The power of PANDAA: enabling resistance testing for better HIV care

Dr Iain MacLeod, along with Dr David Raiser, from Aldatu Biosciences, discuss how the use of their innovative PANDAA technology could facilitate drug resistance testing in HIV patients, transforming the quality of patient care, especially in developing countries.

The war against Human Immunodeficiency Virus (HIV) is a global struggle. Worryingly, over 74 countries, particularly South Africa, have experienced an increased rate of new HIV infections between 2005 and 2015.

Despite this grim statistic, more HIV patients than ever are receiving antiretroviral drug (ARV) therapy. These drugs suppress the HIV life cycle, not only ensuring that patients live longer and healthier lives, but also minimising further HIV transmission. By the end of 2015, 18 million of the 38 million people worldwide suffering from HIV had access to ARV treatment.

HIV Resistance Against ARVs
However, HIV is fighting back. Different strains have developed resistance to specific ARV drugs, rendering those drugs ineffective as treatment. Annually, ARV treatment failure occurs in 10% of patients on average, which has severe consequences when allowed to persist: patients continue to suffer from this debilitating disease until alternative medication is administered. Drug resistance testing is necessary to identify effective alternative ARVs, but these tests are simply too expensive for most developing countries. Without resistance testing, patients can remain on ineffective ARVs for up to 18 months, during which time they are also more likely to transmit their HIV to others. Furthermore, prescribing ineffective medication does not make economic sense.

Overall, PANDAA technology offers a cost-effective, simpler and quicker process compared to other genetic sequencing technologies.

Clearly, drug resistance testing plays a critical role in HIV patient care management: if resistance was consistently monitored and identified early, patients could be more readily switched to effective ARVs and have better health outcomes.

Aldatu Biosciences: A Public Benefit Mission
The main aim of Aldatu Biosciences (founded by Dr David Raiser and Dr Iain MacLeod in 2014) is to increase access to drug resistance testing through the use of their ground-breaking technology, PANDAA (Pan Degenerate Amplification and Adaptation).

Drs Iain MacLeod and David Raiser in the Aldatu lab

As a result of his experiences working at the Botswana Harvard AIDS Institute and witnessing first-hand the need for better drug resistance testing options, Dr MacLeod was inspired to design PANDAA — a simple, sensitive, and inexpensive way to detect drug-resistant HIV. Aldatu Biosciences’ first application of the technology to a specific disease, HIV, led to the development of their lead product PANDAA HIV6. Although developing countries have the highest HIV rates, they often lack the funds and resources needed to deploy comprehensive HIV management strategies that include drug resistance testing. Local laboratories dedicated to HIV testing are over-burdened and struggle to cope with demand. A simple, affordable solution for HIV drug resistance testing is badly needed in these settings, and PANDAA HIV6 could be the answer.

PANDAA — an Enabling Technology
Typically speaking, the number of diagnostic techniques that utilise quantitative real-time polymerase chain reaction (qPCR) molecular probes has been unrivalled in modern genomics. However, with highly polymorphic pathogens, such as HIV, traditional qPCR’s greatest strength, its sensitivity and specificity, becomes a weakness. Traditional qPCR cannot tolerate changes in the probe binding sequence, which commonly present at regions near resistance-conferring mutations within the HIV genome. This is referred to as “secondary sequence variation”. As a result, probes designed to detect resistance mutations are unable to bind, resulting in high false negative rates during experimentation — the results show no resistance although resistance is present. So, in other words, HIV’s high genomic variability has limited the development of molecular probes for HIV drug resistance genotyping.

PANDAA testing in action

This is where PANDAA technology comes in. Using proprietary reagent design techniques, PANDAA maintains the superior sensitivity of qPCR without losing any of its specificity. The primers it uses include special design features that strip out secondary sequence variation in the probe binding sequence, while leaving the drug resistance information intact. Not only that, but by harnessing an existing low-cost technology (qPCR), PANDAA provides a technique for HIV drug resistance genotyping that is affordable where it is most needed.

Moving forward, Dr MacLeod believes that PANDAA technology could be used to detect drug resistance in other debilitating diseases including influenza, hepatitis B and MDR-TB.

PANDAA has been specifically designed to meet the clinical and economic needs of healthcare settings with limited resources. The technology provides a vital advancement in HIV drug resistance diagnostics that can radically improve HIV patient care delivery and management in lower- and middle-income countries.

Advantages of PANDAA Over Current Technology
Current technology used for drug resistance testing involves collecting long sequences of the HIV genome, using methods such as Sanger or Next Generation Sequencing. However, only around 1% of the sequenced genome is of clinical importance i.e., provides information on the resistance level of the HIV strain. PANDAA-enabled qPCR allows for a more efficient approach called ‘focused genotyping’, wherein only those genomic positions that are relevant to a particular therapeutic decision are queried in a given test. For example, PANDAA HIV6 collects information on just six different mutation positions in the HIV genome, and in doing so, it can detect resistance in more than 99% of HIV-infected patients in Africa who are failing their WHO-recommended first line ARV therapy regimen. In essence, focused genotyping allows for simplified and cost-efficient testing product design, which has been employed for PANDAA HIV6.

Current Sanger sequencing-based methods used to detect the presence of drug-resistant variants have a relatively low sensitivity at 80–85%. As such, they are only able to detect variants if they are present within 15–20% or more of the total virus population of a patient. PANDAA, on the other hand, can detect these variants with 99% sensitivity and in a fraction (roughly 3%) of the time. The technology’s ‘sample-in/answer-out’ format limits the sample handling time and means that less-skilled users can perform the tests — both of which are important considerations within resource-limited environments. PANDAA tests also do not require any new machinery to be purchased, as the technology utilises existing equipment found in centralised testing laboratories. Aldatu’s current focus is to offer the tests in a thermostable format. This will mean samples do not have to be kept in cold storage, avoiding the typically high costs involved with cold chain transfer storage and transportation.

Future Uses
PANDAA technology is an innovative technology that could potentially be life-saving, especially in developing countries where resources are limited. Currently, Aldatu Biosciences are focusing on targeting HIV patients in Southern Africa who are failing their first-line treatment, especially pregnant women who could transmit HIV to the next generation. Moving forward, Dr MacLeod believes that PANDAA technology could be used to detect drug resistance in other debilitating diseases, including influenza, hepatitis B and MDR-TB. PANDAA technology is a revolutionary step for clinical diagnostics and will undoubtedly transform the lives of many people all over the world.