Dacarbazine

Role of Cytochrome P450 Enzymes (CYP1A1, CYP1A2, CYP2E1) in N-Demethylation

The N-demethylation of this compound, a crucial step in its activation, is catalyzed by specific cytochrome P450 isoforms. Studies have shown that CYP1A1, CYP1A2, and CYP2E1 are involved in this process in human liver microsomes. nih.govaacrjournals.org CYP1A2 is considered the predominant enzyme catalyzing hepatic this compound metabolism, while CYP2E1 contributes at higher substrate concentrations. nih.govaacrjournals.org CYP1A1 may play a role in extrahepatic metabolism of this compound. nih.govaacrjournals.org

Data on the kinetics of this compound N-demethylation by recombinant human CYP enzymes indicate varying efficiencies:

Note: Vmax values are reported differently across sources; both are included where available. nih.govaacrjournals.org

Inhibition studies using chemical inhibitors and antibodies have further supported the role of these CYP isoforms in this compound N-demethylation. nih.govaacrjournals.org

Formation of Active Metabolites: Diazomethane and Methyl Diazonium Ion

Following N-demethylation catalyzed by CYP enzymes, this compound is converted to 5-[3-hydroxymethyl-3-methyl-triazen-1-yl]-imidazole-4-carboxamide (HMMTIC). mdpi.comresearchgate.net HMMTIC is unstable and loses formaldehyde to form 5-[3-methyl-triazen-1-yl]-imidazole-4-carboxamide (MTIC). mdpi.comresearchgate.net MTIC is a key intermediate that rapidly decomposes to yield 5-aminoimidazole-4-carboxamide (AIC) and a highly reactive methylating species, which is believed to be the methyl diazonium ion (or diazomethane hydroxide). patsnap.comnih.govresearchgate.net The methyl diazonium ion is considered the primary cytotoxic species responsible for DNA alkylation. pharmacology2000.compatsnap.com

Generation of 5-aminoimidazole-4-carboxamide (AIC) as a Metabolite

5-aminoimidazole-4-carboxamide (AIC) is a major metabolite of this compound and is excreted in the urine. pharmacology2000.comdrugbank.com AIC is formed from the breakdown of MTIC. nih.govresearchgate.net Studies using radioactive labeling have confirmed that AIC is derived from the this compound molecule. hemonc.org While AIC is a significant metabolite, it does not possess chemotherapeutic activity. nih.gov

Alkylation at O6 and N7 Positions of Guanine

A major mechanism of this compound's action involves the alkylation of guanine residues in DNA. patsnap.comcuni.cz The primary sites of methylation are the N7 and O6 positions of guanine. patsnap.comnih.gov While N7-methylguanine is the most frequently formed adduct, O6-methylguanine is considered particularly important for the cytotoxic and mutagenic effects of this compound. researchgate.netnih.gov Alkylation at the O6 position can lead to mispairing with thymine during DNA replication, potentially causing G:C to A:T transitions. patsnap.comcuni.cz

Induction of DNA Cross-Linking and Double Strand Breaks

This compound is known to alkylate and cross-link DNA. selleckchem.comrnceus.com The formation of DNA adducts, particularly O6-methylguanine, can trigger the DNA mismatch repair system. patsnap.com Persistent O6-methylguanine lesions can overwhelm the repair system, leading to futile cycling and the generation of DNA double-strand breaks. patsnap.comnih.gov These double-strand breaks are a significant form of cytotoxic damage that impedes cancer cell proliferation and can induce apoptosis. selleckchem.comfrontiersin.org

Disruption of DNA Replication and Transcription

The primary mechanism by which MTIC exerts its cytotoxic effect is through the formation of methylcarbonium ions. patsnap.combccancer.bc.ca These highly reactive ions attack nucleophilic groups in DNA, leading to alkylation. patsnap.combccancer.bc.ca Alkylation of DNA disrupts the normal structure and function of the DNA helix, interfering with crucial cellular processes such as DNA replication and transcription. patsnap.comcsic.esresearchgate.net

Specific sites of alkylation include the O6 and N7 positions of guanine residues in DNA. patsnap.com The formation of O6-methylguanine is considered a major cytotoxic lesion induced by MTIC. patsnap.comnih.gov This modification can lead to mispairing of guanine with thymine during DNA replication, triggering the DNA mismatch repair (MMR) system. patsnap.com Persistent O6-methylguanine lesions can overwhelm the MMR system, resulting in futile repair cycles and ultimately activating cell death pathways, including apoptosis. patsnap.com While N7-alkylation of guanine is a more frequent event, O6-alkylation is generally considered more cytotoxic. nih.gov

The presence of DNA adducts formed by alkylation interferes with the DNA replication machinery, causing errors in DNA synthesis and leading to cell cycle arrest. patsnap.com This disruption of DNA function is a critical factor in the antineoplastic activity of this compound.

Inhibition of DNA Synthesis via Purine Analog Activity

Role of O6-methylguanine-DNA methyltransferase (MGMT)

O6-methylguanine-DNA methyltransferase (MGMT), also known as methylguanine methyltransferase, is a key enzyme that repairs DNA damage, specifically the methylation at the O6 position of guanine. nih.govfrontiersin.org This methylation is a primary site of this compound-induced DNA adduction. patsnap.com MGMT removes the alkyl group from O6-methylguanine by transferring it to an internal cysteine residue within the enzyme, in a stoichiometric and auto-inactivating reaction. nih.gov High levels of MGMT in both normal and tumor cells are associated with resistance to triazenes, including this compound. patsnap.comnih.gov Consequently, tumors with low MGMT activity are generally more responsive to this compound treatment. patsnap.com Studies have shown a correlation between the methylation status of the MGMT promoter and sensitivity to this compound, with cells having a methylated (and thus silenced) MGMT promoter being more sensitive to the drug. researchgate.netsemanticscholar.org

Impact of Dysregulated DNA Repair in Tumor Cells

Dysregulated DNA repair mechanisms are a hallmark of cancer and can significantly impact the efficacy of DNA-damaging agents like this compound. patsnap.comfrontiersin.org While dysregulation can sometimes lead to increased susceptibility to DNA-damaging drugs, enhanced activity of certain repair pathways can also promote resistance. frontiersin.orgnih.gov For instance, increased expression of DNA repair enzymes is a mechanism of resistance to this compound. patsnap.com The mismatch repair (MMR) system, which typically corrects base mismatches and insertion-deletion loops, is also involved in the susceptibility of cells to triazenes. nih.gov However, rather than suppressing the cytotoxic effects, the MMR system can promote them by attempting to repair the O6-methylguanine lesions, leading to futile cycling and activation of cell death pathways when overwhelmed. patsnap.comnih.gov The base excision repair (BER) pathway also plays a role in repairing this compound-induced DNA damage, and robust BER activities have been associated with resistance. nih.gov

Apoptosis Induction

Apoptosis, or programmed cell death, is a primary modality of cell death induced by this compound and other DNA-alkylating agents. researchgate.netptfarm.pl this compound-induced DNA alkylation triggers the intrinsic apoptosis pathway. researchgate.net Studies have demonstrated that this compound treatment leads to typical characteristics of apoptotic cell death, including nuclear condensation and fragmentation, as well as mitochondrial fragmentation. researchgate.netajmb.org Flow cytometry assays have indicated an accumulation of treated cells in the sub-G1 phase of the cell cycle, which is indicative of apoptosis. ajmb.orgnih.gov Activation of apoptosis has been confirmed by techniques such as Annexin V staining and the detection of increased levels of active caspase-3 and cleaved poly (ADP-ribose) polymerase (PARP). nih.govnih.gov The intrinsic apoptotic pathway is mediated by mitochondria and is typically activated by intracellular signals such as DNA damage. nih.gov The release of cytochrome c is an indicator of the intrinsic pathway activation. nih.gov

Data from studies investigating the effects of this compound on apoptosis:

Necrotic Cell Death

While apoptosis is a primary mode of cell death induced by this compound, necrotic cell death can also occur. nih.govptfarm.pl Necrosis is characterized by different morphological changes compared to apoptosis, including loss of cell membrane integrity. ptfarm.pl Studies using staining methods like acridine orange/propodium iodide (AO/PI) can differentiate between apoptotic and necrotic cells based on their staining characteristics. ajmb.orgajmb.org Higher doses of this compound have been observed to cause an intensification of the necrotic process in some cell lines. ptfarm.pl

Cell Cycle Delay at G2/M Phase

This compound treatment can induce cell cycle arrest, particularly at the G2/M phase. aacrjournals.orgmdpi.comptfarm.pl This arrest is a consequence of the DNA damage caused by the drug's alkylating activity. patsnap.commdpi.com The formation of DNA adducts interferes with the proper progression of the cell cycle, halting cells before they can enter mitosis (M phase) from the G2 phase. patsnap.comaacrjournals.org Studies have shown that this compound can cause cell arrest in both the S and G2/M phases in melanoma cell lines, with higher concentrations often leading to a more pronounced block in the G2/M phase. ptfarm.pl The integrity of the G2/M cell-cycle checkpoint may be important for the cytotoxicity of alkylating agents like this compound. aacrjournals.org

Reactive Oxygen Species (ROS) Formation

A key aspect of this compound-induced oxidative stress is the formation of reactive oxygen species (ROS). sbmu.ac.irnih.govresearchgate.net Studies using isolated rat hepatocytes have demonstrated that the addition of this compound results in the generation of ROS, such as hydrogen peroxide (H₂O₂) and superoxide anion radicals (O₂•⁻), before cytotoxicity is observed. nih.gov This ROS generation appears to be involved in the mechanism of this compound-induced cytotoxicity. sbmu.ac.irnih.gov Antioxidants and ROS scavengers have been shown to prevent this compound-induced cytotoxicity and ROS formation in these studies. sbmu.ac.irnih.govresearchgate.net It is suggested that H₂O₂ can cross the lysosomal membrane and react with lysosomal Fe²⁺ to form hydroxyl radicals (•OH) via the Haber-Weiss reaction, leading to lysosomal membrane disruption and the release of digestive enzymes, ultimately causing cell death. sbmu.ac.irnih.gov

Upregulation of Interleukin-8 (IL-8) and Vascular Endothelial Growth Factor (VEGF)

Research has indicated that this compound can cause the transcriptional up-regulation of Interleukin-8 (IL-8) and Vascular Endothelial Growth Factor (VEGF) in melanoma cells. aacrjournals.orgnih.gov This leads to the overexpression and secretion of these proteins. aacrjournals.orgnih.gov Studies using melanoma cell lines like SB-2 and MeWo have shown that this compound treatment increases luciferase activity driven by the IL-8 and VEGF promoters, indicating increased gene transcription. aacrjournals.orgnih.gov For instance, luciferase activity driven by the IL-8 and VEGF promoters was up-regulated by 1.5–2-fold and 1.6–3.5-fold, respectively, in SB2 and MeWo melanoma cell lines. aacrjournals.orgnih.gov The mitogen-activated protein kinase (MAPK) pathway appears to partially regulate IL-8 activation, while it is not involved in VEGF promoter regulation. aacrjournals.orgnih.gov Electrophoretic mobility shift analysis (EMSA) has revealed increased binding activity of activator protein 1 (AP-1, specifically c-Jun) and nuclear factor-kappaB (NF-κB) after this compound treatment, suggesting their involvement in the transcriptional up-regulation of these cytokines. aacrjournals.org Elevated levels of IL-8 and VEGF secretion have been associated with increased resistance to this compound in melanoma cell lines. aacrjournals.orgnih.gov

Data Tables

Based on the research findings discussed, here are examples of static data representations:

Table 1: Fold Up-regulation of IL-8 and VEGF Promoter Activity by this compound in Melanoma Cell Lines

Note: Data derived from research findings on transcriptional up-regulation. aacrjournals.orgnih.gov

Table 2: Effect of this compound Concentration on Cell Cycle Arrest in Mouse Melanoma Cells

Note: Data derived from research findings on cell cycle arrest. ptfarm.pl

Autocrine IL-8 Cytokine Stimulation

Autocrine stimulation by the cytokine Interleukin-8 (IL-8) has been implicated in the re-activation of the MAPK pathway, contributing to resistance in this compound-conditioned BRAF V600E cells. researchgate.netnih.gov Elevated levels of IL-8 mRNA have been observed in cell populations desensitized to this compound. nih.gov

Alternative MAPK Signaling via COT and CRAF

Resistance to this compound can also emerge through the activation of alternative MAPK signaling pathways involving COT (also known as MAP3K8) and CRAF (also known as RAF1). researchgate.netnih.gov Overexpression of COT and CRAF can lead to the reactivation of MEK/ERK signaling, even in the presence of BRAF inhibition. ascopubs.orgmdpi.com While the exact mechanisms for the increase in CRAF expression or the switch to ARAF or CRAF are not fully known, this alternative signaling contributes to drug resistance. openaccessjournals.com

IL8/PDGFRβ-Dependent Bypass Signaling via AKT

Desensitization to this compound, particularly in melanoma cells that have developed resistance to BRAF inhibitors, can be caused by canonical MAPK-independent survival signaling. researchgate.netnih.gov This bypass signaling is dependent on IL-8 and PDGFRβ (Platelet-Derived Growth Factor Receptor beta) and proceeds via the AKT pathway. researchgate.netresearchgate.netnih.gov Elevated expression of both IL-8 and PDGFRβ has been noted in resistant cells, leading to the hyperactivation of the PI3K/AKT pathway, which promotes cell survival. researchgate.netnih.govspandidos-publications.commdpi.com

Induction of NKG2D Ligand Expression on Melanoma Cells

This compound treatment elicits the expression of ligands for the immunoreceptor NKG2D on melanoma cells. researchgate.netnih.govresearchgate.netnih.gov NKG2D ligands are typically poorly expressed on normal cells but are upregulated by tumor cells or virus-infected cells. nih.gov This upregulation can be triggered or enhanced by DNA-damaging agents like this compound, a process regulated by DNA-damage checkpoint pathways such as those initiated by ATM and ATR protein kinases. nih.govfrontiersin.org Studies have shown that this compound is capable of triggering or enhancing the expression of NKG2D ligands on both murine and human tumor cells. nih.gov

Activation of Natural Killer (NK) Cells

Engagement of NKG2D on NK cells by its ligands on tumor cells leads to the activation of NK cells. researchgate.netnih.govresearchgate.netnih.gov This activation allows for enhanced tumor-cell killing. researchgate.net Studies in mouse models have shown that NK cells are indispensable for the therapeutic effect of this compound. researchgate.net NK cell activation mechanisms, metabolism, and proliferative capacity can be dysregulated by the tumor microenvironment, but the induction of NKG2D ligands by this compound helps promote NK cell cytotoxicity. mdpi.com

Release of Interferon-gamma (IFN-γ)

Activation of NK cells by this compound-induced NKG2D ligands leads to the release of Interferon-gamma (IFN-γ). researchgate.netnih.govresearchgate.netnih.gov IFN-γ is a crucial cytokine in the anti-tumor immune response. Serum IFNγ levels have been observed to increase during therapy, particularly after the addition of this compound in combination treatments. nih.govtandfonline.com This induction of IFNγ-stimulated genes correlating with increased serum IFNγ has been predictive of better clinical outcomes in some studies. nih.govtandfonline.com

Upregulation of Major Histocompatibility Complex Class I (MHC Class I)

IFN-γ released by activated NK cells in turn upregulates Major Histocompatibility Complex Class I (MHC Class I) expression on tumor cells. researchgate.netnih.govresearchgate.netnih.gov MHC Class I molecules are essential for presenting tumor antigens to T lymphocytes. frontiersin.org Upregulation of MHC Class I expression on tumor cells is necessary for their recognition by T cells. researchgate.net this compound may condition for the success of immunotherapy by upregulating MHC-I expression. researchgate.net

Enhanced Recognition by Cytotoxic CD8+ T Lymphocytes (CTLs)

The upregulation of MHC Class I expression on tumor cells, facilitated by IFN-γ release, favors their recognition by cytotoxic CD8+ T lymphocytes (CTLs). researchgate.netnih.govresearchgate.netnih.gov This increased antigen presentation makes tumor cells more susceptible to lysis by tumor-specific CD8+ T cells. nih.govnih.gov Studies have indicated that this compound can sensitize tumor cells to CD8+ T-cell mediated pathways. researchgate.net The antitumor effect of this compound has been shown to rely on CD8+ T cells in mouse models. researchgate.net Furthermore, this compound treatment before peptide vaccination has been shown to enlarge the T-cell repertoire diversity of tumor-reactive CD8+ T cells in melanoma patients, accompanied by high avidity and tumor reactivity. aacrjournals.orgnih.govnih.gov

Here is a summary of the key immunomodulatory effects of this compound:

Cytogenetic Assays: Chromosomal Aberrations and Micronuclei

This compound has been shown to induce genotoxic damage in vitro. Studies using human lymphocytes have demonstrated a significant increase in both micronucleus and chromosomal aberration frequency following exposure to this compound. tandfonline.com Micronuclei formation is considered an indicator of genetic toxicity. nih.gov Chromosomal aberrations observed include various types of anomalies. tandfonline.com These findings indicate that this compound can interact with the genetic material of normal cells, in addition to targeting cancer cells. tandfonline.com

Cell Viability, Proliferation, and Apoptosis Assays

The effects of this compound on cell viability, proliferation, and apoptosis have been investigated in various cancer cell lines, including melanoma cells. This compound has been shown to inhibit the growth of melanoma cell lines in a dose-dependent manner. core.ac.ukeuropeanreview.org Studies using MTT assays have quantified the reduction in cell viability upon treatment with this compound. sums.ac.irsums.ac.irthieme-connect.comnih.goviiarjournals.org

This compound can induce apoptosis in melanoma cells. tandfonline.comsums.ac.irsums.ac.irthieme-connect.comnih.gov For instance, studies have shown that this compound treatment at various concentrations caused apoptosis in CD117+ melanoma cells. thieme-connect.com The combination of this compound with other agents, such as metformin or PTD4-apoptin protein, has been shown to significantly increase apoptosis compared to this compound alone in melanoma cell lines. core.ac.uksums.ac.irsums.ac.irnih.govresearchgate.net

Interactive Table 1: Effect of this compound and Metformin on B16-F10 Cell Viability

Data derived from observations in Source sums.ac.irsums.ac.ir. Specific numerical data points varied with dose and time in the source.

Interactive Table 2: Apoptosis Induction by this compound and Combination Therapy in B16-F10 Cells

Data derived from observations in Source nih.govresearchgate.net. Specific numerical data points for this compound alone were not precisely quantified in the snippet.

Effects on Nuclear Division Index

The Nuclear Division Index (NDI) is a measure used in cytogenetic studies to assess cell cycle progression. Studies have shown that this compound can cause a delay in the cell cycle, which is reflected in the Nuclear Division Index, particularly at higher concentrations. tandfonline.com This indicates that this compound interferes with the normal progression of the cell cycle in exposed cells.

Modulation of Gene and Protein Expression

This compound has been shown to modulate the expression of various genes and proteins in cancer cells. Studies in melanoma cell lines have indicated that this compound treatment can transcriptionally induce the expression of Interleukin 8 (IL-8) and Vascular Endothelial Growth Factor (VEGF). aacrjournals.org This induction has been suggested as a potential mechanism of escape from chemotherapy in melanoma cells. aacrjournals.org The activation of the MAPK pathway, specifically increased activity of ERK, has been observed following this compound treatment, which may be responsible for the upregulation of IL-8. aacrjournals.org

Furthermore, this compound has been found to up-regulate the expression of miRNA-200 family members in melanoma cells, which is associated with the inhibition of cell proliferation and induction of apoptosis. europeanreview.org this compound treatment has also been shown to modulate the levels of apoptosis-related proteins such as Bim, Bak, BAX, and Bad. europeanreview.org In human melanoma A375 cells, this compound treatment influenced the mRNA levels of genes associated with reactive oxygen species (ROS), such as SOD1 and CAT. spandidos-publications.com It also affected the expression of vitamin D-associated genes like VDR and CYP family members. spandidos-publications.com

Silencing of Dicer, an enzyme involved in miRNA processing, has been shown to enhance this compound resistance in melanoma cells by inhibiting the expression of ADSL (Adenylosuccinate lyase). aging-us.com Dicer silencing was correlated with increased mRNA and protein expression of cancer stem cell-related transcription factors including Oct-4, Nanog, KLF4, and SOX2. aging-us.com

Mouse Models of Melanoma and Other Cancers

Mouse models, particularly those involving melanoma xenografts or syngeneic melanoma cells, have been widely used to study the in vivo effects of this compound. In a xenograft model using a human melanoma cell line, nanoemulsion formulations of this compound showed greater efficacy in reducing tumor size compared to suspension preparations. acs.org Topical application of nanoemulsions resulted in up to 10-fold greater percentage reductions in tumor size. acs.org Intramuscular injection of the nanoemulsion also showed comparable efficacy. acs.org

Studies in B16-F10 mouse melanoma models have investigated the antineoplastic effects of this compound, including its immunogenic properties. tandfonline.com In this model, the antineoplastic effects of this compound in vivo were found to be dependent on the immune system, specifically on natural killer (NK) cells and CD8+ T cells. tandfonline.com this compound was observed to upregulate the expression of NKG2D ligands on tumor cells, leading to the activation of these immune cells. tandfonline.com

Combination therapies involving this compound have also been evaluated in mouse models. The combination of celecoxib and this compound demonstrated much greater effectiveness than either drug alone in inhibiting both melanoma growth and metastasis in B16-F10 mouse models. nih.govresearchgate.net

Interactive Table 3: Median Survival Rates in Melanoma Mouse Model

Data derived from Source nih.govresearchgate.net.

Interactive Table 4: Average Number of Metastatic Tumors in Murine Lungs

Data derived from Source nih.govresearchgate.net.

In a dedifferentiated solitary fibrous tumor (SFT) xenograft model in SCID mice, this compound showed high antitumor activity, resulting in approximately 95% tumor volume inhibition. aacrjournals.orgresearchgate.netnih.gov This activity was comparable to that of temozolomide in this model. aacrjournals.orgresearchgate.netnih.gov No tumor regrowth was observed up to 100 days after the end of treatment with this compound in this model, suggesting a cytotoxic effect. aacrjournals.orgresearchgate.netnih.gov

Studies have also investigated the genotoxic effects of this compound in vivo in mice. This compound induced micronuclei in bone marrow cells, with the frequency increasing linearly with dose in the range of 0-125 mg/kg. lifesciencesite.comnih.govoup.com It also induced chromosomal aberrations in bone marrow cells. lifesciencesite.com

The combination of this compound with PTD4-apoptin protein in a mouse B16-F1 melanoma model showed a significantly increased antitumor effect compared to single treatments. core.ac.uk A combined treatment with a reduced dose of this compound and PTD4-apoptin revealed similar antitumor activities to the full dose of this compound alone. core.ac.uk

While in vitro studies showed additive effects between this compound and imexon in human A375 melanoma cells, the combination was not effective in reducing tumor growth in an in vivo mouse model. iiarjournals.orgiiarjournals.org

Assessment of Genotoxicity and Mutagenicity

Preclinical studies have consistently shown that this compound possesses genotoxic potential. It has been demonstrated to be mutagenic in bacteria and has produced evidence of both clastogenicity (causing structural chromosome changes) and mutagenicity in mammalian cells in vitro pfizer.com. Furthermore, this compound has shown clastogenicity in mouse bone marrow in vivo pfizer.com. These findings are consistent with its mechanism of action as a methylating cytostatic drug pfizer.com. Due to its pharmacological properties, this compound shows mutagenic, carcinogenic, and teratogenic effects detectable in experimental test systems hpra.iemedac.eu.

Tumor Growth and Metastasis Inhibition

Research has investigated this compound's ability to inhibit tumor growth and metastasis, often in combination with other agents. Studies in mice with metastatic melanoma have shown that cotreatment with this compound and statins significantly inhibited tumor growth and metastasis researchgate.net. This inhibition was observed to occur via the suppression of the RhoA/RhoC/LIM domain kinase/serum response factor/c-Fos pathway and enhanced expression of p53, p21, p27, cleaved caspase-3, and cleaved poly(ADP-ribose) polymerase 1 in vivo researchgate.net. This cotreatment also significantly improved the survival rate in metastasis-bearing mice researchgate.net. For instance, treatment with this compound plus 100 mg/kg simvastatin or fluvastatin prevented metastasis-associated death in a percentage of mice researchgate.net.

Another study evaluating the combination of this compound with the pro-oxidant imexon in human A375 melanoma cells found additive growth inhibition in vitro. However, this combination was not effective in reducing human A375 melanoma tumors in vivo iiarjournals.org.

In a Lewis lung carcinoma (3LL) model in mice, treatment with this compound alone or in combination with cyclophosphamide significantly reduced tumor growth and the number of metastases nih.gov. These effects were linked to changes in the biochemical and immunological properties of the treated 3LL cells, including reduced α6 integrin expression, increased susceptibility to natural immunity in vivo, and increased immunogenicity nih.gov.

Immunomodulatory Profiling in Vivo

This compound has been explored for its potential immunomodulatory effects in vivo. Preclinical studies suggest that many chemotherapeutic drugs, including this compound, can exert immunostimulatory effects researchgate.net. This can occur by inhibiting immunosuppressive cells, activating effector cells, or increasing immunogenicity and T-cell infiltration researchgate.net.

Research indicates that this compound can induce local activation of natural killer (NK) and T cells researchgate.net. This is supported by findings that this compound treatment elicits the expression of ligands for the immunoreceptor NKG2D on melanoma cells researchgate.net. The engagement of these ligands by NKG2D on NK cells leads to their activation, resulting in enhanced tumor-cell killing and the release of IFN-γ researchgate.net. Subsequently, IFN-γ upregulates major histocompatibility complex class I expression on tumor cells, which favors their recognition by cytotoxic CD8+ T lymphocytes (CTLs) researchgate.net.

A study investigating the immunomodulatory effects of sorafenib and this compound on melanoma in patients found that sorafenib had a direct immunomodulatory effect on lymphocytes and an indirect effect on the tumor microenvironment, helping to reduce the immunosuppressive network amegroups.org. Future studies are encouraged to focus on the immunomodulatory effects of this compound on the tumor microenvironment, which could potentially restore sensitivity to immune checkpoint-based therapy researchgate.net.

Nanoparticle-Based Delivery Systems

Nanoparticle-based delivery systems have emerged as a promising approach for delivering this compound. These systems can potentially increase solubility, stability, and bioavailability, decrease toxicities, and provide site-specific drug delivery frontiersin.org.

Studies have explored various nanoparticle platforms for this compound delivery:

Stearic acid nanostructured nanoparticles: A cream formulation with this compound-loaded stearic acid nanostructured nanoparticles showed an encapsulation efficiency of around 70% and spherical particles between 10 and 20 nm in size that could penetrate cancer cells frontiersin.org. This formulation demonstrated a higher rate of drug release compared to a drug suspension frontiersin.org.

Nanoemulsions: this compound nanoemulsions have been investigated for topical application in mouse xenograft models of human melanoma frontiersin.orgresearchgate.net. A nanoemulsion formulation was found to reduce tumor size in a xenograft mouse epidermoid carcinoma model compared to a this compound suspension researchgate.net.

Polymeric nanoparticles: Biodegradable polymeric nanoparticles, such as those based on methoxy poly(ethylene glycol)-poly(lactide) (MPEG-PLA), have been developed for controlled delivery of this compound internationalscholarsjournals.org. These nanoparticles, with a size of around 144.2 ± 7.8 nm, showed an encapsulation efficiency of 70.1 ± 2.3% and were suitable for vascular administration internationalscholarsjournals.org. In vitro and in vivo studies demonstrated that DTIC-loaded MPEG-PLA NPs could induce more apoptosis in cancer cells and showed enhanced antitumor activity compared to free DTIC internationalscholarsjournals.org. Star-shaped block polymer nanoparticles based on Cholic acid-poly(lactide-co-glycolide)-b-polyethylene glycol (CA-PLGA-b-PEG) have also been constructed for this compound encapsulation and targeted therapy of malignant melanoma researchgate.netnih.gov.

Mesoporous silica nanoparticles (MSNPs): MSNPs have been utilized as drug delivery vehicles for various anticancer therapeutics, including this compound japsonline.com.

Nanosponges: this compound-loaded nanosponges incorporated into a hydrogel have been developed for targeted melanoma treatment researchgate.netresearcher.life. Optimized nanosponges showed controlled particle size (338.6 nm) and high entrapment efficiency (92.2%) researchgate.net.

These nanoparticle systems aim to improve the therapeutic index of this compound by enhancing its delivery to tumor sites while minimizing exposure to healthy tissues nih.gov.

Targeted Drug Delivery

Targeted drug delivery strategies aim to deliver this compound specifically to cancer cells, increasing its efficacy and reducing systemic toxicity researchgate.netnih.gov. This is often achieved by functionalizing nanoparticles with ligands that bind to receptors overexpressed on cancer cells nih.govmdpi.com.

For instance, DTIC-loaded CA-PLGA-b-PEG nanoparticles have been modified with the nucleic acid aptamer AS1411 to achieve active targeted therapy of malignant melanoma researchgate.netnih.gov. These targeted nanoparticles demonstrated enhanced cytotoxicity to melanoma cells in vitro and excellent anti-tumor effects in vivo with no obvious side effects researchgate.netnih.gov.

Mesoporous silica nanoparticles functionalized with aptamers are also being explored to facilitate targeted drug delivery by specifically binding to target molecules on the surface of cancer cells japsonline.com.

Controlled Release Mechanisms

Controlled release mechanisms are being incorporated into novel this compound formulations to maintain therapeutic drug levels at the tumor site over an extended period and minimize systemic exposure researchgate.netmdpi.com.

Polymeric nanoparticles, such as MPEG-PLA NPs, have demonstrated a sustained release profile for this compound internationalscholarsjournals.org. The release pattern was biphasic, with an initial burst release followed by a slower, continuous release internationalscholarsjournals.org. Approximately 43.3% of DTIC was released within 24 hours, and nearly 90% was released in 7 days internationalscholarsjournals.org.

Metal-organic frameworks (MOFs), such as MIL-100(Fe), have also been investigated for their potential in controlling the release of this compound researchgate.net. Coating the surface of MIL-100(Fe) with polyethylene glycol (PEGylation) significantly affected controlled drug release, extending the complete release time researchgate.net.

Injectable hydrogel formulations with high viscosity can also prolong drug retention time and achieve slow release compared to other carriers mdpi.com. This compound-loaded nanosponges incorporated into a hydrogel base using HPMC showed sustained drug release (82% over 12 hours) and effective skin permeation in ex vivo studies researchgate.net.

These controlled release strategies, often combined with targeted delivery, aim to optimize the therapeutic window of this compound and improve patient outcomes researchgate.netmdpi.com.

6.3.4. Overcoming Solubility and Stability Challenges

This compound (DTIC) is an important chemotherapeutic agent, but its clinical application is often hampered by inherent physicochemical limitations, namely low aqueous solubility and significant instability. These challenges necessitate the development of strategies to enhance its biopharmaceutical profile during preclinical and translational research.

This compound exhibits low and pH-dependent aqueous solubility, making formulation for administration challenging. researchgate.netacs.orgnih.govnih.gov It is also known to be photosensitive, undergoing rapid degradation when exposed to light. researchgate.netacs.orgnih.govnih.govacs.org The primary degradation product is 2-azahypoxanthine, which is considered inactive and potentially linked to adverse effects. researchgate.netacs.orgnih.govnih.gov Degradation kinetics are influenced by factors such as light exposure, pH, and temperature. researchgate.netnih.gov Acid hydrolysis can also contribute to its degradation. neuroquantology.com

To address these limitations, various preclinical strategies have been explored to improve this compound's solubility and stability:

Modification of Solid Forms: Research has focused on developing novel solid forms, such as salts and cocrystals, through crystal engineering techniques. By reacting this compound with aliphatic carboxylic acids, new multicomponent crystals have been successfully prepared, including salts, cocrystals, and salt-cocrystals. acs.orgnih.gov These new solid forms have demonstrated substantial enhancements in solubility and dissolution rates compared to the native drug. Studies have reported up to a 19-fold increase in solubility and intrinsic dissolution rates that are 1.3 to 22-fold faster. acs.orgnih.gov This improved dissolution behavior contributes positively to reducing the photodegradation of this compound in solution. acs.orgnih.gov

Inclusion Complexes with Cyclodextrins: The formation of inclusion complexes with cyclodextrins, particularly β-cyclodextrin (βCD), has been investigated as a method to increase this compound's aqueous solubility and stability. researchgate.netnih.govresearchgate.netmdpi.commdpi.com this compound can be included within the non-polar internal cavity of βCD, forming inclusion complexes. mdpi.com This inclusion has been shown to enhance both aqueous solubility and stability. researchgate.netnih.govresearchgate.netmdpi.com Studies evaluating different cyclodextrins have indicated that stable 1:1 inclusion complexes are formed, with HE-β-CD demonstrating the highest thermodynamic stability among those tested. mdpi.com

Nanoparticle Formulations: Nanotechnology approaches have been explored to improve the delivery, solubility, and stability of this compound. Nanoemulsion preparations have been developed, which can render the lipid-soluble this compound water-soluble. google.comresearchgate.net These nanoemulsions have shown improved anti-cancer efficacy in preclinical xenograft models compared to this compound suspensions. google.comresearchgate.net Solid Lipid Nanoparticles (SLNs) have been investigated, particularly for topical delivery applications, demonstrating increased skin permeation and improved efficacy in a tumor model. frontiersin.org SLNs can enhance aqueous solubility and extend physiological stability. d-nb.info Lipid nanoformulations have also been explored for the co-delivery of this compound with other therapeutic agents, aiming to improve the solubility and stability of the combined therapy. d-nb.info Furthermore, Lipid Polymer Hybrid Nanoparticles (LPHNs) have been developed for topical delivery, exhibiting favorable characteristics such as particle size, encapsulation efficiency, and zeta potential. researchgate.net The integration of this compound with plasmonic gold nanoparticles (AuNPs) within a β-cyclodextrin complex has been studied for developing laser-triggered controlled release systems. researchgate.netnih.govmdpi.com

Optimized Storage and Handling: While a fundamental aspect of managing instability, optimizing storage and handling conditions is crucial in preclinical studies to maintain drug integrity. This compound is sensitive to light, and protection from light using measures such as amber glass vials, aluminum foil wrapping, or opaque tubing is essential to reduce degradation and the formation of 2-azahypoxanthine. researchgate.net Refrigeration at 2-6°C can also extend the stability of reconstituted and diluted solutions. researchgate.net

These preclinical investigations into modified solid forms, inclusion complexes, and various nanoparticle formulations highlight promising strategies to overcome the inherent solubility and stability challenges of this compound, potentially leading to improved therapeutic outcomes.

ABVD Regimen in Hodgkin's Lymphoma

The ABVD regimen is a widely used first-line treatment for Hodgkin's lymphoma. cancerresearchuk.orgdrugs.com This combination includes doxorubicin (Adriamycin), bleomycin, vinblastine, and this compound. cancerresearchuk.orgdrugs.com ABVD replaced the older MOPP protocol due to its comparable efficacy and generally lower toxicity. ascopubs.orgwikipedia.org Studies have shown ABVD to be highly effective, particularly in early-stage disease, with reported 12-year freedom from progression rates of approximately 87% in early-stage cases. drugs.comnih.gov The ABVD regimen is typically administered in cycles, with drugs given intravenously on specific days within each cycle. cancerresearchuk.orgdrugs.com

MAID Regimen in Sarcoma

This compound is also used as part of the MAID regimen for the treatment of sarcoma. wikipedia.org While the search results mention the MAID regimen, specific details regarding its components and efficacy in sarcoma were not extensively provided in the context of this compound combinations. However, ifosfamide is a component of regimens for soft tissue sarcomas, and this compound is also used for this indication, suggesting a potential combination. taylorandfrancis.comwikipedia.org

Combination with Cisplatin, Carmustine, Tamoxifen

Here is a summary of findings from a study on the combination of this compound, carmustine, cisplatin, and tamoxifen:

Combination with Temozolomide

A study in dogs with relapsed or refractory lymphoma compared treatment with temozolomide or this compound in combination with an anthracycline (doxorubicin or dactinomycin). nih.govavma.org The results showed that 72% of dogs treated with the temozolomide-anthracycline combination and 71% of dogs treated with the this compound-anthracycline combination had a complete or partial response. nih.govavma.org The median duration of response was 40 days for the temozolomide group and 50 days for the this compound group, with no significant differences in response rate, duration of response, or survival time between the two combinations in this animal model. nih.govavma.org

Here is a summary of findings from a study comparing temozolomide or this compound with an anthracycline in dogs with lymphoma:

Combination with Paclitaxel

Combinations involving this compound and paclitaxel have also been investigated, particularly in the context of metastatic melanoma. Paclitaxel is a taxane that acts by stabilizing microtubules, interfering with cell division. wikipedia.orgnih.gov A clinical trial treated patients with metastatic melanoma who had previously received chemotherapy, including a combination of this compound, carmustine, cisplatin, and tamoxifen, with paclitaxel plus tamoxifen. cancerconnect.com In this study, 5 out of 21 evaluable patients (24%) responded to the treatment with paclitaxel and tamoxifen, with one complete response. cancerconnect.com This suggests that a combination including paclitaxel may have activity in patients whose disease progressed after this compound-based regimens. While this specific study did not combine this compound directly with paclitaxel, it highlights the exploration of paclitaxel in a patient population previously treated with this compound-containing regimens.

BRAF/MEK Inhibitors (e.g., Vemurafenib/PLX4032, Sorafenib)

BRAF and MEK inhibitors target specific kinases in the MAPK signaling pathway, which is frequently mutated in melanoma. Vemurafenib (PLX4032) is a BRAF inhibitor wikipedia.orgmims.comnih.gov. Sorafenib is a multikinase inhibitor that targets BRAF, as well as VEGFR and PDGFR guidetopharmacology.orghznu.edu.cnwikipedia.orgmims.comamegroups.orgnih.gov.

Studies have investigated the combination of this compound with BRAF/MEK inhibitors. For instance, sorafenib, a multikinase inhibitor including BRAF, has been explored in combination with this compound in patients with advanced melanoma and sarcomas. A phase II trial investigating sorafenib and this compound in leiomyosarcoma, synovial sarcoma, and malignant peripheral nerve sheath tumors reported a disease control rate of 46% nih.gov. In a phase II trial involving patients with metastatic melanoma, the combination of sorafenib and this compound showed a median progression-free survival of 3.6 months, with a partial response rate of 16.7% and a stable disease rate of 43.3% in the initial stage analysis ascopubs.org. Pharmacokinetic studies of sorafenib in combination with this compound have indicated that concomitant administration may lead to decreased this compound exposure but increased exposure to its metabolite, 5-amino-imidazole-4-carboxamide (AIC) d-nb.info.

Anti-angiogenic Agents (e.g., Bevacizumab)

Anti-angiogenic agents target the formation of new blood vessels that tumors need to grow. Bevacizumab is a monoclonal antibody that targets vascular endothelial growth factor (VEGF) nih.govunil.chtorvergata.itncpsb.org.cncore.ac.uk. Combining this compound with anti-angiogenic agents aims to inhibit tumor blood supply and potentially enhance the delivery and efficacy of chemotherapy. Research has explored the use of bevacizumab in various cancer treatments, and its combination with chemotherapy regimens is a subject of ongoing investigation.

Statins (e.g., Simvastatin, Fluvastatin)

Statins are primarily known for their lipid-lowering effects, but some studies have suggested potential anti-cancer properties. Simvastatin and Fluvastatin are examples of statins wikipedia.orgwikipedia.orgwikidata.orgalfa-chemistry.commims.comciteab.comnih.govnih.govnih.govresearchgate.net. Research into the combination of statins with chemotherapy agents like this compound is an area of interest for potentially improving treatment outcomes.

Curcumin Analogs (e.g., DM-1)

Curcumin, a compound found in turmeric, and its analogs have been investigated for their potential anti-cancer properties. DM-1 is a synthetic curcumin analog plos.orgeuropeanreview.orgplos.orgresearchgate.netnih.gov. Studies have explored the use of DM-1 in combination with this compound as a strategy to inhibit melanoma progression. One study demonstrated that the curcumin analog DM-1 showed promising in vivo tumor reductions via intrinsic apoptosis in both monotherapy and in combination with this compound researchgate.net. Another study described the anti-inflammatory effect of DM-1 and its modulation of gene expression in cell lines nih.gov.

Metformin

Metformin, a biguanide primarily used to treat type 2 diabetes, has also shown potential anti-cancer effects in various studies probes-drugs.orgnih.govnih.govscribd.comguidetomalariapharmacology.org. Its potential mechanisms of action in cancer include activation of the AMP-activated protein kinase (AMPK) pathway and inhibition of the mTOR pathway. The combination of metformin with this compound is an area of emerging research, exploring whether metformin can enhance the cytotoxic effects of this compound or overcome resistance.

Immune Checkpoint Inhibitors (e.g., Pembrolizumab)

Combinations of this compound with immune checkpoint inhibitors (ICIs), such as pembrolizumab, have been investigated, particularly in the context of classical Hodgkin lymphoma (cHL) and metastatic melanoma. A phase II trial is studying the effects of doxorubicin hydrochloride, pembrolizumab, vinblastine, and this compound in treating patients with cHL cancer.gov. This combination aims to leverage the distinct mechanisms of action of chemotherapy and immunotherapy to improve treatment outcomes cancer.gov. In this trial, patients with untreated cHL receive doxorubicin hydrochloride, vinblastine, and this compound intravenously, along with pembrolizumab intravenously cancer.gov.

Table 1. Efficacy of this compound with or without Prior Pembrolizumab in Metastatic Melanoma

*Data derived from a retrospective study researchgate.net.

Interleukin-2 (IL-2) and Interferon-alpha (IFN-α)

The combination of this compound with biological response modifiers such as Interleukin-2 (IL-2) and Interferon-alpha (IFN-α), often referred to as biochemotherapy, has been investigated for the treatment of advanced metastatic melanoma capes.gov.brscilit.com. These regimens aim to combine the cytotoxic effects of chemotherapy with the immunomodulatory effects of cytokines capes.gov.br.

Table 2. Response Rates of this compound-Based Biochemotherapy Regimens in Metastatic Melanoma

*Note: Response rates may vary depending on the specific study design, patient population, and treatment schedule.

Disruption of DNA Repair Mechanisms

The effectiveness of this compound, an alkylating agent, is closely linked to DNA repair mechanisms within cancer cells patsnap.comnih.gov. This compound's active metabolite methylates DNA, primarily at the O⁶-guanine position, leading to DNA mismatch and cell death patsnap.comnih.gov. However, DNA repair enzymes, such as O⁶-methylguanine-DNA methyltransferase (MGMT) and those involved in the base excision repair (BER) pathway, can remove these modifications, thereby reducing the drug's efficacy and contributing to resistance patsnap.comnih.gov. High levels of MGMT expression have been associated with reduced sensitivity to this compound patsnap.com.

Strategies to overcome this resistance mechanism include inhibiting key DNA repair pathways. For instance, inhibiting poly (ADP-ribose) polymerase (PARP), an enzyme involved in BER, has been explored to circumvent this compound resistance by blocking the repair of N⁷-guanine and N³-adenine methylation-induced damage nih.gov. Studies have supported the use of PARP inhibitors in combination with alkylating agents like this compound to enhance cell death nih.gov. Histone deacetylase (HDAC) inhibitors have also been shown to enhance the cytotoxic effects of this compound by disrupting DNA repair mechanisms mdpi.com. HDAC inhibitors can modulate DNA repair mechanisms, increasing the sensitivity of melanoma cells to chemotherapy mdpi.com.

Modulation of Oncogenic Pathways

Aberrant activity in key oncogenic signaling pathways contributes to the development of resistance to various cancer therapies, including this compound mdpi.comresearchgate.net. HDACs, for example, influence pathways such as MAPK, PI3K/AKT, and WNT/β-catenin, promoting cancer cell survival and proliferation even in the presence of chemotherapy mdpi.comresearchgate.net.

Cross-resistance between this compound and targeted therapies, such as BRAF inhibitors, has been observed and is linked to the modulation of these pathways researchgate.net. Resistance to BRAF inhibitors can involve the re-activation of the MAPK pathway or activation of alternative signaling via AKT researchgate.net. Conversely, resistance to this compound can also involve canonical MAPK-independent survival through bypass signaling via AKT researchgate.net.

Targeting specific growth factors and their associated pathways can also contribute to overcoming this compound resistance. For example, targeting Nodal, a growth factor, in conjunction with this compound has shown synergistic anti-cancer effects in metastatic melanoma models nih.gov. This combination demonstrated a synergistic increase in cell killing at concentrations that had limited effects as monotherapy nih.gov.

Induction of Ferroptosis

While the direct induction of ferroptosis by this compound is not extensively detailed in the provided search results, there is an indication that this compound treatment can contribute to a metabolic imbalance in melanoma cells, affecting processes such as iron metabolism nih.gov. Ferroptosis is a form of regulated cell death characterized by iron-dependent lipid peroxidation. Modulating cellular metabolism and iron handling pathways are areas of research being explored to overcome drug resistance in various cancers. Further research may elucidate the specific relationship between this compound, metabolic alterations, and the induction of ferroptosis as a strategy to enhance its cytotoxic effects or overcome resistance.

Melatonin and Melatonin Nanoparticles

Melatonin, a powerful antioxidant, has demonstrated cytoprotective and genoprotective effects in various studies. researchgate.netresearchgate.net Research has explored the potential of melatonin and melatonin-loaded nanoparticles to reduce this compound-induced genotoxicity and oxidative stress. In vitro studies using Hep G2 cells have shown that both melatonin and its nanoparticles can reduce DNA damage and oxidative stress induced by this compound. researchgate.netresearchgate.net Melatonin nanoparticles, in particular, have shown a greater protective effect against this compound-induced genotoxicity and oxidative stress compared to melatonin alone in Hep G2 cells. researchgate.netresearchgate.net

Resveratrol

Resveratrol, a natural polyphenol with antioxidant and antigenotoxic properties, has also been investigated for its ability to mitigate this compound-induced genotoxicity. lifesciencesite.comnih.gov Studies in mice have shown that administration of resveratrol can improve the genotoxic effects of this compound. lifesciencesite.com Resveratrol treatment has been found to significantly reduce the increased averages of chromosomal aberrations and micronucleated peripheral blood reticulocytes induced by this compound. lifesciencesite.com It can also significantly elevate the averages of polychromatic erythrocytes (PCEs) and percentages of mitotic index that were reduced by this compound, suggesting an improvement in mitotic processes. lifesciencesite.com

Amifostine

Amifostine is a cytoprotective agent that is converted to an active metabolite, WR-1065, which is believed to protect noncancerous cells against the toxic effects of DNA-binding chemotherapeutic agents, including alkylating agents. wikipedia.orgmims.com WR-1065 acts as a scavenger of free radicals and binds and detoxifies reactive metabolites. mims.com Studies have evaluated the genoprotective effect of amifostine on this compound-induced genotoxicity. In vitro studies using HepG2 cells have shown that amifostine can prevent DNA damage induced by this compound, as indicated by a significant decrease in tail length in the comet assay. nih.govresearchgate.netnih.gov The protective effect of amifostine has been observed to be concentration-dependent. nih.govresearchgate.net These findings suggest that amifostine could potentially be included in chemotherapy protocols containing this compound to help prevent the formation of secondary cancers. nih.govscience.govresearchgate.netnih.gov