Z-flipon variants reveal the many roles of Z-DNA and Z-RNA in health and disease.

Identifying roles for Z-DNA remains challenging given their dynamic nature. Here, we perform genome-wide interrogation with the DNABERT transformer algorithm trained on experimentally identified Z-DNA forming sequences (Z-flipons). The algorithm yields large performance enhancements (F1 = 0.83) over existing approaches and implements computational mutagenesis to assess the effects of base substitution on Z-DNA formation. We show Z-flipons are enriched in promoters and telomeres, overlapping quantitative trait loci for RNA expression, RNA editing, splicing, and disease-associated variants. We cross-validate across a number of orthogonal databases and define BZ junction motifs. Surprisingly, many effects we delineate are likely mediated through Z-RNA formation. A shared Z-RNA motif is identified in SCARF2, SMAD1, and CACNA1 transcripts, whereas other motifs are present in noncoding RNAs. We provide evidence for a Z-RNA fold that promotes adaptive immunity through alternative splicing of KRAB domain zinc finger proteins. An analysis of OMIM and presumptive gnomAD loss-of-function datasets reveals an overlap of Z-flipons with disease-causing variants in 8.6% and 2.9% of Mendelian disease genes, respectively, greatly extending the range of phenotypes mapped to Z-flipons.

An Open-Label Study of the Safety and Efficacy of Tag-7 Gene-Modified Tumor Cells-Based Vaccine in Patients with Locally Advanced or Metastatic Malignant Melanoma or Renal Cell Cancer.

LESSONS LEARNED: This study showed that carefully selected patients with locally advanced and metastatic forms of malignant melanoma and renal cell carcinoma could potentially have long-term disease control with a tag-7 gene-modified tumor cells-based vaccine. Randomized clinical trials in patients whose tumors produce low amounts of immunosuppressive factors are needed to confirm this hypothesis in both the adjuvant and metastatic settings. BACKGROUND: Immunotherapy may produce long-lasting effects on survival and toxicity. The magnitude of efficacy may be dependent on immune factors. We analyzed the results of a phase I/II study of a tag-7 gene-modified tumor cells-based vaccine (GMV) in patients with malignant melanoma (MM) or renal cell carcinoma (RCC) with biomarker analysis of immunosuppressive factors (ISFs) production by their tumor cells. METHODS: From 2001 to 2014, 80 patients received GMV: 68 with MM and 12 with RCC. Treatment in the metastatic setting included 61 patients (MM, 51; RCC, 10), and treatment in the adjuvant setting (after complete cytoreduction) included 19 patients (MM, 17; RCC, 2). Twenty-six patients were stage III (33%), and 54 (67%) were stage IV. The patients' tumor samples were transferred to culture, transfected with tag-7 gene, and inactivated by radiation. The produced product was injected subcutaneously every 3 weeks until progression or 2 years of therapy. ISFs were measured in the supernatants of the tumor cell cultures and used as predictive factors. RESULTS: No major safety issues or grade 5 adverse events (AEs) were seen. One grade 4 and two grade 3 AEs were registered. No AEs were registered in 89.4% of treatment cycles. No delayed AE was found. The 5-year overall survival (OS) in the intention-to-treat population was 25.1%. There were no differences between MM OS and RCC OS (log rank, p = .44). Median OS in the metastatic setting was 0.7 years and in the adjuvant setting was 3.1 years. Classification trees were built on the basis of ISF production (Fig. 1). The median OS was 6.6 years in the favorable prognosis (FP) group (major histocompatibility complex class I polypeptide-related sequence A [MICA] level ≤582 pg/mL, n = 15) and 4.6 months in the unfavorable (UF) group (MICA level >582 pg/mL, n = 12; p < .0001). No significant differences were found between classification trees based on ISFs (transforming growth factor ß1 [TGF-ß1], interleukin-10 [IL-10], and vascular endothelial growth factor [VEGF]). In patients with stage III-IV MM with FP, median OS was 2.3 years, with 31% patients alive at 10 years (Fig. 2) in the UF group (0.4 years; log rank, p = 1.94E-5). No FP patients received modern immunotherapy. CONCLUSION: GMV showed high results in carefully selected patients with low ISF (TGF-ß1, IL-10, and VEGF) production. The method should be further investigated in patients with FP.

Cancer/testis antigens expression during cultivation of melanoma and soft tissue sarcoma cells.

BACKGROUND: Autologous dendritic cells (DC) loaded with tumor-associated antigens (TAAs) are a promising approach for anticancer immunotherapy. Polyantigen lysates appear to be an excellent source of TAAs for loading onto the patient's dendritic cells. Cancer/testis antigens (CTA) are expressed by a wide range of tumors, but are minimally expressed on normal tissues, and could serve as a universal target for immunotherapy. However, CTA expression levels can vary significantly in patients with the same tumor type. We proposed that patients who do not respond to DC-based therapy may have distinct features of the CTA expression profile on tumor cells. PATIENTS AND METHODS: We compared the gene expression of the principal families CTA in 22 melanoma and 27 soft tissue and bone sarcomas cell lines (STBS), received from patients and used for DC vaccine preparation. RESULTS: The majority (47 of 49, 95.9%) cell lines showed CTA gene activity. The incidence of gene expression of GAGE, NYESO1, MAGEA1, PRAME's was significantly different (adj. p < 0.05) between melanoma and sarcoma cell lines. The expression of the SCP1 gene was detected neither in melanoma cells nor in the STBS cells. Clustering by the gene expression profile revealed four different expression patterns. We found three main patterns types: hyperexpression of multiple CTA, hyperexpression of one CTA with almost no expression of others, and no expression of CTA. All clusters types exist in melanoma and sarcoma cell lines. We observed dependence of killing efficacy from the PRAME (rho = 0.940, adj. p < 0.01) expression during real-time monitoring with the xCELLigence system of the interaction between melanoma or sarcoma cells with the T-lymphocytes activated by the lysate of selected allogenous melanoma cell lines with high expression of CTA. CONCLUSION: Our results demonstrate that one can use lysates from allogeneic melanoma cell lines as a source of CTA for DC load during the production of anticancer vaccines for the STBS treatment. Patterns of CTA expression should be evaluated as biomarkers of response in prospective clinical trials.

THE IMPACT OF COVID-19 PANDEMIC ON IMPROVING THE LEGAL REGULATION OF PROTECTION OF HUMAN RIGHT TO HEALTH.

OBJECTIVE: The aim: Theoretical and methodological substantiation of the impact of COVID-19 on the implementation of state policy on the protection of human right to health in terms of improving the legal framework in the field of demographic security. PATIENTS AND METHODS: Materials and methods: The main research materials are the norms of the International Covenant on Economic, Social and Cultural Rights, the Conventions for the Protection of Human Rights and Fundamental Freedoms and the legal framework of the countries that have adopted temporary quarantine measures. This research is based on empiricaland analytical data from WHO, Bloomberg's financial information provider. During the research, the following methods have been used: statistical, system-structural analysis, content-analysis, comparison, grouping and forecasting. CONCLUSION: Conclusions: Under the conditions of pandemic, attention should be paid to strengthening both administrative and criminal liability for violating quarantine, which will serve as a prerequisite for improving the legal mechanism of combating threats to the country's demographic security. The protection of the right to health requires the state to create conditions to prevent the risk of occupational diseases among health care workers and others involved in the response to COVID-19.

The Effect of Polyphenolic Composition BP-C3 on the Efficacy and Hematological Toxicity of Cyclophosphamide in the Chemotherapy of Mice Bearing Soft Tissue Sarcomas Induced by Benzo[a]pyrene.

This study aimed to evaluate the effect of lignin-derived polyphenolic composition BP-C3 on the efficacy and hematological toxicity of cyclophosphamide (CPA). Male and female Swiss-H derived mice bearing benzo[a]pyrene-induced soft tissue sarcomas were treated with CPA 300 mg/kg, BP-C3 75 mg/kg, or a combination. Tumor growth inhibition in male mice treated with CPA, BP-C3, or a combination of CPA and BP-C3 was significant and corresponded to 78%, 45%, and 82%, respectively, on day 21 after CPA administration on day 0. In female mice, tumor growth inhibition was 58%, -11%, and 35% when treated with CPA, BP-C3, or a combination of CPA and BP-C3, respectively. CPA administration resulted in significant hematological toxicity evidenced by a decreased white blood cell count on day 4 (2.43 ± 1.77 × 109/L in male mice and 1.19 ± 0.71 × 109/L in female mice) and anemia development on day 7 (6.55 ± 1.74 × 1012/L in male mice and 5.89 ± 2.24 × 1012/L in female mice). The red blood cell count measured on day 7 in animals treated with the combination of BP-C3 and CPA constituted 7.12 ± 1.17 × 1012/L and 7.36 ± 2.07 × 1012/L for male and female mice, respectively. The results of our study demonstrate the antitumor activity of BP-C3 in male mice bearing soft tissue sarcomas. Neither the antitumor activity nor the hematological toxicity of CPA were significantly influenced by BP-C3. A less pronounced effect of CPA on RBC count is demonstrated when this agent is given jointly with BP-C3.

Primary cultures of human colon cancer as a model to study cancer stem cells.

The principal cause of death in cancer involves tumor progression and metastasis. Since only a small proportion of the primary tumor cells, cancer stem cells (CSCs), which are the most aggressive, have the capacity to metastasize and display properties of stem cells, it is imperative to characterize the gene expression of diagnostic markers and to evaluate the drug sensitivity in the CSCs themselves. Here, we have examined the key genes that are involved in the progression of colorectal cancer and are expressed in cancer stem cells. Primary cultures of colorectal cancer cells from a patient's tumors were studied using the flow cytometry and cytological methods. We have evaluated the clinical and stem cell marker expression in these cells, their resistance to 5-fluorouracil and irinotecan, and the ability of cells to form tumors in mice. The data shows the role of stem cell marker Oct4 in the resistance of primary colorectal cancer tumor cells to 5-fluorouracil.

Cell technologies in immunotherapy of cancer.

Tumor growth is accompanied by active immune reactions even on the early stages. Vaccine therapy implies the use of single antigen or combination of antigens, either with or without adjuvants, for the modulation of immune response. N.N. Petrov Institute of Oncology joined the field of antitumor vaccine therapy and related cellular technologies in 1998. The following activities are held: (1) Optimization of the preparation of autologous and allogeneic antitumor vaccines and development of tumor cell culture bank for the experiments on allogeneic vaccination. (2) Clinical evaluation of autologous vaccine therapy by (a) bone marrow precursors of dendritic cells (DCs), which are loaded with tumor lysates; (b) genetically modified tumor cells; (c) intact tumor cells used in combination with various adjuvants (BCG, IL-1beta, and IL-1beta combined with low doses of cyclophosphamide) in patients with disseminated melanoma, metastatic kidney cancer, and colorectal cancer. Total 117 patients have received non-modified vaccine (48 patients: 2-6 intracutaneous BCG injections; 54 patients: 4-6 intracutaneous IL-1beta injections; 15 patients: up to 6 injections of IL-1beta in combination with low doses of cyclophosphamide). Clinical trial of genetically modified vaccine included 59 patients (clinical results: I PR (partial response) / 8 SD (disease stabilization)--melanoma, 2 PR/ 2 MR (minimal response) / 3 SD--renal cancer). Vaccine prepared from tumor cell-activated DC bone marrow precursors was administered to 18 patients (clinical results: 2 MR and 6 SD).