Exploring the immunomodulatory potential of Brahmi (Bacopa monnieri) in the treatment of invasive ductal carcinoma.

Bacopa monnieri (L) Wettst, commonly known as Brahmi, stands as a medicinal plant integral to India's traditional medical system, Ayurveda, where it is recognized as a "medhya rasayana"-a botanical entity believed to enhance intellect and mental clarity. Its significant role in numerous Ayurvedic formulations designed to address conditions such as anxiety, memory loss, impaired cognition, and diminished concentration underscores its prominence. Beyond its application in cognitive health, Brahmi has historically been employed in Ayurvedic practices for the treatment of inflammatory diseases, including arthritis. In contemporary biomedical research, Bacopa monnieri can attenuate the release of pro-inflammatory cytokines TNF-α and IL-6 in animal models. However, there remains a paucity of information regarding Bacopa's potential as an anticancer agent, warranting further investigation in this domain. Based on previous findings with Brahmi (Bacopa monnieri), the current study aims to find out the role of Brahmi plant preparation (BPP) in immunomodulatory actions on IDC. Employing a specific BPP concentration, we conducted a comprehensive study using MTT assay, ELISA, DNA methylation analysis, Western blotting, ChIP, and mRNA profiling to assess BPP's immunomodulatory properties. Our research finding showed the role of BPP in augmenting the action of T helper 1 (TH1) cells which secreted interferon-γ (IFN-γ) which in turn activated cytotoxic T-lymphocytes (CTL) to kill the cells of IDC (*p < 0.05). Moreover, we found out that treatment with BPP not only increased the activities of tumor-suppressor genes (p53 and BRCA1) but also decreased the activities of oncogenes (Notch1 and DNAPKcs) in IDC (*p < 0.05). BPP had an immense significance in controlling the epigenetic dysregulation in IDC through the downregulation of Histone demethylation & Histone deacetylation and upregulation of Histone methylation and Histone acetylation (*p < 0.05). Our Chromatin immunoprecipitation (ChIP)-qPCR data showed BPP treatment increased percentage enrichment of STAT1 & BRCA1 (*p < 0.05) and decreased percentage enrichment of STAT3, STAT5 & NF ΚB (*p < 0.05) on both TBX21 and BRCA1 gene loci in IDC. In addition, BPP treatment reduced the hypermethylation of the BRCA1-associated-DNA, which is believed to be a major factor in IDC (*p < 0.05). BPP not only escalates the secretion of type 1 specific cytokines but also escalates tumor suppression and harmonizes various epigenetic regulators and transcription factors associated with Signal Transducer and Activator of Transcription (STAT) to evoke tumor protective immunity in IDC.

Depletion of enhancer zeste homolog 2 (EZH2) directs transcription factors associated with T cell differentiation through epigenetic regulation of Yin Yang 1(YY1) in combating non-small cell lung cancer (NSCLC).

Non-Small Cell Lung Cancer (NSCLC) is the leading cause of death in all countries alike. In the current study, we have found out that Histone H3Lys4trimethylation is abnormal on YY1 in CD4+T Helper (TH) cells of NSCLC patients which is evident by Histone H3Lys27 trimethylation mediated via EZH2. We investigated the status of Yin Yang 1 (YY1) and the involvement of certain transcription factors that lead to tumorigenesis after depleting endogenous EZH2 in vitro by CRISPR/Cas9 in the CD4+TH1-or-TH2-polarized cells isolated initially as CD4+TH0 cells from the PBMC of the control subjects and patients suffering from NSCLC. After depletion of endogenous EZH2, RT-qPCR based mRNA expression analysis showed that there was an increase in the expression of TH1 specific genes and a decrease in the expression of TH2 specific genes in NSCLC patients CD4+TH cells. We can conclude that this group of NSCLC patients may have the tendency at least in vitro to elucidate adaptive/protective immunity through the depletion of endogenous EZH2 along with the reduction in the expression of YY1. Moreover, depletion of EZH2 not only suppressed the CD4+CD25+FOXP3+Regulatory T cells (Treg) but also it aided the generation of CD8+Cytotoxic T Lymphocytes (CTL) which were involved in killing of the NSCLC cells. Thus the transcription factors involved in EZH2 mediated T cell differentiation linked to malignancies offers us an appealing avenue of targeted therapeutic intervention for NSCLC.

Diospyros malabarica fruit preparation mediates immunotherapeutic modulation and epigenetic regulation to evoke protection against non-small cell lung cancer (NSCLC).

ETHNOPHARMACOLOGICAL RELEVANCE: Diospyros malabarica is an ethnomedicinal plant with hypoglycaemic, anti-bacterial, and anti-cancer properties and it belongs to the Ebenaceae family which is well known for its medicinal uses since ancient times and application of its bark and unripened fruit has been significantly mentioned in Ayurvedic texts. The Diospyros malabarica species which is known as the Gaub in Hindi and Indian Persimmon in English is native to India, however, it is distributed throughout the tropics. AIM OF THE STUDY: As Diospyros malabarica fruit preparation (DFP) possesses medicinal values, the study aims to evaluate its role as natural, non-toxic, and cost-effective dendritic cells (DCs) maturing immunomodulatory agent and also as an epigenetic regulator to combat Non-small cell lung cancer (NSCLC) which is a type of lung cancer whose treatment options such as chemotherapy, radiation therapy, etc. are accompanied with some adverse side effects. Thus, immunotherapeutic strategies are in high demand to evoke tumor protective immunity against NSCLC without causing such side effects. MATERIALS AND METHODS: Peripheral Mononuclear Cells (PBMCs) derived monocytes of normal subjects and NSCLC patients were utilized to generate DCs matured with either LPS (LPSDC) or DFP (DFPDC). Mixed Lymphocyte Reaction (MLR) was carried out with the differentially matured DCs co-culturing T cells and cytotoxicity of lung cancer cells (A549) was measured through LDH release assay and cytokine profiling was carried out via ELISA respectively. PBMCs of normal subjects and NSCLC patients have transfected separately in vitrowith CRISPR-activation plasmid of p53 and CRISPR-Cas9 knockout plasmid of c-Myc to analyze epigenetic mechanism(s) in the presence and absence of DFP. RESULTS: Diospyros malabarica fruit preparation (DFP) treated DC upregulates the secretion of T helper (TH)1 cell specific cytokines (IFN-γ and IL-12) and signal transducer and activator of transcription molecules (STAT1 and STAT4). Furthermore, it also downregulates the secretion of TH2-specific cytokines (IL-4 and IL-10). Diospyros malabarica fruit preparation (DFP) enhances p53 expression by reducing methylation levels at the CpG island of the promoter region. Upon c-Myc knockout, epigenetic markers such as H3K4Me3, p53, H3K14Ac, BRCA1, and WASp were enhanced whereas H3K27Me3, JMJD3, and NOTCH1 were downregulated. CONCLUSION: Diospyros malabarica fruit preparation (DFP) not only increases the expression of type 1 specific cytokines but also augments tumor suppression modulating various epigenetic markers to evoke tumor protective immunity without any toxic activities.

Neem Leaf Glycoprotein in immunoregulation of cancer.

Cancer is a disease having global consequences. Though several new strategies and treatments have been developed so far, they often come with malicious side effects and this paved ways for demand of naturally extracted/driven product as potent anti-cancer agent owing to their reduced toxicity and side effects. One such common Indian household plant Neem (Azadirachta Indica) and its extract have variegated immunomodulatory effects as anti-cancer agent. Neem Leaf Glycoprotein (NLGP) modifies immune cells present in the tumor surroundings as well as in the peripheral system, rather than directly attacking the cancer cells. NLGP acts as a natural immunomodulator showing several functions like sustained tumor growth regulation by stimulating central and effector memory cells as a vaccination adjuvant, normalization of angiogenic activities, controls hypoxia, improves immune evasion techniques as well as suppresses the activity of several immunological cells (Tregs, myeloid-derived suppressor cells, and tumor-associated macrophages) which promote tumor growth and metastasis in the tumor microenvironment (TME). NLGP prioritises type1 immune-microenvironment which consists of T-bet+IFN-γ-producing group 1 innate lymphoid cell (ILC) (ILC1 and natural killer cells), CD8+ cytotoxic T cells (TC1), and CD4+ T helper1 (Th1) cells. In this review we aim to summarize detailed activity of NLGP in cancer immunoregulation.