On a scale of 1 to 5 (1=definitely not feasible; 3=too uncertain to guess, 5= highly feasible), do you think it will be feasible to rely solely on non-animal methods (NAMs) for safety testing of chemicals, pesticides, pharmaceuticals by 2035? (please feel free to explain)

Complex biological processes can only be studied in living organisms. Modals are only as good as the information used to program them.

The technology is here. Introduction to new practices, training and education of scientists need to be implemented.

Reliable non-animal methods still need to be developed in the coming years. For example, mathematical models that can predict effects of complex chemical mixtures are still lacking.

Animal models had many many similarities with humans. Unless we study animal models, it would be challenging to get exclusively suggestive data. The biochemical, enzymatic, and physiologic activities may not be controlled in tissue or other models as this may not represent the whole body system.

It will not be feasible. This is because animals have similarities in terms of their genetic makeup to the man. So testing of chemicals etc using non-animals alone may lead to exclusion of results that could be obtained if animals were used for the research.

It is going to be a political decision. As the humankind lived for a long time without any toxicity testing (except trial - error), it will live also with solely non-animal testing methods. However, not without consequences such as losses of reliability (and safety) and a huge cost increase.

Not only single chemicals, drugs and foods will still require animal tests, but we need to consider the impact of possible of factorial groupings of them. Curtailing the use of animals should be applied on projects with insuficient societal merit, or on ill-founded innovation.

Animals have different physiology from humans and often side effects in humans could not be determined by animal testing. However, much work is needed to design suitable non-animal testing particularly for safety studies.

I do not agree to rely upon NAMs resources.

I believe artificial intelligence/machine learning will help us predict the effect of a chemical on mammalian cells.

YES

The NAMs would be better for action on bio-physics properties of susbstance but one need to confirm the biological action which is cumulative effect of its biological, chemical and physical properties more so variation of which Animals undergoes in day to day life.

Animal testing will be necessary to advance science. Some parts cannot be replaced, such as the immune response and blood circulation for testing. The generation of knockout mice is also still needed in current cancer research.

Many activities point into this direction. Further work is needed to convince the scientific community including regulators of the merits of NAMs. These need to be developed further and standardized. It needs to be defined which NAM exactly should be used to adress which question. Regulators should be asked to define which NAM outcome they are ready to use in their decision making.

as above

We already know that one cannot translate from cell systems to humans with any accuracy. Humans are a highly, complex multi-integrated organism that cannot be represent by non-animal approaches.

Animal models still provide valuable insights that non-animal methods can not provide. For example, organ development. It is not feasible to completely remove animal methods.

Currently, testing on laboratory animals of the toxicity of chemical species or drugs is still too widespread to believe attainable the goal of carrying out all the experimentation on NAM by 2035

I think it relatively not feasible to rely solely on non- animal testing

We should strive to move all testing to NAMs, more should be debated/discussed about a given cutoff date.

Yes, but we need investment into the AOPs and NAMs. We also need to understand what we are validating, its not always about comparing animal tests with NAMs.

Why not each physiological changes can be observed using respective enzymatic level detection

Testing on the target species is difficult to avoid: Alternative to pesticides to cite just this example, are necessary to protect crops and human from pest insects. Seeking alternatives require testing these alternatives on the target species before to apply them in nature. Regarding pharmaceuticals and other chemicals targeted at humans, these should be tested only on humans only and thus alternatives to animal testing must be found.

I believe it would be possible well before then with full commitment.

I silico or non-living systems are way off and lack alot of credibility

I don't think so because of we face alot of Resistance from some scientists

Validating human clinical trials without carrying out previous animal tests is not correct in my opinion. The anatomical and functional similarity of several mammals used in scientific research with humans, allows adjusting the criteria of clinical trials to increase the chance of performing them within the therapeutic window of the studied compound.

NAMs still have huge gaps that cannot fully depict a chemical's fate in biological systems.

2035 is not too far away and while we have made good progress with in vitro models as a scientific community more integrated and prolonged effort (both integration of agencies/industy and academic research/industrial research as well as integration of those in vitro tissue systems) is needed to make this goal a reality. The integration of tissues towards capturing systemic as well as multi-tissue effects is critical yet very complex for relying solely on alternative methods. While I am enthusiastic about the end goal, I am not entirely sure about the timeline. Still, any effort towards replacing/reducing animal use is commendable for two reasons a) we don't only want to keep solving animals' problems, we also want to solve human problems b) we don't want to waste money and keep going back to the drawing board.

There are too many instances for crosstalk between systems to fully model in a dish .

I support the direction of the above-quoted memo of the EPA’s Administrator in 2019 concerning animal study requests, but "completely eliminating...mammal study requests...by 2035" may or may not be justified by 2035. The need for organismic testing for product safety can at the present state of knowledge not yet be rigorously excluded for any precisely defined time point in future. Lets remain modest in order to allow care for human safety and not excusively for animal well-being.

Si tratta di composti che in massima parte non coinvolgono gli aspetti per i quali un organismo intero è insostituibile per la valutazione di tossicità,stabilità,bioaccumulabilità

there will still be tests that single cells or other cultivated materials cannot replace, such as knocking out genes in mice or brain reactions.

In vitro and non-mammalian systems will take decades to be fully validated

This answer is case specific. If there is sufficient existing evidence for a thorough read-across, the mode of action is reasonably understood, and the NAMs are comprehensive and reliable, then non-vertebrate testing and registration is possible by 2035.

Support for additional research and application in case studies is necessary, in addition to the development of regulatory guidelines and policies.

I am confident that many drug targets will be associated with a good surrogate marker/model

Toxicity can be tested in fruit flies

Elimination of animal testing is likely to lead to overly conservative decision making and may still miss chemical agents at potential risks to humans.

HT assays, advanced modelling and big data (from all different kinds of studies) will be evolved in a way that we can simply achieve at least the same level of certainty that animal testing provides.

The current ex vivo or in vitro organ-on-a-chip methods are not verified. They are extremely heterogeneous from lab to lab. There are no current standards or guidelines. Unless there are standards and guidelines in place, and models are standardized, we are not ready to stop animal testing, in particular for pharmaceuticals. Perhaps chemicals and peticide testing can be eliminated first.

Cell line models were preferred to understand the molecular mechanisms of any toxicant and pharmaceuticals.

A transition period is needed in which NAMs and animal tests are used simultaneously.

Given the high lack of reproducibility in the biomedical literature, it will be very difficult to establish many AOPs with sufficient confidence to replace many in vivo adverse effects using in vitro assays. In addition, there are some adverse effects that occur only rarely that will take significant investments in money and time to develop AOPs, for example, a fetal aortic arch aneurism, it won't be practical to develop in vitro models for every adverse effect. We can reduce the number of animals used in tox testing for general adverse effects, but for adverse effects through complex biological mechanisms such as developmental toxicity and neurobehavioral toxicity this will take years beyond 2035

At the same time, we can measure omics at many levels much more precisely and more reproducible that these can ack as model system at least equally well. Given we can sufficiently stabilize these models (which I believe we can), then we can correlated their output with the overall toxicity and safety of chemicals. The results do not have to replicate what we found with animals; they have to predict the human safety.

The lack of a strong scientific basis for the 100x factor used in toxicology safety assessment shows how rudimentary the understanding of toxicology is.

It is possible, but we need openmindness, investment and willing to reach the goal

The use of animals is ubiquitous. If the western scientists adopt the use of non-animal methods, legislative component could be a problem world wide.

Non- animal testing includes sophisticated tests using human cells and tissues (in vitro methods), advanced computer-modeling techniques (in silico models), and studies with human volunteers. These and other non-animal methods make applying animal test results to humans possible. However, animal methods are necessary for safety testing of some chemicals, pesticides and pharmaceuticals, and such testing is unethical if conducted on humans.

I worked for EPA when the comptox initiative began some so years ago. I participated in some "validation" studies and except for some of the endocrine disruptor comparisons, the comptox modeling simply were of marginal value. Indeed, there remains to be a comprehensive assessment of drugs/chemicals of which we have substantial understanding across the comptox test methods and adverse outcome pathway models, select animal models and humans (based on long history of use). There is considerable resistance to doing this and I don't know why. Their sexiness alone is not enough for me to see some chemical or drug ultimately sieve through to the mouths of children or ailing individuals. One would not create a climate model by taking the temperature, windspeed, and rainfall in one's neighborhood, no matter how detailed those measurements are. Biology and climate are inherently complex systems with feedbacks and unknowns which require more study. In the case of health, we at least have animals model test systems to add to our understanding.

Alternative endpoints such as skin sensitivity have already been developed, adopted and used by the Agency and for these my score would be 5. For most target organ endpoints such as liver, kidney, lung etc. the biology is fairly well understood and great progress to apply this knowledge to develop alternative tests including multicellular and differentiated human cell based 3-D models. I have great confidence that alternative tests will be developed, evaluated and found to be sufficiently reliable to inform regulatory decisions by 2035 too. However, for more complex multifactorial endpoints whose biology is not as well understood such as cancer, and birth defects barring major, unexpected scientific breakthroughs I do not have as much confidence that acceptable alternatives will be able to be developed, evaluated and deemed to have sufficient confidence that their results will be used by regulators to inform all decisions by 2035. I do have high confidence that this will be the case by 2050 or perhaps before

Not at all, there is a lot of factors that are lost with non-animal testing that only animal testing can give you. There is no way at our current level of understanding that you can eliminate animal testing and also protect the public from harmful chemicals at the same time. It is a nice idea, but it will shift the test subject from controlled animal experiments to uncontrol humans as there will be much data lost, making public health safety decisions much less informed.

This is certainly a highly ambitious goal; however, the growing level of commitment and engagement within the scientific community around development of NAMs gives me a lot of hope that we will make tremendous progress. That being said, it is difficult to say we will definitely reach this objective within the time-frame.

Anyone who actually understands developmental biology, toxicology, endocrinology, etc. knows that we don't know anywhere near enough biology to do good safety testing for the foreseeable future based solely on NAMs.

As mentioned above, I do not see any chance that alternative methods will be able (in this short time-frame) to replace complex toxicity tests such as carcinogenicity, reproductive tox, or even simple sub-acute to sub-chronic repeat-dose tox studies. Even replacement of simpler tests like acute tox or irritation took decades to validate and implement. I cannot see how we (the tox community) will be able to succeed in completely replace animal testing with VALIDATED (!) non-animal methods by 2035.

Difficult because of unavailability of alternative solutions

Determining the health safety of new chemicals is a prerequisite for approval posed by the same regulatory agencies, including EPA. Cell cultures can provide useful information about the mechanisms, but cannot ensure safety for the living organisms and the environment. By skipping the animal testing phase, the burden of proving a new chemical safe will be on humans.

There has been significant progress made in computational toxicology and in vitro methodologies, and advances will continue. But whether this exact benchmark is accomplished, is difficult to predict. What is important is that high level management directives are a critical driver for supporting and advancing the science, as well as the incremental applications of new technologies to decision-making over time. To have confidence in moving toward this goal, the Agency needs to provide leadership in steering the efforts to reduce the reliance on animal testing. Additionally, similar progress needs to occur with alternative methodologies for both human health and ecological risk assessments. And finally, the stakeholder and international communities need to have a level of comfort that the alternatives do represent the best science to ensure safety and are for the public good. This will depend on partnerships, the research conducted in concert with regulatory dialogue, and involvement of and sincere interactions with stakeholders.

Yes for some chemical categories and endpoints, and no or questionable for others

Ideally, we would have perfect information about physiology so that NAMs could perfectly recapitulate the complexity of anatomy and physiology. We do not yet have that information.

Biology is too complex to rely exclusively on in vitro and in silico analyses to predict toxic outcomes. For instance, being able to model an entire immune system including tissue resident subsets and how toxicants may affect host responses is not feasible. While 3D organoid systems are building toward this, there is so much cross-talk between organ systems that we might miss significant toxicities which can be captured with animal models.

While I strongly support the mission to reduce animal studies, complete elimination would impact our ability to determine human health risks of the countless compounds introduced into our products and environment. Without this crucial evidence, it will be more difficult to set exposure limits or remove toxic compounds from use.

NAMs are great for screening. But for some chemicals I do still think it could make sense to supplement with animal data especially for long term studies.

Safety testing should use the appropriated model this can be an in vitro test if completely validated. If not animal testing is still required.

So much still needed, e.g. in vivo biokinetics (ADME) is a huge challenge for non-animal testing. Chronic toxicity testing remains a big testing hurdle for short-term alternatives. Individual variance in response is another major obstacle in need of innovation. Defining domains of applicability for alternatives needs to be addressed.

For the same reasons noted above, I just dont think this concept is ready for prime time. Perhaps a global harmonization effort, similar to ICH to align on and agree to which animal tests can be formally replaced with NAMs would be appropriate, instead of such a disjointed effort across agencies and geographies. These animals tests apply to food, drugs, chemicals and medical devices alike.

I hope this will be possible, but I am not completely sure if the NAMs will be able to replace animals in all endpoints.

Safety testing for certain endpoints such as neurotoxicity, neurodegeneration, developmental immune toxicity etc still relies on animals, if performed at all. The fact that for some of these endpoint a general lack of knowledge seems to paralize progression renders me reluctant to think it will be feasible to rely solely on non-animal testing.

The interaction of the genome (of any and all living species) with the environment is far too complex to expect accurate predictions in 13 years.

There will be some remaining needs for animals in case of novel targets for pesticides and pharmaceuticals or for new chemistries that are not represented by the present inventory. I guess that testing may be more sucessfully avoided for chemicals and pesticides (usually low exposures), but there will still be some need for pharmaceuticals

There are lots of interactions apically than what we observe in non-animal methods and may not be true in real-world cases.

The law is going to have to change because many regulatory decisions require whole animal data. Unless that changes we're going to need that animal data to maximize public safety.

Hubris!! We do not understand physiology well enough to do this. 13 years will not change that.

In silicon has a long way to go. There are people who dont believe in vaccines, after all.

Possible

Animal models of diseases have proven as are the best way to understand and extend the results to human being and influence of drugs and chemicals.

In my opinion, still the NAMs methods are in the development phase, relying solely on them is not advisable.

It is already being done for some chemicals, but as a screening method. I doubt that ALL animal testing will be eliminated. But it is important to remember that animal testing isn't that accurate due to inter-species differences. It is merely the best we have available but the correlations are not that good.

I think great progress can be made and get to nearly all non-animal testing methods. Whether the testing will be able to completely eliminate animal methods for all areas is potentially a bit farther off than 2035 - but great progress can be made.

Significant reduction of animal testing is most likely and already taking place.

Many advancements have been made over the past two decades - I am more hesitant though extending them to pharmaceuticals.

Technology has evolved to the extent that it would be feasible to rely on NAMs

NO, I'm not sure it will be feasible to rely solely on non-animal methods for safety testing chemicals because studying the response of biological organisms is very important to defining mechanisms, and toxic and negative effects of pollutants on living organisms.

See Q2

toxicity analysis will be entirely possible without use of animals by 2030

Not feasible and in accurate data will be gathered from the NAMs

2035 is tomorrow in term of research, especially for human safety. It will take more than 10 years to validate new models and 10 years to see if it's mimic perfectly what we have with animals

See before

See above.

All in vitro test are not applicable to whole animal model results and tests using cell or organoid models are some times only usable to study toxicity.

More research is needed, however, there are several experiments that are replacing animal methods (such as cotton strip assay, colonization of artificial substrates...). Moreover, other methods may include mollecular testing (e.g. comet assay) instead of animal bioassays.

The depth and breadth of knowledge generated by testing drugs and vaccines in live animals is simply unprecedented and it may be very challenging for any artificial system to compete with this.

Some preliminary hazard assessment yes: no more. See answer above. There appears to be more politics than science here.