 and IgG2 bind the FcγRII (CD32A) to a similar extent. Our study compares the ability of T cells, engineered with a novel low-affinity CD32A131R -chimeric receptor (CR), and those engineered with the low-affinity CD16158F -CR T cells, in eliminating EGFR positive epithelial cancer cells (ECCs) in combination with cetuximab or panitumumab. After T-cell transduction, the percentage of CD32A131R -CR T cells was 74 ± 10%, whereas the percentage of CD16158F -CR T cells was 46 ± 15%. Only CD32A131R -CR T cells bound panitumumab. CD32A131R -CR T cells combined with the mAb 8.26 (anti-CD32) and CD16158F -CR T cells combined with the mAb 3g8 (anti-CD16) eliminated colorectal carcinoma (CRC), HCT116FcγR+ cells, in a reverse ADCC assay in vitro. Crosslinking of CD32A131R -CR on T cells by cetuximab or panitumumab and CD16158F -CR T cells by cetuximab induced elimination of triple negative breast cancer (TNBC) MDA-MB-468 cells, and the secretion of interferon gamma and tumor necrosis factor alpha. Neither cetuximab nor panitumumab induced Fcγ-CR T antitumor activity against Kirsten rat sarcoma (KRAS)-mutated HCT116, nonsmall-cell-lung-cancer, A549 and TNBC, MDA-MB-231 cells. The ADCC of Fcγ-CR T cells was associated with the overexpression of EGFR on ECCs. In conclusion, CD32A131R -CR T cells are efficiently redirected by cetuximab or panitumumab against breast cancer cells overexpressing EGFR.

Tumor Infiltration by OX40+ Cells Enhances the Prognostic Significance of CD16+ Cell Infiltration in Colorectal Cancer.

OBJECTIVES: Analysis of tumor immune infiltration has been suggested to outperform tumor, node, metastasis staging in predicting clinical course of colorectal cancer (CRC). Infiltration by cells expressing OX40, a member of the tumor necrosis factor receptor family, or CD16, expressed by natural killer cells, monocytes, and dendritic cells, has been associated with favorable prognosis in patients with CRC. We hypothesized that assessment of CRC infiltration by both OX40+ and CD16+ cells might result in enhanced prognostic significance. METHODS: Colorectal cancer infiltration by OX40 and CD16 expressing cells was investigated in 441 primary CRCs using tissue microarrays and specific antibodies, by immunohistochemistry. Patients' survival was evaluated by Kaplan-Meier and log-rank tests. Multivariate Cox regression analysis, hazard ratios, and 95% confidence intervals were also used to evaluate prognostic significance of OX40+ and CD16+ cell infiltration. RESULTS: Colorectal cancer infiltration by OX40+ and CD16+ cells was subclassified into 4 groups with high or low infiltration levels in all possible combinations. High levels of infiltration by both OX40+ and CD16+ cells were associated with lower pT stage, absence of peritumoral lymphocytic (PTL) inflammation, and a positive prognostic impact. Patients bearing tumors with high infiltration by CD16+ and OX40+ cells were also characterized by significantly longer overall survival, as compared with the other groups. These results were confirmed by analyzing an independent validation cohort. CONCLUSIONS: Combined infiltration by OX40+ and CD16+ immune cells is an independent favorable prognostic marker in CRC. The prognostic value of CD16+ immune cell infiltration is significantly improved by the combined analysis with OX40+ cell infiltration.

Breast Cancer Is Associated with Increased HLA-DRB1*11:01 and HLA-DRB1*10:01 Allele Frequency in a Population of Patients from Central Italy.

The relationship between HLA-DRB1 allele polymorphism and breast cancer (BC) development is still unclear and needs further investigation. To address this issue, we analyzed HLA-DRB1 allele frequency (AF) by sequence-based typing (SBT) in 47 patients from central Italy with BC and 156 sex and age-matched healthy controls. Two hundred ninety-seven individuals from the same region were utilized as historical controls. Pearson's chi-square analysis with Yate's correction or Fisher's Exact test with Bonferroni's correction, as appropriate, were used to compare HLA-DRB1 AF differences in patients and controls. A total of 36 HLA-DRB1 alleles were identified. A detailed study showed that HLA-DRB1*11:01 and HLA-DRB1*10:01 alleles are significantly associated with increased BC risk. In particular, HLA-DRB1*11:01 AF was significantly higher in patients with BC than in healthy females and historical controls, even following Bonferroni's correction (stage I-II BC patients vs historical controls p<0.00; stage III-IV BC patients vs female healthy controls p=0.025 and historical controls p<0.00). The HLA-DRB1*10:01 allele was also positively associated with BC as evidenced by a significantly higher AF in patients with BC than in healthy controls (BC patients stage I-II vs historical controls corrected p =0.01). These results suggest that both HLA-DRB1*11:01 and HLA-DRB1*10:01 AF could represent interesting markers in patients at risk of developing BC.

Role of KIR and CD16A genotypes in colorectal carcinoma genetic risk and clinical stage.

BACKGROUND: NK cell cytotoxicity is regulated by the types of the interaction between killer immunoglobulin-like receptors (KIRs) and human leukocyte antigen (HLA) class I ligands on target cells and the different binding affinity of the Fcγ receptor IIIA (CD16A) for IgG-coated tumor cells. Thus, it is conceivable that KIR and CD16A gene contents may contribute to the function of NK cells by modulating an immune response in the colorectal carcinoma (CRC) microenvironment. This hypothesis is supported by recent evidence suggesting that NK cells improve the clinical course of CRC patients by enhancing the anti-CRC effect of CD8 + T cells. This information provides the rationale to test the hypothesis whether the independent KIR segregation and specificity, as well as CD16A gene polymorphisms, have an impact on CRC. METHODS: Using polymerase chain reaction-sequence-specific primers (PCR-SSP) and sequence-based typing (SBT), we investigated KIR/HLA-C complex and CD16A (48H/R/L,158V/F) gene polymorphisms in 52 CRC patients and 61 local healthy controls (LCTRs). RESULTS: The allele frequency (AF) of at least five activating KIR (aKIRs) of the B haplotype (p = 0.036, OR 0.204), KIR2DL2 (p = 0.047, OR 0.2616), and KIR2DS2 genes (5.8 vs LCTR 13.8 % and vs. Fasano's CTR 16.3 %, p = 0.05, OR 0.3145), in the absence of their cognate HLA-C1 ligands, were significantly associated with a reduced genetic risk of CRC. In contrast, CD16A-48H polymorphism was positively associated with an increased genetic risk of CRC (p = 0.05, OR 2.761). The latter was also found to be correlated with advanced stages of disease [III and IV (p = 0.03, OR 3.625)]. CONCLUSIONS: Our data suggest that the analysis of aKIRs and KIR2DL2 gene and CD16A-48H may be of interest for the identification of individuals at reduced and increased genetic risk of CRC, respectively.

T lymphocytes engineered to express a CD16-chimeric antigen receptor redirect T-cell immune responses against immunoglobulin G-opsonized target cells.

BACKGROUND AIMS: Chimeric antigen receptors (CARs) designed for adoptive immunotherapy need to achieve two functions: antigen recognition and triggering of the lytic machinery of reprogrammed effector cells. Cytotoxic T cells have been engineered with FcγRIII (CD16) chimeric molecules to be redirected against malignant cells by monoclonal antibodies (mAbs). These cells have been proven to mediate granule-dependent cellular cytotoxicity, but it is not clear whether they can also kill malignant cells by a granule-independent mechanism of cell cytotoxicity. METHODS: We engineered a CD16A-CAR equipped with the extracellular CD16A, the hinge spacer and the transmembrane region of CD8, and the ζ-chain of the T-cell receptor/CD3 complex in tandem with the CD28 co-stimulatory signal transducer module. The CD16A-CAR was expressed and functionally tested in the MD45 cell line, a murine T-cell hybridoma with a defective granular exocytosis pathway but capable of killing target cells by a Fas ligand-mediated lysis. RESULTS: Our results indicate that in vitro cross-linking of CD16A-CAR on MD45 cells by the Fc fragment of mAb opsonized tumor cells induced interleukin-2 release and granule-independent cellular cytotoxicity. CONCLUSIONS: We conclude that strategies aimed to implement the therapeutic functions of mAbs used in the clinic with T-dependent immune responses driven by engineered T cells expressing FcγR-CAR can boost the antitumor efficacy of mAbs used in the clinic.

A cluster of Geotrichum clavatum (Saprochaete clavata) infection in haematological patients: a first Italian report and review of literature.

Invasive fungal infections, usually Aspergillus and Candida, represent a major cause of morbidity and mortality in patients with malignant haematological diseases, but in the last years rare fungal infections have more frequently been reported. Here, we report the clinical history of three patients affected with haematological malignancies who developed an infection caused by Geotrichum (G.) clavatum. Two out of three patients were affected by acute myeloid leukaemia (AML), and one by mantle cell lymphoma (MCL). All patients received cytarabine-based chemotherapeutic regimens and developed G. clavatum infection within 3 weeks from therapy initiation. In all cases, G. clavatum was isolated from central venous catheter and peripheral blood cultures. In vitro susceptibility test confirmed an intrinsic resistance to echinocandins and, in all cases, visceral localisations (spleen, liver and lung) were documented by total body computed tomography (CT) scan. A prolonged antifungal therapy with high doses liposomal amphotericin-B was necessary to obtain fever resolution. Only the patient with MCL died while the other two AML recovered, and one of them after received an allogeneic stem cell transplantation. We consecutively reviewed all published cases of infection caused by G. clavatum. Our experience and literature review indicate that G. clavatum can cause invasive infection in haematological patients, mainly in those with acute leukaemia.

Liver protein profiles in insulin receptor-knockout mice reveal novel molecules involved in the diabetes pathophysiology.

Liver has a principal role in glucose regulation and lipids homeostasis. It is under a complex control by substrates such as hormones, nutrients, and neuronal impulses. Insulin promotes glycogen synthesis, lipogenesis, and lipoprotein synthesis and inhibits gluconeogenesis, glycogenolysis, and VLDL secretion by modifying the expression and enzymatic activity of specific molecules. To understand the pathophysiological mechanisms leading to metabolic liver disease, we analyzed liver protein patterns expressed in a mouse model of diabetes by proteomic approaches. We used insulin receptor-knockout (IR(-/-)) and heterozygous (IR(+/-)) mice as a murine model of liver metabolic dysfunction associated with diabetic ketoacidosis and insulin resistance. We evaluated liver fatty acid levels by microscopic examination and protein expression profiles by orthogonal experimental strategies using protein 2-DE MALDI-TOF/TOF and peptic nLC-MS/MS shotgun profiling. Identified proteins were then loaded into Ingenuity Pathways Analysis to find possible molecular networks. Twenty-eight proteins identified by 2-DE analysis and 24 identified by nLC-MS/MS shotgun were differentially expressed among the three genotypes. Bioinformatic analysis revealed a central role of high-mobility group box 1/2 and huntigtin never reported before in association with metabolic and related liver disease. A different modulation of these proteins in both blood and hepatic tissue further suggests their role in these processes. These results provide new insight into pathophysiology of insulin resistance and hepatic steatosis and could be useful in identifying novel biomarkers to predict risk for diabetes and its complications.

HLA-DRB1*13:01 allele in the genetic susceptibility to colorectal carcinoma.

Increasing evidence suggests that HLA-DRB1 alleles reduce or increase the risk of developing ulcerative colitis-associated colorectal carcinoma (CRC) tumors. However, the role of HLA-DRB1 locus on the susceptibility to develop CRC tumor, in the absence of a history of inflammatory bowel diseases (IBDs), is unclear. The aim of our study was to determine whether HLA-DRB1 alleles are associated with IBD-independent CRC tumor. HLA-DRB1 allele polymorphisms were identified by sequence-based typing method in 53 CRC patients and 57 sex- and age-matched healthy Caucasian controls. Pearson's chi-squared analysis with Yate's correction or Fisher's exact test with Bonferroni's correction, as appropriate, were used to compare the allele frequency (AF) differences of HLA-DRB1 in patients and controls. A total of 29 HLA-DRB1 alleles were recognized. A detailed study of these alleles allowed to identify DRB1*13:01 and DRB1*11:01 alleles that were significantly associated with an increased and reduced risk to develop CRC tumor, respectively. AF of DRB1*13:01, in CRC patients, was significantly higher than that of healthy controls, even following Bonferroni's correction (p = 0.029). In contrast, the presence of the DRB1*11:01 allele was negatively associated with CRC tumor as evidenced by the significantly lower AF in CRC patients than that of healthy controls (p = 0.005). However, following Bonferroni's correction, the AF of DRB*11:01 lost its statistical significance. These results suggest that HLA-DRB1*13:01 allele could be a potential marker for predicting genetic susceptibility to CRC tumor. In contrast, the protective role of DRB1*11:01 remains unclear.

Natural humoral immune response to ribosomal P0 protein in colorectal cancer patients.

BACKGROUND: Tumor associated antigens are useful in colorectal cancer (CRC) management. The ribosomal P proteins (P0, P1, P2) play an important role in protein synthesis and tumor formation. The immunogenicity of the ribosomal P0 protein in head and neck, in breast and prostate cancer patients and the overexpression of the carboxyl-terminal P0 epitope (C-22 P0) in some tumors were reported. METHODS: Sera from 72 colorectal tumor patients (67 malignant and 5 benign tumors) were compared with 73 healthy donor sera for the presence of antibodies to CEA, EGFR, ErbB2 and ribosomal P proteins by western blotting or ELISA. Expression of the C-22 P0 epitope on tissues and colon cancer cells was determined by immunoperoxidase staining and indirect immunofluorescence/western blotting, respectively, employing MAb 2B2. Biological effects of MAb 2B2 on colon cancer cells were assessed by the Sulforhodamine B cell proliferation assay, trypan blue exclusion test and cleaved caspase-3 detection. Fisher's exact test was used to compare the number of auto-antibodies positive patients with healthy donors. Variation in the C-22 P0 expression, and in the number of apoptotic cells was evaluated by Student's t-test. Variation in cell survival and cell death was evaluated by Newman-Keuls test. RESULTS: No significant humoral response was observed to CEA, EGFR and ErbB2 in CRC patients. Conversely, 7 out of 67 CRC patient sera reacted to ribosomal P proteins. The prevalence of P proteins auto-antibodies in CRC patients was significant. Five patients showed restricted P0 immunoreactivity, while two patients reacted simultaneously to all P proteins. The C-22 P0 epitope was homogenously expressed both in malignant tumors and the adjacent mucosa, but the intensity of expression was higher in the tumor. Starved colon cancer cells showed a higher C-22 P0 epitope plasma membrane expression compared to control cells. MAb 2B2 inhibited colon cancer cell growth and induced cell death in a dose dependent manner. CONCLUSIONS: Our study shows a spontaneous humoral immune response to ribosomal P0 protein in CRC patients and the inhibition of in vitro cancer cell growth after C-22 P0 epitope targeting. The ribosomal P0 protein might be a useful immunological target in CRC patients.

Minimal residual disease as biomarker for optimal biologic dosing of ARA-C in patients with acute myeloid leukemia.

We assessed by flow cytometry minimal residual disease (MRD) in patients with acute myeloid leukemia (AML) given standard-dose (SDAC) and high-dose ARA-C (HDAC) regimens. Of 163 patients enrolled, 130 (median age, 45 years; range, 18-59 years) qualified for analysis, all achieving complete remission after treatment with SDAC (n = 78) or HDAC (n = 52) plus etoposide and daunorubicin. Consolidation consisted of intermediate-dose ARA-C and daunorubicin. MRD negativity was significantly more frequent in the SDAC vs. HDAC arm after both induction (37% vs. 15%, P = 0.007) and consolidation (44% vs. 18%, P = 0.002). Respective median residual leukemic cell counts with SDAC and HDAC use were 1.5 × 10(-3) and 4 × 10(-3) (P = 0.033) after induction and 5.7 × 10(-4) and 2.9 × 10(-3) (P = 0.008) after consolidation. Based on ARA-C schedule and post-consolidation MRD status, the four patient groups (SDAC-MRD(-) , HDAC-MRD(-) , SDAC-MRD(+) , and HDAC-MRD(+) ) displayed 5-year overall survival rates of 60%, 33%, 24%, and 42% (P = 0.007), respectively, with 24%, 35%, 74%, and 48% (P < 0.0001) respective cumulative incidence of relapse estimates. MRD may serve as a biomarker for optimal biologic dosing of ARA-C, and SDAC regimen appears to yield more frequent MRD negativity.

Prognostic and therapeutic implications of minimal residual disease detection in acute myeloid leukemia.

The choice of either induction or postremission therapy for adults with acute myeloid leukemia is still largely based on the "one size fits all" principle. Moreover, pretreatment prognostic parameters, especially chromosome and gene abnormalities, may fail in predicting individual patient outcome. Measurement of minimal residual disease (MRD) is nowadays recognized as a potential critical tool to assess the quality of response after chemotherapy and to plan postremission strategies that are, therefore, driven by the individual risk of relapse. PCR and multiparametric flow cytometry have become the most popular methods to investigate MRD because they have been established as sensitive and specific enough to allow MRD to be studied serially. In the present review, we examine the evidence supporting the appropriateness of incorporating MRD detection into the AML risk assessment process. A comprehensive prognostic algorithm, generated by combining pretreatment cytogenetics/genetics and posttreatment MRD determination, should promote advances in development of personalized therapeutic approaches.

High sensitivity of flow cytometry improves detection of occult leptomeningeal disease in acute lymphoblastic leukemia and lymphoblastic lymphoma.

Conventional cytology (CC) of cerebrospinal fluid (CSF) fails to demonstrate malignant cells in up to 45 % of patients with acute lymphoblastic leukemia or lymphoblastic lymphoma (ALL/LL) in whom occult leptomeningeal disease is present. Flow cytometry (FCM) is considered more sensitive than CC, but clinical implications of CC negativity/CC positivity are not yet established. CSF samples from 38 adult patients with newly diagnosed ALL/LL were examined. Five (13 %) and nine (24 %) specimens were CC positive-FC positive (FCM(pos)/CC(pos)) and CC negative-FC positive (CC(neg)/FCM(pos)), respectively. The remaining 24 (63 %) samples were double negative (CC(neg)/FCM(neg)) (p = 0.001). CC(neg)/FCM(pos) patients showed a significantly shorter overall survival (OS) compared to CC(neg)/FCM(neg) ones. In multivariate analysis, the status of single FCM positivity was demonstrated to affect independently duration of OS (p = 0.005). In conclusion, FCM significantly improves detection of leptomeningeal occult localization in ALL/LL and appears to anticipate an adverse outcome. Further prospective studies on larger series are needed to confirm this preliminary observation.

Local therapy in glioma: An evolving paradigm from history to horizons (Review).

Despite the implementation of multimodal treatments after surgery, glioblastoma (GBM) remains an incurable disease, posing a significant challenge in neuro-oncology. In this clinical setting, local therapy (LT), a developing paradigm, has received significant interest over time due to its potential to overcome the drawbacks of conventional therapy options for GBM. The present review aimed to trace the historical development, highlight contemporary advances and provide insights into the future horizons of LT in GBM management. In compliance with the Preferred Reporting Items for Systematic Review and Meta-Analysis Protocols criteria, a systematic review of the literature on the role of LT in GBM management was conducted. A total of 2,467 potentially relevant articles were found and, after removal of duplicates, 2,007 studies were screened by title and abstract (Cohen's κ coefficient=0.92). Overall, it emerged that 15, 10 and 6 clinical studies explored the clinical efficiency of intraoperative local treatment modalities, local radiotherapy and local immunotherapy, respectively. GBM recurrences occur within 2 cm of the radiation field in 80% of cases, emphasizing the significant influence of local factors on recurrence. This highlights the urgent requirement for LT strategies. In total, three primary reasons have thus led to the development of numerous LT solutions in recent decades: i) Intratumoral implants allow the blood-brain barrier to be bypassed, resulting in limited systemic toxicity; ii) LT facilitates bridging therapy between surgery and standard treatments; and iii) given the complexity of GBM, targeting multiple components of the tumor microenvironment through ligands specific to various elements could have a synergistic effect in treatments. Considering the spatial and temporal heterogeneity of GBM, the disease prognosis could be significantly improved by a combination of therapeutic strategies in the era of precision medicine.

Langerhans cell histiocytosis: current advances in molecular pathogenesis.

Langerhans cell histiocytosis (LCH) is a rare and clinically heterogeneous hematological disease characterized by the accumulation of mononuclear phagocytes in various tissues and organs. LCH is often characterized by activating mutations of the mitogen-activated protein kinase (MAPK) pathway with BRAFV600E being the most recurrent mutation. Although this discovery has greatly helped in understanding the disease and in developing better investigational tools, the process of malignant transformation and the cell of origin are still not fully understood. In this review, we focus on the newest updates regarding the molecular pathogenesis of LCH and novel suggested pathways with treatment potential.

Acute Lymphoblastic Leukemia Immunotherapy Treatment: Now, Next, and Beyond.

Acute lymphoblastic leukemia (ALL) is a blood cancer that primarily affects children but also adults. It is due to the malignant proliferation of lymphoid precursor cells that invade the bone marrow and can spread to extramedullary sites. ALL is divided into B cell (85%) and T cell lineages (10 to 15%); rare cases are associated with the natural killer (NK) cell lineage (<1%). To date, the survival rate in children with ALL is excellent while in adults continues to be poor. Despite the therapeutic progress, there are subsets of patients that still have high relapse rates after chemotherapy or hematopoietic stem cell transplantation (HSCT) and an unsatisfactory cure rate. Hence, the identification of more effective and safer therapy choices represents a primary issue. In this review, we will discuss novel therapeutic options including bispecific antibodies, antibody-drug conjugates, chimeric antigen receptor (CAR)-based therapies, and other promising treatments for both pediatric and adult patients.

Impact of HLA Class I Antigen, Killer Inhibitory Receptor, and FCGR3A Genotypes on Breast Cancer Susceptibility and Tumor Stage.

BACKGROUND: The identification in breast cancer (BC) of novel genetic biomarkers regulating natural killer (NK) cell function, including the HLA, KIR, and CD16A (FCGR3A), may be still a challenge. OBJECTIVE: We aimed to evaluate whether the combined effect of these polymorphisms has an impact on BC susceptibility and progression. METHODS: 47 BC Italian patients and healthy individuals (39 females and 66 males/females) were genotyped by Sanger sequencing (HLA-C exon 2-4 and FCGR3A-158V/F, 48L/R/H) and PCR-SSP typing (KIR genes). RESULTS: HLA-C gene allele analysis showed the group C1, with HLA-C*07:02:01 allele, to be significantly associated with tumor progression (16.7% vs. 4.0%, p=0.04, OR=4.867), and instead, group C2, with HLA-C*05:01:01, was protective against disease susceptibility (0.0% vs. 7.2%, p=0.019, OR=0.087). In addition, we highlighted a significant reduction of the KIR2DS4ins in BC patients (pcorr.=0.022) and an increased combined presence of KIR2DL1 and KIR2DS1 genes in advanced BC patients compared to earlier stages (66.7% vs. 19.2%, p=0.002). The concurrent lack of KIR2DL2 and KIR2DS4 genes in the presence of HLA-C2 alleles was significantly associated with increased susceptibility to BC (p=0.012, OR=5.020) or with lymph node involvement (p=0.008, OR=6.375). Lastly, we identified different combinations of the FCGR3A-48/158 variants and KIR genes in BC patients compared to controls. CONCLUSION: Our findings suggest that in the development of BC probably exists a disorder of the NK innate immunity influenced by KIR/HLA-C gene content and FCGR3A-158 polymorphisms and that the combined analysis of these biomarkers might help predict genetic risk scores for tailored screening of BC patients in therapy.

Contribution of A-to-I RNA editing, M6A RNA Methylation, and Alternative Splicing to physiological brain aging and neurodegenerative diseases.

Aging is a physiological and progressive phenomenon in all organisms' life cycle, characterized by the accumulation of degenerative processes triggered by several alterations within molecular pathways. These changes compromise cell fate, resulting in the loss of functions in tissues throughout the body, including the brain. Physiological brain aging has been linked to structural and functional alterations, as well as to an increased risk of neurodegenerative diseases. Post-transcriptional RNA modifications modulate mRNA coding properties, stability, translatability, expanding the coding capacity of the genome, and are involved in all cellular processes. Among mRNA post-transcriptional modifications, the A-to-I RNA editing, m6A RNA Methylation and Alternative Splicing play a critical role in all the phases of a neuronal cell life cycle and alterations in their mechanisms of action significantly contribute to aging and neurodegeneration. Here we review our current understanding of the contribution of A-to-I RNA editing, m6A RNA Methylation, and Alternative Splicing to physiological brain aging process and neurodegenerative diseases.