Andres Stucky

Histone acetylation: novel target for the treatment of acute lymphoblastic leukemia

Acute lymphoblastic leukemia (ALL) has been generally considered a genetic disease (disorder) with an aggressive tumor entity of highly proliferative malignant lymphoid cells. However, in recent years, significant advances have been made in the elucidation of the ALL-associated processes. Thus, we understand that histone acetylation is involved in the permanent changes of gene expression controlling ALL developmental outcomes. In this article, we will focus on histone acetylation associated with ALL, their implications as biomarkers for prognostic, and their preclinical and clinical applications.

Increased expression of CX43 on stromal cells promotes leukemia apoptosis

Connexin 43 (Cx43) induced apoptosis has been reported in solid tumors, but the effect of Cx43 expressed by bone marrow stromal cells (BMSC) in leukemia has not been fully investigated. Manipulating Cx43 expression could be a potential therapeutic strategy for leukemia. Here, we investigate the effect of Cx43 expressed by BMSCs (human Umbilical Cord Stem Cells over-expressed CX43, Cx43-hUCSC) on leukemia cells. When co-cultured with Cx43-hUCSC, leukemia cells show significant lower growth rate with increasing apoptosis activity, and more leukemia cells enter S phase. Functional assays of fluorescence recovery after photo bleaching (FRAP) showed improved gap junctional intercellular communication (GJIC) on leukemia cells when co-cultured with Cx43-hUCSC (p < 0.01). In a mouse minimal disease model, the mean survival time and mortality rate were significantly improved in mice transplanted with Cx43-hUCSC. Our results indicate that Cx43 expressed by BMSC induces apoptosis on leukemia cells. Small molecules or other pharmaceutical approaches for modulating Cx43 expression in BMSCs could be used for delaying relapse of leukemia.

Human-derived normal mesenchymal stem/stromal cells in anticancer therapies

The tumor microenvironment (TME) not only plays a pivotal role during cancer progression and metastasis, but also has profound effects on therapeutic efficacy. Stromal cells of the TME are increasingly becoming a key consideration in the development of active anticancer therapeutics. However, dispute concerning the role of stromal cells to fight cancer continues because the use of mesenchymal stem/stromal cells (MSCs) as an anticancer agent is dependent on the specific MSCs subtype, in vitro or in vivo conditions, factors secreted by MSCs, types of cancer cell lines and interactions between MSCs, cancer cells and host immune cells. In this review, we mainly focus on the role of human-derived normal MSCs in anticancer therapies. We first discuss the use of different MSCs in the therapies for various cancers. We then focus on their anticancer mechanism and clinical application.

CCR5 Governs DNA damage repair and breast cancer stem cell expansion

The functional significance of the chemokine receptor CCR5 in human breast cancer epithelial cells is poorly understood. Here, we report that CCR5 expression in human breast cancer correlates with poor outcome. CCR5+ breast cancer epithelial cells formed mammospheres and initiated tumors with >60-fold greater efficiency in mice. Reintroduction of CCR5 expression into CCR5-negative breast cancer cells promoted tumor metastases and induced DNA repair gene expression and activity. CCR5 antagonists Maraviroc and Vicriviroc dramatically enhanced cell killing mediated by DNA-damaging chemotherapeutic agents. Single-cell analysis revealed CCR5 governs PI3K/Akt, ribosomal biogenesis, and cell survival signaling. As CCR5 augments DNA repair and is reexpressed selectively on cancerous, but not normal breast epithelial cells, CCR5 inhibitors may enhance the tumor-specific activities of DNA damage response-based treatments, allowing a dose reduction of standard chemotherapy and radiation.Significance: This study offers a preclinical rationale to reposition CCR5 inhibitors to improve the treatment of breast cancer, based on their ability to enhance the tumor-specific activities of DNA-damaging chemotherapies administered in that disease. Cancer Res; 78(7); 1657-71. ©2018 AACR.

Genetic mutations associated with metastatic clear cell renal cell carcinoma

Metastasis is the major cause of death among cancer patients, yet early detection and intervention of metastasis could significantly improve their clinical outcomes. We have sequenced and analyzed RNA (Expression) and DNA (Mutations) from the primary tumor (PT), tumor extension (TE) and lymphatic metastatic (LM) sites of patients with clear cell renal cell carcinoma (CCRCC) before treatment. Here, we report a three-nucleotide deletion near the C-region of Plk5 that is specifically associated with the lymphatic metastasis. This mutation is un-detectable in the PT, becomes detectable in the TE and dominates the LM tissue. So while only a few primary cancer cells carry this mutation, the majority of metastatic cells have this mutation. The increasing frequency of this mutation in metastatic tissue suggests that this Plk5 deletion could be used as an early indicator of CCRCC metastasis, and be identified by low cost PCR assay. A large scale clinical trial could reveal whether a simple PCR assay for this mutation at the time of nephrectomy could identify and stratify high-risk CCRCC patients for treatments.

Microfluidic enrichment of plasma cells improves treatment of multiple myeloma

Cytogenetic alterations form the basis for risk stratification for multiple myeloma (MM) and guide the selection of therapy; however, current pathology assays performed on bone marrow samples can produce false‐negatives due to the unpredictable distribution and rarity of MM cells. Here, we report on a microfluidic device used to facilitate CD45 depletion to enhance the detection of cytogenetic alterations in plasma cells (PCs). Bone marrow samples from 48 patients with MM were each divided into two aliquots. One aliquot was subjected to classic flow cytometry and fluorescent in situ hybridization (FISH). The other first went through CD45+ cell depletion, further enriched by microfluidic size selection. The enriched samples were then analyzed using flow cytometry and FISH and compared to those analyzed using the classic method only. Unlike the traditional method, the microfluidic device removed the CD45+ leukocytes and specifically selected PCs from the remaining white blood cells. Therefore, the microfluidic method (MF‐CD45‐TACs) significantly increased the percentage of CD38+/CD138+ cells to 37.7 ± 20.4% (P < 0.001) from 10.3 ± 8.5% in bone marrow. After the MF‐CD45‐TAC enrichment, the detection rate of IgH rearrangement, del(13q14), del(17p), and 1q21 gains, rose to 56.3% (P < 0.001), 37.5% (P < 0.001), 22.9% (P < 0.001), and 41.7% (P = 0.001), respectively; all rates of detection were significantly increased compared to the classically analyzed samples. In this clinical trial, this microfluidic‐assisted assay provided a precise detection of cytogenetic alterations in PCs and improved clinical outcomes.

Single-cell genomic analysis of head and neck squamous cell carcinoma

Head and neck squamous cell carcinoma (HNSCC) incidence or rates have increased dramatically recently with little improvement in patient outcomes. There is an unmet need in HNSCC to develop reliable molecular markers capable of evaluating patient risks and advising treatments. This review focuses on recent developments in single-cell molecular analysis of cancer, and its applications for HNSCC diagnosis and treatments. For proof of concept, we examined gene expression levels of 62 patients with HNSCC, and correlate the gene expression profiles to single-cell gene expression profiles obtained from a pilot single-cell study of CCR5-positive breast carcinoma cells. The single-cell molecular analyses complemented the lysate data and reveals heterogeneity of oncogenesis pathways with the cancer cell population. Our single-cell molecular analysis indicated that molecular heterogeneity exists in HNSCC and should be addressed in treatment strategy of HNSCC. Single-cell molecular technology can have significant impact on diagnosis, therapeutic decision making, and prognosis of HNSCC.Head and neck squamous cell carcinoma (HNSCC) incidence or rates have increased dramatically recently with little improvement in patient outcomes. There is an unmet need in HNSCC to develop reliable molecular markers capable of evaluating patient risks and advising treatments. This review focuses on recent developments in single-cell molecular analysis of cancer, and its applications for HNSCC diagnosis and treatments. For proof of concept, we examined gene expression levels of 62 patients with HNSCC, and correlate the gene expression profiles to single-cell gene expression profiles obtained from a pilot single-cell study of CCR5-positive breast carcinoma cells. The single-cell molecular analyses complemented the lysate data and reveals heterogeneity of oncogenesis pathways with the cancer cell population. Our single-cell molecular analysis indicated that molecular heterogeneity exists in HNSCC and should be addressed in treatment strategy of HNSCC. Single-cell molecular technology can have significant impact on diagnosis, therapeutic decision making, and prognosis of HNSCC.

VirTect: a computational method for detecting virus species from RNA-Seq and its application in head and neck squamous cell carcinoma

Next generation sequencing (NGS) provides an opportunity to detect viral species from RNA-seq data on human tissues, but existing computational approaches do not perform optimally on clinical samples. We developed a bioinformatics method called VirTect for detecting viruses in neoplastic human tissues using RNA-seq data. Here, we used VirTect to analyze RNA-seq data from 363 HNSCC (head and neck squamous cell carcinoma) patients and identified 22 HPV-induced HNSCCs. These predictions were validated by manual review of pathology reports on histopathologic specimens. Compared to two existing prediction methods, VirusFinder and VirusSeq, VirTect demonstrated superior performance with many fewer false positives and false negatives. The majority of HPV carcinogenesis studies thus far have been performed on cervical cancer and generalized to HNSCC. Our results suggest that HPV-induced HNSCC involves unique mechanisms of carcinogenesis, so understanding these molecular mechanisms will have a significant impact on therapeutic approaches and outcomes. In summary, VirTect can be an effective solution for the detection of viruses with NGS data, and can facilitate the clinicopathologic characterization of various types of cancers with broad applications for oncology. Significance Statement We developed a new bioinformatics tool, and reported the new inside of HPV carcinogenesis mechanism in HPV-induced head and neck squamous cell carcinoma (HNSCC). This novel bioin-formatics tool and the new knowledge of HPV-induced HNSCC will facilitate the development of target therapies for treating HNSCC.

Single-cell transcriptomes reveal the mechanism for a breast cancer prognostic gene panel

The clinical benefits of the MammaPrint® signature for breast cancer is well documented; however, how these genes are related to cell cycle perturbation have not been well determined. Our single-cell transcriptome mapping (algorithm) provides details into the fine perturbation of all individual genes during a cell cycle, providing a view of the cell-cycle-phase specific landscape of any given human genes. Specifically, we identified that 38 out of the 70 (54%) MammaPrint® signature genes are perturbated to a specific phase of the cell cycle. The MammaPrint® signature panel derived its clinical prognosis power from measuring the cell cycle activity of specific breast cancer samples. Such cell cycle phase index of the MammaPrint® signature suggested that measurement of the cell cycle index from tumors could be developed into a prognosis tool for various types of cancer beyond breast cancer, potentially improving therapy through targeting a specific phase of the cell cycle of cancer cells.

Gene Manipulation with Micro RNAs at Single-Human Cancer Cell

Micro RNAs (miRNAs) are small RNAs processed from longer precursor RNA transcripts that can fold back on themselves to form Watson-Crick paired hairpin structures. Once processed from the longer molecule, the small RNA is much too short to code for proteins but can play other very important roles, like gene regulation. The phenomenon of RNA interference was initially observed by Napoli and Jorgensen in transgenic petunia flowers, where gene suppression was observed after introducing a transgene of chalcone synthase (CHS) belonging to the flavonoid biosynthesis pathway. miRNAs were first discovered for their roles in development but it has quickly become evident that they have causal roles in cancer as well. miRNA can also be used to manipulate genes for the investigation of carcinogenesis. Single-cell transcriptome profiling studies in our laboratory suggest that carcinogenesis often is the result of the malfunction of multiple members of a molecular pathway. Here, we describe a protocol to manipulate multiple cancer-related genes in a single human cell to investigate how multiple genes interact during carcinogenesis.