Bo R. Rueda, PhD

Massachusetts General Hospital

Academic Titles

Associate Professor, Obstetrics, Gynecology and Reproductive Biology, Harvard Medical School

Research Program Affiliation

Member, Gynecologic Cancers

Research Abstract

The Rueda lab is involved in basic, translational and clinical research focused on women’s reproductive health. The lab specializes in reproductive tract and cancer biology, with an emphasis on the investigation of exogenous or endogenous factors that contribute to infertility, benign gynecologic diseases, malignant transformation of gynecologic tissues, and mechanisms driving drug resistance in recurrent gynecologic cancers. A secondary initiative of the Rueda laboratory has been to increase the number of samples collected by our fertility and gynecologic biorepository. The samples are needed for developing pre-clinical models for our translational investigations. In addition, it serves as a valuable resource for developing diagnostics for early detection of disease and or biomarkers to inform options for targeted therapy or treatment response.

Research Projects

PETAL (Plastics Exposure & Tubal-associated Lesions): Environmental contamination by plastics and their eventual breakdown into micro- and nanoplastics (MNPs) presents a global climate and health crisis. MNPs can enter the human body through ingestion, inhalation, or physical contact and are found in circulation and in various human organs, including the lungs, brain, gut, and reproductive tract. MNPs may negatively impact human health through physical interactions and/or cellular uptake, leading to inflammatory signaling, DNA damage, and potential carcinogenic exposure. MNPs have been found in follicular fluid (FF), which is rich in hormones, cytokines, reactive oxygen species, and more. FF normally supports the oocyte, signals follicular rupture for ovulation, and promotes subsequent wound healing. Incessant ovulation, and thus the repeated exposure to FF, has been linked to high-grade serous ovarian cancer (HGSOC), the most common form of ovarian cancer, by promoting inflammation and inducing DNA damage that may lead to eventual chromosomal instability. HGSOC has recognized precursor lesions originating from fallopian tube epithelial (FTE) cells, including p53 signatures and Serous Tubal Intraepithelial Carcinomas, that are more prevalent in patients harboring BRCA mutations. We hypothesized that MNPs found in FF can cause nucleotide or organelle damage in FTE, potentially leading to the generation of HGSOC precursor lesions. Thus, our objective is to interrogate the presence of micro- and nanoplastics in FF and determine their potential to promote aberrant change in fallopian tube epithelial cells that might lead to precursor lesions.

Determining the role of the retrotransposon LINE-1 (L1) in Ovarian Carcinogenesis: High-grade serous ovarian cancer (HGSOC) is one of the most lethal gynecologic malignancies, largely due to its silent progression and late clinical presentation. Studies have shown that a majority of HGSOC initiates in the secretory epithelium of the distal fallopian tube rather than on the ovarian surface. Despite these advances, the earliest events that trigger malignant transformation in the tube remain poorly understood. Repeated ovulation has been strongly associated with increased risk of HGSOC, suggesting that physiological processes linked to ovulation may inadvertently damage the fallopian tube epithelium over time. During ovulation, human follicular fluid (hFF), enriched with inflammatory mediators, reactive oxygen species, and proteolytic enzymes, is released near the fimbriae of the tube. Exposure to these factors can induce DNA damage and replication stress, which are alterations characteristic of early precursor lesions. Our preliminary studies have suggested that this stress environment may activate L1 retrotransposons, a mobile genetic element normally silenced in healthy adult tissues. Aberrant L1 expression is associated with the formation of DNA double-strand breaks, chromosomal instability, and impairment of p53 tumor-suppressor function, all of which are recognized hallmarks of early serous tubal carcinogenesis. These observations have led to the hypothesis that ovulation-associated derepression of L1 contributes causally to the initiation of or progression of tubal precursor lesions linked to malignant transformation. To investigate this hypothesis, advanced experimental model systems, including fallopian tube epithelial patient-derived organoids, immortalized tubal epithelial cell lines, and genetic engineering to control L1 expression, are being used. These models and modifications allow controlled evaluation of genome integrity, retrotransposon activity, and cellular phenotypic shifts following exposure to human follicular fluid. High-resolution genomic analyses and ultrasensitive protein-detection technologies are being employed to define the timing and consequences of L1 activation at the earliest stages of transformation. This aspect is being conducted in collaboration with Dr. Kathy Burns at the Dana-Farber Cancer Institute and Dr. Martin Taylor at Brown University. By elucidating how physiologic reproductive processes influence genomic instability in the fallopian tube, this research aims to identify molecular events that precede cancer development. The discovery of L1-driven genomic alterations may ultimately support new strategies for prevention and early detection, enabling clinical interception of ovarian cancer before lethality becomes inevitable.

Identifying and impeding inherent and acquired mechanisms of BCL-XL-mediated drug resistance in endometrial cancer: Uterine cancer has recently surpassed ovarian cancer in lethality, becoming the deadliest gynecologic cancer in the United States. This is due, in part, to an increase in uterine cancer deaths associated with the more aggressive subtypes. Although uterine serous cancer (USC) is considered rare compared to the more common endometrioid subtype, it accounts for 40% of all uterine cancer deaths due to its metastatic potential and high likelihood of developing drug-resistant recurrent disease. Inhibitors of apoptosis, a programmed cell death process, have long been suspected in the genesis, progression, and drug resistance of many solid tumors. While the antiapoptotic Bcl-2 protein family members are postulated to contribute to drug resistance in USC, their role is not yet known. Solid tumors reportedly depend more on BCL-X Long (BCL-XL) than on other antiapoptotic BCL-2 proteins. Unfortunately, drugs targeting BCL-XL are known to have severe toxicities. Consequently, a PROteolysis TArgeting Chimera (PROTAC) against BCL-XL, currently being tested in ovarian cancer, is being tested in in vitro and in vivo preclinical models of USC. Using established USC cell lines and USC patient-derived organoids (PDOs), we are testing the effect of a PROTAC targeting BCL-XL as monotherapy and/or in combination with standard-of-care cytotoxic agents, biologics, and antibody-drug conjugates (ADCs). The endpoint analyses included MTT, Cell Titer Glo (CTG), flow cytometry, and BCL-XL by immunoblotting. Drug synergy was determined using a zero-interaction potency model. To validate our in vitro findings, we are conducting in vivo experiments using a mouse xenograft model. We aim to demonstrate that combining a BCL-XL targeting PROTAC with current treatment strategies has the potential to overcome recurrent, inherent, or acquired drug-resistant USC.

Defining mechanisms of ADC resistance in high-grade endometrial cancer: High-grade uterine cancer, unlike many other solid tumors, is on the rise. Moreover, the incidence of mortality rate is increasing, which is attributed, in part, to limited options for a durable treatment of recurrent drug-resistant disease. Antibody-drug conjugates (ADCs) are a rapidly emerging treatment strategy for advanced recurrent endometrial cancer. ADCs are designed to exploit antibody specificity to bind an antigen on a tumor cell and deliver a cytotoxic or other bioactive payload that would be intolerable if delivered systemically. ADCs are known to be highly effective in other solid tumor types; mounting evidence suggests that tumors can develop resistance. Our objective is to be proactive and investigate how high-grade endometrial cancers might develop resistance to ADCs, with a focus on therapies targeting the HER2 and FLRα receptors. Although ADCs, such as trastuzumab deruxtecan (TDXd), show promising activity in HER2-positive endometrial cancers, the mechanisms of resistance in endometrial cancer remain undefined. While no ADC has been explicitly approved for this disease, several new ADCs are in the pipeline. Consequently, we plan to use our growing number of patient-derived organoids with varying HER2 or FLRa expression to elucidate the cellular mechanisms underlying ADC resistance in high-grade endometrial cancer, encompassing alterations in antigen presentation, receptor density, signaling, ADC uptake/internalization, extracellular vesicle export, drug efflux pumps, and impaired apoptosis.

Defining how extracellular vesicles facilitate the cell-to-cell transfer of stem-like properties driving drug resistance: It is well accepted that ovarian cancer stem cells (CSCs) can seed recurrent drug-resistant disease. Similarly, it has been shown that non-CSCs can acquire CSC-like phenotypes in response to treatment. Understanding how this process is mediated is critical for informing how it might be prevented. We are testing the hypothesis that ovarian CSC and/or drug-resistant tumor cells confer stem-like properties via extracellular vesicles (EVs). Specifically, we are investigating how EVs might mediate Enhancer of Zeste Homolog 2 (EZH2) signaling to promote a phenotypic change in drug-sensitive, non-CSCs. To accomplish this, we utilized paired PARP inhibitor-sensitive and -resistant ovarian cancer cell lines, EZH2 knockdown lines, and patient-derived organoids (PDOs) originating from recurrent high-grade serous ovarian cancer. Small EVs isolated from drug-sensitive, CSC and/or drug-resistant enriched cultures, PARP inhibitor (olaparib) resistant lines, or drug-treated (olaparib or carboplatin) lines were cultured with treatment naïve or sensitive lines for defined time points. We are assessing the impact of small EV exposure by assessing cell number, metabolic activity, viability, sphere and colony-forming capacity, ALDH activity, DNA damage, and changes in associated signaling pathways. To date, we have found that EVs from CSC-enriched or drug-resistant cell fractions communicate CSC-like phenotypes to more sensitive tumor cells via EZH2 canonical and non-canonical signaling pathways, promoting stemness. Our data suggest that EV-mediated activation of EZH2 signaling represents a targetable mechanism contributing to stemness-associated drug resistance in ovarian cancer. We are currently exploring targeted strategies to disrupt this EV-mediated communication.

Identification of Liquid Biomarker and Diagnostics for early detection and monitoring of gynecologic malignancies: The Rueda Lab, along with their collaborators, Drs. Castro, Im, and Lee from the Center for Systems Biology at MGH, and Dr. Oladapo, a member of the VCRB and MGB Cancer Cener are are heavily invested in using the blood samples in our biorepository to develop diagnostic platforms for the detection of early onset and recurrence of gynecologic cancers, as well as identifying novel companion biomarkers to inform which patients might benefit most from specific treatment strategies. The team's focus has been on circulating extracellular vesicles (EVs), small lipid-bound nanoparticles, shuttling cargo into and out of cells, and mediating cell-to-cell signaling. The EVs contain proteins, lipids, and nucleic acids (i.e., lncRNA, miRNA, mRNA, and DNA). The team is developing strategies to detect, isolate, and enrich for tumor-derived EVs, aiming to improve the detection of gynecologic cancers and serve as companion biomarkers to inform treatment. Define mechanisms that contribute to the genesis, progression or pathology of benign gynecologic diseases: In addition to in depth studies on gynecologic malignances we also focus on benign diseases that impact reproductive aged women. Specifically, we have and continue to be focused on endometriosis and leiomyoma. These non-malignant diseases can have a devastating negative impact on women’s health and quality of life. Despite the prevalence of the disease very little progress have been made in long term solutions with the exception of surgical removal of the of uterus. We have used mouse models as well as primary human tissues to assess the mechanisms by which specific cell signaling factors positively or negatively impact the development, progression and/or pathological properties associated with these diseases.