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Rational combination therapies for metastatic cancer

Periodic Reporting for period 2 - CombaTCancer (Rational combination therapies for metastatic cancer)

Reporting period: 2019-09-01 to 2021-02-28

"In their lifetime, 1 in 2 men and 1 in 3 women are diagnosed with invasive cancer. The perpetuation of a tumor hinges largely on cancer cell-intrinsic signals maintaining tumor growth and the tumor’s ability to evade destruction by the immune system. These dependencies are exploited by targeted therapies, which inhibit cancer cell-intrinsic signaling pathways, and immunotherapies, which unleash an immune response against cancer cells. Targeted therapies and immunotherapies have shown remarkable success in subgroups of patients, but therapy resistance and low response rates pose daunting challenges. The combination of different therapies is a promising path to overcome low response rates and acquired resistance. However, rational combination and optimal sequencing of therapies are hampered by a lack of molecular insight into resistance mechanisms and the interplay of therapies. The overall aim of ""CombaTCancer"" is to (1) understand how resistant cells emerge and (2) to gain insights into how oncogenic pathways, and consequently their inhibition using targeted therapies, impact the immune cells in the tumor microenvironment (TME). Our vision and mission is to provide a scientific basis for guiding rational combination therapies and to identify the ideal sequence of therapies to achieve durable responses in cancer patients."
"Tumors consist of a mixture of clones with different biological properties. In a Darwinian fashion, environmental pressures such as immunity or therapies select for the fittest clones. During these evolutionary selection processes, which can be strongly influenced by the residual tumor mass (Obenauf et al, Nature, 2015), clones change their abundance, identity, and cellular properties. A long-standing question in cancer biology is whether resistance mechanisms (1) are already present in a minority of cancer cells before therapy (‘pre-existing, resistant clones’), (2) develop in ‘resistance-prone clones’ which possess intrinsic properties that either provide a survival benefit (‘drug-tolerant persisters’) or facilitate the rapid adaptation to the new selective pressure, or (3) may stochastically develop during therapy in any clone. The functional characterization of founder clones and direct comparison to their post-selection counterparts can give new insights into this question. However, methods for efficient isolation of viable founder clones in heterogeneous cell populations prior to evolutionary selection processes have not been available.

To better understand the emergence of resistant cells within the framework of ""CombaTCancer"", we developed a novel functional lineage tracing tool that acts as a molecular time-machine termed CaTCH (CRISPRa tracing of clones in heterogeneous cell populations) (Umkehrer et al., Nature Biotechnology, 2020). CaTCH combines precise mapping of the lineage history of millions of cells with the ability to isolate any given clone alive from a complex population based on genetic barcodes. Thus, CaTCH enables the retrospective isolation and analysis of founding clones from heterogeneous cell populations before evolutionary selection with a remarkable resolution of 1:50.000 (0.002%) and purity (98%, ~20,000-fold enrichment). In first applications, we used CaTCH to provide insights into resistance to targeted cancer therapies in vivo. In our model, we find that most clones can acquire resistance to combined RAF/MEK inhibitor therapy, indicating that resistance to this clinically relevant regimen is a universally achievable state. We found a de novo KrasG12R mutation in a resistant clone, but not in its treatment-naïve counterpart. Our results provide experimental evidence that mutations are acquired during drug treatment, which is difficult to prove using prior methods, and challenges the current notion that mutations conferring drug-resistance generally pre-exist before therapy. We and several other collaborators are currently exploiting the technology in a wide range of topic related to cancer therapy, metastasis and stem cell behaviour.

To functionally dissect tumors and their microenvironment at different stages of cancer progression, including an active therapy-response and therapy-resistance, we have established novel clinically relevant human and murine model systems that recapitulate key stages of disease progression (Haas et al. in revision). Moreover, we are utilizing genome-wide CRISPR screens to assess mechanisms of immune evasion and sensitization to immunotherapies (Holstein et al. in preparation). For cancer types with no known genetic vulnerabilities, we are investigating whether lineage-specific dependencies could be used for treatment, in particular epigenetic regulators, which govern cell fate, provide unexplored therapeutic entry points. We have recently identified Lysine-specific histone demethylase 1A (LSD1, also known as KDM1A) as a new therapeutic target in MCC in vitro and in vivo (Leiendecker, Jung et al., EMBO MM, 2020)."
Our work within the framework of CombaTCancer may not only provide new therapeutic entry points for metastatic cancer but also give fundamental insights into physiological processes, such as immunity and gene regulation.
Artistic interpretation of the CaTCH approach developed as part of the ERC grant CombaTCancer