How to use tumor organs for tumor drug screening
2020-06-07
Methods and reasons for the use of tumor like organs in the in vitro tumor drug screening program were reviewed.

Why use organ like screening for cancer drugs?

Before we go directly to how to use tumor like organs for drug screening, a key question to be answered is why do we use organs like organs - why not continue to use the standard in vitro drug screening platform currently used?


The main answer is clinical relevance. Typical drug development relies largely on immortalized cell lines for high throughput screening (HTS) and testing target binding and cell activity. Although this method has many advantages, it can lead to cell line genetic drift, poor translatability and disease correlation. These high rates of drug attrition have been observed in all tumors.

Although more predictive models have been developed using clinical materials, such as in vitro primary culture or in vivo patient xenograft (PDX) tumor models, these models are not suitable for HTS. For subsequent studies, the primary culture was limited by the number of available tissues and amplification.

PDX is stably cultured in vivo and provides a high level of translation platform for later drug development, including through mouse clinical trials (MCT). However, the time and cost of PDX development are too high, and these models have not been effectively translated for high-throughput in vitro use.

There is an urgent need for early robust and scalable screening platform to provide clinical relevant translatable data to guide drug development, patient selection and concomitant diagnosis (CDX) development as soon as possible. Organoids have the potential to fill this gap and provide in vitro preclinical models needed to overcome current screening challenges.

Tumor like organs

We have written extensively in previous announcements on the background and development of organoids and oncoids. In short, organ like organs from patients are produced in vitro from stem cells or progenitor cells grown in 3D. Organoids have the ability of self-organization and self-renewal, which leads to the development of "micro organs" in culture dishes. These organoids contain multiple differentiated cell lineages, which are significantly similar to the organs of origin in vivo.

Pioneering work by hubrecht organoid Technology (hub) has produced robust solutions for the development of organoids from adult stem cells from normal and diseased tissues. Tumor like organs of hub reflect the genomic, morphological and pathophysiological characteristics of their parent tumors. This provides a highly clinically relevant 3D in vitro tumor model for drug development.

The tumor like organs were cryopreserved to generate the model biological sample bank, and then expanded after resuscitation without losing the original identity of the organ like. This means that organoids are a good tool for understanding the molecular mechanisms of cancer and developing new therapies. Tumor like organs can also predict the treatment response of patients, so it can provide a more relevant in vitro model than the standard 2D in vitro platform.

Drug screening using tumor like organs

Establishing the largest tumor organ library to reproduce the humanization of tumors

To start using tumor like organs for tumor drug screening, the first step is to develop a large number of organ like models covering a wide range of cancer types, similar to the large number of cell lines preserved in HTS or PDX currently available for clinical trials in mice.

Historically, tumor like organs were directly derived from patients' tumors (patient derived organoids or PDOS). Although this is still a highly effective technique for producing closely related models for screening, developing the large number of models required can be time-consuming because the patient's tumor tissue must be obtained from a variety of different cancers.

A new method is to develop PDX derived organ like (pdxo) from a large number of well annotated PDX models in vivo. This provides a unique opportunity to amplify the full range of patient derived organ like models available, including a wider range of cancer indications with a range of mutation characteristics and pharmacological responses. This includes tumors that are not responsive or drug-resistant.

Pdxo is built from cancer stem cells found in PDX tumors, just as PDOS originate from patients' tumors. Based on the common genomic diversity, tumor heterogeneity, histopathology and drug response, mature pdxo and parental PDX are bioequivalent. Pdxo also maintained the stable genomic, morphological and pathophysiological characteristics of PDX tumors during multiple passages.

Therefore, this highlights that pdxo is a unique in vitro platform, which is characterized by the gold standard patient correlation of PDX models, the scalability of organoids from traditional patient sources, and the development efficiency of thousands of PDX deposits generated. This combination provides an ideal platform for large-scale drug screening in patient-related models, including combination strategy testing, biomarker identification and CDX development.

Pdxo screening workflow

Like other typical HTS screening, pdxo drug screening is an automated, highly reproducible and robust system designed to improve efficiency. The additional advantage of PDX screening in early drug development is high predictability.

From the main biological sample bank of cryopreservation model, pdxo was amplified into a working biological sample bank for HTS. These pdxo biobanks have been tested for Mycoplasma and analyzed for STR through sufficient storage, recovery and growth profiling analysis, so that the model can be reused. Specific organoids can be selected from the online organoid biological sample database. Then the selected pdxo was amplified and inoculated into the porous plate for drug screening.

Filtering works in a way similar to 2D filtering:

384 hole format
In general, the IC50 values of eight compounds are used to generate
There were positive and negative controls on each plate.
The test agent was incubated with organoids for 5 days, and the end point was celltier glo & reg; activity reading or morphological assessment. For multiple agents and multiple tumor models, it may take up to five weeks for the model to be amplified and incorporated into the study for IC50 readings and analysis. Endpoint readings can be extended to other physiological parameters, such as apoptosis or high-resolution imaging.

Main advantages of tumor drugs in drug screening

There are many advantages to the use of organoids in cancer drug screening, some of which are described above. Screening includes organoids that increase patient relevance due to retention of original and phenotypic features and 3D structure of tumors, all of which are known to influence drug response.

Tumor organ drug screening also provides high-quality readings, such as high signal-to-noise ratio and low intra plate variation, demonstrating the robustness of the platform.

Pdxo screening can be used to evaluate compounds and their combinations in a large number of tumor models. The research can then be transferred directly to in vivo modeling using the matched PDX model.

Differences between tumor like organs and standard 2D drug screening

Some differences from standard 2D screening need to be considered when planning organ screening.

As mentioned above, tumor like organs can be cryopreserved and then resuscitated for HTS incorporation. This may result in a slightly longer lead time than standard in vitro cell lines.

The current 384 format used for organ like screening is not as high as the standard cell line HTS. This is partly due to the amplification potential of organoids, because the culture and inoculation standards of organoids are different from 2D monolayer culture. For example, the size, density and morphology of organoids in 3D culture, as well as passage steps, take more time.

Therefore, it is necessary to have experience and professional knowledge of organ like culture in the design of screening scheme and genomic, histopathological and pharmacological analysis of tumor like organs.

Based on the clinical relevance of tumor like organs, it is clear that other in vitro screening tools cannot match the organoid screening data. In early drug development, the availability of such large-scale screening is exciting.

Conclusion
The lack of a preclinical model system that can reflect the primary patient's tumor is a major setback in early oncology drug development. The use of tumor like organs in vitro in large-scale drug screening can help overcome this problem and provide a robust, scalable and predictable 3D system to revitalize the current development workflow.

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