The project began with installing state-of-the-art equipment, recruiting staff, and obtaining one of the largest collections of cell lines in academia. Our first milestone was to disseminate guidelines for good practice in studying the effects of acidosis on cancer cells. These instructions enabled well-controlled experiments, including our milestone identification of genes that are essential for surviving acidity. For this, we performed a screen to test which genes confer resistance to acidosis, identifying NDUFS1, part of a complex required for mitochondrial respiration. We showed that blocking respiration can selectively eliminate the survival advantage of acid-resistant cells, causing a stable disruption to acid-driven selection. We explained why respiration is essential in terms of its robustness under acidic conditions, which is in stark contrast to the acid-sensitivity of the other major energy and resource harnessing pathway, glycolysis. Additionally, we described how the pH-regulatory apparatus of cells adapts to acidic conditions through a novel mechanism that degrades acid-loading transporters, a process that normally operates to prevent excessively alkaline conditions inside cells. Further work classified cancer cells according to their acid-sensitivity, and associated resistant lines with the expression of CEACAM5 and CEACAM6 on the surface membrane. The significance of this marker is in acting as a moiety for targeting drugs specifically to acid-resistant cells, the ones that are most likely to survive selection by acidity. Our work also described how cancer cells interact through connections called gap junctions, which confers a novel level of resilience by allowing neighbours to compensate for each other’s deficiencies. This discovery challenged the paradigm that individual cells are the substrates of selection, and made a case for cell-networks. We described methods for extracting cells according to metabolic activity, and characterised an intrinsic rhythm-generator that causes cell populations to show metabolic heterogeneity, a survival advantage when resources are limited. Our collaborative work with Moffitt Cancer Center described how lymph nodes are acidic and how this attenuates T cells, a type of immune cell important for killing cancer cells. This discovery provided new insight into the mechanisms by which acidic tumours evade immune surveillance, and how this can be overcome. We developed a method to study oxygen transport by red cells, deployed it to study red cell quality in blood banks and anaemias, and developed algorithms to understand its impact for cancer oxygenation and pH. Our molecular investigations identified new pathways in which acids influence gene expression, ranging from rare diseases to general mechanisms relevant also to tissue development. These findings were disseminated through 23 scientific publications, including reviews and conference presentations.
