As traditional medicinal fungi, Ganoderma lucidum and Sanghuangporus vaninii are widely used in the treatment of tumor related diseases and cancer adjuvant therapy with potent anticancer effects. However, the anticancer effect and mechanism of action of their compound extract remain unclear.

To investigate the anticancer effect of Ganoderma lucidum and Sanghuangporus vaninii compound extract and explore the underlying mechanism.

First, MTT assay was performed to investigate the effect of 8 different extracts on tumor cell viability. Moreover, the synergistic effect of Ganoderma lucidum spore and Sanghuangporus vaninii was evaluated by Chou-Talalay method. Subsequently, the fractional extractions were conducted to further isolate anti-tumor active components. Next, network pharmacology combined with transcriptomics was used to explore the potential mechanisms underlying the anticancer effect of compound extract. Finally, the mechanism of action was verified using in vitro and in vivo models.

Among all 8 extracts, Ganoderma lucidum spore and Sanghuangporus vaninii compound ethanol extract (GSEE) showed the most significant cell viability inhibitory effect on cancer cells, especially colorectal cancer (CRC) cells, which was even better than combination of Sanghuangporus vaninii ethanol extract (SVEE) and Ganoderma lucidum spore ethanol extract (GLEE). The ethyl acetate fraction of GSEE (GSEAE) was screened as the anti-tumor active fraction of GSEE and could suppress CRC proliferation in vitro and in vivo. The CYP24A1-mediated Vitamin D receptor (VDR) pathway and TERT-mediated Wnt signaling pathway were identified as the main mechanisms of GSEAE against CRC. Multiple CRC models confirmed that GSEAE suppressed CRC metastasis, arrested cell cycle and induced mitochondrial apoptosis of CRC cells via VDR pathway and Wnt signaling pathway.

Collectively, our data suggest that compound extract GSEAE exerts anti-CRC effects via CYP24A1-mediated VDR pathway and TERT-mediated Wnt signaling pathway.

Vitamin D3 plays a crucial regulatory role in steroid hormone production, but the specific mechanism remains not fully understood. In this study, we investigated the expression and distribution patterns of Vitamin D receptor (VDR), vitamin D3 metabolic enzymes (CYP2R1, CYP27B1 and CYP24A1), mitochondrial dynamics and biogenesis (proteins and genes), and steroidogenic enzymes in the scent glands of muskrats during the breeding and non-breeding periods. VDR, vitamin D3 metabolic enzymes, mitochondrial dynamics and biogenesis-related proteins, and steroidogenic enzymes were immunolocalized in the scent glandular cells in both breeding and non-breeding seasons, with stronger immunostaining in the breeding season. The mRNA expression levels of Cyp27b1, Cyp24a1, Vdr, Mfn1, Opa1, Vdac, Tfam, Pgc1b, Star, Cyp11a1, Cyp17a1, and Cyp19a1 were higher in the scent glands during the breeding season than those of the non-breeding season. 1,25(OH)₂D₃ concentration were positively correlated with the mean mRNA expression levels of mitochondrial dynamics and biogenesis marker genes and steroidogenic enzymes in the scent glands. The concentrations of circulating testosterone (T) and 17β-estradiol (E2), and 1,25(OH)₂D₃ of the scent glands were also significantly higher in the breeding season. Additionally, the addition of 1,25(OH)2D3 to the primary scent glandular cells in vitro increased the expression levels of mitochondrial dynamics and biogenesis-related genes and steroidogenic enzymes in the scent glands of muskrats. These findings suggested that 1,25(OH)₂D₃ might promote the secretion of steroid hormones by upregulating the mitochondrial dynamics and biogenesis in the scent glands of muskrats.

The active form of vitamin D, calcitriol (1,25(OH)2D3), is produced from 25(OH)D3 via enzymes encoded by CYP2R1, CYP27A1, and CYP27B1. Polymorphisms in these genes may alter vitamin D metabolism and increase cardiovascular disease risk. This preliminary study investigated these polymorphisms in 27 patients with cardiovascular disease and 26 healthy volunteers using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP), while measuring 25(OH)D3 and 1,25(OH)2D3 concentrations by UPLC-MS/MS and ELISA, respectively. Among patients, those with the GT genotype of rs10877012 (CYP27B1) had higher 25(OH)D3 levels compared to other genotypes. Additionally, this polymorphism was associated with lower 1,25(OH)2D3 in TT homozygotes, suggesting reduced CYP27B1 activity. Furthermore, the TT genotype of rs6709815 (CYP27A1) was three times more prevalent in cardiac patients than in healthy controls, possibly indicating increased susceptibility to the disease. Although these findings suggest a genetic influence on vitamin D metabolism in cardiovascular disease, larger and more comprehensive studies are needed to confirm these associations.

Prostate cancer is one of the most common malignancies in men and represents a major challenge for modern medicine. In recent years, there has been growing interest in the role of vitamins in the pathogenesis of this disease, as well as their potential impact on prevention and supportive treatment. Studies conducted so far suggest that certain vitamins may exhibit antioxidant and anti-inflammatory properties, thus supporting the immune response, which may influence cancer risk and treatment efficacy. This article aims to present the current state of knowledge on the role of vitamins in prostate cancer, focusing on their potential importance both in prevention and as an adjunct to therapy. Vitamins A, D, E, C, and B-group compounds may influence prostate cancer development and progression through mechanisms such as antioxidant activity, regulation of cell proliferation, apoptosis, and immune modulation. Despite promising insights from basic research, clinical studies remain inconclusive, and the effects of vitamin supplementation depend on factors like dosage, form, and individual variability. Therefore, a balanced diet rich in natural vitamins is recommended, while further research is needed to clarify their therapeutic and preventive roles in prostate cancer.

Vitamin D plays a crucial role in the regulation of the immune system, with immunomodulatory effects that are key in the prevention of colorectal cancer (CRC). Over the past decades, research has shown that this steroid hormone impacts much more than bone health, significantly influencing immune responses. Vitamin D enhances immune organ functions such as the spleen and lymph nodes, and boosts T-cell activity, which is essential in defending the body against tumors. Additionally, vitamin D mitigates inflammatory responses closely linked to cancer development, reducing the inflammation that contributes to CRC. It acts via vitamin D receptors (VDRs) expressed on immune cells, modulating immune responses. Adequate vitamin D levels influence gene expression related to inflammation and cell proliferation, inhibiting tumor development. Vitamin D also activates mechanisms that suppress cancer cell survival, proliferation, migration, and metastasis. Low levels of vitamin D have been associated with an increased risk of CRC, with deficiency correlating with higher disease incidence. Lifestyle factors, such as a diet high in red meat and calories but low in fiber, fruits, and vegetables, as well as physical inactivity, contribute significantly to CRC risk. Insufficient calcium and vitamin D intake are also linked to disease occurrence and poorer clinical outcomes. Maintaining optimal vitamin D levels and adequate dietary intake is crucial in preventing CRC and improving patient prognosis. This review explores the role of vitamin D in immune regulation and summarizes findings from randomized clinical trials assessing the effects of vitamin D supplementation on CRC outcomes.

Achieving a pCR serves as a biomarker indicating enhanced overall survival for breast cancer patients undergoing NST. Vitamin D enhances the antitumor effect of chemotherapeutics as demonstrated in cancer cells and animal models. In this prospective randomized clinical study, we aim to investigate the effect of oral vitamin D supplementation during neoadjuvant systemic therapy (NST) on pathologic complete response (pCR).

Between June 2019 and June 2023, an oral form of 50,000 IU vitamin D3 (cholecalciferol) replacement was administered once a week during NST for the study group.

There were 114 (50.2%) cases in the study group and 113 (49.8%) in the control group (totally 227 cases). Factors that positively influenced pCR were higher clinical T stage, higher AJCC clinical stage, Estrogen receptor negativity, progesterone receptor negativity, human epidermal growth factor receptor 2 positivity, high Ki-67 expression (≥ 20%), hormone negative molecular subtypes, and vitamin D intake in univariate analysis. In the multivariate analysis, factors significantly affecting pCR were vitamin D intake (OR: 2.33, 95% CI 1.20-4.53; p = 0.013), hormone receptor negativity (OR: 2.22, 95% CI 1.11-4.43; p = 0.024), and Ki-67 ≥ 20% (OR: 3.27, 95% CI 1.03-10.34; p = 0.044).

This is the first and only study to compare the effect of oral vitamin D supplementation on pCR during NST. Vitamin D supplementation during NST has a significant effect on pCR in breast cancer patients. Although this effect is not significant for axillary pCR, there is an almost significant correlation.

ClinicalTrials.gov (Identifier: NCT03986268).

Pancreatic cancer remains one of the most aggressive malignancies with limited treatment options and poor survival rates. Vitamin D deficiency has been suggested to influence cancer progression and survival outcomes in various malignancies.

This study aimed to investigate the association between Vitamin D deficiency and disease progression as well as survival in patients diagnosed with pancreatic cancer.

A retrospective cohort study was conducted, including 151 patients diagnosed with pancreatic cancer between 2012 and 2022. Serum Vitamin D levels at the time of diagnosis were measured. Disease progression was evaluated through radiological assessments and clinical reports. Survival outcomes, including overall survival (OS) and progression-free survival (PFS), were analyzed using Kaplan-Meier survival curves and Cox proportional hazards regression models.

Of the 151 patients, 84 (56 %) were identified as Vitamin D deficient at the time of diagnosis. The deficient group exhibited a significantly higher frequency of advanced-stage disease (stages III and IV) compared to the non-deficient group (p < 0.05). During the follow-up period, 66 (78.6 %) of Vitamin d-deficient patients and 56 (84.8 %) of non-deficient patients experienced disease progression (p = 0.51). Moreover, Kaplan-Meier analysis showed a non-significant trend toward shorter median PFS (8.95 months vs. 9.27 months, p = 0.51) and OS (17.64 months vs. 19.05 months, p = 0.616) in the Vitamin d-deficient group.

Vitamin D deficiency is prevalent among patients with pancreatic cancer and appears to be associated with more advanced disease at diagnosis. Although a trend toward poorer survival outcomes was observed, the association between Vitamin D deficiency and OS/PFS did not reach statistical significance. Additional prospective studies are needed to confirm these findings and explore potential benefits of Vitamin D supplementation in pancreatic cancer management.

Pancreatic ductal adenocarcinoma (PDAC) remains one of the most feared diseases worldwide owing to its poor prognosis, negligible therapeutic advances, and high mortality. Herein, multifunctional enzyme-responsive micelles for the controlled delivery of paclitaxel (PTX) were prepared to circumvent its current clinical challenges. Accordingly, two enzyme-responsive structural units composed of Vitamin D3 (VD3) conjugated with polyethylene glycol of different molecular weights (600 Da and 2000 Da) were synthesized and characterized using different analytical methods. By applying the solvent evaporation method, these bioactive structural units self-assembled into sub-100 nm VD3 micelles with minimal batch-to-batch variation, monomodal particle size distribution, and high encapsulation efficiency. The enzyme-triggered disassembly of PTX-loaded VD3 micelles was demonstrated by release studies in the presence of a high esterase content typically featured by PDAC cells. PTX-loaded VD3 micelles also exhibited prominent cell internalization and induced a considerable cytotoxic synergistic effect against human PDAC cells (BxPC-3 cells) in 2D and 3D cell culture models compared with free PTX. The PTX-loaded VD3 micelles were hemocompatible and stable after long-term storage in the presence of biorelevant media, and showed higher efficiency to inhibit the tumor growth compared to the approved clinical nanoformulation (Abraxane®) in an in ovo tumor model. The findings reported here indicate that VD3S-PEG micelles may have a promising role in PDAC therapy, since VD3 could act not only as a hydrophobic core of the micelles but also as a therapeutic agent that provides synergetic therapeutic effects with the encapsulated PTX.

Background/Objectives: Vitamin D is classically associated with calcium and phosphate homeostasis, but recent research has expanded its role to include several new roles such as immune regulation, inflammation, and potential anti-cancer properties. The vitamin D receptor (VDR) is expressed in over 400 tissues, including those of the head and neck, implying a potential link between vitamin D and head and neck cancers (HNCs). Given the need for newer and better therapeutic approaches, this systematic review aims to synthesize existing clinical evidence on the relationship between vitamin D status and clinical outcomes in HNC patients. Methods and Results: A comprehensive literature search, across multiple databases including PubMed, Google Scholar and Science Direct, identified 187,642 studies related to vitamin D and cancer, from which 16 studies met the inclusion criteria. The inclusion criteria were English-language, full-text original research (2015-2025) on vitamin D's role in HNC progression and treatment, focusing on human studies. The findings indicate that vitamin D deficiency is highly prevalent among HNC patients, with rates ranging from 47% to 95%, particularly in advanced-stage cancers and those undergoing intensive treatment. Inverse association between vitamin D levels and HNC risk was reported, with higher serum 25(OH)D levels linked to a 30-32% reduction in cancer risk. Additionally, higher vitamin D levels correlated with improved survival rates and reduced recurrence, though some findings lacked statistical significance. Deficiencies were associated with higher rates of malnutrition and postoperative complications, reinforcing vitamin D's role in nutritional stability and surgical recovery. Conclusions: This systematic review highlights how common and significant vitamin D deficiency is among head and neck cancer (HNC) patients, exploring its possible role in cancer risk, prognosis, survival, treatment-related side effects, malnutrition, and post-surgical complications. The evidence suggests that while higher vitamin D levels are linked to better survival and fewer treatment-related issues, the benefits seem to level off beyond a certain point, indicating a more complex relationship. Additionally, vitamin D supplementation appears to help reduce chemoradiation side effects like mucositis, skin toxicity, dysphagia, and pain, ultimately improving patients' quality of life during treatment.

Vitamin D was discovered as the cure for nutritional rickets, a disease of bone growth arising from inadequate intestinal calcium absorption, and for much of the 20th century, it was studied for its critical role in calcium homeostasis. However, we now recognize that the vitamin D receptor and vitamin D metabolic enzymes are expressed in numerous tissues unrelated to calcium homeostasis. Notably, vitamin D signaling can induce cellular differentiation and cell cycle arrest. Moreover, the vitamin D receptor and the enzyme CYP27B1, which produces the hormonal form of vitamin D, 1,25-dihydroxyvitamin D (1,25D), are expressed throughout the immune system. In addition, CYP27B1 expression in immune cells is regulated by physiological inputs independent of those controlling its expression in calcium homeostatic tissues. These observations have driven the development of 1,25D-like secosteroidal analogs and nonsecosteroidal analogs to separate the effects of vitamin D on cell differentiation and function from its calcemic activities. Notably, some of these analogs have had considerable success in the clinic in the treatment of inflammatory and immune-related disorders. In this review, we described in detail the mechanisms of vitamin D signaling and the physiological signals controlling 1,25D synthesis and catabolism, with a focus on the immune system. We also surveyed the effects of 1,25D and its analogs on the regulation of immune system function and their implications for human immune-related disorders. Finally, we described the potential of vitamin D analogs as anticancer therapeutics, in particular, their use as adjuncts to cancer immunotherapy. SIGNIFICANCE STATEMENT: Vitamin D signaling is active in both the innate and adaptive arms of the immune system. Numerous vitamin D analogs, developed primarily to minimize the dose-limiting hypercalcemia of the active form of vitamin D, have been used widely in preclinical and clinical studies of immune system regulation. This review presents a description of the mechanisms of action of vitamin D signaling, an overview of analog development, and an in-depth discussion of the immunoregulatory roles of vitamin D analogs.

Numerous studies over the past century have reported an inverse correlation between lifetime solar ultraviolet radiation (UV) exposure and all-cancer incidence and mortality. For decades, this relationship was hypothesized to reflect the action of photosynthesized vitamin D, though subsequent clinical trials have failed to demonstrate the expected anti-cancer properties. Rather than a consequence of vitamin D, I hypothesize that this inverse correlation reflects the immune stimulatory action of UV-derived skin neoantigens. Over time, such UV-mediated immune education drives immune repertoire diversification and superior adaptive immune responses to infectious disease and cancer. This hypothesis would explain the strong positive selection for light skin pigmentation following the out-of-Africa migration among humans inhabiting northerly latitude regions, and the longstanding racial disparities in cancer and infectious disease observed in North America. It suggests that the skin comprises an important reservoir of anti-cancer T cells that may be harnessed for anti-cancer therapy, and that skin mutation burden (SMB) may serve as a predictive biomarker of immunotherapy response. I propose a novel, non-invasive method for quantifying SMB as a biomarker.

Colitis-associated colorectal cancer (CAC), a serious complication of ulcerative colitis (UC), is associated with a poor prognosis. The vitamin D receptor (VDR) is recognized for its protective role in UC and CAC through the maintenance of intestinal barrier integrity and the regulation of inflammation. This study demonstrates a significant reduction in m6A-related genes, particularly methyltransferase like 14 (METTL14), in UC and CAC patients and identifies an association between METTL14 and VDR. In the azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced mouse model, vitamin D treatment increases METTL14 expression and reduces tumor burden, while Vdr-knockout mice exhibit lower METTL14 levels and increased tumorigenesis. In vitro, the VDR agonist calcipotriol upregulates METTL14 in NCM460 cells, with this effect attenuated by VDR knockdown. VDR knockdown in DLD-1 colon cancer cells decreases METTL14 expression and promotes proliferation, which is reversed by METTL14 overexpression. Mechanistic studies reveal that VDR regulates METTL14 expression via promoter binding, modulating key target genes such as SOX4, DROSH, and PHLPP2. This study highlights the role of the VDR-METTL14 axis as a protective mechanism in CAC and suggests its potential as a therapeutic target for preventing and treating CAC.

The incorporation of silicon bioisosteres into pharmacological structures has been used as a strategy to improve the therapeutic potential of drugs. However, no secosteroidal silicon-containing VDR ligands have been developed. Here we report the design, synthesis, and biological activity of six analogues of the natural hormone 1,25-dihydroxyvitamin D3 (1,25D3), which incorporate a silicon atom as a side chain-C25 isostere. The analogues were synthesized by the Wittig-Horner approach starting from Inhoffen-Lythgoe diol. The crystal structures of the complexes formed by the sila-analogues with the ligand binding domain of VDR revealed additional interactions of the sila-containing side chains that stabilize the VDR active conformation. These sila-analogues show similar VDR binding and transcriptional activity in comparison with the natural hormone 1,25D3, but with significantly less hypercalcemic activity. The new analogues, when combined with chemotherapy, significantly decrease cell proliferation.

The aim of this research was to design and synthesize new lipid conjugates of 7-DHC that could serve as a new storage form of esterified provitamin D3, increasing the reservoir of this biomolecule in the epidermis and enabling controlled production of vitamin D3 even during periods of sunlight deficiency. Acylglycerol and glycerophospholipid containing succinate-linked provitamin D3 at the sn-2 position of the glycerol backbone were synthesized from dihydroxyacetone (DHA) and sn-glycerophosphocholine (GPC), respectively. The three-step synthesis of 1,3-dipalmitoyl-2-(7-dehydrocholesterylsuccinoyl)glycerol involved the esterification of DHA with palmitic acid, reduction of the carbonyl group, and conjugation of the resulting 1,3-dipalmitoylglycerol with 7-dehydrocholesterol hemisuccinate (7-DHC HS). The use of NaBH3CN as a reducing agent was crucial to avoid acyl migration and achieve the final product with 100% regioisomeric purity. For the preparation of 1-palmitoyl-2-(7-dehydrocholesterylsuccinoyl)-sn-glycero-3-phosphocholine, a two-step process was applied, involving the esterification of GPC at the sn-1 position with palmitic acid, followed by the conjugation of 1-palmitoyl-sn-glycero-3-phosphocholine with 7-DHC HS. Alongside the main product, a small amount of its regioisomer with provitamin D3 linked at the sn-1 position and palmitic acid at the sn-2 position was detected, indicating acyl migration from the sn-1 to the sn-2 position in the intermediate 1-palmitoyl-sn-glycerophosphocholine. The synthesized novel lipids were fully characterized using spectroscopic methods. They can find applications as novel lipid-based prodrugs as additives to sunscreen creams.

Vitamin D (Vit D) has gained significant attention in health research recently as a result of its potential protective effects against various cellular damages. This study aimed to investigate the ability of vitamin D to mitigate deoxyribonucleic acid (DNA) fragmentation in liver cells and bone marrow cytotoxicity induced by chloramphenicol (CAP). Sixty male albino mice were divided into six groups: control, chloramphenicol-treated (250 and 500 mg/kg body weight, 5 days per week for 4 weeks), vitamin D-treated (800 IU/kg body weight, 5 days per week for 4 weeks) and vitamin D plus chloramphenicol-treated groups. Results of DNA fragmentation test revealed that oral treatment with low and high doses of CAP significantly increased the frequency of DNA fragmentation in liver cells in comparison with the control, whereas oral treatment with vitamin D alone or plus low and high doses of chloramphenicol significantly reduced the genotoxicity in liver cells in comparison with the control group. Micronucleus analysis showed that CAP treatment at low and high doses significantly increased micronuclei formation and cytotoxicity in bone marrow cells. However, vitamin D significantly reduced the micronuclei formation in bone marrow cells of mice treated with chloramphenicol. Vitamin D alone showed no significant difference in the frequency of micronuclei and bone marrow cytotoxicity compared to the control group. Accordingly, further research exploring the mechanisms underlying the protective effects of vitamin D and investigating optimal dosing regimens is warranted. Also, clinical studies evaluating the efficacy of vitamin D supplementation to mitigate the adverse effects of chloramphenicol in human patients are recommended.

Multiple sclerosis (MS) is a demyelinating and inflammatory disease of the central nervous system (CNS) in need of a curative treatment. MS research has recently focused on the development of pro-remyelinating treatments and neuroprotective therapies. Here, we aimed at favoring remyelination and reducing neuro-inflammation in a cuprizone mouse model of brain demyelination using nanomedicines. We have selected lipid nanocapsules (LNC) coated with the cell-penetrating peptide transactivator of translation (TAT), loaded with either a pro-remyelinating compound, calcitriol (Cal-LNC TAT), or an anti-inflammatory bioactive lipid, prostaglandin D2-glycerol ester (PGD2-G) (PGD2-G-LNC TAT). Following the characterization of these formulations, we showed that Cal-LNC TAT in combination with PGD2-G-LNC TAT increased the mRNA expression of oligodendrocyte differentiation markers both in the CG-4 cell line and in primary mixed glial cell (MGC) cultures. However, while the combination of Cal-LNC TAT and PGD2-G-LNC TAT showed promising results in vitro, no significant impact, in terms of remyelination, astrogliosis, and microgliosis, was observed in vivo in the corpus callosum of cuprizone-treated mice following intranasal administration. Thus, although calcitriol's beneficial effects have been abundantly described in the literature in the context of MS, here, we show that the different doses of calcitriol tested had a negative impact on the mice well-being and showed no beneficial effect in the cuprizone model in terms of remyelination and neuro-inflammation, alone and when combined with PGD2-G-LNC TAT.

Several studies have indicated that 1α,25-hydroxyvitamin D [1α,25(OH)2D3] inhibits the proliferation and metastasis of cancer cells through suppressing epithelial-mesenchymal transition. However, its influence on the translocation of β-catenin remains unclear. In the present study, ovarian cancer stem-like cells (CSCs), including side population (SP) and CD44+/CD117+, were isolated from mouse ovarian surface epithelial (MOSE) cells with malignant transformation. The findings revealed that 1α,25(OH)2D3 obviously reduced the sphere-forming ability, as well as Notch1 and Klf levels. Moreover, the limiting dilution assay demonstrated that 1α,25(OH)2D3 effectively hindered the tumorigenesis of ovarian CSCs in vitro. Notably, treatment with 1α,25(OH)2D3 led to a substantial increase in the cell population of CD44+/CD117+ forming one tumor from ≤ 100 to 445 in orthotopic transplanted model, indicating a pronounced suppression of stemness of ovarian CSCs. Additionally, 1α,25(OH)2D3 robustly promoted the translocation of β-catenin from the nuclear to the cytoplasm through directly binding to VDR, which resulted in decreased levels of c-Myc and CyclinD1 within late MOSE cells. Taken together, these results strongly supported the role of 1α,25(OH)2D3 in inhibiting stem-like properties in ovarian cancer cells by restraining nuclear translocation of β-catenin, thereby offering a promising target for cancer therapeutics.

Childhood acute lymphoblastic leukemia (ALL) is a common pediatric cancer. The heterogeneous characterization of B cells in ALL progression poses new challenges to researchers. We used single-cell sequencing to explore the critical role of B cells in regulating the ALL immune microenvironment.

We collected the single cell (sc) RNA-seq data of ALL and health sample from the gene expression omnibus (GEO) database, the "Seurat" and "harmony" R package was used for quality control and scRNA-seq analysis, in which the CellMarker2.0 database was used for cell type annotation. Subsequently, the FindAllMarkers function was used to identify the differentially expressed genes (DEGs) among various cell types and the DAVID database was applied for the biological process of DEGs. Then, the "inferCNV" package was used for copy number variation, regulons and cell communication were performed by SCENIC tool and CellChat package. The role of the target gene in regulating ALL progression was assessed using RT-qPCR, Transwell and scratch healing assays.

We identified nine mainly cell clusters after scRNA-seq analysis, in which the B cells had higher infiltration proportion in the ALL samples and were sub-clustered into five cell sub-groups. The B cells 1 is closely associated with cell proliferation and stemness (TNFAIP3 and KDM5B), and the significant CNV of amplification occurred on chr6 and chr21 that supported stemness of B cells1. RXRB is a key transcription factor mediated the proliferation of B cells 1, which in turn suppressed hematopoietic stem cells (HSCs) proliferation and promoted cytotoxic NK/T cells activation through diverse cell communication ways. One of the key regulators of B cells is MYC, which promotes the migration and invasive ability of cell line leukemia cell lines.

This study reveals the stemness characteristics of B cells and their critical role in ALL progression, a finding that provides new potential directions for the development of targeted therapies against ALL.

Liver fibrosis is characterized by the excessive accumulation of extracellular matrix proteins, which can lead to cirrhosis and liver cancer. Metabolic dysfunction-associated steatosis liver diseases are common causes of liver fibrosis, sharing a similar pathogenesis with carbon tetrachloride (CCl₄) exposure. This process involves the activation of hepatic stellate cells (HSCs) into myofibroblasts. However, the detailed mechanism and effective treatment strategies require further investigation. In this study, we uncovered a negative correlation between VDR expression and YAP within HSCs. Subsequently, we demonstrated that VDR exerted a downregulatory influence on YAP transcriptional activity in HSCs. Intriguingly, activation VDR effectively inhibited the culture induced activation of primary HSCs by suppressing the transcriptional activity of early YAP. Furthermore, in vivo results manifested that hepatic-specific deletion of YAP/TAZ ameliorates CCl4-induced liver fibrosis, and nullified the antifibrotic efficacy of VDR. Importantly, a YAP inhibitor rescued the exacerbation of liver fibrosis induced by hepatic-specific VDR knockout. Moreover, the combined pharmacological of VDR agonist and YAP inhibitor demonstrated a synergistic effect in diminishing CCl4-induced liver fibrosis, primary HSCs activation and hepatic injury in vivo. These effects were underpinned by their collective ability to inhibit HSC activation through AMPK activation, consequently curbing ATP synthesis and HSCs proliferation. In conclusion, our results not only revealed the inhibition of VDR on YAP-activated liver stellate cells but also identified a synergistic effect of VDR agonist and YAP inhibitor in an AMPKα-dependent manner, providing a practical foundation for integration of multi-targeted drugs in the therapy of CCl4-induced hepatic fibrosis.

Vitamin D3, which originates from cholesterol, exerts its influence on immune cells and potentially cancer cells via the metabolite 1,25-dihydroxycholecalciferol (1,25(OH)2D3), impacting their proliferation, differentiation, and apoptosis. An umbrella review was conducted to evaluate the potential protective effect of vitamin D3 intake and serum levels on the incidence and mortality of cancer.

A systematic search was conducted in MEDLINE, Cochrane Central Register of Controlled Trials, and EMBASE databases from their inception to October 1, 2023. We included meta-analyses of observational or randomized clinical trials that compared interventions (vitamin D3 intake) or blood levels in a healthy population, with cancer incidence or mortality as outcomes. The grading of evidence certainty followed established criteria, including strong, highly suggestive, suggestive, weak, or not significant.

A total of 71 systematic reviews were included. Strong evidence indicated that vitamin D3 supplementation reduced total cancer mortality (odds ratio [OR], 0.9 [95% CI, 0.87-0.92]; P < 0.01). In the context of site-specific cancers, there exists highly suggestive evidence pointing towards the potential prevention of head and neck, breast, colorectal, lung, and renal cell cancers through the intake of vitamin D3. Furthermore, strong evidence suggests that maintaining sufficient levels of vitamin D3 may effectively lower the risk of renal cell and thyroid cancer (OR = 0.76 [95%CI 0.64-0.88]).

There is significant evidence that vitamin D3 intake may reduce the incidence of some cancers. Routine assessments to ensure sufficient levels of vitamin D3 and administering supplements to address deficiencies may serve as crucial preventive measures for healthcare systems.

Inhibition of human carcinomas has previously been linked to vitamin D due to its effects on cancer cell proliferation, migration, angiogenesis, and apoptosis induction. The anticancer activity of vitamin D has been confirmed by several studies, which have shown that increased cancer incidence is associated with decreased vitamin D and that dietary supplementation of vitamin D slows down the growth of xenografted tumors in mice. Vitamin D inhibits the growth of cancer cells by the induction of apoptosis as well as by arresting the cells at the G0/G1 (or) G2/M phase of the cell cycle. Aim and Key Scientific Concepts of the Review: The purpose of this article is to thoroughly review the existing information and discuss and debate to conclude whether vitamin D could be used as an agent to prevent/treat cancers. The existing empirical data have demonstrated that vitamin D can also work in the absence of vitamin D receptors (VDRs), indicating the presence of multiple mechanisms of action for this sunshine vitamin. Polymorphism in the VDR is known to play a key role in tumor cell metastasis and drug resistance. Although there is evidence that vitamin D has both therapeutic and cancer-preventive properties, numerous uncertainties and concerns regarding its use in cancer treatment still exist. These include (a) increased calcium levels in individuals receiving therapeutic doses of vitamin D to suppress the growth of cancer cells; (b) hyperglycemia induction in certain vitamin D-treated study participants; (c) a dearth of evidence showing preventive or therapeutic benefits of cancer in clinical trials; (d) very weak support from proof-of-principle studies; and (e) the inability of vitamin D alone to treat advanced cancers. Addressing these concerns, more potent and less toxic vitamin D analogs have been created, and these are presently undergoing clinical trial evaluation. To provide key information regarding the functions of vitamin D and VDRs, this review provided details of significant advancements in the functional analysis of vitamin D and its analogs and VDR polymorphisms associated with cancers.

Colorectal Cancer (CRC) has been molecularly classified into several subtypes according to tumor, stromal, and immune components. Here, we investigated whether the preventive effect of vitamin D on CRC varies with subtypes defined by Vitamin D receptor (VDR) expression in tumors and their surrounding stroma, along with the association of somatic mutations in CRC.

In a population-based prospective study of 22,743 Japanese participants, VDR expression levels in tumors and their surrounding stroma were defined in 507 cases of newly diagnosed CRC using immunohistochemistry. Hazard ratios of CRC and its subtypes according to dietary vitamin D intake were estimated using multivariable Cox proportional hazards models.

Dietary vitamin D intake was not associated with CRC or its subtypes defined by VDR expression in tumors. However, an inverse association was observed for CRC with high VDR expression in the stroma (the highest tertile vs the lowest tertile: 0.46 [0.23-0.94], Ptrend = 0.03), but not for CRC with low VDR expression in the stroma (Pheterogeneity = 0.02). Furthermore, CRC with high VDR expression in the stroma had more somatic TP53 and BRAF mutations and fewer APC mutations than those with low VDR expression in the stroma.

This study provides the first evidence that the preventive effect of vitamin D on CRC depends on VDR expression in the stroma rather than in the tumors. CRC with high VDR expression in the stroma is likely to develop through a part of the serrated polyp pathway, which tends to occur with BRAF but not with APC mutations.

The assessment of sarcoma during clinical practice is primarily based on imaging examination, with no effective biomarkers available. Although it has been established that 1,25(OH)2D3 is abnormally expressed in patients with sarcoma, it remains unclear whether 1,25(OH)2D3 level could be used as an evaluation marker in these patient population.

This real-world study investigated 1,25(OH)2D3 level and its association with clinical features in sarcoma patients. Data on 1,25(OH)2D3, parathyroid hormone, calcium, and calcitonin were collected from 331 patients with sarcoma, while the imaging results and the variation in 1,25(OH)2D3 among 213 patients with sarcoma before and after treatment was further analyzed.

We found that the serum 1,25(OH)2D3 level was predominantly decreased in patients with sarcoma, with a mean of 45.68 nmol/L. 1,25(OH)2D3 was significantly correlated with the gender and age of sarcoma patients, with more substantial reductions in women and younger patients. Among sarcoma patients, those with progressive disease exhibited a 7.08 nmol/L (-13.73%) decrease in serum 1,25(OH)2D3 levels compared to baseline, while patients with non-progressive disease showed a 1.11 nmol/L (+ 7.0%) increase.

The variation of serum 1,25(OH)2D3 can predict the disease status of patients with sarcoma. Decreased serum 1,25(OH)2D3 levels are indicative of disease progression in sarcoma patients, suggesting its potential for application as a prognostic marker for disease assessment in this patient population.

This study examined the link between serum 25-hydroxyvitamin D (25(OH)D) and mortality in patients with colon cancer. Using a clinical database from the University of California, serum 25(OH)D measures were averaged for the time following diagnosis until either the time of death or 5 years had elapsed. Analytical methods included the use of Generalized Additive Models (GAM), logistic regression, and Cox proportional hazards models to examine non-linear relationships and the impact of 25(OH)D on 5-year mortality. This study assessed 1,602 patients with colon cancer having a median 25(OH)D of 31.8 ng/mL and a 5-year mortality rate of 22.7%. A significant association between higher post-diagnosis vitamin D levels and decreased 5-year mortality was observed. This association persisted after adjusting for disease severity and significant demographic confounders, in both a logistic regression model for 5-year mortality (OR = 0.79, 95% CI: 0.66-0.92, p < 0.001) and a cox proportional hazards model for survival (HR = 0.94, CI: 0.88-0.99, p = 0.048). GAM illustrated a steep increase in survival probability up to a plateau, suggesting a threshold effect beyond roughly 50.0 ng/mL. This study found a potential protective role of vitamin D in the survival of colon cancer patients, supporting the correction of levels below 25 ng/mL but ideally above 50 ng/mL.

1,25(OH)2D3 is a fat-soluble vitamin, involved in regulating Ca2+ homeostasis in the body. Its storage in adipose tissue depends on the fat content of the body. Obesity is the result of abnormal lipid deposition due to the prolonged positive energy balance and increases the risk of several cancer types. Furthermore, it has been associated with vitamin D deficiency and defined as a low 25(OH)2D3 blood level. In addition, 1,25(OH)2D3 plays vital roles in Ca2+-Pi and glucose metabolism in the adipocytes of obese individuals and regulates the expressions of adipogenesis-associated genes in mature adipocytes.

The present contribution focused on the VDR mediated mechanisms interconnecting the obese condition and cancer proliferation due to 1,25(OH)2D3-deficiency in humans. This contribution also summarizes the identification and development of molecular targets for VDR-targeted drug discovery.

Several studies have revealed that cancer development in a background of 1,25(OH)2D3 deficient obesity involves the VDR gene. Moreover, 1,25(OH)2D3 is also known to influence several cellular processes, including differentiation, proliferation, and adhesion. The multifaceted physiology of obesity has improved our understanding of the cancer therapeutic targets. However, currently available anti-cancer drugs are notorious for their side effects, which have raised safety issues. Thus, there is interest in developing 1,25(OH)2D3-based therapies without any side effects.

This retrospective cohort study explores the relationship between vitamin D levels and 5-year mortality risk among 1,549 colon cancer patients seen at University of California health centers between 2012 and 2019, with a particular focus on the mediating role of comorbidities. Methods leveraged structural equation modeling to assess both direct and indirect pathways linking vitamin D to mortality risk. This analysis revealed a protective direct effect of higher vitamin D levels against mortality risk. Additionally, this study uncovered an indirect pathway, demonstrating that vitamin D lowers mortality risk by mitigating comorbidity, which subsequently influence mortality risk. Study results indicate that approximately 9.2% of the beneficial effect of vitamin D on mortality risk is attributable to its capacity to reduce comorbidity burden. In disaggregated and confounder-adjusted structural modeling, there were significant indirect effects for 25(OH)D on mortality risk through its effects on depression and obesity but not on anxiety, diabetes, or chronic kidney disease. These results suggest that the protective effects of vitamin D in colon cancer etiology appear to be through direct action on cancer progression, though patients who also suffer from depression and obesity would especially benefit from achieving adequate levels of serum vitamin D.

This study aimed to evaluate the prevalence and risk of malignant neoplasm in primary hyperparathyroidism (PHPT) patients. Potentially eligible studies were retrieved from PubMed and Embase databases from inception to November 2023 using search strategy consisting of terms for "Primary hyperparathyroidism" and "Malignant neoplasm". Eligible study must report prevalence of malignant neoplasm among patients with PHPT or compare the risk of malignant neoplasm between patients with PHPT and comparators. Point estimates with standard errors were extracted from each study and combined using the generic inverse variance method.A total of 11,926 articles were identified. After two rounds of systematic review, 50 studies were included. The meta-analysis revealed that pooled prevalence rates of overall cancer was 0.19 (95%CI: 0.13-0.25; I2 94%). The two most prevalent types of malignancy among patients with PHPT ware papillary thyroid cancer (pooled prevalence: 0.07; 95%CI: 0.06-0.08; I2 85%) and breast cancer (pooled prevalence: 0.05; 95%CI: 0.03-0.07; I2 87%). Subgroup analysis of studies focusing on patients undergoing parathyroidectomy reported a fourfold higher prevalence of papillary thyroid cancer than the remaining studies (0.08 versus 0.02). The meta-analysis of cohort studies found a significant association between PHPT and overall cancer with the pooled risk ratio of 1.28 (95%CI: 1.23-1.33; I2 66.9%).We found that the pooled prevalence of malignant neoplasm in PHPT was 19%, with papillary thyroid cancer and breast cancer being the most prevalent types. The meta-analysis of cohort studies showed that patient with PHPT carried an approximately 28% increased risk of malignancy.

Melanoma, originating through malignant transformation of melanin-producing melanocytes, is a formidable malignancy, characterized by local invasiveness, recurrence, early metastasis, resistance to therapy, and a high mortality rate. This review discusses etiologic and risk factors for melanoma, diagnostic and prognostic tools, including recent advances in molecular biology, omics, and bioinformatics, and provides an overview of its therapy. Since the incidence of melanoma is rising and mortality remains unacceptably high, we discuss its inherent properties, including melanogenesis, that make this disease resilient to treatment and propose to use AI to solve the above complex and multidimensional problems. We provide an overview on vitamin D and its anticancerogenic properties, and report recent advances in this field that can provide solutions for the prevention and/or therapy of melanoma. Experimental papers and clinicopathological studies on the role of vitamin D status and signaling pathways initiated by its active metabolites in melanoma prognosis and therapy are reviewed. We conclude that vitamin D signaling, defined by specific nuclear receptors and selective activation by specific vitamin D hydroxyderivatives, can provide a benefit for new or existing therapeutic approaches. We propose to target vitamin D signaling with the use of computational biology and AI tools to provide a solution to the melanoma problem.

Colorectal cancer (CRC) is one of the most life-threatening neoplasias in terms of incidence and mortality worldwide. Vitamin D deficiency has been associated with an increased risk of CRC. 1α,25-Dihydroxyvitamin D3 [1,25(OH)2D3], the most active vitamin D metabolite, is a pleiotropic hormone that, through its binding to a transcription factor of the nuclear receptor superfamily, is a major regulator of the human genome. 1,25(OH)2D3 acts on colon carcinoma and stromal cells and displays tumor protective actions. Here, we review the variety of molecular mechanisms underlying the effects of 1,25(OH)2D3 in CRC, which affect multiple processes that are dysregulated during tumor initiation and progression. Additionally, we discuss the epidemiological data that associate vitamin D deficiency and CRC, and the most relevant randomized controlled trials of vitamin D3 supplementation conducted in both healthy individuals and CRC patients.

The causes of infertility have remained an important challenge. The relationship between VDR gene polymorphisms and infertility has been reported, with controversial findings.

We aimed to determine this relationship by conducting a systematic review and meta-analysis.

The study was started with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) declaration and the final draft was registered as a protocol in PROSPERO (ID: CRD42023416535). The international electronic databases including PubMed (Medline), Scopus, Web of Sciences, and Cumulative Index to Nursing and Allied Health Literature (CINHAL) were searched until January 30, 2023, by using appropriate keywords. The quality of the final studies was assessed using the NOS Checklist for case-control studies. The odds ratios (ORs) for each of the genetic models were pooled, and a subgroup analysis based on geographical region and types of infertility was carried out by the MetaGenyo online tool.

Case-control studies including 18 and 2 studies about infertility in women and men, respectively, and 4 miscarriage studies were entered into the meta-analysis. The VDR gene TaqI polymorphism was associated with infertility susceptibility in women in the allele contrast [OR = 1.2065, 95% CI (1.0846-1.3421); P = 0.0005], Recessive model [OR = 1.3836, 95% CI (1.1197-1.7096); P = 0.002], Dominant model [OR = 1.2146, 95% CI (0.0484-1.4072); P = 0.009], Homozygote [OR = 1.4596, 95% CI (1.1627-1.8325); P = 0.001], and TT vs. Tt [OR = 1.2853, 95% CI (1.0249-1.6117); P = 0.029. ApaI and FokI gene polymorphisms were found to be significantly protective SNPs against women and men infertility in the Dominant model [OR = 0.8379, 95% CI (0.7039- 0.9975); P = 0.046] and Recessive model [OR = 0.421, 95% CI (0.1821-0.9767); P = 0.043], respectively. Sub-group meta-analysis showed a protection association of ApaI in dominant [OR = 0.7738, 95% CI = 0.6249-0.9580; P = 0.018] and AA vs. aa [OR = 0.7404, 95 CI% (0.5860-0.9353) P = 0.011725] models in PCOS subgroup, however, a negative association with idiopathic infertility was found in AA vs. Aa [OR = 1.7063, 95% CI (1.1039-2.6375); P = 0.016187] and Aa vs. aa [OR = 0.6069, 95% CI (0.3761-0.9792); P = 0.040754]. TaqI SNP was significantly associated with infertility in the African population and BsmI was associated with the disease mostly in the Asian population.

This meta-analysis showed that the TaqI polymorphism may be linked to women's infertility susceptibility. However, ApaI and FokI might be the protective SNPs against infertility in Women and men, respectively.

Vitamin D binding protein (DBP) might increase substantially after ovarian stimulation and hence could be associated with IVF/ICSI outcomes because it determines the fraction of free bioavailable 25(OH) vitamin D. In this study, we aim to determine whether DBP is associated with E2 level after ovarian stimulation and IVF/ICSI outcomes.

Post-hoc analysis of a prospective observational cohort.

Single-center study.

2569 women receiving embryo transfer.

None.

The main outcomes were oocyte and embryo quality as well as pregnancy outcomes.

DBP concentration correlates with E2 on hCG day (=day of inducing ovulation with hCG; correlation coefficient r = 0.118, P<0.001) and E2 x-fold change to baseline level (r = 0.108, P<0.001). DBP is also positively correlated with total 25(OH)D (r = 0.689, R2 = 0.475, P<0.001) and inversely with free 25(OH)D (r=-0.424, R2=0.179, P<0.001), meaning that E2-stimulated DBP synthesis results in a decrease of free 25(OH)D during ovarian stimulation. However, such alteration does not affect IVF/ICSI outcomes when considering confounding factors, such as the number and quality of oocytes nor embryo quality as well as pregnancy outcomes.

DBP concentration correlates with the degree of E2 increase after ovarian stimulation. DBP is also positively correlated with total 25(OH)D and inversely with free 25(OH)D, suggesting that the proportion of free 25(OH)D decreases during ovarian stimulation caused by E2-stimulated DBP synthesis. However, such alteration does not affect clinical IVF/ICSI outcomes.

Vitamin D3 is a fat-soluble secosteroid predominantly synthesized in the skin or delivered with a diet. Nevertheless, recently it is considered more as a hormone than a vitamin due to its pleiotropic function within the organism ensured by widely distributed vitamin D receptors and metabolic enzymes. Besides the main role in calcium and phosphorus homeostasis, vitamin D3 was shown to regulate many cellular and metabolic processes in normal and cancerous tissues within the immune system, the cardiovascular system, the respiratory system and the endocrine system. The ovary is an important extraskeletal tissue of vitamin D3 action and local metabolism, indicating its role in the regulation of ovarian functions upon physiological and pathological conditions. This chapter reviews firstly the updated information about vitamin D3 metabolism and triggered intracellular pathways. Furthermore, the basic information about ovarian physiology and several aspects of vitamin D3 effects within the ovary are presented. Finally, the special attention is paid into possible mechanism of vitamin D3 action within ovarian pathologies such as premature ovarian failure, polycystic ovary syndrome, and ovarian cancer, considering its clinical application as alternative therapy.

Vitamin D is a fat-soluble steroid hormone that was initially known only for regulating calcium and phosphorus levels and maintaining bone health. However, it was later discovered that many organs express vitamin D metabolizing enzymes and have a ligand for vitamin D, which regulates the expression of an extensive assortment of genes. As a result, vitamin D is indispensable for the proper function of organs, and its deficiency is believed to be a critical factor in symptoms and disorders such as cardiovascular diseases, autoimmune diseases, and cancers. The significance of vitamin D in reproductive tissues was recognized later, and studies have revealed its crucial role in male and female fertility, as well as proper reproductive function during pregnancy. Vitamin D deficiency has been identified as a risk factor for infertility, gonadal cancers, pregnancy complications, polycystic ovary syndrome, and endometriosis. However, data investigating the association between vitamin D levels and reproductive disorders, including endometriosis, have encountered inconsistencies. Therefore, the present study aims to review existing research on the effect of vitamin D on proper reproductive function, and the role of deficiency in reproductive diseases and specifically focuses on endometriosis.

A role for vitamin D in immune modulation and in cancer has been suggested. In this work, we report that mice with increased availability of vitamin D display greater immune-dependent resistance to transplantable cancers and augmented responses to checkpoint blockade immunotherapies. Similarly, in humans, vitamin D-induced genes correlate with improved responses to immune checkpoint inhibitor treatment as well as with immunity to cancer and increased overall survival. In mice, resistance is attributable to the activity of vitamin D on intestinal epithelial cells, which alters microbiome composition in favor of Bacteroides fragilis, which positively regulates cancer immunity. Our findings indicate a previously unappreciated connection between vitamin D, microbial commensal communities, and immune responses to cancer. Collectively, they highlight vitamin D levels as a potential determinant of cancer immunity and immunotherapy success.

Vitamin D is a promising anticancer agent for the prevention and treatment of several cancers, including melanoma. Low 25-hydroxyvitamin D levels, a routinely used marker for vitamin D, have been suggested as one of the factors in the development and progression of melanoma. The parent vitamin D needs activation by cytochrome P450 (CYP) enzymes to exert its actions via the vitamin D receptor (VDR). This review discusses the role of vitamin D in melanoma and how CYP-mediated metabolism can potentially affect the actions of vitamin D. Through interacting with the retinoid X receptor, VDR signaling leads to anti-inflammatory, antioxidative, and anticancer actions. Calcitriol, the dihydroxylated form of vitamin D3, is the most active and potent ligand of VDR. CYP27A1, CYP27B1, and CYP2R1 are involved in the activation of vitamin D, whereas CYP24A1 and CYP3A4 are responsible for the degradation of the active vitamin D. CYP24A1, the primary catabolic enzyme of calcitriol, is overexpressed in melanoma tissues and cells. Several drug classes and natural health products can modulate vitamin D-related CYP enzymes and eventually cause lower levels of vitamin D and its active metabolites in tissues. Although the role of vitamin D in the development of melanoma is yet to be fully elucidated, it has been proposed that melanoma prevention may be significantly aided by increased vitamin D signaling. Furthermore, selective targeting of the catabolic enzymes responsible for vitamin D degradation could be a plausible strategy in melanoma therapy. Vitamin D signaling can be improved by utilizing dietary supplements or by modulating CYP metabolism. A positive association exists between the intake of vitamin D supplements and improved prognosis for melanoma patients. Further investigation is required to determine the function of vitamin D supplementation and specific enzyme targeting in the prevention of melanoma.

The use of doxorubicin (Dox) in the treatment of breast cancer negatively affects the intestines and other tissues. Many studies have proven that probiotics and vitamin D3 have antitumor and intestinal tissue-protecting properties. To achieve effectiveness and minimize side effects, the current study aims to administer Dox together with probiotics (Lactobacillus acidophilus and Lactobacillus casei) and vitamin D3. Forty-two female BALB/c inbred mice were divided into six groups: Group 1 (Control), Group 2 (Dox), Group 3 (Dox and probiotics), Group 4 (Dox and vitamin D3), Group 5 (Dox, probiotics, and vitamin D3), and Group 6 (probiotics and vitamin D3). The 4T1 mouse carcinoma cell line was injected into the mammary fat pad of each mouse. Gene expression was examined using quantitative real-time PCR. The treated groups (except group 6) showed significantly reduced tumor volume and weight compared to the control group (P < 0.05, P < 0.01). Probiotics/vitamin D3 with Dox reduced chemotherapy toxicity and a combination of supplements had a significant protective effect against Dox (P < 0.05, 0.01, 0.001). The treated groups (except 6) had significantly higher expression of Bax/Caspase 3 genes and lower expression of Bcl-2 genes than the control group (P < 0.05, 0.01). Coadministration of Dox with probiotics and vitamin D3 showed promising results in reducing tumor size, protecting intestinal tissue and influencing gene expression, suggesting a strategy to enhance the effectiveness of breast cancer treatment while reducing side effects.