HIV-1 Coreceptor Tropism, Proviral DNA

Tropism refers to the type of cytokine coreceptor used by HIV-1 when infecting the host cell. The viruses in most (>80%) treatment-naïve patients use the CCR5 (R5) coreceptor.¹ Conversely, the viruses in up to 50% of treatment-experienced patients use either the CXCR4 (X4) coreceptor or both coreceptors (ie, R5 and X4).² Viruses that use both coreceptors are called dual-mixed (D/M) viruses.

A coreceptor tropism test determines whether a patient exclusively harbors R5-tropic virus or has X4-tropic or D/M virus. Patients who exclusively harbor R5-tropic virus can be treated with CCR5 antagonists such as maraviroc (Selzentry^®). CCR5 antagonists block R5 viruses from binding to the CCR5 coreceptor and infecting cells. CCR5 antagonists are ineffective in patients with X4 or D/M virus. Thus, coreceptor tropism testing can help determine patient eligibility for CCR5 antagonist therapy.

A coreceptor tropism test should be performed when the use of a CCR5 antagonist is being considered.^3,4 Coreceptor tropism testing might also be considered for patients who exhibit virologic failure while taking a CCR5 inhibitor.^3,4 See also Question 6.

We offer two tests for genotypic tropism testing:

1. HIV-1 Coreceptor Tropism, Ultradeep Sequencing (test codes 94014(X), 94015(X) or 94016(X)): determines tropism of HIV-1 RNA; used when the plasma viral load is ³1000 copies/mL
   For more information, go to HIV-1 Coreceptor Tropism Ultradeep Sequencing | Quest Diagnostics.
2. HIV-1 Coreceptor Tropism, Proviral DNA (test code 91299[X]): used when the plasma viral load is <1000 copies/mL

A proviral DNA tropism test determines the tropism of HIV-1 DNA that has integrated into the host genome of infected T-lymphocytes. Proviral HIV-1 DNA persists despite suppressive antiviral therapy and therefore is present when plasma viral RNA is low or undetectable. Whole-blood or peripheral blood mononuclear cells (PBMC), rather than plasma, should be submitted for this test.

Although HIV-1 X4 sequences are more commonly found in proviral DNA than RNA, several studies have found a high degree of correlation between proviral DNA tropism and RNA tropism. For example:

• Soulie et al determined proviral DNA tropism in a genotype assay and compared results to archived RNA tropism data from treatment-experienced patients with undetectable plasma viral loads. The authors saw no change in viral tropism over a median of 4 years in 92.9% (119/128) of these patients and concluded genotypic proviral DNA tropism could be used to determine viral tropism.⁵
• Verhofstede et al found a concordance of 95.2% when comparing HIV-1 proviral PBMC DNA tropism to plasma RNA tropism determined from simultaneously collected sample types (n=165 patients). Similarly, concordance was 88.1% when current proviral DNA tropism was compared to pretreatment RNA tropism data from 126 patients.⁶

U.S. guidelines indicate that proviral testing can be used to determine tropism in patients with undetectable plasma viral load, but they note that the clinical utility of this approach has not yet been determined.³ The European tropism testing guidelines recommend proviral DNA tropism testing for patients with a low (<1000 copies/mL) or undetectable plasma viral load when they are being considered for maraviroc therapy.⁷

Proviral DNA tropism testing may be appropriate for successfully treated patients seeking to switch to a new regimen that includes a CCR5 antagonist.

We first extract DNA from whole blood. PCR amplification of DNA in the V3 loop region of the HIV-1 envelope gene is performed in triplicate and the replicates are pooled for ultradeep sequencing (UDS).  Tropism is then determined by bioinformatic analysis of the V3 loop sequence.

The turnaround time is approximately one week.

X4 Virus Detected

CXCR4 (X4)                                   DETECTED

Net Tropism Assessment                DM/X4

MVC Activity Anticipated                 NO

X4 Virus Not Detected

CXCR4 (X4)                                   NOT DETECTED

Net Tropism Assessment                R5

MVC Activity Anticipated                 YES