Publications by authors named "Kasey L Couts"

12 Publications

  • Page 1 of 1

A Novel Regimen for Treating Melanoma: MCL1 Inhibitors and Azacitidine.

Pharmaceuticals (Basel) 2021 Jul 30;14(8). Epub 2021 Jul 30.

Department of Dermatology, School of Medicine, University of Colorado Anschutz Medical Campus, Mail Stop 8127, Aurora, CO 80045, USA.

Although treatment options for melanoma patients have expanded in recent years with the approval of immunotherapy and targeted therapy, there is still an unmet need for new treatment options for patients that are ineligible for, or resistant to these therapies. BH3 mimetics, drugs that mimic the activity of pro-apoptotic BCL2 family proteins, have recently achieved remarkable success in the clinical setting. The combination of BH3 mimetic ABT-199 (venetoclax) plus azacitidine has shown substantial benefit in treating acute myelogenous leukemia. We evaluated the efficacy of various combinations of BH3 mimetic + azacitidine in fourteen human melanoma cell lines from cutaneous, mucosal, acral and uveal subtypes. Using a combination of cell viability assay, BCL2 family knockdown cell lines, live cell imaging, and sphere formation assay, we found that combining inhibition of MCL1, an anti-apoptotic BCL2 protein, with azacitidine had substantial pro-apoptotic effects in multiple melanoma cell lines. Specifically, this combination reduced cell viability, proliferation, sphere formation, and induced apoptosis. In addition, this combination is highly effective at reducing cell viability in rare mucosal and uveal subtypes. Overall, our data suggest this combination as a promising therapeutic option for some patients with melanoma and should be further explored in clinical trials.
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http://dx.doi.org/10.3390/ph14080749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8399604PMC
July 2021

Melanoma Metastases to the Adrenal Gland Are Highly Resistant to Immune Checkpoint Inhibitors.

J Natl Compr Canc Netw 2021 Aug 4. Epub 2021 Aug 4.

3Center for Rare Melanomas.

Background: Adrenal gland metastases (AGMs) are common in advanced-stage melanoma, occurring in up to 50% of patients. The introduction of immune checkpoint inhibitors (ICIs) has markedly altered the outcome of patients with melanoma. However, despite significant successes, anecdotal evidence has suggested that treatment responses in AGMs are significantly lower than in other metastatic sites. We sought to investigate whether having an AGM is associated with altered outcomes and whether ICI responses are dampened in the adrenal glands.

Patients And Methods: We retrospectively compared ICI responses and overall survival (OS) in 68 patients with melanoma who were diagnosed with an AGM and a control group of 100 patients without AGMs at a single institution. Response was determined using RECIST 1.1. OS was calculated from time of ICI initiation, anti-PD-1 initiation, initial melanoma diagnosis, and stage IV disease diagnosis. Tumor-infiltrating immune cells were characterized in 9 resected AGMs using immunohistochemical analysis.

Results: Response rates of AGMs were significantly lower compared with other metastatic sites in patients with AGMs (16% vs 22%) and compared with those without AGMs (55%). Patients with AGMs also had significantly lower median OS compared with those without AGMs (3.1 years vs not reached, respectively). We further observed that despite this, AGMs exhibited high levels of tumor-infiltrating immune cells.

Conclusions: In this cohort of patients with melanoma, those diagnosed with an AGM had lower ICI response rates and OS. These results suggest that tissue-specific microenvironments of AGMs present unique challenges that may require novel, adrenal gland-directed therapies or surgical resection.
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http://dx.doi.org/10.6004/jnccn.2020.7800DOI Listing
August 2021

Clinical and molecular features of subungual melanomas are site-specific and distinct from acral melanomas.

Melanoma Res 2020 12;30(6):562-573

Division of Medical Oncology, Department of Medicine.

Subungual melanomas (SUM) arise beneath the nails of the hands and feet, and account for 0.7-3.5% of all malignant melanomas. Most studies include SUM in the category of acral melanoma, but understanding the specific features of SUM is critical for improving patient care. In this study, we performed a site-specific comparison of the clinical and molecular features between 54 cases of SUM and 78 cases of nonsubungual acral melanoma. Compared to patients with acral melanoma, patients with SUM were younger at diagnosis, had a higher prevalence of primary melanomas on the hand, and had more frequent reports of previous trauma at the tumor site. SUM was deeper than acral melanoma at diagnosis, which correlated with an increased frequency of metastases. Analysis of common melanoma driver genes revealed KIT and KRAS mutations were predominantly found in SUM, whereas BRAF and NRAS mutations occurred almost exclusively in acral melanoma. We also discovered molecular differences in the cell cycle pathway, where CDK4/CCND1 amplifications were more frequent in SUM and CDKN2A/B loss occurred mostly in acral melanoma, and in the PI3K/mTOR pathway, where RICTOR amplification and TSC1 K587R mutations were exclusively in SUM and PTEN loss and AKT1 mutations were exclusively in acral melanoma. Comparison of hand versus foot tumors revealed more frequent ulceration of SUM foot tumors, which correlated with more distal metastases and poorer overall survival. In summary, we find SUM are both clinically and molecularly distinct from acral melanoma, and our data suggest KIT, CDK4/6, and mTOR inhibitors may be particularly relevant and effective treatments for patients with SUM.
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http://dx.doi.org/10.1097/CMR.0000000000000688DOI Listing
December 2020

MCL1 inhibitors S63845/MIK665 plus Navitoclax synergistically kill difficult-to-treat melanoma cells.

Cell Death Dis 2020 06 8;11(6):443. Epub 2020 Jun 8.

Department of Dermatology, University of Colorado Anschutz Medical Campus, School of Medicine, Mail Stop 8127, Aurora, CO, 80045, US.

Current treatment for patients with metastatic melanoma include molecular-targeted therapies and immune checkpoint inhibitors. However, a subset of melanomas are difficult-to-treat. These melanomas include those without the genetic markers for targeted therapy, non-responsive to immunotherapy, and those who have relapsed or exhausted their therapeutic options. Therefore, it is necessary to understand and explore other biological processes that may provide new therapeutic approaches. One of most appealing is targeting the apoptotic/anti-apoptotic system that is effective against leukemia. We used genetic knockdown and pharmacologic approaches of BH3 mimetics to target anti-apoptotic BCL2 family members and identified MCL1 and BCLXL as crucial pro-survival members in melanoma. We then examined the effects of combining BH3 mimetics to target MCL1 and BCLXL in vitro and in vivo. These include clinical-trial-ready compounds such as ABT-263 (Navitoclax) and S63845/S64315 (MIK655). We used cell lines derived from patients with difficult-to-treat melanomas. In vitro, the combined inhibition of MCL1 and BCLXL resulted in significantly effective cell killing compared to single-agent treatment (p < 0.05) in multiple assays, including sphere assays. The combination-induced cell death was independent of BIM, and NOXA. Recapitulated in our mouse xenograft model, the combination inhibited tumor growth, reduced sphere-forming capacity (p < 0.01 and 0.05, respectively), and had tolerable toxicity (p > 0.40). Taken together, this study suggests that dual targeting of MCL1 and BCLXL should be considered as a treatment option for difficult-to-treat melanoma patients.
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http://dx.doi.org/10.1038/s41419-020-2646-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7280535PMC
June 2020

High-Dimensional Analysis of Postsplenectomy Peripheral Immune Cell Changes.

Immunohorizons 2020 02 18;4(2):82-92. Epub 2020 Feb 18.

Department of Surgery, School of Medicine, University of Colorado Anschutz Medical Campus, Aurora, CO 80045;

Although the consequences of splenectomy are well understood in mice, much less is known about the immunologic changes that occur following splenectomy in humans. We sought to characterize the circulating immune cell populations of patients before and after elective splenectomy to determine if these changes are related to postsplenectomy survival outcomes. Retrospective clinical information was collected from 95 patients undergoing elective splenectomy compared with 91 patients undergoing pancreaticoduodenectomy (Whipple procedure). We further analyzed peripheral blood from five patients in the splenectomy group, collected before and after surgery, using single-cell cytometry by time-of-flight mass spectrometry. We compared pre- and postsplenectomy data to characterize both the major and minor immune cell populations in significantly greater detail. Compared with patients undergoing a Whipple procedure, splenectomized patients had significant and long-lasting elevated counts of lymphocytes, monocytes, and basophils. Cytometry by time-of-flight mass spectroscopy analysis demonstrated that the elevated lymphocytes primarily consisted of naive CD4 T cells and a population of activated CD25CD56CD4 T cells, whereas the elevated monocyte counts were mainly mature, activated monocytes. We also observed a significant increase in the expression of the chemokine receptors CCR6 and CCR4 on several cellular populations. Taken together, these data indicate that significant immunological changes take place following splenectomy. Whereas other groups have compared splenectomized patients to healthy controls, this study compared patients undergoing elective splenectomy to those undergoing a similar major abdominal surgery. Overall, we found that splenectomy results in significant long-lasting changes in circulating immune cell populations and function.
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http://dx.doi.org/10.4049/immunohorizons.1900089DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7476217PMC
February 2020

IL-6 and IL-8 Are Linked With Myeloid-Derived Suppressor Cell Accumulation and Correlate With Poor Clinical Outcomes in Melanoma Patients.

Front Oncol 2019 8;9:1223. Epub 2019 Nov 8.

Division of Surgical Oncology, Department of Surgery, University of Colorado Anschutz Medical Campus, Aurora, CO, United States.

We sought to identify tumor-secreted factors that altered the frequency of MDSCs and correlated with clinical outcomes in advanced melanoma patients. We focused our study on several of the many factors involved in the expansion and mobilization of MDSCs. These were identified by measuring circulating concentrations of 13 cytokines and growth factors in stage IV melanoma patients ( = 55) and healthy controls ( = 22). Based on these results, we hypothesized that IL-6 and IL-8 produced by melanoma tumor cells participate in the expansion and recruitment of MDSCs and together would be predictive of overall survival in melanoma patients. We then compared the expression of IL-6 and IL-8 in melanoma tumors to the corresponding plasma concentrations and the frequency of circulating MDSCs. These measures were correlated with clinical outcomes. Patients with high plasma concentrations of either IL-6 (40%) or IL-8 (63%), or both (35%) had worse median overall survival compared to patients with low concentrations. Patients with low peripheral concentrations and low tumoral expression of IL-6 and IL-8 showed decreased frequencies of circulating MDSCs, and patients with low frequencies of MDSCs had better overall survival. We have previously shown that IL-6 is capable of expanding MDSCs, and here we show that MDSCs are chemoattracted to IL-8. Multivariate analysis demonstrated an increased risk of death for subjects with both high IL-6 and IL-8 (HR 3.059) and high MDSCs (HR 4.265). Together these results indicate an important role for IL-6 and IL-8 in melanoma patients in which IL-6 potentially expands peripheral MDSCs and IL-8 recruits these highly immunosuppressive cells to the tumor microenvironment. This study provides further support for identifying potential therapeutics targeting IL-6, IL-8, and MDSCs to improve melanoma treatments.
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http://dx.doi.org/10.3389/fonc.2019.01223DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6857649PMC
November 2019

BRAF fusions identified in melanomas have variable treatment responses and phenotypes.

Oncogene 2019 02 25;38(8):1296-1308. Epub 2018 Sep 25.

Division of Medical Oncology, Department of Medicine, University of Colorado Denver, Aurora, CO, USA.

Oncogenic BRAF fusions have emerged as an alternate mechanism for BRAF activation in melanomas and other cancers. A number of BRAF fusions with different 5' gene partners and BRAF exon breakpoints have been described, but the effects of different partners and breakpoints on cancer phenotypes and treatment responses has not been well characterized. Targeted RNA sequencing was used to screen 60 melanoma patient-derived xenograft (PDX) models for BRAF fusions. We identified three unique BRAF fusions, including a novel SEPT3-BRAF fusion, occurring in four tumors (4/60, 6.7%), all of which were "pan-negative" (lacking other common mutations) (4/18, 22.2%). The BRAF fusion PDX models showed variable growth rates and responses to MAPK inhibitors in vivo. Overexpression of BRAF fusions identified in our study, as well as other BRAF fusions previously identified in melanomas, resulted in a high degree of variability in 2D proliferation and 3D invasion between the different fusions. While exogenously expressed BRAF fusions all responded to MAPK inhibition in vitro, we observed potential differences in signaling and feedback mechanisms. In summary, BRAF fusions are actionable therapeutic targets, however there are significant differences in phenotypes, treatment responses, and signaling which may be clinically relevant.
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http://dx.doi.org/10.1038/s41388-018-0514-7DOI Listing
February 2019

BH3 mimetics induce apoptosis independent of DRP-1 in melanoma.

Cell Death Dis 2018 09 5;9(9):907. Epub 2018 Sep 5.

University of Colorado Anschutz Medical Campus, School of Medicine, Department of Dermatology, Mail Stop 8127, Aurora, CO, 80045, USA.

Despite the recent advancement in treating melanoma, options are still limited for patients without BRAF mutations or in relapse from current treatments. BH3 mimetics against members of the BCL-2 family have gained excitement with the recent success in hematological malignancies. However, single drug BH3 mimetic therapy in melanoma has limited effectiveness due to escape by the anti-apoptotic protein MCL-1 and/or survival of melanoma-initiating cells (MICs). We tested the efficacy of the BH3 mimetic combination of A-1210477 (an MCL-1 inhibitor) and ABT-263 (a BCL-2/BCL-XL/BCL-W inhibitor) in killing melanoma, especially MICs. We also sought to better define Dynamin-Related Protein 1 (DRP-1)'s role in melanoma; DRP-1 is known to interact with members of the BCL-2 family and is a possible therapeutic target for melanoma treatment. We used multiple assays (cell viability, apoptosis, bright field, immunoblot, and sphere formation), as well as the CRISPR/Cas9 genome-editing techniques. For clinical relevance, we employed patient samples of different mutation status, including some relapsed from current treatments such as anti-PD-1 immunotherapy. We found the BH3 mimetic combination kill both the MICs and non-MICs (bulk of melanoma) in all cell lines and patient samples irrespective of the mutation status or relapsed state (p < 0.05). Unexpectedly, the major pro-apoptotic proteins, NOXA and BIM, are not necessary for the combination-induced cell death. Furthermore, the combination impedes the activation of DRP-1, and inhibition of DRP-1 further enhances apoptosis (p < 0.05). DRP-1 effects in melanoma differ from those seen in other cancer cells. These results provide new insights into BCL-2 family's regulation of the apoptotic pathway in melanoma, and suggest that inhibiting the major anti-apoptotic proteins is sufficient to induce cell death even without involvement from major pro-apoptotic proteins. Importantly, our study also indicates that DRP-1 inhibition is a promising adjuvant for BH3 mimetics in melanoma treatment.
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http://dx.doi.org/10.1038/s41419-018-0932-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6125485PMC
September 2018

ALK Inhibitor Response in Melanomas Expressing Fusions and Alternate Isoforms.

Mol Cancer Ther 2018 01 20;17(1):222-231. Epub 2017 Oct 20.

Division of Medical Oncology, Department of Medicine, University of Colorado Anschutz Medical Campus, Aurora, Colorado.

Oncogenic fusions occur in several types of cancer and can be effectively treated with ALK inhibitors; however, fusions and treatment response have not been characterized in malignant melanomas. Recently, a novel isoform of ( ) was reported in 11% of melanomas but the response of melanomas expressing to ALK inhibition has not been well characterized. We analyzed 45 melanoma patient-derived xenograft models for mRNA and protein expression. expression was identified in 11 of 45 (24.4%) melanomas. Ten melanomas express wild-type (wt) and/or and one mucosal melanoma expresses multiple novel fusion variants. Melanoma cells expressing different variants were tested for response to ALK inhibitors. Whereas the melanoma expressing were sensitive to ALK inhibitors and , the melanomas expressing wt or were not sensitive to ALK inhibitors. In addition, a patient with mucosal melanoma expressing was treated with an ALK/ROS1/TRK inhibitor (entrectinib) on a phase I trial but did not respond. Our results demonstrate fusions occur in malignant melanomas and respond to targeted therapy, whereas melanomas expressing do not respond to ALK inhibitors. Targeting fusions is an effective therapeutic option for a subset of melanoma patients, but additional clinical studies are needed to determine the efficacy of targeted therapies in melanomas expressing wt or .
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http://dx.doi.org/10.1158/1535-7163.MCT-17-0472DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5752582PMC
January 2018

Whole-exome sequencing identifies recurrent SF3B1 R625 mutation and comutation of NF1 and KIT in mucosal melanoma.

Melanoma Res 2017 06;27(3):189-199

aDepartment of Medicine, Division of Medical Oncology bDepartment of Pathology cDepartment of Surgery, School of Medicine dDepartment of Biostatistics and Informatics, Colorado School of Public Health eUniversity of Colorado Cancer Center fDepartment of Dermatology, University of Colorado Anschutz Medical Campus, Aurora, Colorado, USA.

Mucosal melanomas are a rare subtype of melanoma, arising in mucosal tissues, which have a very poor prognosis due to the lack of effective targeted therapies. This study aimed to better understand the molecular landscape of these cancers and find potential new therapeutic targets. Whole-exome sequencing was performed on mucosal melanomas from 19 patients and 135 sun-exposed cutaneous melanomas, with matched peripheral blood samples when available. Mutational profiles were compared between mucosal subgroups and sun-exposed cutaneous melanomas. Comparisons of molecular profiles identified 161 genes enriched in mucosal melanoma (P<0.05). KIT and NF1 were frequently comutated (32%) in the mucosal subgroup, with a significantly higher incidence than that in cutaneous melanoma (4%). Recurrent SF3B1 R625H/S/C mutations were identified and validated in 7 of 19 (37%) mucosal melanoma patients. Mutations in the spliceosome pathway were found to be enriched in mucosal melanomas when compared with cutaneous melanomas. Alternative splicing in four genes were observed in SF3B1-mutant samples compared with the wild-type samples. This study identified potential new therapeutic targets for mucosal melanoma, including comutation of NF1 and KIT, and recurrent R625 mutations in SF3B1. This is the first report of SF3B1 R625 mutations in vulvovaginal mucosal melanoma, with the largest whole-exome sequencing project of mucosal melanomas to date. The results here also indicated that the mutations in SF3B1 lead to alternative splicing in multiple genes. These findings expand our knowledge of this rare disease.
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http://dx.doi.org/10.1097/CMR.0000000000000345DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5470740PMC
June 2017

Understanding melanoma stem cells.

Melanoma Manag 2015;2(2):179-188

Department of Dermatology, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, USA ; Denver Veterans Affairs Medical Center, Denver, CO 80220, USA.

Tumors are incredibly diverse and contain many different subpopulations of cells. The cancer stem cell (CSC) subpopulation is responsible for many aspects of tumorigenesis and has been shown to play an important role in melanoma development, progression, drug resistance and metastasis. However, it is becoming clear that tumor cell populations are dynamic and can be influenced by many factors, such as signals from the tumor microenvironment and somatic evolution. This review will present the current understanding of CSCs and the challenges of identifying and characterizing this dynamic cell population. The known characteristics and functions of melanoma stem cells, and the potential for therapeutic targeting of these cells in melanoma, will be discussed.
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http://dx.doi.org/10.2217/mmt.15.4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4649940PMC
January 2015

Functional proteomics identifies targets of phosphorylation by B-Raf signaling in melanoma.

Mol Cell 2009 Apr;34(1):115-31

Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO 80309-0215, USA.

Melanoma and other cancers harbor oncogenic mutations in the protein kinase B-Raf, which leads to constitutive activation and dysregulation of MAP kinase signaling. In order to elucidate molecular determinants responsible for B-Raf control of cancer phenotypes, we present a method for phosphoprotein profiling, using negative ionization mass spectrometry to detect phosphopeptides based on their fragment ion signature caused by release of PO(3)(-). The method provides an alternative strategy for phosphoproteomics, circumventing affinity enrichment of phosphopeptides and isotopic labeling of samples. Ninety phosphorylation events were regulated by oncogenic B-Raf signaling, based on their responses to treating melanoma cells with MKK1/2 inhibitor. Regulated phosphoproteins included known signaling effectors and cytoskeletal regulators. We investigated MINERVA/FAM129B, a target belonging to a protein family with unknown category and function, and established the importance of this protein and its MAP kinase-dependent phosphorylation in controlling melanoma cell invasion into three-dimensional collagen matrix.
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http://dx.doi.org/10.1016/j.molcel.2009.03.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2735263PMC
April 2009
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