COLORECTAL CANCER PROGRAMS (CRC)
The laboratory has two CRC programs - a retrospective targeted next-generation sequencing study of histologically verified CRC tissues from Filipino patients, and a prospective study of a more restrictive and homogeneous group of sporadic young-onset CRC patients 45 years and under, with no identified familial predisposition to CRC and with a microsatellite stable (MSS) profile. In recent years, increased incidence of CRC in the young has been observed globally, with estimates ranging from 2 to 8.5% annually. Here, in the Philippines, incidence has been noted at 17%, which is substantially higher than most countries. This includes both the sporadic cases as well as those with identified hereditary predisposition to CRC.
Based on related studies elsewhere, the mutational signature of this subset of CRC patients is different from that of late-onset cases and there appears to be differences as well between ethnic groups. Both programs focused on mutations in the KRAS, NRAS, BRAF, PIK3CA and PTEN genes. The novel mutations were/are being functionally characterized to determine if they are harmless polymorphisms, activating mutations, and/or mutations conferring an aggressive metastatic phenotype
circRNAs
Exosomes, lncRNAs, miRNAs, circRNAs and ceRNAs
The laboratory has major interests in the regulatory roles of long non-coding RNAs (lncRNAs), microRNAs (miRNAs), circular RNAs (circRNAs) and competing endogenous RNAs (ceRNAs) in cancer. We have shown, for instance, that miR-92a directly targets NF2 and affects its tumor suppressive function. The lab has also identified and validated a novel ceRNA network that includes the mRNAs coding for the tumor suppressor PTEN and the methylation- associated proteins DNMT3B and TET3.
Other ncRNA projects thesis students are working on include the effects of naturally occurring miRSNPs on the tumor suppressive role of the lncRNA MEG3; the effects of 3'UTR miRSNPs on the oncogenic or tumor suppressive roles of EGFR pathway genes; the effects of the cigarette-smoke upregulated SCAL1 lncRNA on cytoskeletal remodelling, other oncogenic phenotypes, and ROS detoxification; functional characterization of the novel e-cigarette-upregulated lncRNAs VALT-1 and SLBP-DT; the circPVT1/let-7/NRAS regulatory axis; the pseudogene INGX; and the lncRNAs CRNDE, TUSC7, HULC and DANCR, among others.
Lastly, the lab has an ongoing project on differential sorting of exosome cargo. In particular, we are looking at the effect of KRAS mutational status on sorting of lung-cancer associated lncRNAs in exosomes released by lung adenocarcinoma cells.
CONFIRMATORY AND ORTHOGONAL ASSAYS TO ELIMINATE ARTEFACTUAL DRUG BIOACTIVITIES
The laboratory participates in a nationwide drug discovery program that seeks to leverage the Philippines' rich biodiversity. An important component of the program is the incorporation of confirmatory orthogonal and/or secondary assays in the workflow. These assays are able to remove false positives and artefactual bioactivities by replicating the exact same biology revealed by a primary assay in an independent assay of a different readout, e.g., fluorescence, luminescence, or imaging as alternatives to a colorimetric readout. Most of these assays are cell-based, thus providing the necessary physiological context. The use of high-content analysis and phenotypic screening, enabled by a high-content imaging system, also allows high throughput conversion of visual observations to quantitative data. In practical terms, failing early by identifying these potential dead-ends will mean considerable savings in time and money and will help re-channel efforts towards legitimate drug leads.
BRIDGING EFFICACY AND SAFETY: IND-ENABLING SUITE OF ADME-TOX ASSAYS
The laboratory is part of a 5-component program that offers ADMET assays to help identify the best iteration of lead compounds in the national drug discovery pipeline. The suite of assays constitute the bulk of ADMET package that will be required by regulatory authorities for an investigational new drug application (IND). The lab does cell-based assays including Caco-2 cell permeability and P-gp transporter inhibition assays; CYP450 inhibition, induction and reaction phenotyping assays, as well as hepatotoxicity, nephrotoxicity, hERG cardiotoxicity, neuronal toxicity, endothelial toxicity, skeletal muscle toxicity, and Integrated Discrete Multiple Organ Culture (IDMOC) toxicity assays. The laboratory works closely with the physico-chemical properties and mass spec groups at the Institute of Chemistry. Selected in vivo ADME assays, are also being done by a collaborating lab and will provide valuable information such as drug oral bioavailability, exposure, distribution, clearance, and duration of exposure for a drug and its metabolites. Another collaborating lab is in charge of doing imaging mass spectrometry, to confirm the identity of a drug or its metabolites, as well as their spatial distribution in tissue sections such as those of liver (main site of metabolism) and kidney (main route of drug excretion).
X-linked Dystonia Parkinonism (XDP)
X-linked Dystonia Parkinsonism is a hereditary, middle-onset (third or fourth decade of life) neurological disorder affecting mostly males whose maternal roots can be traced to Panay Island in the Philippines. The disease manifests firstly as Dystonia, followed by a combination of Dystonia and Parkinsonism, before finally being fully characterized by Parkinsonism.
Quite understandably, there are only a handful of laboratories working on XDP, given its narrow ethnic and geographic distribution. Most of the pioneering work on the disease were done here in the Philippines by Dr. Lillian Lee et al in the ‘80s and 90’s, with help from collaborators in the US, Japan and Germany whose interests encompass Dystonia, Parkinson’s and other movement disorders. Efforts towards understanding its molecular pathogenesis, however, have been confined to these three countries. Most efforts take on global approaches using techniques such as RNA-Seq, CHIP-Seq, microarray and other tools that yield disease signatures followed by pathway analysis. More focused functional studies on TAF-1/N-TAF1 and other genes within the disease locus are very limited.
This project aims to fill in some gaps in our understanding of the molecular basis of XDP, with middle-onset neuronal loss and downregulation of N-TAF1 as the guiding clues.
