Severe Hepatotoxicity of Mithramycin Therapy Caused by Altered Expression of Hepatocellular Bile Transporters.

Mithramycin demonstrates preclinical anticancer activity, but its therapeutic dose is limited by the development of hepatotoxicity that remains poorly characterized. A pharmacogenomics characterization of mithramycin-induced transaminitis revealed that hepatotoxicity is associated with germline variants in genes involved in bile disposition: ABCB4 (multidrug resistance 3) rs2302387 and ABCB11 [bile salt export pump (BSEP)] rs4668115 reduce transporter expression (P < 0.05) and were associated with ≥grade 3 transaminitis developing 24 hours after the third infusion of mithramycin (25 mcg/kg, 6 hours/infusion, every day ×7, every 28 days; P < 0.0040). A similar relationship was observed in a pediatric cohort. We therefore undertook to characterize the mechanism of mithramycin-induced acute transaminitis. As mithramycin affects cellular response to bile acid treatment by altering the expression of multiple bile transporters (e.g., ABCB4, ABCB11, sodium/taurocholate cotransporting polypeptide, organic solute transporter α/ß) in several cell lines [Huh7, HepaRG, HepaRG BSEP (-/-)] and primary human hepatocytes, we hypothesized that mithramycin inhibited bile-mediated activation of the farnesoid X receptor (FXR). FXR was downregulated in all hepatocyte cell lines and primary human hepatocytes (P < 0.0001), and mithramycin inhibited chenodeoxycholic acid- and GW4046-induced FXR-galactose-induced gene 4 luciferase reporter activity (P < 0.001). Mithramycin promoted glycochenodeoxycholic acid-induced cytotoxicity in ABCB11 (-/-) cells and increased the overall intracellular concentration of bile acids in primary human hepatocytes grown in sandwich culture (P < 0.01). Mithramycin is a FXR expression and FXR transactivation inhibitor that inhibits bile flow and potentiates bile-induced cellular toxicity, particularly in cells with low ABCB11 function. These results suggest that mithramycin causes hepatotoxicity through derangement of bile acid disposition; results also suggest that pharmacogenomic markers may be useful to identify patients who may tolerate higher mithramycin doses. SIGNIFICANCE STATEMENT: The present study characterizes a novel mechanism of drug-induced hepatotoxicity in which mithramycin not only alters farnesoid X receptor (FXR) and small heterodimer partner gene expression but also inhibits bile acid binding to FXR, resulting in deregulation of cellular bile homeostasis. Two novel single-nucleotide polymorphisms in bile flow transporters are associated with mithramycin-induced liver function test elevations, and the present results are the rationale for a genotype-directed clinical trial using mithramycin in patients with thoracic malignancies.

Gene expression dynamics following mithramycin treatment: A possible model for post-chemotherapy cognitive impairment.

Chemotherapy-induced cognitive changes is a major burden on a substantial number of cancer survivors. The mechanism of this sequel is unknown. In this study, we followed long-term effects of early in life mithramycin (MTR) treatment on behaviour and on the normal course of alterations of gene expression in brain. Between post-natal days (PND) 7 and 10, male rats were divided into 2 groups, 1 receiving MTR (0.1 mg/kg s.c. per day) and the other receiving saline. At PND11, frontal cortex tissue samples were dissected from 4 rats from each group. At PND 65 the remaining rats underwent behavioural tests after which all the rats were decapitated and their prefrontal cortex incised. Rats treated transiently with MTR early in life, showed impairments in spatial working memory and anxious-like behaviour in adulthood. The immediate molecular effect of MTR was expressed in a limited number of altered genes of different unconnected trajectories, which were simultaneously distorted by the drug. In contrast, 3 months later we observed a change in the expression of more than 1000 genes that converged into specific cellular processes. Time-dependent gene expression dynamics of several genes was significantly different between treated and untreated rats. The differences in the total number of altered genes and in gene expression trends, immediately and long after MTR treatment cessation, suggest the evolution of a new cellular homeostatic set point, which can lead to behavioural abnormalities following chemotherapy treatment.

Chemotherapy for pain: reversing inflammatory and neuropathic pain with the anticancer agent mithramycin A.

ABSTRACT: The persistence of inflammatory and neuropathic pain is poorly understood. We investigated a novel therapeutic paradigm by targeting gene networks that sustain or reverse persistent pain states. Our prior observations found that Sp1-like transcription factors drive the expression of TRPV1, a pain receptor, that is blocked in vitro by mithramycin A (MTM), an inhibitor of Sp1-like factors. Here, we investigate the ability of MTM to reverse in vivo models of inflammatory and chemotherapy-induced peripheral neuropathy (CIPN) pain and explore MTM's underlying mechanisms. Mithramycin reversed inflammatory heat hyperalgesia induced by complete Freund adjuvant and cisplatin-induced heat and mechanical hypersensitivity. In addition, MTM reversed both short-term and long-term (1 month) oxaliplatin-induced mechanical and cold hypersensitivity, without the rescue of intraepidermal nerve fiber loss. Mithramycin reversed oxaliplatin-induced cold hypersensitivity and oxaliplatin-induced TRPM8 overexpression in dorsal root ganglion (DRG). Evidence across multiple transcriptomic profiling approaches suggest that MTM reverses inflammatory and neuropathic pain through broad transcriptional and alternative splicing regulatory actions. Mithramycin-dependent changes in gene expression following oxaliplatin treatment were largely opposite to and rarely overlapped with changes in gene expression induced by oxaliplatin alone. Notably, RNAseq analysis revealed MTM rescue of oxaliplatin-induced dysregulation of mitochondrial electron transport chain genes that correlated with in vivo reversal of excess reactive oxygen species in DRG neurons. This finding suggests that the mechanism(s) driving persistent pain states such as CIPN are not fixed but are sustained by ongoing modifiable transcription-dependent processes.

The effect of cyclophosphamide and other drugs on the incidence of pulmonary metastases in mice.

A study was made of the effect of various cytotoxic drugs on the ability of i.v.-injected KHT sarcoma cells to form lung colonies in syngeneic C3H mice. Some enhancement of the number of lung colonies following an i.v. injection was seen following pretreatment of the mice with actinomycin D and mithramycin, while pretreatment with vinblastine, bleomycin, methotrexate, cytosine arabinoside, or 5-fluorouracil had little or no effect on lung colony formation. Pretreatment of the mice with cyclophosphamide, however, greatly increased lung colony formation (by a factor of approximately 100). This enhancement in lung colony formation was maximal when the drug was given 24 hr prio to the injection of tumor cells, but was seen as early as 2 hr and persisted as long as 8 weeks prior to the tumor cell injection. The degree of enhancement of lung colony formation was related to the dose of cyclophosphamide and was present in weaning as well as adult mice. This enhancement was not significantly reversed by anticoagulation with either aspirin or warfarin. Immunosuppression by whole-body irradiation did not affect the number of lung colonies seen in cyclophosphamide-treated mice. The mechanism by which cyclophosphamide enhances metastatic tumor growth within the lung is not known. The major effect, however, does not appear to be mediated either by specific immunological or clotting factors.

Phase II trial of plicamycin and hydroxyurea in acute myelogenous leukemia.

A total of 23 patients with high-risk acute myelogenous leukemia (AML) at diagnosis (2 patients), relapsing AML (14) or resistant AML (6) were treated with 25 micrograms/kg i.v. plicamycin every other day for 3 weeks and 500-4,000 mg hydroxyurea per day p. o. according to the WBC count. Aplasia was observed in only two patients. Severe extrahematologic toxicity included sepsis (four cases), vomiting (four patients), toxic hepatitis (three cases), and fibrinopenia (one patient). No partial or complete responses were observed. The 95% confidence interval limit of the overall response rate (CR + PR) was 0-14%.

Chemotherapeutic stress mediated by certain antitumor antibiotics induces an atypical CD69+ surface phenotype in peripheral T-lymphocytes.

Surface antigen CD69 is a Type II integral membrane protein that is generally considered a cell activation marker expressed very early in the normal lymphocyte activation cascade. The conformation of this surface antigen suggests a putative role in transmembrane signal transduction, yet the precise function of this surface antigen has not been clearly elucidated. We had previously reported robust atypical CD69 expression in peripheral T-lymphocytes as concentration-dependent, phenotypic responses to actinomycin D-induced chemotherapeutic stress in the absence of secondary stimulation. Additional antitumor antibiotics were evaluated for inductive potential, and the incidence and respective magnitudes of this chemotherapeutic stress-induced shift in lymphocytic CD69 expression were assessed. Results indicated that atypical CD69 expression is a common response to chemotherapy drug-induced stress. Differences in the respective percentages of CD69 + T-lymphocytes, and the resulting numbers of CD69 surface antigens ultimately expressed by these cells, were documented following in vitro drug exposure. The effective drug concentrations required to mediate detectable shifts in the CD69+ phenotype differed among the selected drugs, as well, suggesting a concentration-dependent induction mechanism putatively related to drug modality. Static CD69 expression responses in CD3+ peripheral T-lymphocytes were also documented, which further suggests that the different intracellular modalities do not mediate proportional T-lymphocyte responses through elevated CD69 expression.

Nephrotoxicity from chemotherapy: prevention and management.

Among the most devastating effects of antineoplastic chemotherapy is nephrotoxicity. This article discusses the effects of plicamycin, methotrexate, mitomycin C, nitrosoureas, 5-azacytidine, cisplatin, ifosfamide, carboplatin, and gallium nitrate on kidney function when these drugs are used either alone or in combination with each other and/or other agents. The importance of adequate hydration during chemotherapy in preventing and reversing drug-induced nephrotoxicity is also discussed.

Acquired dysfibrinogenemia secondary to mithramycin toxicity.

A 58-year-old black woman with IgD multiple myeloma developed a hemorrhagic diathesis within 48 hours after receiving mithramycin (20 micrograms/kg/day) for therapy of hypercalcemia. Her coagulation studies were characterized by prolonged prothrombin, partial thromboplastin, thrombin, and reptilase clotting times. Her plasma and partially purified fibrinogen were inhibitory to the clotting of normal plasma and fibrinogen. The patient's isolated fibrinogen showed a normal rate of fibrinopeptide release, but her fibrin monomer aggregation was markedly abnormal. These studies document the development of a dysfibrinogenemia secondary to mithramycin toxicity.

[Nephrotoxicity of antitumor chemotherapy]. / Néphrotoxicité des chimiothérapies antitumorales.

The renal toxicity of antitumoral drugs is an increasingly disturbing problem. These drugs are now prescribed in an ever wider variety of cases, and delayed renal reactions, previously unknown, are revealed by the longer survivals obtained. For a number of years, patients whose cancer had been cured have been placed under haemodialysis on account of drug-induced renal failure. The renal toxicity of cisplatinum, nitrosoureas and methotrexate is well-known, but mitomycin C is also capable of inducing permanent renal failure; the delayed toxicity of this drug explains that it has long been underestimated. This example emphasizes the need for close co-operation between oncologists, nephrologists and pharmacologists in order to determine, for each patient, the most effective treatment with the minimum of side effects.

[Acute fatal hepatorenal failure during treatment with mithramycin]. / Insuffisance hépato-rénale aiguë mortelle au cours d'un traitement par la mithramycine.

A case of fatal hepato-renal failure occurring during mithramycin treatment is reported. A 64 year-old female patient was admitted to hospital in a state of acute renal failure. She also presented with hypercalcaemia and bilateral pulmonary metastases. She had been operated on 10 years previously of a parathyroid cancer. Despite treatment with mithramycin (total dose 8.25 mg) and haemodialysis, the hypercalcaemia returned; it was then decided to remove the secretory lung metastases (parathormone 420 micrograms X ml-1). 48 hours before surgery, the patient was again given 1.25 mg mithramycin. Immediately after surgery, she developed hepatic failure with massive cell destruction and anuria. The patient died 48 h after the operation. The hepatic and renal complications of mithramycin are discussed.

[Myeloid crisis of chronic myelogenous leukemia showing dramatic response to mithramycin and hydroxyurea combination].

After 4 years of chronic phase, a 22-year-old female with Ph1 (+) chronic myelogenous leukemia developed myelomonocytic crisis. On admission, her Hb was 9.9 g/dl, Plt 4.1 x 10(4)/microliters, WBC 138,000/microliters with 16.5% blasts. Bone marrow contained 38% blasts. She received a combination chemotherapy of mithramycin and hydroxyurea, as reported by Koller et al. Dose of mithramycin was reduced to 20 micrograms/kg. Following 1st and 2nd infusions of mithramycin, severe nasal bleeding was seen. Prednisolone 10 mg/day was given from the 3rd dose of mithramycin with apparent hemostatic effects. Calcium gluconate 3 g/day was administered concomitantly. Her disease responded promptly to this treatment and hematological remission was achieved.