Objective: Chemotherapy-based regimens remain the standard first-and second-line treatment options for patients with driver gene-negative non-small cell lung cancer (NSCLC). However, in real-world settings, certain patients cannot tolerate chemotherapy or opt to decline it. Immune checkpoint inhibitors (ICIs) constitute the preferred chemotherapy-free alternative. To enhance patient prognosis, this study aimed to examine the efficacy of ICIs combined with anlotinib in real-world scenarios. Methods: This prospective, multicenter, real-world study evaluated the efficacy and safety of ICIs combined with anlotinib in patients with advanced NSCLC. Patients undergoing first-or second-line treatment were enrolled. The primary endpoint was progression-free survival (PFS), while the secondary endpoints included overall survival (OS), objective response rate (ORR), disease control rate (DCR), and safety. Results: In total, 242 patients were enrolled from 28 centers. The median PFS for the entire cohort was 7.8 [95% confidence interval (95% CI), 7.0-9.5] months, OS events occurred in 112 (46.3%) patients, with a current median OS of 17.0 (95% CI, 15.1-19.4) months. The ORR and DCR were 36.0% (95% CI, 30.2%-42.2%) and 97.9% (95% CI, 95.3%-99.1%), respectively. The median PFS was 9.8 (95% CI, 7.4-12.5) months for first-line therapy and 6.9 (95% CI, 6.0-8.3) months for second-line therapy. Treatment-related adverse events (AEs) occurred in 198 (81.8%) patients, with grade 3-4 AEs reported in 22 (9.1%) patients. Conclusions: This multicenter, real-world study demonstrates that the anlotinib-ICI combination regimen exhibits clinically meaningful efficacy and tolerability as a chemotherapy-free alternative for advanced NSCLC, offering viable evidence to guide treatment for patients who are unsuitable for conventional chemotherapy.

PURPOSE:Cervical cancer (CC), a common female malignancy, has been linked to alterations in DNA methylation. This study employed an integrated "dry-wet lab" strategy combining bioinformatics, machine learning, and experimental validation to identify novel methylation biomarkers for CC. METHODS:Methylome and transcriptome data from the TCGA and GEO cohorts (n=349 discovery, n=414 validation) were analyzed to identify differentially methylated CpGs. The top candidates were validated by pyrosequencing, methylation-specific PCR, and quantitative assays. Diagnostic models were developed, and functional studies were performed for the target markers. RESULTS:Eighteen differentially methylated CpGs were identified, with five top candidates (three in the ZSCAN18 promoter) showing diagnostic potential. ZSCAN18 promoter methylation levels and positivity rates were significantly greater in CC tissues than in normal tissues (p<0.05), reaching 77.8% (21/27) in ThinPrep cytology test (TCT) samples. The ridge regression diagnostic model achieved an AUC of 0.9421 in the validation cohort. Similarly, ZSCAN18 overexpression suppressed CC cell proliferation (p<0.05). CONCLUSIONS:This study established a rapid, effective and systematic systemic research strategy to screen novel methylation markers for CC. ZSCAN18 promoter methylation correlates with cervical lesion severity, and the diagnostic model enhances the diagnostic ability. These findings highlight the dual role of ZSCAN18 as a diagnostic marker and potential therapeutic target.

The role of B cells in the anti-tumor immune response remains controversial. An increase in the number of B cells in the peripheral blood of some tumor patients has been associated with poor immunotherapy efficacy. However, the mechanism leading to the generation of these cells is not well-described. Using a fibrosarcoma model, we show that intraperitoneal administration of a xenogeneic antigen in tumor-bearing mice evokes large increases in antigen-specific serum immunoglobulin formation compared to tumor-naïve mice. An inability of tumor-bearing mice to induce enhanced antibody production after myeloid cell depletion suggests the antibody responses are CD11b+ myeloid cell-dependent. In vitro, CD11b+ myeloid cells promoted B cell proliferation, activation, and survival. High levels of tumor necrosis factor (TNF)-α were produced by CD11b+ cells, and TNF-α blockade inhibited B cell responses. CD11b+ cells appear to be important promoters of B cell responses and targeting B cells may increase the efficacy of immunotherapy in tumor-bearing hosts.

Background Anti-angiogenic drugs increase anti-tumor efficacy of immune checkpoint inhibitors (ICIs). However, the optimal dose of anti-angiogenic drugs remains unclear. Methods We retrospectively analyzed efficacy and safety data from patients diagnosed with advanced or metastatic non-small cell lung cancer (NSCLC) that received PD-1 blockade with low-doses of anlotinib, a highly selective receptor tyrosine kinase inhibitor mainly targeting vascular endothelial growth factor receptors, as second or later line therapy. The primary endpoint was progression-free survival (PFS). Secondary endpoints included overall survival (OS), overall response rate (ORR), disease control rate (DCR), and safety profile. Univariate and multivariate analyses were used to identify prognostic factors. Results A total of 40 eligible patients were included. The median PFS was 11.4 months. The median OS of the entire cohort was 27.0 months. ORR was achieved in 16 patients (40.0%) and DCR was maintained in 33 patients (82.5%). The overall incidence of adverse events (AEs) was 52.5%, and the most common all grade AE was gastrointestinal reactions, which occurred in four patients (10.0%). Treatment-related grade 3/4 toxicity was observed in one patient (2.5%). Conclusions Low-dose anlotinib may be an effective and well-tolerated anti-angiogenesis partner for combination therapy with ICIs in second-line and later settings for advanced NSCLC.

Identification of reliable biomarkers to predict efficacy of immune checkpoint inhibitors and to monitor relapse in cancer patients receiving this therapy remains one of the main objectives of cancer immunotherapy research. We found that the pretreatment B cell number in the peripheral blood differed significantly between responders and non-responders to anti-PD-1-based immunotherapy. Patients with various cancer types achieving a clinical response had a significantly lower number of B cells compared with those with progressive disease. Patients who progressed from partial response to progressive disease exhibited a gradually increased number of circulating B cells. Our findings suggest that B cells represent a promising biomarker for anti-PD-1-based immunotherapy responses and inhibit the effect of PD-1 blockade immunotherapy. Thus, preemptive strategies targeting B cells may increase the efficacy of PD-1 blockade immunotherapy in patients with solid tumors.

Background: Advanced pancreatic cancer (PC) has very poor prognosis with present treatments, thus necessitating continued efforts to find improved therapeutic approaches. Both preclinical and preliminary clinical data indicate that cytokine-induced killer (CIK) cells are an effective tool against various types of solid tumors. Here, we conducted a study to determine whether CIK cell-based therapy (CBT) can improve the outcomes of advanced PC. Methods: Eighty-two patients with advanced PC, whose predicted survival time was longer than 3 months, were analyzed retrospectively. Of all the patients, 57 individuals were receiving chemotherapy, while the remaining 25 individuals were treated with CBT. Results: The overall survival analysis was based on 48 deaths in the 57 patients in the chemotherapy group (84.2 %) and 18 deaths in the 25 patients in the CBT group (72.0 %). In the CBT group, the median overall survival time was 13.5 months, as compared to 6.6 months in the chemotherapy group (hazard ratio for death, 0.39; 95 % confidence interval, 0.23 to 0.65; p < 0.001). The survival rate was 88.9 % in the CBT group versus 54.2 % in the chemotherapy group at 6 months, 61.1 % versus 12.5 % at 12 months, and 38.9 % versus 4.2 % at 18 months. The disease control rate was 68.0 % in the CBT group and 29.8 % in the chemotherapy group (p < 0.001). Conclusions: These results from this retrospective analysis appeared to imply that CBT might prolong survival in these high-risk PC patients. Prospective study is needed to corroborate this observation.

Adoptive immunotherapy using cytokine-induced killer (CIK) cells is a promising cancer treatment, but its efficacy is restricted by various factors, including the accumulation of myeloid-derived suppressor cells (MDSCs). In this study, we determine whether chemotherapeutic drugs that reduce MDSC levels enhance the efficacy of CIK cell therapy in the treatment of solid tumors. Fifty-three patients were included in this study; 17 were diagnosed with metastatic renal cell carcinoma (MRCC), 10 with advanced pancreatic cancer (PC), and 26 with metastatic melanoma (MM). These patients were divided into two groups: CIK cell therapy alone and CIK cell therapy combined with chemotherapy. Combining CIK cell therapy and chemotherapy increased 1-year survival rates and median survival times in MRCC and PC patients, but not in MM patients. The disease control rate did not differ between treatment groups for MRCC or MM patients, but was higher in PC patients receiving combined treatment than CIK cell treatment alone. These data suggest that addition of MDSC-decreasing chemotherapy to CIK cell therapy improves survival in MRCC and PC patients.

Metastatic renal cell carcinoma (MRCC) is one of the malignancies that are sensitive to immunotherapy. However, the underlying immune inhibitory factors such as myeloid-derived suppressor cells (MDSCs) might restrain the efficacy of immunotherapy. The present study investigates the clinical efficacy of cytokine-induced killer (CIK) cell therapy in patients with MRCC and explores whether the levels of peripheral MDSCs are associated with the prognosis of patients receiving this therapy. Twenty-nine patients with measurable MRCC were treated with an adoptive transfer of autologous CIK cells, followed by 5 consecutive days of interleukin-2 administration. The tumor response and 1-year survival were observed. The proportion of MDSCs in the peripheral blood was detected, and the correlation of MDSCs with prognosis was analyzed. Of 29 evaluable patients, no complete responses were seen; 4 patients exhibited a partial response (13.8%), 18 patients displayed stable disease (62.1%), and 7 patients showed progressive disease (24.1%). Twenty patients (69.0%) were alive 14.8-41.4 months at the time of the last follow-up (median follow-up=20.2 mo). The 1-year survival was 82.8% (24/29). Peripheral blood MDSCs were elevated in almost all MRCC patients and decreased after CIK-cell infusion. Subgroup analysis indicated that patients with a relatively low proportion of MDSCs exhibited prolonged survival. In conclusion, our data suggest that transfusion of autologous CIK cells can induce regression of MRCC, and MDSCs can serve as a potential marker for the prognosis of patients receiving a CIK-based therapy.

目的 评价化疗联合自体CIK(cytokine-induced killer)细胞治疗转移性肾癌的临床疗效及安全性.方法 19例转移性肾癌患者接受至少1周期的5-氟尿嘧啶和(或)吉西他滨(5-fluorouracil/gemcitabine,5-Fu/Gem)化疗,化疗结束后接受4～8次CIK细胞治疗,细胞回输后应用3天d1～d3白细胞介素2(interleukin-2,IL-2)和胸腺肽α-1.结果 19例患者中1例部分缓解(PR),13例稳定(SD),5例进展(PD),疾病总拉制率(CP+ PR+ SD)为73.68％.评价为PD的5例患者无进展生存期(PFS)为(4.43±2.85)月;其余14例患者PFS未到,中位PFS已达11.64月.结论 CIK细胞联合化疗治疗转移性肾癌疗效确切,疾病控制率得到明显提高,患者带瘤生存时间明显廷长.

The aim of the study was to investigate the sensitivity of AHH-1 human lymphoblastoid cells to radiation and its relevance to intracellular events, specifically alteration in cellular energy-producing systems. AHH-1 human lymphoblastoid cells were irradiated with 6 Gy of gamma radiation, and then were collected at the indicated time points. Parallel studies were conducted to assess the effects of radiation on the cell proliferation and apoptotic index. Mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) production were monitored. A marked decrease of cell viability was observed as early as 12 h postirradiation and fraction of apoptotic cells was highest at 24 It. Intracellular ROS generation measured with 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) appeared to be highest as early as 30 min postirradiation and resumed to normal level at 6 h. Unexpectedly, the fluorescence intensity of Rhodamine 123 for measuring NIMP did not change during the first 3 h afterradiation and exhibited an aberrant increase at 6 It. The results suggestthat AHH-1 cells are sensitive to radiation-induced apoptosis and ROS generation is an early phase in the apoptosis process. Moreover, the results might cast doubts on those studies using Rhodamine 123 which hypothesized that the fall in NIMP is one of the early events of apoptosis. (c) 2007 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.

AIM:To investigate the changes of lymphocyte subpopulations, especially CD4 + CD25 + T regulatory cells in Smad3 -/-mice. METHODS:Hematological changes and changes of lymphocyte subpopulations were detected in Smad3 -/-mice using cell counter and flow cytometry, respectively, and compared to their littermate controls. RESULTS:The numbers of neutrophils and lymphocytes in peripheral blood were significantly increased in Smad3 -/-mice compared to littermate controls.CD19 + expressing cells in blood and spleen, and CD8 + T cells in thymus were all markedly decreased in Smad3 -/-mice.More important, Smad3 -/-mice had an increased population of CD4 + CD25 + T cells in peripheral lymphoid tissues, including thymus, spleen, and lymph nodes. CONCLUSION:These observations suggest that the changes of lymphocyte subpopulations might play a role in susceptibility to inflammation of Smad3 -/-mice.

Bcl-xL belongs to a family of proteins which inhibit apoptosis in a number of stimuli including ionizing radiation. To better understand the effects and mechanisms of Bcl-xL on the apoptosis of lymphocytes and provide experimental basis to treat immune injury induced by radiation, we used normal human lymphoblastoid AHH-1 cells that were engineered to overexpress Bcl-xL proteins. Our results showed that overexpressed Bcl-xL reduced time-dependent increase of apoptosis induced by ionizing radiation. Reactive oxygen species (ROS) generation and Bax protein expression in the transfected AHH1-Bcl-xL cells were also lower compared to parental AHH-1 cells. Unexpectedly, the fluorescence intensity of Rhodomine 123 (Rh 123) for measuring mitochondrial membrane potential (MMP) did not change at all detected time points. These results possess a vital significance for insights into a new strategy for gene therapy of radiation-induced immune injury.

Apoptosis or programmed cell death is an active form of cell death which is essential for tissue homeostasis. Many proteins are involved in the molecular signal transduction of apoptosis. The caspase enzymes, a family of specific cysteine proteases, play a central role in cell death machinery. In this review, we mainly discuss the current understanding of several pathways to activate caspases and some key proteins related to these pathways.