IRX4204 Induces Senescence and Cell Death in HER2-positive Breast Cancer and Synergizes with Anti-HER2 Therapy.

PURPOSE: Rexinoids, agonists of nuclear retinoid X receptor (RXR), have been used for the treatment of cancers and are well tolerated in both animals and humans. However, the usefulness of rexinoids in treatment of breast cancer remains unknown. This study examines the efficacy of IRX4204, a highly specific rexinoid, in breast cancer cell lines and preclinical models to identify a biomarker for response and potential mechanism of action. EXPERIMENTAL DESIGN: IRX4204 effects on breast cancer cell growth and viability were determined using cell lines, syngeneic mouse models, and primary patient-derived xenograft (PDX) tumors. In vitro assays of cell cycle, apoptosis, senescence, and lipid metabolism were used to uncover a potential mechanism of action. Standard anti-HER2 therapies were screened in combination with IRX4204 on a panel of breast cancer cell lines to determine drug synergy. RESULTS: IRX4204 significantly inhibits the growth of HER2-positive breast cancer cell lines, including trastuzumab and lapatinib-resistant JIMT-1 and HCC1954. Treatment with IRX4204 reduced tumor growth rate in the MMTV-ErbB2 mouse and HER2-positive PDX model by 49% and 44%, respectively. Mechanistic studies revealed IRX4204 modulates lipid metabolism and induces senescence of HER2-positive cells. In addition, IRX4204 demonstrates additivity and synergy with HER2-targeted mAbs, tyrosine kinase inhibitors, and antibody-drug conjugates. CONCLUSIONS: These findings identify HER2 as a biomarker for IRX4204 treatment response and demonstrate a novel use of RXR agonists to synergize with current anti-HER2 therapies. Furthermore, our results suggest that RXR agonists can be useful for the treatment of anti-HER2 resistant and metastatic HER2-positive breast cancer.

A novel retinoic acid receptor-γ agonist antagonizes immune checkpoint resistance in lung cancers by altering the tumor immune microenvironment.

All-trans-retinoic acid (ATRA), the retinoic acid receptors (RARs) agonist, regulates cell growth, differentiation, immunity, and survival. We report that ATRA-treatment repressed cancer growth in syngeneic immunocompetent, but not immunodeficient mice. The tumor microenvironment was implicated: CD8+ T cell depletion antagonized ATRA's anti-tumorigenic effects in syngeneic mice. ATRA-treatment with checkpoint blockade did not cooperatively inhibit murine lung cancer growth. To augment ATRA's anti-tumorigenicity without promoting its pro-tumorigenic potential, an RARγ agonist (IRX4647) was used since it regulates T cell biology. Treating with IRX4647 in combination with an immune checkpoint (anti-PD-L1) inhibitor resulted in a statistically significant suppression of syngeneic 344SQ lung cancers in mice-a model known for its resistance to checkpoints and characterized by low basal T cell and PD-L1 expression. This combined treatment notably elevated CD4+ T-cell presence within the tumor microenvironment and increased IL-5 and IL-13 tumor levels, while simultaneously decreasing CD38 in the tumor stroma. IL-5 and/or IL-13 treatments increased CD4+ more than CD8+ T-cells in mice. IRX4647-treatment did not appreciably affect in vitro lung cancer growth, despite RARγ expression. Pharmacokinetic analysis found IRX4647 plasma half-life was 6 h in mice. Yet, RARα antagonist (IRX6696)-treatment with anti-PD-L1 did not repress syngeneic lung cancer growth. Together, these findings provide a rationale for a clinical trial investigating an RARγ agonist to augment check point blockade response in cancers.

Preserving and enhancing mitochondrial function after stroke to protect and repair the neurovascular unit: novel opportunities for nanoparticle-based drug delivery.

The neurovascular unit (NVU) is composed of vascular cells, glia, and neurons that form the basic component of the blood brain barrier. This intricate structure rapidly adjusts cerebral blood flow to match the metabolic needs of brain activity. However, the NVU is exquisitely sensitive to damage and displays limited repair after a stroke. To effectively treat stroke, it is therefore considered crucial to both protect and repair the NVU. Mitochondrial calcium (Ca2+) uptake supports NVU function by buffering Ca2+ and stimulating energy production. However, excessive mitochondrial Ca2+ uptake causes toxic mitochondrial Ca2+ overloading that triggers numerous cell death pathways which destroy the NVU. Mitochondrial damage is one of the earliest pathological events in stroke. Drugs that preserve mitochondrial integrity and function should therefore confer profound NVU protection by blocking the initiation of numerous injury events. We have shown that mitochondrial Ca2+ uptake and efflux in the brain are mediated by the mitochondrial Ca2+ uniporter complex (MCUcx) and sodium/Ca2+/lithium exchanger (NCLX), respectively. Moreover, our recent pharmacological studies have demonstrated that MCUcx inhibition and NCLX activation suppress ischemic and excitotoxic neuronal cell death by blocking mitochondrial Ca2+ overloading. These findings suggest that combining MCUcx inhibition with NCLX activation should markedly protect the NVU. In terms of promoting NVU repair, nuclear hormone receptor activation is a promising approach. Retinoid X receptor (RXR) and thyroid hormone receptor (TR) agonists activate complementary transcriptional programs that stimulate mitochondrial biogenesis, suppress inflammation, and enhance the production of new vascular cells, glia, and neurons. RXR and TR agonism should thus further improve the clinical benefits of MCUcx inhibition and NCLX activation by increasing NVU repair. However, drugs that either inhibit the MCUcx, or stimulate the NCLX, or activate the RXR or TR, suffer from adverse effects caused by undesired actions on healthy tissues. To overcome this problem, we describe the use of nanoparticle drug formulations that preferentially target metabolically compromised and damaged NVUs after an ischemic or hemorrhagic stroke. These nanoparticle-based approaches have the potential to improve clinical safety and efficacy by maximizing drug delivery to diseased NVUs and minimizing drug exposure in healthy brain and peripheral tissues.

Targeting the Retinoid X Receptor Pathway Prevents and Ameliorates Murine Chronic Graft-Versus-Host Disease.

Most allogeneic hematopoietic stem cell transplant (allo-HSCT) recipients receive peripheral blood stem cell grafts resulting in a 30%-70% incidence of chronic graft-versus-host disease (cGVHD), a major cause of mortality and morbidity in long-term survivors. While systemic steroids remain the standard of care for first-line therapy, patients may require long-term administration, and those with steroid-resistant or refractory cGVHD have a worse prognosis. Although durable and deep responses with second-line therapies can be achieved in some patients, there remains an urgent need for new therapies. In this study, we evaluated the efficacy of IRX4204, a novel agonist that activates RXRs and is in clinical trials for cancer treatment to prevent and treat cGVHD in two complementary murine models. In a major histocompatibility complex mismatched, non-sclerodermatous multiorgan system model with bronchiolitis obliterans, IRX4204 prevented and reversed cGVHD including associated pulmonary dysfunction with restoration of germinal center T-follicular helper: T-follicular regulatory cell balance. In a minor histocompatibility antigen disparate sclerodermatous model, IRX4204 treatment significantly prevented and ameliorated skin cGVHD by reducing Th1 and Th17 differentiation due to anti-inflammatory properties. Together, these results indicate that IRX4204 is a promising therapeutic option to treat cGVHD with bronchiolitis obliterans or sclerodermatous manifestations.

Predicting Metastasis in Melanoma by Enumerating Circulating Tumor Cells Using Photoacoustic Flow Cytometry.

BACKGROUND AND OBJECTIVES: Enumerating circulating tumor cells has been used as a method of monitoring progression of various cancers. Various methods for detecting circulating melanoma cells (CMCs) have been reported, but none has had sufficient sensitivity to determine if the presence of rare CMCs in the blood of Stage I-III melanoma patients predicts if those patients eventually develop metastatic disease. STUDY DESIGN: We quantified CMCs in serial blood samples from 38 early stage melanoma patients to determine if CMC numbers predict development of metastatic melanoma. CMCs were enumerated using a photoacoustic flow cytometric detection system that uses a laser to induce high frequency acoustic signals in pigmented CMCs. RESULTS: We observed that detection of greater than 2 CMCs/ml of blood from patients with Stage I-III melanoma predicts metastatic disease. Of the 11 patients we studied who had two or fewer CMCs detected at all time points tested, none progressed to metastatic disease over a mean follow-up of 1288 days. In contrast, 18 of the 27 patients (67%) having more than 2 CMCs/ml at one or more time points progressed to metastatic disease over a mean follow-up of 850 days. CONCLUSIONS: Photoacoustic flow cytometry can detect rare CMCs in the blood of Stage I-III melanoma patients and detectionof these cells is predictive of subsequent development of metastatic disease. Lasers Surg. Med.

Photoacoustic detection of circulating melanoma cells in late stage patients.

Melanoma is the deadliest skin cancer and is responsible for over 7000 deaths in the US annually. The spread of cancer, or metastasis, is responsible for these deaths, as secondary tumors interrupt normal organ function. Circulating tumor cells, or those cells that spread throughout the body from the primary tumor, are thought to be responsible for metastasis. We developed an optical method, photoacoustic flow cytometry, in order to detect and enumerate circulating melanoma cells (CMCs) from blood samples of patients. We tested the blood of Stage IV melanoma patients to show the ability of the photoacoustic flow cytometer to detect these rare cells in blood. We then tested the system on archived blood samples from Stage III melanoma patients with known outcomes to determine if detection of CMCs can predict future metastasis. We detected between 0 and 66 CMCs in Stage IV patients. For the Stage III study, we found that of those samples with CMCs, 2 remained disease free and 5 developed metastasis. Of those without CMCs, 6 remained disease free and 1 developed metastasis. We believe that photoacoustic detection of CMCs provides valuable information for the prediction of metastasis and we postulate a system for more accurate prognosis.

Selective brain penetrable Nurr1 transactivator for treating Parkinson's disease.

Parkinson's disease (PD) is one of the most common movement disorders, and currently there is no effective treatment that can slow disease progression. Preserving and enhancing DA neuron survival is increasingly regarded as the most promising therapeutic strategy for treating PD. IRX4204 is a second generation retinoid X receptor (RXR) agonist that has no cross reactivity with retinoic acid receptors, farnesoid X receptor, liver X receptors or peroxisome proliferator-activated receptor PPARγ. We found that IRX4204 promotes the survival and maintenance of nigral dopaminergic (DA) neurons in a dose-dependent manner in primary mesencephalic cultures. Brain bioavailability studies demonstrate that IRX4204 can cross the blood brain barrier and reach the brain at nM concentration. Oral administration of IRX4204 can activate nuclear receptor Nurr1 downstream signaling in the substantia nigra (SN) andattenuate neurochemical and motor deficits in a rat model of PD. Our study suggests that IRX4204 represents a novel, potent and selective pharmacological means to activate cellular RXR-Nurr1 signaling and promote SN DA neuron survival in PD prevention and/or treatment.

Recombinant human relaxin in the treatment of systemic sclerosis with diffuse cutaneous involvement: a randomized, double-blind, placebo-controlled trial.

OBJECTIVE: A phase II randomized controlled trial of recombinant human relaxin suggested that a dosage of 25 microg/kg/day was safe and clinically effective in improving skin disease and reducing functional disability in scleroderma (systemic sclerosis; SSc). We undertook a large randomized, double-blind, placebo-controlled clinical trial to compare placebo with 10 microg/kg/day and 25 microg/kg/day recombinant human relaxin, given for 24 weeks in patients with stable, diffuse, moderate-to-severe SSc. METHODS: Men and women ages 18-70 years with diffuse cutaneous SSc (dcSSc) were administered recombinant human relaxin (10 microg/kg/day or 25 microg/kg/day) or placebo for 24 weeks as a continuous subcutaneous infusion. There was a followup safety visit at week 28. RESULTS: The primary outcome measure, the modified Rodnan skin thickness score, was similar among the 3 groups at baseline and at weeks 4, 12, and 24. Secondary outcomes such as functional disability were similar in all 3 groups, while the forced vital capacity decreased significantly in the relaxin groups. The discontinuation of both doses of relaxin at week 24 led to statistically significant declines in creatinine clearance and serious renal adverse events (defined as doubling of serum creatinine, renal crisis, or grade 3 or 4 essential hypertension) in 7 patients who had received relaxin therapy but in none who had received placebo. CONCLUSION: Recombinant relaxin was not significantly better than placebo in improving the total skin score or pulmonary function or in reducing functional disability in patients with dcSSc. In addition, relaxin was associated with serious renal adverse events, the majority of which occurred after stopping the infusion. If relaxin is used therapeutically for any conditions other than scleroderma, close monitoring of blood pressure and renal function must be performed.