Project Summary/abstract: HPV associated cancers have increased significantly (>16%) in USA, as per the
latest data. Resistance to the current chemotherapy leads to a high recurrence rate and patient death. This
resistance has largely been attributed to inter-and intra- tumor heterogeneity. The identification of new
therapeutic agents is urgently needed. The hallmark of HPV cancer is a highly expressed E6/E7 genes of high-
risk HPV types. Their expression is necessary for the survival of cervical cancer cell lines. These viral oncogenes
are known to dysregulate host DNA damage responses, apoptosis, metabolism, mitotic checkpoint, and immune
responses. We propose that inhibitors intervening one or more of these targets in combination with current
chemotherapeutic agent, cisplatin, would increase efficacy and reduce morbidity and mortality. NCI encourages
drug testing in patient derived cervical tumor xenografts (PDXs) in mice, rather than cancer cell lines. The
establishment and inhibitor screening in these model is time consuming, labor intensive and costly, limiting the
scope of testing the candidate inhibitors or their combination to predict patient-specific treatment. We have
established a novel method to culture cervical cancer as organoid raft cultures as an in vitro tumor model. The
organoids can be used to initiate PDX in SCID mice. Conversely, PDX can be grown as organoids. Here, we
propose to combine the PDX model with the ease and economy of organoid raft cultures for initial evaluation of
new candidate inhibitors. In the first Aim, we propose to validate similarities in phenotypic, genetic, and molecular
characteristics among organoids, organoid-derived and patient-derived PDXs in SCID mice and the founder
tumors. Significant and precious time and costs will be saved in preclinical investigation of new therapeutic
strategies, if the organoid raft culture displays high similarity to the PDX and parental tumors. The HPV E6/E7
oncoproteins dysregulate the cell cycle regulation, the DNA damage and Spindle assembly pathways and
metabolism, basis for viral carcinogenesis. The second aim is to evaluate a panel of 6 inhibitors against novel
protein targets (Hec1, KIF-11, Aurora A, TTK, HSP90B1, and O-GlcNac Transferase) in the above named
pathways. Our preliminary study showed that each was critically required for the survival of cervical cancer cell
lines in submerged cultures or HPV transformed cells in vivo. We will also test Vorinostat, a pan HDAC inhibitor,
which we found to be promising in our preliminary study. These inhibitors alone or in combination with cisplatin,
which is the most commonly used chemotherapeutic agent to treat cervical cancer, will be evaluated in vitro in
organoid raft cultures of PDX derived from four independent cervical cancers and in corresponding PDXs in vivo
for comparison. The results are expected to validate organoid raft culture as an economical and expedient
method of drug evaluation prior to validating in vivo. Additionally we will establish new cc PDXs for future
evaluation of agents. These studies will reveal whether the incorporation of these new pathway-specific inhibitors
increased the efficacy of current chemotherapy and are candidates for further preclinical investigation.